Sample records for saa mrna levels

  1. Characterization of rat serum amyloid A4 (SAA4): A novel member of the SAA superfamily

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Rossmann, Christine; Windpassinger, Christian; Brunner, Daniela

    2014-08-08

    Highlights: • The full length rat SAA4 (rSAA4) mRNA was characterized by rapid amplification of cDNA ends. • rSAA4 mRNA has 1830 bases including a GA dinucleotide tandem repeat in the 5′UTR. • Three consecutive C/EBP promoter elements are crucial for transcription of rSAA4. • rSAA4 is abundantly expressed in the liver on mRNA and protein level. - Abstract: The serum amyloid A (SAA) family of proteins is encoded by multiple genes, which display allelic variation and a high degree of homology in mammals. The SAA1/2 genes code for non-glycosylated acute-phase SAA1/2 proteins, that may increase up to 1000-fold duringmore » inflammation. The SAA4 gene, well characterized in humans (hSAA4) and mice (mSaa4) codes for a SAA4 protein that is glycosylated only in humans. We here report on a previously uncharacterized SAA4 gene (rSAA4) and its product in Rattus norvegicus, the only mammalian species known not to express acute-phase SAA. The exon/intron organization of rSAA4 is similar to that reported for hSAA4 and mSaa4. By performing 5′- and 3′RACE, we identified a 1830-bases containing rSAA4 mRNA (including a GA-dinucleotide tandem repeat). Highest rSAA4 mRNA expression was detected in rat liver. In McA-RH7777 rat hepatoma cells, rSAA4 transcription was significantly upregulated in response to LPS and IL-6 while IL-1α/β and TNFα were without effect. Luciferase assays with promoter-truncation constructs identified three proximal C/EBP-elements that mediate expression of rSAA4 in McA-RH7777 cells. In line with sequence prediction a 14-kDa non-glycosylated SAA4 protein is abundantly expressed in rat liver. Fluorescence microscopy revealed predominant localization of rSAA4-GFP-tagged fusion protein in the ER.« less

  2. Paradoxical effects of SAA on lipoprotein oxidation suggest a new antioxidant function for SAA[S

    PubMed Central

    Jayaraman, Shobini; Haupt, Christian

    2016-01-01

    Oxidative stress and inflammation, which involve a dramatic increase in serum amyloid A (SAA) levels, are critical in the development of atherosclerosis. Most SAA circulates on plasma HDL particles, altering their cardioprotective properties. SAA-enriched HDL has diminished anti-oxidant effects on LDL, which may contribute to atherogenesis. We determined combined effects of SAA enrichment and oxidation on biochemical changes in HDL. Normal human HDLs were incubated with SAA, oxidized by various factors (Cu2+, myeloperoxidase, H2O2, OCl−), and analyzed for lipid and protein modifications and biophysical remodeling. Three novel findings are reported: addition of SAA reduces oxidation of HDL and LDL lipids; oxidation of SAA-containing HDL in the presence of OCl− generates a covalent heterodimer of SAA and apoA-I that resists the release from HDL; and mild oxidation promotes spontaneous release of proteins (SAA and apoA-I) from SAA-enriched HDL. We show that the anti-oxidant effects of SAA extend to various oxidants and are mediated mainly by the unbound protein. We propose that free SAA sequesters lipid hydroperoxides and delays lipoprotein oxidation, though much less efficiently than other anti-oxidant proteins, such as apoA-I, that SAA displaces from HDL. These findings prompt us to reconsider the role of SAA in lipid oxidation in vivo. PMID:27744369

  3. Measurement of serum amyloid A1 (SAA1), a major isotype of acute phase SAA.

    PubMed

    Xu, Yuanyuan; Yamada, Toshiyuki; Satoh, Takahiko; Okuda, Yasuaki

    2006-01-01

    Serum amyloid A (SAA), a plasma precursor of reactive amyloid deposits, is a multigene product. SAA1 and SAA2, with primary structures that are 93% identical (98 of 104 amino acids), behave as acute phase proteins, as demonstrated by their increasing levels in plasma. Heretofore, it has been understood that SAA1 predominates and functions as an isotype in plasma. However, accurate measurements differentiating the two isotypes have not been reported. In this study, using monoclonal antibodies specific for SAA1, we developed an enzyme-linked immunosorbent assay (ELISA) for SAA1. The levels and ratios of SAA1 in total SAA (TSAA) were investigated in healthy subjects and patients with rheumatoid arthritis (RA). The SAA1/TSAA ratio was 74 +/- 12% and 77 +/- 12% in healthy subjects and RA patients, respectively. In RA patients, the ratios were not influenced by SAA1 genotype, which has been proposed to affect plasma SAA values. The kinetics of SAA1 in inflamed patients undergoing hemodialysis was found to be parallel with total SAA and C-reactive protein. Finally, this study confirmed that SAA1 is a major isotype of acute phase SAA and may determine total SAA values. This specific assay could be used in the evaluation of SAA behavior in several clinical conditions.

  4. Quantification of SAA1 and SAA2 in lung cancer plasma using the isotype-specific PRM assays.

    PubMed

    Kim, Yeoun Jin; Gallien, Sebastien; El-Khoury, Victoria; Goswami, Panchali; Sertamo, Katriina; Schlesser, Marc; Berchem, Guy; Domon, Bruno

    2015-09-01

    The quantification of plasma proteins using the high resolution and accurate mass (HR/AM)-based parallel reaction monitoring (PRM) method provides an immediate benefit over the conventional SRM-based method in terms of selectivity. In this study, multiplexed PRM assays were developed to analyze isotypes of serum amyloid A (SAA) proteins in human plasma with a focus on SAA1 and SAA2. Elevated plasma levels of these proteins in patients diagnosed with lung cancer have been reported in previous studies. Since SAA1 and SAA2 are highly homologous, the available immunoassays tend to overestimate their concentrations due to cross-reactivity. On the other hand, when mass spectrometry (MS)-based assays are used, the presence of the several allelic variants may result in a problem of underestimation. In the present study, eight peptides that represent the target proteins at three different levels: isotype-specific (SAA1α,  SAA 1β,  SAA1γ,  SAA2α,  SAA2β), protein-specific (SAA1 or SAA2), and pan SAA (SAA1 and SAA2) were chosen to differentiate SAAs in lung cancer plasma samples using a panel of PRM assays. The measurement of specific isotypes, leveraging the analytical performance of PRM, allowed to quantify the allelic variants of both target proteins. The isotypes detected were corroborated with the genetic information obtained from the same samples. The combination of SAA2α and SAA2β assays representing the total SAA2 concentration demonstrated a superior analytical outcome than the previously used assay on the common peptide when applied to the detection of lung cancer. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Evaluating the impact of virtualization characteristics on SaaS adoption

    NASA Astrophysics Data System (ADS)

    Tomás, Sara; Thomas, Manoj; Oliveira, Tiago

    2018-03-01

    Software as a service (SaaS) is a service model in which the applications are accessible from various client devices through internet. Several studies report possible factors driving the adoption of SaaS but none have considered the perception of the SaaS features and the organization's context. We propose an integrated research model that combines the process virtualization theory (PVT), the technology-organization-environment (TOE) framework and the institutional theory (INT). PVT seeks to explain whether processes are suitable for migration into virtual environments via an information technology-based mechanism as SaaS. The TOE framework seeks to explain the effects of the intra-organizational factors, while INT seeks to explain the effects of the inter-organizational factors on the technology adoption. This research addresses a gap in the SaaS adoption literature by studying the internal perception of the technical features of SaaS and technology, organization, and environment perspectives. Additionally, the integration of PVT, the TOE framework, and INT contributes to the information system (IS) discipline, deepening the applicability and strengths of these theories.

  6. A Systematic Process for Developing High Quality SaaS Cloud Services

    NASA Astrophysics Data System (ADS)

    La, Hyun Jung; Kim, Soo Dong

    Software-as-a-Service (SaaS) is a type of cloud service which provides software functionality through Internet. Its benefits are well received in academia and industry. To fully utilize the benefits, there should be effective methodologies to support the development of SaaS services which provide high reusability and applicability. Conventional approaches such as object-oriented methods do not effectively support SaaS-specific engineering activities such as modeling common features, variability, and designing quality services. In this paper, we present a systematic process for developing high quality SaaS and highlight the essentiality of commonality and variability (C&V) modeling to maximize the reusability. We first define criteria for designing the process model and provide a theoretical foundation for SaaS; its meta-model and C&V model. We clarify the notion of commonality and variability in SaaS, and propose a SaaS development process which is accompanied with engineering instructions. Using the proposed process, SaaS services with high quality can be effectively developed.

  7. Research on SaaS and Web Service Based Order Tracking

    NASA Astrophysics Data System (ADS)

    Jiang, Jianhua; Sheng, Buyun; Gong, Lixiong; Yang, Mingzhong

    To solve the order tracking of across enterprises in Dynamic Virtual Enterprise (DVE), a SaaS and web service based order tracking solution was designed by analyzing the order management process in DVE. To achieve the system, the SaaS based architecture of data management on order tasks manufacturing states was constructed, and the encapsulation method of transforming application system into web service was researched. Then the process of order tracking in the system was given out. Finally, the feasibility of this study was verified by the development of a prototype system.

  8. A Mechanism of Modeling and Verification for SaaS Customization Based on TLA

    NASA Astrophysics Data System (ADS)

    Luan, Shuai; Shi, Yuliang; Wang, Haiyang

    With the gradually mature of SOA and the rapid development of Internet, SaaS has become a popular software service mode. The customized action of SaaS is usually subject to internal and external dependency relationships. This paper first introduces a method for modeling customization process based on Temporal Logic of Actions, and then proposes a verification algorithm to assure that each step in customization will not cause unpredictable influence on system and follow the related rules defined by SaaS provider.

  9. Connecting long-tail scientists with big data centers using SaaS

    NASA Astrophysics Data System (ADS)

    Percivall, G. S.; Bermudez, L. E.

    2012-12-01

    Big data centers and long tail scientists represent two extremes in the geoscience research community. Interoperability and inter-use based on software-as-a-service (SaaS) increases access to big data holdings by this underserved community of scientists. Large, institutional data centers have long been recognized as vital resources in the geoscience community. Permanent data archiving and dissemination centers provide "access to the data and (are) a critical source of people who have experience in the use of the data and can provide advice and counsel for new applications." [NRC] The "long-tail of science" is the geoscience researchers that work separate from institutional data centers [Heidorn]. Long-tail scientists need to be efficient consumers of data from large, institutional data centers. Discussions in NSF EarthCube capture the challenges: "Like the vast majority of NSF-funded researchers, Alice (a long-tail scientist) works with limited resources. In the absence of suitable expertise and infrastructure, the apparently simple task that she assigns to her graduate student becomes an information discovery and management nightmare. Downloading and transforming datasets takes weeks." [Foster, et.al.] The long-tail metaphor points to methods to bridge the gap, i.e., the Web. A decade ago, OGC began building a geospatial information space using open, web standards for geoprocessing [ORM]. Recently, [Foster, et.al.] accurately observed that "by adopting, adapting, and applying semantic web and SaaS technologies, we can make the use of geoscience data as easy and convenient as consumption of online media." SaaS places web services into Cloud Computing. SaaS for geospatial is emerging rapidly building on the first-generation geospatial web, e.g., OGC Web Coverage Service [WCS] and the Data Access Protocol [DAP]. Several recent examples show progress in applying SaaS to geosciences: - NASA's Earth Data Coherent Web has a goal to improve science user experience using

  10. Universal SaaS platform of internet of things for real-time monitoring

    NASA Astrophysics Data System (ADS)

    Liu, Tongke; Wu, Gang

    2018-04-01

    Real-time monitoring service, as a member of the IoT (Internet of Things) service, has a wide range application scenario. To support rapid construction and deployment of applications and avoid repetitive development works in these processes, this paper designs and develops a universal SaaS platform of IoT for real-time monitoring. Evaluation shows that this platform can provide SaaS service to multiple tenants and achieve high real-time performance under the situation of large amount of device access.

  11. Sourcing Lifecycle for Software as a Service (SAAS)

    NASA Astrophysics Data System (ADS)

    Santy; Sikkel, K.

    2014-03-01

    In recent years, Software as a Service (SaaS) has changed from curiosity caused concept to an accepted well known concept. A key advantage of this model is that, by cautious engineering, it is possible to influence economy of scale to decrease total cost of ownership compared to on-premises solutions. By using the guideline elaborated in this paper, companies which has interest in implementing SaaS will be led throughout the entire implementation cycle starting before the company decides to implement SaaS until the stage when the company decides to shift from SaaS model to another model or when they shift to another SaaS company

  12. Development and analytical performance evaluation of FREND-SAA and FREND-Hp

    NASA Astrophysics Data System (ADS)

    Choi, Eunha; Seong, Jihyun; Lee, Seiyoung; Han, Sunmi

    2017-07-01

    The FREND System is a portable cartridge reader, quantifying analytes by measuring laser-induced fluorescence in a single-use reagent cartridge. The objective of this study was to evaluate FREND-SAA and FREND-Hp assays. The FREND-SAA and Hp assays were standardized to the WHO and IFCC reference materials. Analytical performance studies of Precision, Linearity, Limits of Detections, Interferences, and Method Comparisons for both assays were performed according to the CLSI guidelines. Both assays demonstrated acceptable imprecision of %CV in three different levels of samples. The linearity of the assays was found to be acceptable (SAA 5 150 mg/L, Hp 30 400 mg/dL). The detection limits were 3.8 mg/L (SAA) and 10.2 mg/dL (Hp). No significant interference and no significant deviation from linearity was found in the both comparison studies. In conclusion, NanoEnTek's FREND-SAA and Hp assays represent rapid, accurate and convenient means to quantify SAA and Hp in human serum on FREND system.

  13. Challenges in Small Screening Laboratories: SaaS to the rescue

    PubMed Central

    Lemmon, Vance P.; Jia, Yuanyuan; Shi, Yan; Holbrook, S. Douglas; Bixby, John L; Buchser, William

    2012-01-01

    The Miami Project to Cure Paralysis, part of the University of Miami Miller School of Medicine, includes a laboratory devoted to High Content Analysis (HCA) of neurons. The goal of the laboratory is to uncover signalling pathways, genes, compounds, or drugs that can be used to promote nerve growth. HCA permits the quantification of neuronal morphology, including the lengths and numbers of axons. HCA screening of various libraries on primary neurons requires a team-based approach, a variety of process steps and complex manipulations of cells and libraries to obtain meaningful results. HCA itself produces vast amounts of information including images, well-based data and cell-based phenotypic measures. Managing experimental workflow and library data, along with the extensive amount of experimental results is challenging. For academic laboratories generating large data sets from experiments using thousands of perturbagens, a laboratory information management system (LIMS) is the data tracking solution of choice. With both productivity and efficiency as driving rationales, the Miami Project has equipped its HCA laboratory with a Software As A Service (SAAS) LIMS to ensure the quality of its experiments and workflows. The article discusses this application in detail, and how the system was selected and integrated into the laboratory. The advantages of SaaS are described. PMID:21631415

  14. SAA drives proinflammatory heterotypic macrophage differentiation in the lung via CSF-1R-dependent signaling

    PubMed Central

    Anthony, Desiree; McQualter, Jonathan L.; Bishara, Maria; Lim, Ee X.; Yatmaz, Selcuk; Seow, Huei Jiunn; Hansen, Michelle; Thompson, Michelle; Hamilton, John A.; Irving, Louis B.; Levy, Bruce D.; Vlahos, Ross; Anderson, Gary P.; Bozinovski, Steven

    2014-01-01

    Serum amyloid A (SAA) is expressed locally in chronic inflammatory conditions such as chronic obstructive pulmonary disease (COPD), where macrophages that do not accord with the classic M1/M2 paradigm also accumulate. In this study, the role of SAA in regulating macrophage differentiation was investigated in vitro using human blood monocytes from healthy subjects and patients with COPD and in vivo using an airway SAA challenge model in BALB/c mice. Differentiation of human monocytes with SAA stimulated the proinflammatory monokines IL-6 and IL-1β concurrently with the M2 markers CD163 and IL-10. Furthermore, SAA-differentiated macrophages stimulated with lipopolysaccharide (LPS) expressed markedly higher levels of IL-6 and IL-1β. The ALX/FPR2 antagonist WRW4 reduced IL-6 and IL-1β expression but did not significantly inhibit phagocytic and efferocytic activity. In vivo, SAA administration induced the development of a CD11chighCD11bhigh macrophage population that generated higher levels of IL-6, IL-1β, and G-CSF following ex vivo LPS challenge. Blocking CSF-1R signaling effectively reduced the number of CD11chighCD11bhigh macrophages by 71% and also markedly inhibited neutrophilic inflammation by 80%. In conclusion, our findings suggest that SAA can promote a distinct CD11chighCD11bhigh macrophage phenotype, and targeting this population may provide a novel approach to treating chronic inflammatory conditions associated with persistent SAA expression.—Anthony, D., McQualter, J. L., Bishara, M., Lim, E. X., Yatmaz, S., Seow, H. J., Hansen, M., Thompson, M., Hamilton, J. A., Irving, L. B., Levy, B. D., Vlahos, R., Anderson, G. P., Bozinovski, S. SAA drives proinflammatory heterotypic macrophage differentiation in the lung via CSF-1R-dependent signaling. PMID:24846388

  15. Associations of SAA1 gene polymorphism with lipid lelvels and osteoporosis in Chinese women.

    PubMed

    Feng, Zheng-Ping; Li, Xiao-Yu; Jiang, Rong; Deng, Hua-Cong; Yang, Mei; Zhou, Qin; Que, Wen-Jun; Du, Jia

    2013-03-22

    The development of osteoporosis is associated with several risk factors, such as genetic polymorphisms and enviromental factors. This study assessed the correlation between SAA1 gene rs12218 polymorphism and HDL-C lelvels and osteoporosis in a population of Chinese women. A total of 387 postmenopausal female patients who were diagnosed with osteoporosis (case group) based on bone mineral density measurements via dual-energy x-ray absorptiometry and 307 females with no osteoporosis (control group) were included in this study. Correlations between SAA1 gene rs12218 polymorphism and osteoporosis and HDL-C level were investigated through the identification of SAA1 gene rs12218 polymorphism genotypes using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The TT genotype of rs12218 was more frequently in osteoporosis patients than in control subjects (P <0.001). And the rs12218 was found to be associated with plasma TG, HDL-C, LDL-C, and BMD levels in osteoporosis patients (P<0.05). The present results indicate that both osteoporosis and lipids levels are associated with the TT genotype of rs12218 in the human SAA1 gene.

  16. Associations of SAA1 gene polymorphism with Lipid lelvels and osteoporosis in Chinese women

    PubMed Central

    2013-01-01

    Background The development of osteoporosis is associated with several risk factors, such as genetic polymorphisms and enviromental factors. This study assessed the correlation between SAA1 gene rs12218 polymorphism and HDL-C lelvels and osteoporosis in a population of Chinese women. Methods A total of 387 postmenopausal female patients who were diagnosed with osteoporosis (case group) based on bone mineral density measurements via dual-energy x-ray absorptiometry and 307 females with no osteoporosis (control group) were included in this study. Correlations between SAA1 gene rs12218 polymorphism and osteoporosis and HDL-C level were investigated through the identification of SAA1 gene rs12218 polymorphism genotypes using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Results The TT genotype of rs12218 was more frequently in osteoporosis patients than in control subjects (P <0.001). And the rs12218 was found to be associated with plasma TG, HDL-C, LDL-C, and BMD levels in osteoporosis patients (P<0.05). Conclusions The present results indicate that both osteoporosis and lipids levels are associated with the TT genotype of rs12218 in the human SAA1 gene. PMID:23522429

  17. The first molluscan acute phase serum amyloid A (A-SAA) identified from oyster Crassostrea hongkongensis: molecular cloning and functional characterization.

    PubMed

    Qu, Fufa; Xiang, Zhiming; Yu, Ziniu

    2014-08-01

    Serum amyloid A (SAA), a major evolutionarily conserved acute-phase protein, participates in many biological processes in eukaryotic cells, including innate immunity. However, little information regarding the relationship between SAA and innate immunity in mollusks is currently available. In this report, the first bivalve SAA (referred to as ChSAA) gene was identified and characterized from the Hong Kong oyster Crassostrea hongkongensis. Its full-length cDNA is 623 bp, including a 5'-UTR of 147 bp, a 3'-UTR of 56 bp containing a poly(A) tail and an open reading frame (ORF) of 420 bp that encodes a polypeptide of 139 amino acids. The predicted amino acid sequence of ChSAA comprises characteristic motifs of the SAA family, including a typical signal peptide and a conserved SAA domain. Comparison and phylogenetic analyses suggested that ChSAA shares a high identity to known acute-phase SAA proteins (A-SAAs). In addition, quantitative real-time PCR analysis revealed that ChSAA is constitutively expressed in all tissues examined, with the highest expression level in the mantle, and that its expression was acutely and significantly up-regulated in hemocytes following challenge by Vibrio alginolyticus (G(-)), Staphylococcus haemolyticus (G(+)) or Saccharomyces cerevisiae (fungus). Furthermore, over-expression of ChSAA via transfection with a ChSAA expression vector led to significantly increased NF-κB activity in HEK293T cells. These results suggest that ChSAA is likely to constitute a member of the A-SAA family involved in anti-pathogen responses in C. hongkongensis. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Circadian integration of glutamatergic signals by little SAAS in novel suprachiasmatic circuits.

    PubMed

    Atkins, Norman; Mitchell, Jennifer W; Romanova, Elena V; Morgan, Daniel J; Cominski, Tara P; Ecker, Jennifer L; Pintar, John E; Sweedler, Jonathan V; Gillette, Martha U

    2010-09-07

    Neuropeptides are critical integrative elements within the central circadian clock in the suprachiasmatic nucleus (SCN), where they mediate both cell-to-cell synchronization and phase adjustments that cause light entrainment. Forward peptidomics identified little SAAS, derived from the proSAAS prohormone, among novel SCN peptides, but its role in the SCN is poorly understood. Little SAAS localization and co-expression with established SCN neuropeptides were evaluated by immunohistochemistry using highly specific antisera and stereological analysis. Functional context was assessed relative to c-FOS induction in light-stimulated animals and on neuronal circadian rhythms in glutamate-stimulated brain slices. We found that little SAAS-expressing neurons comprise the third most abundant neuropeptidergic class (16.4%) with unusual functional circuit contexts. Little SAAS is localized within the densely retinorecipient central SCN of both rat and mouse, but not the retinohypothalamic tract (RHT). Some little SAAS colocalizes with vasoactive intestinal polypeptide (VIP) or gastrin-releasing peptide (GRP), known mediators of light signals, but not arginine vasopressin (AVP). Nearly 50% of little SAAS neurons express c-FOS in response to light exposure in early night. Blockade of signals that relay light information, via NMDA receptors or VIP- and GRP-cognate receptors, has no effect on phase delays of circadian rhythms induced by little SAAS. Little SAAS relays signals downstream of light/glutamatergic signaling from eye to SCN, and independent of VIP and GRP action. These findings suggest that little SAAS forms a third SCN neuropeptidergic system, processing light information and activating phase-shifts within novel circuits of the central circadian clock.

  19. Extending multi-tenant architectures: a database model for a multi-target support in SaaS applications

    NASA Astrophysics Data System (ADS)

    Rico, Antonio; Noguera, Manuel; Garrido, José Luis; Benghazi, Kawtar; Barjis, Joseph

    2016-05-01

    Multi-tenant architectures (MTAs) are considered a cornerstone in the success of Software as a Service as a new application distribution formula. Multi-tenancy allows multiple customers (i.e. tenants) to be consolidated into the same operational system. This way, tenants run and share the same application instance as well as costs, which are significantly reduced. Functional needs vary from one tenant to another; either companies from different sectors run different types of applications or, although deploying the same functionality, they do differ in the extent of their complexity. In any case, MTA leaves one major concern regarding the companies' data, their privacy and security, which requires special attention to the data layer. In this article, we propose an extended data model that enhances traditional MTAs in respect of this concern. This extension - called multi-target - allows MT applications to host, manage and serve multiple functionalities within the same multi-tenant (MT) environment. The practical deployment of this approach will allow SaaS vendors to target multiple markets or address different levels of functional complexity and yet commercialise just one single MT application. The applicability of the approach is demonstrated via a case study of a real multi-tenancy multi-target (MT2) implementation, called Globalgest.

  20. Circadian Integration of Glutamatergic Signals by Little SAAS in Novel Suprachiasmatic Circuits

    PubMed Central

    Atkins, Norman; Mitchell, Jennifer W.; Romanova, Elena V.; Morgan, Daniel J.; Cominski, Tara P.; Ecker, Jennifer L.; Pintar, John E.; Sweedler, Jonathan V.; Gillette, Martha U.

    2010-01-01

    Background Neuropeptides are critical integrative elements within the central circadian clock in the suprachiasmatic nucleus (SCN), where they mediate both cell-to-cell synchronization and phase adjustments that cause light entrainment. Forward peptidomics identified little SAAS, derived from the proSAAS prohormone, among novel SCN peptides, but its role in the SCN is poorly understood. Methodology/Principal Findings Little SAAS localization and co-expression with established SCN neuropeptides were evaluated by immunohistochemistry using highly specific antisera and stereological analysis. Functional context was assessed relative to c-FOS induction in light-stimulated animals and on neuronal circadian rhythms in glutamate-stimulated brain slices. We found that little SAAS-expressing neurons comprise the third most abundant neuropeptidergic class (16.4%) with unusual functional circuit contexts. Little SAAS is localized within the densely retinorecipient central SCN of both rat and mouse, but not the retinohypothalamic tract (RHT). Some little SAAS colocalizes with vasoactive intestinal polypeptide (VIP) or gastrin-releasing peptide (GRP), known mediators of light signals, but not arginine vasopressin (AVP). Nearly 50% of little SAAS neurons express c-FOS in response to light exposure in early night. Blockade of signals that relay light information, via NMDA receptors or VIP- and GRP-cognate receptors, has no effect on phase delays of circadian rhythms induced by little SAAS. Conclusions/Significance Little SAAS relays signals downstream of light/glutamatergic signaling from eye to SCN, and independent of VIP and GRP action. These findings suggest that little SAAS forms a third SCN neuropeptidergic system, processing light information and activating phase-shifts within novel circuits of the central circadian clock. PMID:20830308

  1. Course of serum amyloid A (SAA) plasma concentrations in horses undergoing surgery for injuries penetrating synovial structures, an observational clinical study.

    PubMed

    Haltmayer, Eva; Schwendenwein, Ilse; Licka, Theresia F

    2017-05-22

    Injuries penetrating synovial structures are common in equine practice and often result in septic synovitis. Significantly increased plasma levels of serum amyloid A (SAA) have been found in various infectious conditions in horses including wounds and septic arthritis. Plasma SAA levels were found to decrease rapidly once the infectious stimulus was eliminated. The purpose of the current study was to investigate the usefulness of serial measurements of plasma SAA as a monitoring tool for the response to treatment of horses presented with injuries penetrating synovial structures. In the current study plasma SAA concentrations were measured every 48 hours (h) during the course of treatment. A total of 19 horses with a wound penetrating a synovial structure were included in the current study. Horses in Group 1 (n = 12) (injuries older than 24 h) only needed one surgical intervention. Patients in this group had significantly lower median plasma SAA levels (P = 0.001) between 48 h (median 776 mg/L) and 96 h (median 202 mg/L) after surgery. A significant decrease (P = 0.004) in plasma SAA levels was also observed between 96 h after surgery (median 270 mg/L) and 6 days (d) after surgery (median 3 mg/L). Four horses (Group 2) required more than one surgical intervention. In contrast to Group 1 patients in Group 2 had either very high initial plasma concentrations (3378 mg/L), an increase or persistently high concentrations of plasma SAA after the first surgery (median 2525 mg/L). A small group of patients (n = 3) (Group 3) were admitted less than 24 h after sustaining a wound. In this group low SAA values at admission (median 23 mg/L) and peak concentrations at 48 h after surgery (median 1016 mg/L) were observed followed by a decrease in plasma SAA concentration over time. A decrease in plasma SAA concentrations between two consecutive time points could be associated with positive response to treatment in the current study. Therefore, serial

  2. Serum Amyloid A Production Is Triggered by Sleep Deprivation in Mice and Humans: Is That the Link between Sleep Loss and Associated Comorbidities?

    PubMed Central

    de Oliveira, Edson M.; Visniauskas, Bruna; Tufik, Sergio; Andersen, Monica L.; Chagas, Jair R.; Campa, Ana

    2017-01-01

    Serum amyloid A (SAA) was recently associated with metabolic endotoxemia, obesity and insulin resistance. Concurrently, insufficient sleep adversely affects metabolic health and is an independent predisposing factor for obesity and insulin resistance. In this study we investigated whether sleep loss modulates SAA production. The serum SAA concentration increased in C57BL/6 mice subjected to sleep restriction (SR) for 15 days or to paradoxical sleep deprivation (PSD) for 72 h. Sleep restriction also induced the upregulation of Saa1.1/Saa2.1 mRNA levels in the liver and Saa3 mRNA levels in adipose tissue. SAA levels returned to the basal range after 24 h in paradoxical sleep rebound (PSR). Metabolic endotoxemia was also a finding in SR. Increased plasma levels of SAA were also observed in healthy human volunteers subjected to two nights of total sleep deprivation (Total SD), returning to basal levels after one night of recovery. The observed increase in SAA levels may be part of the initial biochemical alterations caused by sleep deprivation, with potential to drive deleterious conditions such as metabolic endotoxemia and weight gain. PMID:28335560

  3. Human serum amyloid A genes are expressed in monocyte/macrophage cell lines.

    PubMed

    Urieli-Shoval, S; Meek, R L; Hanson, R H; Eriksen, N; Benditt, E P

    1994-09-01

    Serum amyloid A (apoSAA) is a family of proteins found, mainly associated with high density lipoproteins, in the blood plasma of mammals and at least one avian species, the Pekin duck. These proteins are present in small amounts under normal circumstances, but their concentration is capable of rising 100- to 1,000-fold in situations involving tissue injury or infection. Like classic acute phase proteins they are produced in the liver; however, expression of one of the apoSAA genes is known to occur in activated macrophages of mice. We examined three human macrophage precursor cell lines (THP-1, U-937, and HL-60), before and after differentiation with phorbol 12-myristate 13-acetate or 1 alpha,25-dihydroxy-vitamin D3, for apoSAA messenger (m)-RNA expression and found that: 1) induction of steady-state apoSAA mRNA by lipopolysaccharide, interleukin-1, or interleukin-6 required the presence of the synthetic glucocorticoid dexamethasone; 2) the three known active genes, apoSAA1, apoSAA2, and apoSAA4, were induced in THP-1 cells, whereas the pseudogene apoSAA3 was not; 3) differentiated and undifferentiated THP-1 cells expressed apoSAA mRNA, but U-937 cells expressed apoSAA mRNA (low levels) only after phorbol 12-myristate 13-acetate differentiation and HL-60 cells did not express apoSAA mRNA whether differentiated or not; 4) apoSAA protein was detectable immunologically at a low level in lyophilized medium from induced THP-1 cells. Our findings are compatible with the hypotheses that 1) apoSAA gene expression in human monocytes/macrophages in vivo is differentiation dependent; 2) activated macrophages provide a local source of apoSAA at sites of tissue injury or inflammation; 3) apoSAA is induced in tissue macrophages by local stimuli, under conditions that may not evoke the systemic acute phase response.

  4. Forty-Year "Drift" and Change of the SAA

    NASA Technical Reports Server (NTRS)

    Stassinopoulos, Epaminondas G,; Xapsos, Michael A.; Stauffer, Craig A.

    2015-01-01

    A comprehensive study was conducted to evaluate the change in the location and size of the South Atlantic Anomaly (SAA) as a function of time, primarily in relation to trapped Van Allen Belt proton populations. The study was limited to one altitude only (800 km), which is still within the Earth's atmosphere but is removed from the atmospheric cut-off level located at about 100-200 km.

  5. Low-level lasers and mRNA levels of reference genes used in Escherichia coli

    NASA Astrophysics Data System (ADS)

    Teixeira, A. F.; Machado, Y. L. R. C.; Fonseca, A. S.; Mencalha, A. L.

    2016-11-01

    Low-level lasers are widely used for the treatment of diseases and antimicrobial photodynamic therapy. Reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) is widely used to evaluate mRNA levels and output data from a target gene are commonly relative to a reference mRNA that cannot vary according to treatment. In this study, the level of reference genes from Escherichia coli exposed to red or infrared lasers at different fluences was evaluated. E. coli AB1157 cultures were exposed to red (660 nm) and infrared (808 nm) lasers, incubated (20 min, 37 °C), the total RNA was extracted, and cDNA synthesis was performed to evaluate mRNA levels from arcA, gyrA and rpoA genes by RT-qPCR. Melting curves and agarose gel electrophoresis were carried out to evaluate specific amplification. Data were analyzed by geNorm, NormFinder and BestKeeper. The melting curve and agarose gel electrophoresis showed specific amplification. Although mRNA levels from arcA, gyrA or rpoA genes presented no significant variations trough a traditional statistical analysis, Excel-based tools revealed that these reference genes are not suitable for E. coli cultures exposed to lasers. Our data showed that exposure to low-level red and infrared lasers at different fluences alter the mRNA levels from arcA, gyrA and rpoA in E. coli cells.

  6. Impact of STAT/SOCS mRNA Expression Levels after Major Injury

    PubMed Central

    Brumann, M.; Matz, M.; Kusmenkov, T.; Stegmaier, J.; Biberthaler, P.; Kanz, K.-G.; Mutschler, W.; Bogner, V.

    2014-01-01

    Background. Fulminant changes in cytokine receptor signalling might provoke severe pathological alterations after multiple trauma. The aim of this study was to evaluate the posttraumatic imbalance of the innate immune system with a special focus on the STAT/SOCS family. Methods. 20 polytraumatized patients were included. Blood samples were drawn 0 h–72 h after trauma; mRNA expression profiles of IL-10, STAT 3, SOCS 1, and SOCS 3 were quantified by qPCR. Results. IL-10 mRNA expression increased significantly in the early posttraumatic period. STAT 3 mRNA expressions showed a significant maximum at 6 h after trauma. SOCS 1 levels significantly decreased 6 h–72 h after trauma. SOCS 3 levels were significantly higher in nonsurvivors 6 h after trauma. Conclusion. We present a serial, sequential investigation in human neutrophil granulocytes of major trauma patients evaluating mRNA expression profiles of IL-10, STAT 3, SOCS 1, and SOCS 3. Posttraumatically, immune disorder was accompanied by a significant increase of IL-10 and STAT 3 mRNA expression, whereas SOCS 1 mRNA levels decreased after injury. We could demonstrate that death after trauma was associated with higher SOCS 3 mRNA levels already at 6 h after trauma. To support our results, further investigations have to evaluate protein levels of STAT/SOCS family in terms of posttraumatic immune imbalance. PMID:24648661

  7. 24 CFR 3282.309 - Formal and informal presentations of views held by SAAs.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 24 Housing and Urban Development 5 2010-04-01 2010-04-01 false Formal and informal presentations of views held by SAAs. 3282.309 Section 3282.309 Housing and Urban Development Regulations Relating... held by SAAs. (a) When an SAA is the appropriate agency to hold a Formal or Informal Presentation of...

  8. A Novel Method for Mining SaaS Software Tag via Community Detection in Software Services Network

    NASA Astrophysics Data System (ADS)

    Qin, Li; Li, Bing; Pan, Wei-Feng; Peng, Tao

    The number of online software services based on SaaS paradigm is increasing. However, users usually find it hard to get the exact software services they need. At present, tags are widely used to annotate specific software services and also to facilitate the searching of them. Currently these tags are arbitrary and ambiguous since mostly of them are generated manually by service developers. This paper proposes a method for mining tags from the help documents of software services. By extracting terms from the help documents and calculating the similarity between the terms, we construct a software similarity network where nodes represent software services, edges denote the similarity relationship between software services, and the weights of the edges are the similarity degrees. The hierarchical clustering algorithm is used for community detection in this software similarity network. At the final stage, tags are mined for each of the communities and stored as ontology.

  9. Biofortification of riboflavin and folate in idli batter, based on fermented cereal and pulse, by Lactococcus lactis N8 and Saccharomyces boulardii SAA655.

    PubMed

    Chandrasekar Rajendran, S C; Chamlagain, B; Kariluoto, S; Piironen, V; Saris, P E J

    2017-06-01

    Lactococcus lactis N8 and Saccharomyces boulardii SAA655 were investigated for their ability to synthesize B-vitamins (riboflavin and folate) and their functional role as microbial starters in idli fermentation. In this study, ultra-high performance liquid chromatography and microbiological assay were used to determine the total riboflavin and folate content respectively. Increased levels of folate were evident in both L. lactis N8 and S. boulardii SAA655 cultivated medium. Enhanced riboflavin levels were found only in S. boulardii SAA655 grown medium, whereas decreased riboflavin level was found in L. lactis N8 cultivated medium. To evaluate the functional role of microbial starter strains, L. lactis N8 and S. boulardii SAA655 were incorporated individually and in combination into idli batter, composed of wet grounded rice and black gram. For the experiments, naturally fermented idli batter was considered as control. The results indicated that natural idli fermentation did not enhance the riboflavin level and depleted folate levels by half. In comparison with control, L. lactis N8 and S. boulardii SAA655 incorporated idli batter (individually and in combination) increased riboflavin and folate levels by 40-90%. Apart from compensating the folate loss caused by natural fermentation, S. boulardii SAA655 fermented idli batter individually and in combination with L. lactis N8 also showed the highest leavening character. Moreover, the microbial starter incorporation did not significantly influence the pH of idli batter. Incorporation of L. lactis N8 and S. boulardii SAA655 can evidently enhance the functional and technological characteristics of idli batter. UN General Assembly declared 2016 the International Year of pulses emphasizing the importance of legumes as staple food. Furthermore, this is the first experimental report of in situ biofortifcation of riboflavin and folate using microbes in pulse based fermented staple food. The current study suggests possible

  10. Development of a SaaS application probe to the physical properties of the Earth's interior: An attempt at moving HPC to the cloud

    NASA Astrophysics Data System (ADS)

    Huang, Qian

    2014-09-01

    Scientific computing often requires the availability of a massive number of computers for performing large-scale simulations, and computing in mineral physics is no exception. In order to investigate physical properties of minerals at extreme conditions in computational mineral physics, parallel computing technology is used to speed up the performance by utilizing multiple computer resources to process a computational task simultaneously thereby greatly reducing computation time. Traditionally, parallel computing has been addressed by using High Performance Computing (HPC) solutions and installed facilities such as clusters and super computers. Today, it has been seen that there is a tremendous growth in cloud computing. Infrastructure as a Service (IaaS), the on-demand and pay-as-you-go model, creates a flexible and cost-effective mean to access computing resources. In this paper, a feasibility report of HPC on a cloud infrastructure is presented. It is found that current cloud services in IaaS layer still need to improve performance to be useful to research projects. On the other hand, Software as a Service (SaaS), another type of cloud computing, is introduced into an HPC system for computing in mineral physics, and an application of which is developed. In this paper, an overall description of this SaaS application is presented. This contribution can promote cloud application development in computational mineral physics, and cross-disciplinary studies.

  11. Amyloid precursor protein mRNA levels in Alzheimer's disease brain.

    PubMed

    Preece, Paul; Virley, David J; Costandi, Moheb; Coombes, Robert; Moss, Stephen J; Mudge, Anne W; Jazin, Elena; Cairns, Nigel J

    2004-03-17

    Insoluble beta-amyloid deposits in Alzheimer's disease (AD) brain are proteolytically derived from the membrane bound amyloid precursor protein (APP). The APP gene is differentially spliced to produce isoforms that can be classified into those containing a Kunitz-type serine protease inhibitor domain (K(+), APP(751), APP(770), APRP(365) and APRP(563)), and those without (K(-), APP(695) and APP(714)). Given the hypothesis that Abeta is a result of aberrant catabolism of APP, differential expression of mRNA isoforms containing protease inhibitors might play an active role in the pathology of AD. We took 513 cerebral cortex samples from 90 AD and 81 control brains and quantified the mRNA isoforms of APP with TaqMan real-time RT-PCR. After adjustment for age at death, brain pH and gender we found a change in the ratio of KPI(+) to KPI(-) mRNA isoforms of APP. Three separate probes, designed to recognise only KPI(+) mRNA species, gave increases of between 28% and 50% in AD brains relative to controls (p=0.002). There was no change in the mRNA levels of KPI-(APP 695) (p=0.898). Therefore, whilst KPI-mRNA levels remained stable the KPI(+) species increased specifically in the AD brains.

  12. Natural antisense transcript-targeted regulation of inducible nitric oxide synthase mRNA levels.

    PubMed

    Yoshigai, Emi; Hara, Takafumi; Araki, Yoshiro; Tanaka, Yoshito; Oishi, Masaharu; Tokuhara, Katsuji; Kaibori, Masaki; Okumura, Tadayoshi; Kwon, A-Hon; Nishizawa, Mikio

    2013-04-01

    Natural antisense transcripts (asRNAs) are frequently transcribed from mammalian genes. Recently, we found that non-coding asRNAs are transcribed from the 3' untranslated region (3'UTR) of the rat and mouse genes encoding inducible nitric oxide synthase (iNOS), which catalyzes the production of the inflammatory mediator nitric oxide. The iNOS asRNA stabilizes iNOS mRNA by interacting with the mRNA 3'UTR. Furthermore, single-stranded 'sense' oligonucleotides corresponding to the iNOS mRNA sequence were found to reduce iNOS mRNA levels by interfering with mRNA-asRNA interactions in rat hepatocytes. This method was named natural antisense transcript-targeted regulation (NATRE) technology. In this study, we detected human iNOS asRNA expressed in hepatocarcinoma and colon carcinoma tissues. The human iNOS asRNA harbored a sequence complementary to an evolutionarily conserved region of the iNOS mRNA 3'UTR. When introduced into hepatocytes, iNOS sense oligonucleotides that were modified by substitution with partial phosphorothioate bonds and locked nucleic acids or 2'-O-methyl nucleic acids greatly reduced levels of iNOS mRNA and iNOS protein. Moreover, sense oligonucleotides and short interfering RNAs decreased iNOS mRNA to comparable levels. These results suggest that NATRE technology using iNOS sense oligonucleotides could potentially be used to treat human inflammatory diseases and cancers by reducing iNOS mRNA levels. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Calpain expression in lymphoid cells. Increased mRNA and protein levels after cell activation.

    PubMed

    Deshpande, R V; Goust, J M; Chakrabarti, A K; Barbosa, E; Hogan, E L; Banik, N L

    1995-02-10

    Although calpain is ubiquitously present in human tissues and is thought to play a role in demyelination, its activity is very low in resting normal lymphocytes. To determine the nature of calpain expression at the mRNA and protein levels in human lymphoid cells, we studied human T lymphocytic, B lymphocytic, and monocytic lines as well as peripheral blood mononuclear cells. Stimulation of cells with the phorbol ester phorbol myristate acetate and the calcium ionophore A23187 resulted in increased calpain mRNA and protein expression. Calpain mRNA expression is also increased in human T cells stimulated with anti-CD3. A dissociation between the increases of RNA and protein suggested that calpain could be released from the cells; the subsequent experiments showed its presence in the extracellular environment. 5,6-Dichloro-1b-D-ribofuranosylbenzimidazole, a reversible inhibitor of mRNA synthesis, reduced calpain mRNA levels by 50-67% and protein levels by 72-91%. Its removal resulted in resumption of both calpain mRNA and protein synthesis. Cycloheximide, a translational inhibitor, reduced calpain protein levels by 77-81% and calpain mRNA levels by 96% in activated THP-1 cells. Interferon-gamma induced calpain mRNA and protein in U-937 and THP-1 cells. Dexamethasone increased mRNA expression in THP-1 cells. Our results indicate that activation of lymphoid cells results in de novo synthesis and secretion of calpain.

  14. ProSAAS-derived peptides are regulated by cocaine and are required for sensitization to the locomotor effects of cocaine.

    PubMed

    Berezniuk, Iryna; Rodriguiz, Ramona M; Zee, Michael L; Marcus, David J; Pintar, John; Morgan, Daniel J; Wetsel, William C; Fricker, Lloyd D

    2017-11-01

    To identify neuropeptides that are regulated by cocaine, we used a quantitative peptidomic technique to examine the relative levels of neuropeptides in several regions of mouse brain following daily intraperitoneal administration of 10 mg/kg cocaine or saline for 7 days. A total of 102 distinct peptides were identified in one or more of the following brain regions: nucleus accumbens, caudate putamen, frontal cortex, and ventral tegmental area. None of the peptides detected in the caudate putamen or frontal cortex were altered by cocaine administration. Three peptides in the nucleus accumbens and seven peptides in the ventral tegmental area were significantly decreased in cocaine-treated mice. Five of these ten peptides are derived from proSAAS, a secretory pathway protein and neuropeptide precursor. To investigate whether proSAAS peptides contribute to the physiological effects of psychostimulants, we examined acute responses to cocaine and amphetamine in the open field with wild-type (WT) and proSAAS knockout (KO) mice. Locomotion was stimulated more robustly in the WT compared to mutant mice for both psychostimulants. Behavioral sensitization to amphetamine was not maintained in proSAAS KO mice and these mutants failed to sensitize to cocaine. To determine whether the rewarding effects of cocaine were altered, mice were tested in conditioned place preference (CPP). Both WT and proSAAS KO mice showed dose-dependent CPP to cocaine that was not distinguished by genotype. Taken together, these results suggest that proSAAS-derived peptides contribute differentially to the behavioral sensitization to psychostimulants, while the rewarding effects of cocaine appear intact in mice lacking proSAAS. © 2017 International Society for Neurochemistry.

  15. Quantification of VGF- and pro-SAAS-derived peptides in endocrine tissues and the brain, and their regulation by diet and cold stress.

    PubMed

    Chakraborty, Tandra R; Tkalych, Oleg; Nanno, Daniela; Garcia, Angelo L; Devi, Lakshmi A; Salton, Stephen R J

    2006-05-17

    Two novel granin-like polypeptides, VGF and pro-SAAS, which are stored in and released from secretory vesicles and are expressed widely in nervous, endocrine, and neuroendocrine tissues, play roles in the regulation of body weight, feeding, and energy expenditure. Both VGF and pro-SAAS are cleaved into peptide fragments, several of which are biologically active. We utilized a highly sensitive and specific radioimmunoassay (RIA) to immunoreactive, pro-SAAS-derived PEN peptides, developed another against immunoreactive, VGF-derived AQEE30 peptides, and quantified these peptides in various mouse tissues and brain regions. Immunoreactive AQEE30 was most abundant in the pituitary, while brain levels were highest in hypothalamus, striatum, and frontal cortex. Immunoreactive PEN levels were highest in the pancreas and spinal cord, and in brain, PEN was most abundant in striatum, hippocampus, pons and medulla, and cortex. Since both peptides were expressed in hypothalamus, a region of the brain that controls feeding and energy expenditure, double label immunofluorescence studies were employed. These demonstrated that 42% of hypothalamic arcuate neurons coexpress VGF and SAAS peptides, and that the intracellular distributions of these peptides in arcuate neurons differed. By RIA, cold stress increased immunoreactive AQEE30 and PEN peptide levels in female but not male hypothalamus, while a high fat diet increased AQEE30 and PEN peptide levels in female but not male hippocampus. VGF and SAAS-derived peptides are therefore widely expressed in endocrine, neuroendocrine, and neural tissues, can be accurately quantified by RIA, and are differentially regulated in the brain by diet and cold stress.

  16. Low-level lasers alter mRNA levels from traditional reference genes used in breast cancer cells

    NASA Astrophysics Data System (ADS)

    Teixeira, A. F.; Canuto, K. S.; Rodrigues, J. A.; Fonseca, A. S.; Mencalha, A. L.

    2017-07-01

    Cancer is among the leading causes of mortality worldwide, increasing the importance of treatment development. Low-level lasers are used in several diseases, but some concerns remains on cancers. Reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) is a technique used to understand cellular behavior through quantification of mRNA levels. Output data from target genes are commonly relative to a reference that cannot vary according to treatment. This study evaluated reference genes levels from MDA-MB-231 cells exposed to red or infrared lasers at different fluences. Cultures were exposed to red and infrared lasers, incubated (4 h, 37 °C), total RNA was extracted and cDNA synthesis was performed to evaluate mRNA levels from ACTB, GUSB and TRFC genes by RT-qPCR. Specific amplification was verified by melting curves and agarose gel electrophoresis. RefFinder enabled data analysis by geNorm, NormFinder and BestKeeper. Specific amplifications were obtained and, although mRNA levels from ACTB, GUSB or TRFC genes presented no significant variation through traditional statistical analysis, Excel-based tools revealed that the use of these reference genes are dependent of laser characteristics. Our data showed that exposure to low-level red and infrared lasers at different fluences alter the mRNA levels from ACTB, GUSB and TRFC in MDA-MB-231 cells.

  17. Selective probing of mRNA expression levels within a living cell.

    PubMed

    Nawarathna, D; Turan, T; Wickramasinghe, H Kumar

    2009-08-24

    We report on a selective and nondestructive measurement of mRNA (messenger ribonucleic acid) expression levels within a living cell. We first modify an atomic force microscope tip to create a tapered nanoscale coaxial cable. Application of an ac (alternating potential) between the inner and outer electrodes of this cable creates a dielectrophoretic force attracting mRNA molecules toward the tip-end which is pretreated with gene specific primers. We selectively extracted and analyzed both high ( approximately 2500) and extremely low (11 0) copy number mRNA from a living cell mRNA in less than 10 s.

  18. Selective probing of mRNA expression levels within a living cell

    PubMed Central

    Nawarathna, D.; Turan, T.; Wickramasinghe, H. Kumar

    2009-01-01

    We report on a selective and nondestructive measurement of mRNA (messenger ribonucleic acid) expression levels within a living cell. We first modify an atomic force microscope tip to create a tapered nanoscale coaxial cable. Application of an ac (alternating potential) between the inner and outer electrodes of this cable creates a dielectrophoretic force attracting mRNA molecules toward the tip-end which is pretreated with gene specific primers. We selectively extracted and analyzed both high (∼2500) and extremely low (11¯0) copy number mRNA from a living cell mRNA in less than 10 s. PMID:19777090

  19. BIOMARKERS OF ENDOCRINE DISRUPTION AT THE MRNA LEVEL

    EPA Science Inventory

    Denslow, Nancy D., Christopher J. Bowman, Gillian Robinson, H. Stephen Lee, Ronald J. Ferguson, Michael J. Hemmer and Leroy C. Folmar. 1999. Biomarkers of Endocrine Disruption at the mRNA Level. In: Environmental Toxicology and Risk Assessment: Standardization of Biomarkers for ...

  20. TS mRNA levels can predict pemetrexed and raltitrexed sensitivity in colorectal cancer.

    PubMed

    Zhang, Qun; Shen, Jie; Wang, Hao; Hu, Jing; Yu, Lixia; Xie, Li; Wei, Jia; Liu, Baorui; Guan, Wenxian; Qian, Xiaoping

    2014-02-01

    The purpose of the study is to analyze the relationship between tumor thymidylate synthase (TS) mRNA expression levels and raltitrexed/pemetrexed/5-FU sensitivity. We collected freshly removed colorectal tumor specimens from 50 patients. Chemosensitivities to anticancer drugs were evaluated by histoculture drug response assay. We adopted quantitative reverse transcription polymerase chain reaction for TS mRNA detection and immunohistochemical staining for assessing TS expression in tumor tissues. There is a significant relationship between TS mRNA expression levels and in vitro chemosensitivity of freshly removed colorectal tumor specimens to pemetrexed (P < 0.001)/raltitrexed (P = 0.004)/5-FU (P = 0.007). TS mRNA expression levels can predict pemetrexed and raltitrexed sensitivity in colorectal cancer.

  1. Development, food intake, and ethinylestradiol influence hepatic triglyceride lipase and LDL-receptor mRNA levels in rats.

    PubMed

    Staels, B; Jansen, H; van Tol, A; Stahnke, G; Will, H; Verhoeven, G; Auwerx, J

    1990-07-01

    The influence of development and ethinylestradiol on low density lipoprotein (LDL)-receptor mRNA and hepatic triglyceride lipase (HTGL) activity and mRNA levels was studied in rat liver and intestine. Intestinal LDL-receptor mRNA levels are maximal in the perinatal period, whereas liver LDL-receptor and HTGL mRNA levels are highest after weaning in adult life. All mRNA levels reach a maximum between day 15 and 20 when rats still consume a lipid-rich diet, and increase twofold during weaning. Liver and intestinal LDL-receptor mRNA levels are not influenced by ovariectomy, but increase after ethinylestradiol treatment. Liver LDL-receptor mRNA shows a dose-dependent increase after ethinylestradiol and a sevenfold rise in liver LDL-receptor mRNA is attained with a dose of 2000 micrograms/day. Intestinal LDL-receptor mRNA increases slightly more than twofold after ethinylestradiol and this increase is not dose-dependent. Changes in LDL-receptor mRNA are independent of changes in food intake induced by ethinylestradiol treatment, since they are still observed after pair-feeding. The ethinylestradiol-induced increases in LDL-receptor mRNA levels are reflected by decreased serum apoB levels. HTGL mRNA levels increase after ovariectomy and show a dose-dependent decrease after ethinylestradiol. Pair-feeding abolishes the increase seen after ovariectomy, while the estrogen-mediated decrease is attenuated. These alterations in HTGL mRNA are reflected by similar changes in liver HTGL activity.(ABSTRACT TRUNCATED AT 250 WORDS)

  2. Low-level lasers on microRNA and uncoupling protein 2 mRNA levels in human breast cancer cells

    NASA Astrophysics Data System (ADS)

    Canuto, K. S.; Teixeira, A. F.; Rodrigues, J. A.; Paoli, F.; Nogueira, E. M.; Mencalha, A. L.; Fonseca, A. S.

    2017-06-01

    MicroRNA is short non-coding RNA and is a mediator of post-transcriptional regulation of gene expression. In addition, uncoupling proteins (UCPs) regulate thermogenesis, metabolic and energy balance, and decrease reactive oxygen species production. Both microRNA and UCP2 expression can be altered in cancer cells. At low power, laser wavelength, frequency, fluence and emission mode deternube photobiological responses, which are the basis of low-level laser therapy. There are few studies on miRNA and UCP mRNA levels after low-level laser exposure on cancer cells. In this work, we evaluate the micrRNA (mir-106b and mir-15a) and UCP2 mRNA levels in human breast cancer cells exposed to low-level lasers. MDA-MB-231 human breast cancer cells were exposed to low-level red and infrared lasers, total RNA was extracted for cDNA synthesis and mRNA levels by real time quantitative polymerase chain reaction were evaluated. Data show that mir-106b and mir-15a relative levels are not altered, but UCP2 mRNA relative levels are increased in MDA-MB-231 human breast cancer cells exposed to low-level red and infrared lasers at fluences used in therapeutic protocols.

  3. The Generic Spacecraft Analyst Assistant (GenSAA): A tool for automating spacecraft monitoring with expert systems

    NASA Technical Reports Server (NTRS)

    Hughes, Peter M.; Luczak, Edward C.

    1991-01-01

    Flight Operations Analysts (FOAs) in the Payload Operations Control Center (POCC) are responsible for monitoring a satellite's health and safety. As satellites become more complex and data rates increase, FOAs are quickly approaching a level of information saturation. The FOAs in the spacecraft control center for the COBE (Cosmic Background Explorer) satellite are currently using a fault isolation expert system named the Communications Link Expert Assistance Resource (CLEAR), to assist in isolating and correcting communications link faults. Due to the success of CLEAR and several other systems in the control center domain, many other monitoring and fault isolation expert systems will likely be developed to support control center operations during the early 1990s. To facilitate the development of these systems, a project was initiated to develop a domain specific tool, named the Generic Spacecraft Analyst Assistant (GenSAA). GenSAA will enable spacecraft analysts to easily build simple real-time expert systems that perform spacecraft monitoring and fault isolation functions. Lessons learned during the development of several expert systems at Goddard, thereby establishing the foundation of GenSAA's objectives and offering insights in how problems may be avoided in future project, are described. This is followed by a description of the capabilities, architecture, and usage of GenSAA along with a discussion of its application to future NASA missions.

  4. High BIM mRNA levels are associated with longer survival in advanced gastric cancer.

    PubMed

    Wu, Nandie; Huang, Ying; Zou, Zhengyun; Gimenez-Capitan, Ana; Yu, Lixia; Hu, Wenjing; Zhu, Lijing; Sun, Xia; Sanchez, Jose Javier; Guan, Wenxian; Liu, Baorui; Rosell, Rafael; Wei, Jia

    2017-03-01

    Chemotherapy drugs, including 5-fluorouracil (5-FU), oxaliplatin and docetaxel, are commonly used in the treatment of gastric cancer (GC). Apoptosis-relevant genes may be associated with drug resistance. In the present study, the messenger RNA (mRNA) expression levels of B-cell lymphoma 2 interacting mediator of cell death (BIM), astrocyte elevated gene-1 (AEG-1) and AXL receptor tyrosine kinase (AXL) were investigated in 131 advanced GC samples, and the expression levels of these genes were correlated with patients' overall survival (OS). All 131 patients received first-line FOLFOX combination chemotherapy with folinic acid and 5-FU, in which 56 patients were further treated with second-line docetaxel-based chemotherapy. A correlation between the mRNA expression levels of BIM and AEG-1 was observed ( r s =0.30; P=0.002). There was no association between the mRNA expression levels of any of the individual genes analyzed and OS in patients only receiving first-line FOLFOX chemotherapy. In a subgroup of patients receiving docetaxel-based second-line chemotherapy, those with high or intermediate levels of BIM exhibited a median OS of 18.2 months [95% confidence interval (CI), 12.8-23.6], compared with 9.6 months (95% CI, 8.9-10.3) in patients with low BIM levels (P=0.008). However, there was no correlation between the mRNA expression levels of AEG-1 or AXL and OS. The risk of mortality was higher in patients with low BIM mRNA levels than in those with high or intermediate BIM mRNA levels (hazard ratio, 2.61; 95% CI, 1.21-5.62; P=0.010). Therefore, BIM may be considered as a biomarker to identify whether patients could benefit from docetaxel-based second-line chemotherapy in GC.

  5. ATI SAA Annex 3 Button Tensile Test Report I

    NASA Technical Reports Server (NTRS)

    Tang, Henry H.

    2013-01-01

    This report documents the results of a study carried out under Splace Act Agreement SAA-EA-10-004 between the National Aeronautics and Space Administration (NASA) and Astro Technology Incorpporated (ATI). NASA and ATI have entered into this agreement to collaborate on the development of technologies that can benefit both the US government space programs and the oil and gas industry. The report documents the results of a test done on an adnesive system for attaching new monitoring sensor devices to pipelines under Annex III of SAA-EA-10-004: "Proof-of-Concept Design and Testing of a Post Installed Sensing Device on Subsea Risers and Pipelines". The tasks of Annex III are to design and test a proof-of-concept sensing device for in-situ installation on pipelines, risers, or other structures deployed in deep water. The function of the sensor device is to measure various signals such as strain, stress and temperature. This study complements the work done, in Annex I of the SAA, on attaching a fiber optic sensing device to pipe via adhesive bonding. Both Annex I and Annex III studies were conducted in the Crew and Thermal System Division (CTSD) at the Johnson Space Center (JSC) in collaboration with ATI.

  6. Space Shuttle drops down the SAA doses on ISS

    NASA Astrophysics Data System (ADS)

    Dachev, T. P.; Semkova, J.; Tomov, B.; Matviichuk, Yu.; Dimitrov, Pl.; Koleva, R.; Malchev, St.; Reitz, G.; Horneck, G.; De Angelis, G.; Häder, D.-P.; Petrov, V.; Shurshakov, V.; Benghin, V.; Chernykh, I.; Drobyshev, S.; Bankov, N. G.

    2011-06-01

    Long-term analysis of data from two radiation detection instruments on the International Space Station (ISS) shows that the docking of the Space Shuttle drops down the measured dose rates in the region of the South Atlantic Anomaly (SAA) by a factor of 1.5-3. Measurements either by the R3DE detector, which is outside the ISS at the EuTEF facility on the Columbus module behind a shielding of less than 0.45 g cm -2, and by the three detectors of the Liulin-5 particle telescope, which is inside the Russian PEARS module in the spherical tissue equivalent phantom behind much heavier shielding demonstrate that effect. Simultaneously the estimated averaged incident energies of the incoming protons rise up from about 30 to 45 MeV. The effect is explained by the additional shielding against the SAA 30-150 MeV protons, provided by the 78 tons Shuttle to the instruments inside and outside of the ISS. An additional reason is the ISS attitude change (performed for the Shuttle docking) leading to decreasing of dose rates in two of Liulin-5 detectors because of the East-West proton fluxes asymmetry in SAA. The Galactic Cosmic Rays dose rates are practically not affected.

  7. Prognostic impact of mRNA levels of osteopontin splice variants in soft tissue sarcoma patients.

    PubMed

    Hahnel, Antje; Wichmann, Henri; Greither, Thomas; Kappler, Matthias; Würl, Peter; Kotzsch, Matthias; Taubert, Helge; Vordermark, Dirk; Bache, Matthias

    2012-04-02

    It is well known that osteopontin (OPN) plays an important role in tumor progression and that a high OPN expression level in several tumor entities correlates with poor prognosis in cancer patients. However, little is known about the prognostic relevance of the OPN mRNA splice variants. We analyzed the mRNA expression levels of different OPN splice variants in tumor tissue of 124 soft tissue sarcoma (STS) patients. Quantitative real-time PCR (qRT-PCR) was used to analyze the mRNA expression level of three OPN splice variants (OPN-a, -b and -c). The multivariate Cox's proportional hazard regression model revealed that high mRNA expression levels of OPN splice variants are significantly associated with poor prognosis in STS patients (n = 124). Women (n = 68) with high mRNA expression levels of OPN-a and OPN-b have an especially elevated risk of tumor-related death (OPN-a: RR = 3.0, P = 0.01, CI = 1.3-6.8; OPN-b: RR = 3.4, P = 0.01, CI = 1.4-8.2). In particular, we found that high mRNA expression levels of OPN-b and OPN-c correlated with a high risk of tumor-related death in STS patients that received radiotherapy (n = 52; OPN-b: RR = 10.3, P < 0.01, CI = 2.0-53.7; OPN-c: RR = 11.4, P < 0.01, CI = 2.2-59.3). Our study shows that elevated mRNA expression levels of OPN splice variants are negative prognostic and predictive markers for STS patients. Further studies are needed to clarify the impact of the OPN splice variants on prognosis.

  8. mRNA expression levels of hypoxia-induced and stem cell-associated genes in human glioblastoma.

    PubMed

    Bache, Matthias; Rot, Swetlana; Keßler, Jacqueline; Güttler, Antje; Wichmann, Henri; Greither, Thomas; Wach, Sven; Taubert, Helge; Söling, Ariane; Bilkenroth, Udo; Kappler, Matthias; Vordermark, Dirk

    2015-06-01

    The roles of hypoxia-induced and stem cell-associated genes in the development of malignancy and tumour progression are well known. However, there are a limited number of studies analysing the impact of mRNA expression levels of hypoxia-induced and stem cell-associated genes in the tissues of brain tumours and glioblastoma patients. In this study, tumour tissues from patients with glioblastoma multiforme and tumour adjacent tissues were analysed. We investigated mRNA expression levels of hypoxia-inducible factor-1α (HIF-1α), hypoxia-inducible factor-2α (HIF-2α), carbonic anhydrase 9 (CA9), vascular endothelial growth factor (VEGF), glucose transporter-1 (GLUT-1) and osteopontin (OPN), and stem cell-associated genes survivin, epidermal growth factor receptor (EGFR), human telomerase reverse transcriptase (hTERT), Nanog and octamer binding transcription factor 4 (OCT4) using quantitative real-time polymerase chain reaction (qRT-PCR). Our data revealed higher mRNA expression levels of hypoxia-induced and stem cell-associated genes in tumour tissue than levels in the tumour adjacent tissues in patients with glioblastoma multiforme. A strong positive correlation between the mRNA expression levels of HIF-2α, CA9, VEGF, GLUT-1 and OPN suggests a specific hypoxia-associated profile of mRNA expression in glioblastoma multiforme. Additionally, the results indicate the role of stem-cell-related genes in tumour hypoxia. Kaplan-Maier analysis revealed that high mRNA expression levels of hypoxia-induced markers showed a trend towards shorter overall survival in glioblastoma patients (P=0.061). Our data suggest that mRNA expression levels of hypoxia-induced genes are important tumour markers in patients with glioblastoma multiforme.

  9. Pathogenic Serum Amyloid A 1.1 Shows a Long Oligomer-rich Fibrillation Lag Phase Contrary to the Highly Amyloidogenic Non-pathogenic SAA2.2*

    PubMed Central

    Srinivasan, Saipraveen; Patke, Sanket; Wang, Yun; Ye, Zhuqiu; Litt, Jeffrey; Srivastava, Sunit K.; Lopez, Maria M.; Kurouski, Dmitry; Lednev, Igor K.; Kane, Ravi S.; Colón, Wilfredo

    2013-01-01

    Serum amyloid A (SAA) is best known for being the main component of amyloid in the inflammation-related disease amyloid A (AA) amyloidosis. Despite the high sequence identity among different SAA isoforms, not all SAA proteins are pathogenic. In most mouse strains, the AA deposits mostly consist of SAA1.1. Conversely, the CE/J type mouse expresses a single non-pathogenic SAA2.2 protein that is 94% identical to SAA1.1. Here we show that SAA1.1 and SAA2.2 differ in their quaternary structure, fibrillation kinetics, prefibrillar oligomers, and fibril morphology. At 37 °C and inflammation-related SAA concentrations, SAA1.1 exhibits an oligomer-rich fibrillation lag phase of a few days, whereas SAA2.2 shows virtually no lag phase and forms small fibrils within a few hours. Deep UV resonance Raman, far UV-circular dichroism, atomic force microscopy, and fibrillation cross-seeding experiments suggest that SAA1.1 and SAA2.2 fibrils possess different morphology. Both the long-lived oligomers of pathogenic SAA1.1 and the fleeting prefibrillar oligomers of non-pathogenic SAA2.2, but not their respective amyloid fibrils, permeabilized synthetic bilayer membranes in vitro. This study represents the first comprehensive comparison between the biophysical properties of SAA isoforms with distinct pathogenicities, and the results suggest that structural and kinetic differences in the oligomerization-fibrillation of SAA1.1 and SAA2.2, more than their intrinsic amyloidogenicity, may contribute to their diverse pathogenicity. PMID:23223242

  10. Hamp1 mRNA and plasma hepcidin levels are influenced by sex and strain but do not predict tissue iron levels in inbred mice.

    PubMed

    McLachlan, Stela; Page, Kathryn E; Lee, Seung-Min; Loguinov, Alex; Valore, Erika; Hui, Simon T; Jung, Grace; Zhou, Jie; Lusis, Aldons J; Fuqua, Brie; Ganz, Tomas; Nemeth, Elizabeta; Vulpe, Chris D

    2017-11-01

    Iron homeostasis is tightly regulated, and the peptide hormone hepcidin is considered to be a principal regulator of iron metabolism. Previous studies in a limited number of mouse strains found equivocal sex- and strain-dependent differences in mRNA and serum levels of hepcidin and reported conflicting data on the relationship between hepcidin ( Hamp1 ) mRNA levels and iron status. Our aim was to clarify the relationships between strain, sex, and hepcidin expression by examining multiple tissues and the effects of different dietary conditions in multiple inbred strains. Two studies were done: first, Hamp1 mRNA, liver iron, and plasma diferric transferrin levels were measured in 14 inbred strains on a control diet; and second, Hamp1 mRNA and plasma hepcidin levels in both sexes and iron levels in the heart, kidneys, liver, pancreas, and spleen in males were measured in nine inbred/recombinant inbred strains raised on an iron-sufficient or high-iron diet. Both sex and strain have a significant effect on both hepcidin mRNA (primarily a sex effect) and plasma hepcidin levels (primarily a strain effect). However, liver iron and diferric transferrin levels are not predictors of Hamp1 mRNA levels in mice fed iron-sufficient or high-iron diets, nor are the Hamp1 mRNA and plasma hepcidin levels good predictors of tissue iron levels, at least in males. We also measured plasma erythroferrone, performed RNA-sequencing analysis of liver samples from six inbred strains fed the iron-sufficient, low-iron, or high-iron diets, and explored differences in gene expression between the strains with the highest and lowest hepcidin levels. NEW & NOTEWORTHY Both sex and strain have a significant effect on both hepcidin mRNA (primarily a sex effect) and plasma hepcidin levels (primarily a strain effect). Liver iron and diferric transferrin levels are not predictors of Hamp1 mRNA levels in mice, nor are the Hamp1 mRNA and plasma hepcidin levels good predictors of tissue iron levels, at least

  11. Variation of amino acid sequences of serum amyloid a (SAA) and immunohistochemical analysis of amyloid a (AA) in Japanese domestic cats.

    PubMed

    Tei, Meina; Uchida, Kazuyuki; Chambers, James K; Watanabe, Ken-Ichi; Tamamoto, Takashi; Ohno, Koichi; Nakayama, Hiroyuki

    2018-02-02

    Amyloid A (AA) amyloidosis, a fatal systemic amyloid disease, occurs secondary to chronic inflammatory conditions in humans. Although persistently elevated serum amyloid A (SAA) levels are required for its pathogenesis, not all individuals with chronic inflammation necessarily develop AA amyloidosis. Furthermore, many diseases in cats are associated with the elevated production of SAA, whereas only a small number actually develop AA amyloidosis. We hypothesized that a genetic mutation in the SAA gene may strongly contribute to the pathogenesis of feline AA amyloidosis. In the present study, genomic DNA from four Japanese domestic cats (JDCs) with AA amyloidosis and from five without amyloidosis was analyzed using polymerase chain reaction (PCR) amplification and direct sequencing. We identified the novel variation combination of 45R-51A in the deduced amino acid sequences of four JDCs with amyloidosis and five without. However, there was no relationship between amino acid variations and the distribution of AA amyloid deposits, indicating that differences in SAA sequences do not contribute to the pathogenesis of AA amyloidosis. Immunohistochemical analysis using antisera against the three different parts of the feline SAA protein-i.e., the N-terminal, central, and C-terminal regions-revealed that feline AA contained the C-terminus, unlike human AA. These results indicate that the cleavage and degradation of the C-terminus are not essential for amyloid fibril formation in JDCs.

  12. Detoxification of Corncob Acid Hydrolysate with SAA Pretreatment and Xylitol Production by Immobilized Candida tropicalis

    PubMed Central

    Deng, Li-Hong; Tang, Yong; Liu, Yun

    2014-01-01

    Xylitol fermentation production from corncob acid hydrolysate has become an attractive and promising process. However, corncob acid hydrolysate cannot be directly used as fermentation substrate owing to various inhibitors. In this work, soaking in aqueous ammonia (SAA) pretreatment was employed to reduce the inhibitors in acid hydrolysate. After detoxification, the corncob acid hydrolysate was fermented by immobilized Candida tropicalis cell to produce xylitol. Results revealed that SAA pretreatment showed high delignification and efficient removal of acetyl group compounds without effect on cellulose and xylan content. Acetic acid was completely removed, and the content of phenolic compounds was reduced by 80%. Furthermore, kinetic behaviors of xylitol production by immobilized C. tropicalis cell were elucidated from corncob acid hydrolysate detoxified with SAA pretreatment and two-step adsorption method, respectively. The immobilized C. tropicalis cell showed higher productivity efficiency using the corncob acid hydrolysate as fermentation substrate after detoxification with SAA pretreatment than by two-step adsorption method in the five successive batch fermentation rounds. After the fifth round fermentation, about 60 g xylitol/L fermentation substrate was obtained for SAA pretreatment detoxification, while about 30 g xylitol/L fermentation substrate was obtained for two-step adsorption detoxification. PMID:25133211

  13. Pulsed low-level infrared laser alters mRNA levels from muscle repair genes dependent on power output in Wistar rats

    NASA Astrophysics Data System (ADS)

    Trajano, L. A. S. N.; Trajano, E. T. L.; Thomé, A. M. C.; Sergio, L. P. S.; Mencalha, A. L.; Stumbo, A. C.; Fonseca, A. S.

    2017-10-01

    Satellite cells are present in skeletal muscle functioning in the repair and regeneration of muscle injury. Activation of these cells depends on the expression of myogenic factor 5 (Myf5), myogenic determination factor 1(MyoD), myogenic regulatory factor 4 (MRF4), myogenin (MyoG), paired box transcription factors 3 (Pax3), and 7 (Pax7). Low-level laser irradiation accelerates the repair of muscle injuries. However, data from the expression of myogenic factors have been controversial. Furthermore, the effects of different laser beam powers on the repair of muscle injuries have been not evaluated. The aim of this study was to evaluate the effects of low-level infrared laser at different powers and in pulsed emission mode on the expression of myogenic regulatory factors and on Pax3 and Pax7 in injured skeletal muscle from Wistar rats. Animals that underwent cryoinjury were divided into three groups: injury, injury laser 25 Mw, and injury laser 75 mW. Low-level infrared laser irradiation (904 nm, 3 J cm-2, 5 kHz) was carried out at 25 and 75 mW. After euthanasia, skeletal muscle samples were withdrawn and the total RNA was extracted for the evaluation of mRNA levels from the MyoD, MyoG, MRF4, Myf5, Pax3, and Pax7 gene. Pax 7 mRNA levels did not alter, but Pax3 mRNA levels increased in the injured and laser-irradiated group at 25 mW. MyoD, MyoG, and MYf5 mRNA levels increased in the injured and laser-irradiated animals at both powers, and MRF4 mRNA levels decreased in the injured and laser-irradiated group at 75 mW. In conclusion, exposure to pulsed low-level infrared laser, by power-dependent effect, could accelerate the muscle repair process altering mRNA levels from paired box transcription factors and myogenic regulatory factors.

  14. Transcriptional bursting explains the noise–versus–mean relationship in mRNA and protein levels

    DOE PAGES

    Dar, Roy; Shaffer, Sydney M.; Singh, Abhyudai; ...

    2016-07-28

    Recent analysis demonstrates that the HIV-1 Long Terminal Repeat (HIV LTR) promoter exhibits a range of possible transcriptional burst sizes and frequencies for any mean-expression level. However, these results have also been interpreted as demonstrating that cell-tocell expression variability (noise) and mean are uncorrelated, a significant deviation from previous results. Here, we re-examine the available mRNA and protein abundance data for the HIV LTR and find that noise in mRNA and protein expression scales inversely with the mean along analytically predicted transcriptional burst-size manifolds. We then experimentally perturb transcriptional activity to test a prediction of the multiple burst-size model: thatmore » increasing burst frequency will cause mRNA noise to decrease along given burst-size lines as mRNA levels increase. In conclusion, the data show that mRNA and protein noise decrease as mean expression increases, supporting the canonical inverse correlation between noise and mean.« less

  15. Occupational exposure levels of bioaerosol components are associated with serum levels of the acute phase protein Serum Amyloid A in greenhouse workers.

    PubMed

    Madsen, Anne Mette; Thilsing, Trine; Bælum, Jesper; Garde, Anne Helene; Vogel, Ulla

    2016-01-20

    Occupational exposure to particles may be associated with increased inflammation of the airways. Animal experiments suggest that inhaled particles also induce a pulmonary acute phase response, leading to systemic circulation of acute phase proteins. Greenhouse workers are exposed to elevated levels of bioaerosols. The objective of this study is to assess whether greenhouse workers personal exposure to bioaerosol components was associated with serum levels of the acute phase proteins Serum Amyloid A (SAA) and C-reactive protein (CRP). SAA and CRP levels were determined in serum sampled repeatedly from 33 greenhouse workers. Blood was drawn repeatedly on Mondays and Thursdays during work weeks. Acute phase protein levels were compared to levels in a comparison group of 42 people and related to individual exposure levels to endotoxin, dust, bacteria, fungi and β-glucan. Serum levels of SAA and CRP were not significantly different in greenhouse workers and a reference group, or on the two work days. In a mixed model, SAA levels were positively associated with endotoxin exposure levels (p = 0.0007). Results for fungi were not clear. CRP levels were positively associated with endotoxin exposures (p = 0.022). Furthermore, when workers were categorized into three groups based on SAA and CRP serum levels endotoxin exposure was highest in the group with the highest SAA levels and in the group with middle and highest CRP levels. SAA and CRP levels were elevated in workers with asthma. Greenhouse workers did not have elevated serum levels of SAA and CRP compared to a reference group. However, occupational exposure to endotoxin was positively associated with serum levels of the acute phase proteins SAA and CRP. Preventive measures to reduce endotoxin exposure may be beneficial.

  16. TP53 and ATM mRNA expression in skin and skeletal muscle after low-level laser exposure.

    PubMed

    Guedes de Almeida, Luciana; Sergio, Luiz Philippe da Silva; de Paoli, Flavia; Mencalha, Andre Luiz; da Fonseca, Adenilson de Souza

    2017-08-01

    Low-level lasers are widespread in regenerative medicine, but the molecular mechanisms involved in their biological effects are not fully understood, particularly those on DNA stability. Therefore, this study aimed to investigate mRNA expression of genes related to DNA genomic stability in skin and skeletal muscle tissue from Wistar rats exposed to low-level red and infrared lasers. For this, TP53 (Tumor Protein 53) and ATM (Ataxia Telangiectasia Mutated gene) mRNA expressions were evaluated by real-time quantitative PCR (RT-qPCR) technique 24 hours after low-level red and infrared laser exposure. Our data showed that relative TP53 mRNA expression was not significantly altered in both tissues exposed to lasers. For ATM, relative mRNA expression in skin tissue was not significantly altered, but in muscle tissue, laser exposure increased relative ATM mRNA expression. Low-level red and infrared laser radiations alter ATM mRNA expression related to DNA stability in skeletal muscle tissue.

  17. Reduced beta 2-microglobulin mRNA levels in transgenic mice expressing a designed hammerhead ribozyme.

    PubMed Central

    Larsson, S; Hotchkiss, G; Andäng, M; Nyholm, T; Inzunza, J; Jansson, I; Ahrlund-Richter, L

    1994-01-01

    We have generated three artificial hammerhead ribozymes, denoted 'Rz-b', 'Rz-c' and 'Rz-d', with different specificities for exon II of the mouse beta-2-microglobulin (beta 2M) mRNA. In this study we tested for ribozyme mediated reduction of beta 2M mRNA in a cell line and in transgenic mice. Transfections of either of the Rz-b, Rz-c or Rz-d plasmids into a mouse cell-line (NIH/3T3) revealed reductions of beta 2M mRNA substrate in each case. Ribozyme expression in individual transfected clones was accompanied with an up to 80% reduction of beta 2M mRNA levels. Rz-c was selected for a transgenic study. Seven Rz-c transgenic founder animals were identified from which three ribozyme expressing families were established and analysed. Expression of the ribozyme transgene was tested for and detected in lung, kidney and spleen. Expression was accompanied with reduction of the beta 2M mRNA levels of heterozygous (Rz+/-) animals compared to non-transgenic litter mates. The effect was most pronounced in lung with more than 90% beta 2M mRNA reduction in individual mice. In summary, expression of our ribozymes in a cell free system, in a cell-line and in transgenic mice were all accompanied with reductions of beta 2M mRNA levels. Images PMID:8036151

  18. Anabolic androgenic steroid nandrolone decanoate reduces hypothalamic proopiomelanocortin mRNA levels.

    PubMed

    Lindblom, Jonas; Kindlundh, Anna M S; Nyberg, Fred; Bergström, Lena; Wikberg, Jarl E S

    2003-10-03

    Supratherapeutical doses of anabolic androgenic steroids (AASs) have dramatic effects on metabolism in humans, and also inhibit feeding and reduce the rate of body weight gain in rats. In order to test the hypothesis that the AAS metabolic syndrome is accompanied by alterations in the central melanocortin system, we evaluated body weight, food intake and hypothalamic agouti-related protein (AgRP) and proopiomelanocortin (POMC) mRNA levels following administration of different doses of the anabolic androgenic steroid nandrolone decanoate. In order to distinguish changes induced by the steroid treatment per se from those resulting from the reduced food intake and growth rate, we also compared the effect of nandrolone decanoate on AgRP and POMC mRNA expression with both normally fed, and food restricted control groups. We here report that administration of nandrolone specifically reduces arcuate nucleus POMC mRNA levels while not affecting the expression level of AgRP. The effect on POMC expression was not observed in the food restricted controls, excluding the possibility that the observed effect was a mere response to the reduced food intake and body weight. These results raise the possibility that some of the metabolic and behavioural consequences of AAS abuse may be the result of alterations in the melanocortin system.

  19. Increased IL18 mRNA levels in peripheral artery disease and its association with triglyceride and LDL cholesterol levels: a pilot study.

    PubMed

    Deser, Serkan Burc; Bayoglu, Burcu; Besirli, Kazım; Cengiz, Mujgan; Arapi, Berk; Junusbekov, Yerik; Dirican, Ahmet; Arslan, Caner

    2016-06-01

    Peripheral artery disease (PAD) typically refers to lower limb vessel ischemia caused by atherosclerotic stenosis of lower extremity arteries. IL18 is a pleiotropic pro-inflammatory cytokine reported to function as an inflammatory biomarker in cardiovascular diseases. IL18 activity is balanced by high-affinity naturally occurring IL18-binding protein (IL18BP). This study aimed to determine whether IL18, IL18 BP mRNA levels and -137 G/C (rs187238) polymorphism, which was previously associated with IL18 gene transcriptional activity, were associated with PAD etiology. IL18, IL18BP mRNA levels from peripheral blood mononuclear cells and -137 G/C (rs187238) polymorphism were determined by quantitative real-time polymerase chain reaction (qRT-PCR) and RT-PCR, respectively, in 55 PAD patients (26 aorta-iliac, 29 femoro-popliteal) and 61 disease-free controls. IL18 mRNA levels were increased in PAD patients compared with healthy controls (p = 0.09); however, did not reach a statistical significant level, also did not significantly differ between aorta-iliac and femoro-popliteal occlusive PAD subgroups (p = 0.285). However, IL18BP mRNA levels were significantly lower in PAD group compared with controls (p < 0.001). Genotype frequencies of rs187238 polymorphism did not significantly differ between PAD patients and controls (p = 0.385). IL18 mRNA levels were significantly correlated with triglycerides and LDL cholesterol levels in PAD patients (p = 0.003, p = 0.014, respectively). HDL cholesterol levels were negatively correlated with IL18 mRNA levels in controls (p = 0.05). This report is a preliminary study to show an association between IL18, IL18BP mRNA levels and PAD and suggests that the IL18 gene may have a significant relationship with triglyceride and LDL cholesterol levels in PAD patients.

  20. Reduced changes in protein compared to mRNA levels across non-proliferating tissues.

    PubMed

    Perl, Kobi; Ushakov, Kathy; Pozniak, Yair; Yizhar-Barnea, Ofer; Bhonker, Yoni; Shivatzki, Shaked; Geiger, Tamar; Avraham, Karen B; Shamir, Ron

    2017-04-18

    The quantitative relations between RNA and protein are fundamental to biology and are still not fully understood. Across taxa, it was demonstrated that the protein-to-mRNA ratio in steady state varies in a direction that lessens the change in protein levels as a result of changes in the transcript abundance. Evidence for this behavior in tissues is sparse. We tested this phenomenon in new data that we produced for the mouse auditory system, and in previously published tissue datasets. A joint analysis of the transcriptome and proteome was performed across four datasets: inner-ear mouse tissues, mouse organ tissues, lymphoblastoid primate samples and human cancer cell lines. We show that the protein levels are more conserved than the mRNA levels in all datasets, and that changes in transcription are associated with translational changes that exert opposite effects on the final protein level, in all tissues except cancer. Finally, we observe that some functions are enriched in the inner ear on the mRNA level but not in protein. We suggest that partial buffering between transcription and translation ensures that proteins can be made rapidly in response to a stimulus. Accounting for the buffering can improve the prediction of protein levels from mRNA levels.

  1. Enhanced levels of scrapie responsive gene mRNA in BSE-infected mouse brain.

    PubMed

    Dandoy-Dron, F; Benboudjema, L; Guillo, F; Jaegly, A; Jasmin, C; Dormont, D; Tovey, M G; Dron, M

    2000-03-10

    The expression of the mRNA of nine scrapie responsive genes was analyzed in the brains of FVB/N mice infected with bovine spongiform encephalopathy (BSE). The RNA transcripts of eight genes were overexpressed to a comparable extent in both BSE-infected and scrapie-infected mice, indicating a common series of pathogenic events in the two transmissible spongiform encephalopathies (TSEs). In contrast, the serine proteinase inhibitor spi 2, an analogue of the human alpha-1 antichymotrypsin gene, was overexpressed to a greater extent in the brains of scrapie-infected animals than in animals infected with BSE, reflecting either an agent specific or a mouse strain specific response. The levels of spi 2 mRNA were increased during the course of scrapie prior to the onset of clinical signs of the disease and the increase reached 11 to 45 fold relative to uninfected controls in terminally ill mice. Spi 2, in common with four of the other scrapie responsive genes studied, is known to be associated with pro-inflammatory processes. These observations underline the importance of cell reactivity in TSE. In addition, scrg2 mRNA the level of which is enhanced in TSE-infected mouse brain, was identified as a previously unrecognized long transcript of the murine aldolase C gene. However, the level of the principal aldolase C mRNA is unaffected in TSE. The increased representation of the longer transcript in the late stage of the disease may reflect changes in mRNA processing and/or stability in reactive astrocytes or in damaged Purkinje cells.

  2. Getting SaaS-y. Why the sisters of Mercy Health System opted for on-demand portfolio management.

    PubMed

    Carter, Jay

    2011-03-01

    Sisters of Mercy Health System chose the SaaS model as a simpler way to plan, execute, and monitor strategic business initiatives. It also provided something that was easy to use and offered quick time to value.

  3. Changes in the mRNA levels of delayed rectifier potassium channels in human atrial fibrillation.

    PubMed

    Lai, L P; Su, M J; Lin, J L; Lin, F Y; Tsai, C H; Chen, Y S; Tseng, Y Z; Lien, W P; Huang, S K

    1999-01-01

    We measured mRNA levels of delayed rectifier potassium channels in human atrial tissue to investigate the mechanism of the shortening of the atrial effective refractory period and the loss of rate-adaptive shortening of the atrial effective refractory period in human atrial fibrillation. A total of 34 patients undergoing open heart surgery were included. Atrial tissue was obtained from the right atrial free wall, right atrial appendage, left atrial free wall and left atrial appendage, respectively. The mRNA amounts of KVLQT1 (IKs), minK (beta-subunit of IKs), HERG (IKr), and KV1.5 (IKur) were measured by reverse transcription-polymerase chain reaction and normalized to the mRNA amount of GAPDH. We found that the mRNA levels of KV1.5, HERG and KVLQT1 were all significantly decreased in patients with persistent atrial fibrillation for more than 3 months. In contrast, the mRNA level of minK was significantly increased in patients with persistent atrial fibrillation for more than 3 months. We further showed that these changes were independent of the underlying cardiac disease, atrial filling pressure, gender and age. We also found that there was no spatial dispersion of mRNA levels among the four atrial sampling sites. Because the decrease in potassium currents results in a prolonged action potential, the shortening of the atrial effective refractory period in atrial fibrillation should be attributed to other factors. However, the decrease in IKs might contribute, at least in part, to the loss of rate-adaptive shortening of the atrial refractory period.

  4. Abnormal mRNA Expression Levels of Telomere-Binding Proteins Represent Biomarkers in Myelodysplastic Syndromes: A Case-Control Study.

    PubMed

    Liu, Baoshan; Yan, Rongdi; Zhang, Jie; Wang, Bin; Sun, Hu; Cui, Xing

    2017-08-02

    As evidence was shown that abnormal shortening of telomeres begins to accumulate in myelodysplastic syndrome (MDS) patients, this study was conducted to determine the relationship between the mRNA expression levels of telomere-binding proteins (TRF1/TRF2/TIN2/TPP1/POT1/RAP1) and the risk level in MDS. There were 40 patients with MDS and 40 normal controls in this study. Methods including telomere content assays and quantitative reverse transcription-polymerase chain reaction were used to examine the mRNA levels of TRF1/TRF2/TIN2/TPP1/POT1/RAP1 in patients with MDS. Compared to the normal group used as a control, the mRNA expression levels of RAP1/POT1/TPP1 of the patients with MDS were decreased, whereas their levels of TRF1/TRF2 and TIN2 were increased. A positive correlation was found between the TRF1, TRF2, and TIN2 mRNA expression levels and the risk level of the International Prognostic Scoring System (IPSS) and the World Health Organization Prognostic Scoring System (WPSS) criteria; however, a negative correlation was found between RAP1/POT1/TPP1 mRNA expression levels and the risk levels of IPSS and WPSS criteria. Because the reduction of TRF1/TRF2/TIN2 mRNA expression and the increase of RAP1/POT1/TPP1 mRNA expression are closely related to the risk levels of the IPSS and WPSS criteria in MDS, it is thought that these telomere-binding proteins could lead to abnormal telomere length and function, which cause chromosomal abnormalities in MDS. With this evidence, we suggest that those proteins' mRNA expressions could be used as biomarkers for the assessment of the risk degree of MDS patients.

  5. Identification of Relationships Between Interleukin 15 mRNA and Brain-Derived Neurotrophic Factor II mRNA Levels With Formal Components of Temperament in Asthmatic Patients.

    PubMed

    Panek, Michał; Jonakowski, Mateusz; Zioło, Jan; Pietras, Tadeusz; Wieteska, Łukasz; Małachowska, Beata; Mokros, Łukasz; Szemraj, Janusz; Kuna, Piotr

    2017-04-01

    Asthma is a chronic inflammatory and heterogeneous disease developing mostly through allergic inflammation, which modifies the expression of various cytokines and neurotrophins. Previous studies suggest the involvement of interleukin (IL)-15 in the regulation of immune response in asthma. Brain-derived neurotrophic factor (BDNF) II plays an important role as a regulator of development and survival of neurons as well as maintenance of their physiological activity. Chronic stress associated with asthma and elevated IL-15 mRNA and BDNFII mRNA levels may affect the mood and a subjective sensation of dyspnoea-inducing anxiety. Psychopathological variables and numerous cytokine/neurotrophin interactions influence the formation of temperament and strategies of coping with stress. The aim of the study was to identify the role of IL-15 mRNA and BDNFII mRNA expressions and their effect on components of temperament and strategies of coping with stress in asthmatics. A total of 352 subjects (176 healthy volunteers and 176 asthmatic patients) participated in the study. The Formal Characteristic of Behaviour-Temperament Inventory (FCB-TI), Coping Inventory for Stressful Situations (CISS), Beck Depression Inventory, State-Trait Anxiety Inventory, and Borg Rating of Perceived Exertion (RPE) Scale were applied in all the subjects. The expression of IL-15 and BDNFII gene was measured using quantitative real-time polymerase chain reaction (qRT-PCR). Different levels of IL-15 and BDNFII expressions between healthy volunteers and patients were revealed in the study. IL-15 enhanced the BDNFII mRNA expression among patients with bronchial asthma. The depression level negatively correlated with the BDNFII mRNA expression. This neurotrophin modified the temperament variable. BDNFII significantly affected (proportional relationship) the level of briskness in asthmatic patients. BDNFII might influence the level and style of coping with stress (emotion-oriented style). This hypothesis

  6. Expression of the protein serum amyloid A in response to Aspergillus fumigatus in murine models of allergic airway inflammation.

    PubMed

    Moran, Gabriel; Carcamo, Carolina; Concha, Margarita; Folch, Hugo

    2015-01-01

    Serum amyloid A (SAA) is an acute phase protein that is elevated in blood during inflammation. The role of this protein in allergic diseases of airways remains unclear. The objective of this study was to evaluate the SAA in blood, lung and bronchial cells in a murine model of bronchial hypersensitivity to Aspergillus fumigatus. To achieve this purpose, different groups of 5-month-old mice were housed in cages containing hay bedding that was contaminated with A. fumigatus and were kept in an isolation room for 16 days to allow for the induction of allergic airway inflammation. Subsequently, the mice were then exposed once again to Aspergillus spores at 0, 2, 8, 24 and 72 h, and they were bled to acquire serum and sacrificed to obtain bronchoalveolar lavage fluid (BALF) or lung tissues for analysis. SAA levels were measured in lung, serum and BALF by dot blot assay and RT-PCR (reverse transcription polymerase chain reaction). The results indicated that SAA protein levels increased in both serum and lung within 2-24h after mice were exposed to Aspergillus spores. Moreover, the SAA mRNA expression levels in the lungs and BALF cells demonstrated the same trend that was observed for the protein levels through the dot blot assay; in particular, SAA mRNA levels increased within the first hour after mice were exposed to A. fumigatus. In this allergic airway model, we conclude that A. fumigatus can induce an acute inflammatory response in the airways through the stimulation of the SAA protein, increasing its levels in serum, lung tissue and BALF samples during the early hours of exposure of mice that have been sensitised for this fungus. Copyright © 2012 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

  7. Reversible Ca2+-induced fast-to-slow transition in primary skeletal muscle culture cells at the mRNA level

    PubMed Central

    Meißner, Joachim D; Kubis, Hans-Peter; Scheibe, Renate J; Gros, Gerolf

    2000-01-01

    The adult fast character and a Ca2+-inducible reversible transition from a fast to a slow type of rabbit myotube in a primary culture were demonstrated at the mRNA level by Northern blot analysis with probes specific for different myosin heavy chain (MyHC) isoforms and enzymes of energy metabolism. No non-adult MyHC isoform mRNA was detected after 22 days of culture. After 4 weeks of culture the fast MyHCIId mRNA was strongly expressed while MyHCI mRNA was virtually absent, indicating the fast adult character of the myotubes in the primary skeletal muscle culture. The data show that a fast-to-slow transition occurred in the myotubes at the level of MyHC isoform gene expression after treatment with the Ca2+ ionophore A23187. The effects of ionophore treatment were decreased levels of fast MyHCII mRNA and an augmented expression of the slow MyHCI gene. Changes in gene expression started very rapidly 1 day after the onset of ionophore treatment. Levels of citrate synthase mRNA increased and levels of glyceraldehyde 3-phosphate dehydrogenase mRNA decreased during ionophore treatment. This points to a shift from anaerobic to oxidative energy metabolism in the primary skeletal muscle culture cells at the level of gene expression. Withdrawal of the Ca2+ ionophore led to a return to increased levels of MyHCII mRNA and decreased levels of MyHCI mRNA, indicating a slow-to-fast transition in the myotubes and the reversibility of the effect of ionophore on MyHC isoform gene expression. PMID:10673542

  8. Regulation of Bovine Leukemia Virus tax and pol mRNA Levels by Interleukin-2 and -10

    PubMed Central

    Pyeon, Dohun; Splitter, Gary A.

    1999-01-01

    Recently, particular cytokines have been identified to affect progression of a variety of diseases and retrovirus infections. Previously, we demonstrated that interleukin-2 (IL-2), IL-12, and gamma interferon increased in peripheral blood mononuclear cells (PBMCs) from animals with early disease and decreased in PBMCs from animals with late disease stages of bovine leukemia virus (BLV) infection. In contrast, IL-10 increased with disease progression. To examine the effects of these cytokines on BLV expression, BLV tax and pol mRNA and p24 protein were quantified by competitive PCR and immunoblotting, respectively. IL-10 inhibited BLV tax and pol mRNA levels in BLV-infected PBMCs; however, the inhibitory effect of IL-10 was prevented in PBMCs depleted of monocytes and/or macrophages (monocyte/macrophages). To determine whether these factors were secreted or monocyte/macrophage associated, monocyte/macrophage-depleted PBMCs were cultured with isolated monocyte/macrophages in transwells where contact between monocyte/macrophages and nonadherent PBMCs was blocked. BLV tax and pol mRNA levels increased in transwell cultures similar to cultures containing nonseparated cells, and IL-10 addition inhibited the increase of BLV tax and pol mRNA. These results suggest that monocyte/macrophages secrete soluble factor(s) that increases BLV mRNA levels and that secretion of these soluble factor(s) could be inhibited by IL-10. In contrast, IL-2 increased BLV tax and pol mRNA and p24 protein production. Thus, IL-10 production by BLV-infected animals with late stage disease may serve to control BLV mRNA levels, while IL-2 may increase BLV mRNA in the early disease stage. To determine a correlation between cell proliferation and BLV expression, the effect of IL-2 and IL-10 on PBMC proliferation was tested. As anticipated, IL-2 stimulated while IL-10 suppressed antigen-specific PBMC proliferation. The present study, combined with our previous findings, suggests that increased IL-10

  9. Photobiomodulation effects on mRNA levels from genomic and chromosome stabilization genes in injured muscle.

    PubMed

    da Silva Neto Trajano, Larissa Alexsandra; Trajano, Eduardo Tavares Lima; da Silva Sergio, Luiz Philippe; Teixeira, Adilson Fonseca; Mencalha, Andre Luiz; Stumbo, Ana Carolina; de Souza da Fonseca, Adenilson

    2018-04-26

    Muscle injuries are the most prevalent type of injury in sports. A great number of athletes have relapsed in muscle injuries not being treated properly. Photobiomodulation therapy is an inexpensive and safe technique with many benefits in muscle injury treatment. However, little has been explored about the infrared laser effects on DNA and telomeres in muscle injuries. Thus, the aim of this study was to evaluate photobiomodulation effects on mRNA relative levels from genes related to telomere and genomic stabilization in injured muscle. Wistar male rats were randomly divided into six groups: control, laser 25 mW, laser 75 mW, injury, injury laser 25 mW, and injury laser 75 mW. Photobiomodulation was performed with 904 nm, 3 J/cm 2 at 25 or 75 mW. Cryoinjury was induced by two applications of a metal probe cooled in liquid nitrogen directly on the tibialis anterior muscle. After euthanasia, skeletal muscle samples were withdrawn and total RNA extracted for evaluation of mRNA levels from genomic (ATM and p53) and chromosome stabilization (TRF1 and TRF2) genes by real-time quantitative polymerization chain reaction. Data show that photobiomodulation reduces the mRNA levels from ATM and p53, as well reduces mRNA levels from TRF1 and TRF2 at 25 and 75 mW in injured skeletal muscle. In conclusion, photobiomodulation alters mRNA relative levels from genes related to genomic and telomere stabilization in injured skeletal muscle.

  10. Mucin gene mRNA levels in broilers challenged with eimeria and/or Clostridium perfringens.

    PubMed

    Kitessa, Soressa M; Nattrass, Gregory S; Forder, Rebecca E A; McGrice, Hayley A; Wu, Shu-Biao; Hughes, Robert J

    2014-09-01

    The effects of Eimeria (EM) and Clostridium perfringens (CP) challenges on the mRNA levels of genes involved in mucin (Muc) synthesis (Muc2, Muc5ac, Muc13, and trefoil family factor-2 [TFF2]), inflammation (tumor necrosis factor alpha [TNF-alpha] and interleukin-18 [IL-18]), and metabolic processes (cluster of differentiation [CD]36) in the jejunum of broilers were investigated. Two parallel experiments involving 1) EM challenge and 2) EM and CP challenges were conducted. The first experiment was a 2 X 2 study with 12 birds per treatment (N = 48) involving fishmeal substitution (25%) in the diet (FM) and EM challenge. The treatments were: Control (FM-, EM-), Fishmeal (FM+, EM-), EM challenge (FM-, EM+), and fishmeal substitution and EM challenge (FM+, EM+). The second experiment was a 2 X 2 X 2 experiment with six birds per treatment (N = 48) involving fishmeal (FM-, FM+), Eimeria (EM-, EM+), and C perfringens (CP-, CP+). In both arms of the study, male broilers were given a starter diet for the whole period of 16 days, except those assigned to FM+, where 25% of the starter ration was replaced with fishmeal from days 8 to 14. EM inoculation was performed on day 9 and CP inoculation on days 14 and 15. The EM challenge birds were euthanatized for sampling on day 13; postmortem examination and sampling for the Eimeria plus C perfringens challenge arm of the study were on day 16. In the Eimeria challenge arm of the study, fishmeal supplementation significantly suppressed the mRNA levels of TNF-alpha, TFF2, and IL-18 pre-CP inoculation but simultaneously increased the levels of Muc13 and CD36 mRNAs. Birds challenged with Eimeria exhibited increased mRNA levels of Muc13, Muc5ac, TNF-alpha, and IL-18. In the Eimeria and C. perfringens challenge arm, birds exposed to EM challenge exhibited significantly lower mRNA levels of Muc2 and CD36. The mRNA levels of CD36 were also significantly suppressed by CP challenge. Our results showed that the transcription of mucin synthesis

  11. GnRH mRNA levels in male three-spined sticklebacks, Gasterosteus aculeatus, under different reproductive conditions.

    PubMed

    Shao, Yi Ta; Tseng, Yung Che; Chang, Chia-Hao; Yan, Hong Young; Hwang, Pung Pung; Borg, Bertil

    2015-02-01

    In vertebrates, reproduction is regulated by the brain-pituitary-gonad (BPG) axis, where the gonadotropin-releasing hormone (GnRH) is one of the key components. However, very little is known about the possible role of GnRH in the environmental and feedback control of fish reproduction. To investigate this, full-length gnrh2 (chicken GnRH II) and gnrh3 (salmon GnRH) sequences of male three-spined sticklebacks (Gasterosteus aculeatus), which are clustered with the taxa of the same GnRH type as other Euteleostei, were cloned and annotated. gnrh1 is absent in this species. The mRNA levels of gnrh2 and gnrh3 in the sticklebacks' brain were measured under breeding and post-breeding conditions as well as in castrated and sham-operated breeding fish and castrated/sham-operated fish kept under long-day (LD 16:8) and short-day (LD 8:16) conditions. Fully breeding males had considerably higher mRNA levels of gnrh2 and gnrh3 in the thalamus (Th) and in the telencephalon and preoptic area (T+POA), respectively, than post-breeding males. Sham-operated breeding males have higher gnrh3 mRNA levels than the corresponding castrated males. Moreover, higher gnrh2 mRNA levels in the Th and higher gnrh3 mRNA levels in the T+POA and hypothalamus (HypTh) were also found in long-day sham-operated males than in sham-operated fish kept under an inhibitory short day photoperiod. Nevertheless, gnrh2 and gnrh3 mRNA levels were not up-regulated in castrated males kept under long-day photoperiod, which suggests that positive feedbacks on the brain-pituitary-gonad axis are necessary for this response. Copyright © 2014 Elsevier Inc. All rights reserved.

  12. The ectoenzyme-side of matrix metalloproteinases (MMPs) makes inflammation by serum amyloid A (SAA) and chemokines go round.

    PubMed

    De Buck, Mieke; Gouwy, Mieke; Struyf, Sofie; Opdenakker, Ghislain; Van Damme, Jo

    2018-06-02

    During an inflammatory response, a large number of distinct mediators appears in the affected tissues or in the blood circulation. These include acute phase proteins such as serum amyloid A (SAA), cytokines and chemokines and proteolytic enzymes. Although these molecules are generated within a cascade sequence in specific body compartments allowing for independent action, their co-appearance in space and time during acute or chronic inflammation points toward important mutual interactions. Pathogen-associated molecular patterns lead to fast induction of the pro-inflammatory endogenous pyrogens, which are evoking the acute phase response. Interleukin-1, tumor necrosis factor-α and interferons simultaneously trigger different cell types, including leukocytes, endothelial cells and fibroblasts for tissue-specific or systemic production of chemokines and matrix metalloproteinases (MMPs). In addition, SAA induces chemokines and both stimulate secretion of MMPs from multiple cell types. As a consequence, these mediators may cooperate to enhance the inflammatory response. Indeed, SAA synergizes with chemokines to increase chemoattraction of monocytes and granulocytes. On the other hand, MMPs post-translationally modify chemokines and SAA to reduce their activity. Indeed, MMPs internally cleave SAA with loss of its cytokine-inducing and direct chemotactic potential whilst retaining its capacity to synergize with chemokines in leukocyte migration. Finally, MMPs truncate chemokines at their NH 2 - or COOH-terminal end, resulting in reduced or enhanced chemotactic activity. Therefore, the complex interactions between chemokines, SAA and MMPs either maintain or dampen the inflammatory response. Copyright © 2018. Published by Elsevier B.V.

  13. Codon-Resolution Analysis Reveals a Direct and Context-Dependent Impact of Individual Synonymous Mutations on mRNA Level

    PubMed Central

    Chen, Siyu; Li, Ke; Cao, Wenqing; Wang, Jia; Zhao, Tong; Huan, Qing; Yang, Yu-Fei; Wu, Shaohuan; Qian, Wenfeng

    2017-01-01

    Abstract Codon usage bias (CUB) refers to the observation that synonymous codons are not used equally frequently in a genome. CUB is stronger in more highly expressed genes, a phenomenon commonly explained by stronger natural selection on translational accuracy and/or efficiency among these genes. Nevertheless, this phenomenon could also occur if CUB regulates gene expression at the mRNA level, a hypothesis that has not been tested until recently. Here, we attempt to quantify the impact of synonymous mutations on mRNA level in yeast using 3,556 synonymous variants of a heterologous gene encoding green fluorescent protein (GFP) and 523 synonymous variants of an endogenous gene TDH3. We found that mRNA level was positively correlated with CUB among these synonymous variants, demonstrating a direct role of CUB in regulating transcript concentration, likely via regulating mRNA degradation rate, as our additional experiments suggested. More importantly, we quantified the effects of individual synonymous mutations on mRNA level and found them dependent on 1) CUB and 2) mRNA secondary structure, both in proximal sequence contexts. Our study reveals the pleiotropic effects of synonymous codon usage and provides an additional explanation for the well-known correlation between CUB and gene expression level. PMID:28961875

  14. Influence of chronic undernutrition and leptin on GOAT mRNA levels in rat stomach mucosa.

    PubMed

    González, C Ruth; Vázquez, María J; López, Miguel; Diéguez, Carlos

    2008-12-01

    The most unique feature of ghrelin is the acyl-modification of a hydroxyl group of the Ser3 in the N-terminus. The Ser3 is commonly modified by n-octanoic acid in vertebrates being needed for its biological effects, at least in terms of feeding. Therefore, a critical question regarding the role of ghrelin was to characterize the mechanism involved in its acylation. The acyltransferase that catalyzes ghrelin octanoylation has been recently identified and named ghrelin O-acyltransferase (GOAT). The aim of this study was to clarify the physiological implications of GOAT in the regulation of energy balance, by assessing the effect of undernutrition, as well as fasting in adult male rats. We have determined GOAT mRNA expression levels by real time-PCR in the stomach mucosa. Our results show that chronic food restriction led to an increase in GOAT mRNA, particularly following long-term chronic malnutrition (21 days). Furthermore, following 48 h complete fasting, a situation with high-circulating ghrelin levels, we found similar mRNA expression of GOAT in fed and fasted rats; exogenous leptin administration markedly increase GOAT mRNA levels in the stomach mucosa of fasted rats. These findings suggest that increased GOAT mRNA levels may have a role in mediating the physiological responses to chronic undernutrition and could represent an adaptive response to prevent long-lasting alterations in energy balance and body weight homeostasis. Furthermore, our data also offer mechanistic insights into the reason why during fasting acylated ghrelin levels are not increased at a time when a marked increase in an orexigenic signal as important as acylated ghrelin will be expected.

  15. Age Differences of Salivary Alpha-Amylase Levels of Basal and Acute Responses to Citric Acid Stimulation Between Chinese Children and Adults.

    PubMed

    Yang, Ze-Min; Chen, Long-Hui; Zhang, Min; Lin, Jing; Zhang, Jie; Chen, Wei-Wen; Yang, Xiao-Rong

    2015-01-01

    It remains unclear how salivary alpha-amylase (sAA) levels respond to mechanical stimuli in different age groups. In addition, the role played by the sAA gene (AMY1) copy number and protein expression (glycosylated and non-glycosylated) in sAA activity has also been rarely reported. In this study, we analyzed saliva samples collected before and after citric acid stimulation from 47 child and 47 adult Chinese subjects. We observed that adults had higher sAA activity and sAA glycosylated levels (glycosylated sAA amount/total sAA amount) in basal and stimulated saliva when compared with children, while no differences were found in total or glycosylated sAA amount between them. Interestingly, adults showed attenuated sAA activity levels increase over those of children after stimulation. Correlation analysis showed that total sAA amount, glycosylated sAA amount, and AMY1 copy number × total sAA amount were all positively correlated with sAA activity before and after stimulation in both groups. Interestingly, correlation r between sAA levels (glycosylated sAA amount and total sAA amount) and sAA activity decreased after stimulation in children, while adults showed an increase in correlation r. In addition, the correlation r between AMY1 copy number × total sAA amount and sAA activity was higher than that between AMY1 copy number, total sAA amount, and sAA activity, respectively. Taken together, our results suggest that total sAA amount, glycosylated sAA amount, and the positive interaction between AMY1 copy number and total sAA amount are crucial in influencing sAA activity before and after stimulation in children and adults.

  16. Age Differences of Salivary Alpha-Amylase Levels of Basal and Acute Responses to Citric Acid Stimulation Between Chinese Children and Adults

    PubMed Central

    Yang, Ze-Min; Chen, Long-Hui; Zhang, Min; Lin, Jing; Zhang, Jie; Chen, Wei-Wen; Yang, Xiao-Rong

    2015-01-01

    It remains unclear how salivary alpha-amylase (sAA) levels respond to mechanical stimuli in different age groups. In addition, the role played by the sAA gene (AMY1) copy number and protein expression (glycosylated and non-glycosylated) in sAA activity has also been rarely reported. In this study, we analyzed saliva samples collected before and after citric acid stimulation from 47 child and 47 adult Chinese subjects. We observed that adults had higher sAA activity and sAA glycosylated levels (glycosylated sAA amount/total sAA amount) in basal and stimulated saliva when compared with children, while no differences were found in total or glycosylated sAA amount between them. Interestingly, adults showed attenuated sAA activity levels increase over those of children after stimulation. Correlation analysis showed that total sAA amount, glycosylated sAA amount, and AMY1 copy number × total sAA amount were all positively correlated with sAA activity before and after stimulation in both groups. Interestingly, correlation r between sAA levels (glycosylated sAA amount and total sAA amount) and sAA activity decreased after stimulation in children, while adults showed an increase in correlation r. In addition, the correlation r between AMY1 copy number × total sAA amount and sAA activity was higher than that between AMY1 copy number, total sAA amount, and sAA activity, respectively. Taken together, our results suggest that total sAA amount, glycosylated sAA amount, and the positive interaction between AMY1 copy number and total sAA amount are crucial in influencing sAA activity before and after stimulation in children and adults. PMID:26635626

  17. Antioxidants inhibit SAA formation and pro-inflammatory cytokine release in a human cell model of alkaptonuria.

    PubMed

    Spreafico, Adriano; Millucci, Lia; Ghezzi, Lorenzo; Geminiani, Michela; Braconi, Daniela; Amato, Loredana; Chellini, Federico; Frediani, Bruno; Moretti, Elena; Collodel, Giulia; Bernardini, Giulia; Santucci, Annalisa

    2013-09-01

    Alkaptonuria (AKU) is an ultra-rare autosomal recessive disease that currently lacks an appropriate therapy. Recently we provided experimental evidence that AKU is a secondary serum amyloid A (SAA)-based amyloidosis. The aim of the present work was to evaluate the use of antioxidants to inhibit SAA amyloid and pro-inflammatory cytokine release in AKU. We adopted a human chondrocytic cell AKU model to evaluate the anti-amyloid capacity of a set of antioxidants that had previously been shown to counteract ochronosis in a serum AKU model. Amyloid presence was evaluated by Congo red staining. Homogentisic acid-induced SAA production and pro-inflammatory cytokine release (overexpressed in AKU patients) were evaluated by ELISA and multiplex systems, respectively. Lipid peroxidation was evaluated by means of a fluorescence-based assay. Our AKU model allowed us to prove the efficacy of ascorbic acid combined with N-acetylcysteine, taurine, phytic acid and lipoic acid in significantly inhibiting SAA production, pro-inflammatory cytokine release and membrane lipid peroxidation. All the tested antioxidant compounds were able to reduce the production of amyloid and may be the basis for establishing new therapies for AKU amyloidosis.

  18. Antioxidants inhibit SAA formation and pro-inflammatory cytokine release in a human cell model of alkaptonuria

    PubMed Central

    Spreafico, Adriano; Millucci, Lia; Ghezzi, Lorenzo; Geminiani, Michela; Braconi, Daniela; Amato, Loredana; Chellini, Federico; Frediani, Bruno; Moretti, Elena; Collodel, Giulia; Bernardini, Giulia

    2013-01-01

    Objective. Alkaptonuria (AKU) is an ultra-rare autosomal recessive disease that currently lacks an appropriate therapy. Recently we provided experimental evidence that AKU is a secondary serum amyloid A (SAA)-based amyloidosis. The aim of the present work was to evaluate the use of antioxidants to inhibit SAA amyloid and pro-inflammatory cytokine release in AKU. Methods. We adopted a human chondrocytic cell AKU model to evaluate the anti-amyloid capacity of a set of antioxidants that had previously been shown to counteract ochronosis in a serum AKU model. Amyloid presence was evaluated by Congo red staining. Homogentisic acid-induced SAA production and pro-inflammatory cytokine release (overexpressed in AKU patients) were evaluated by ELISA and multiplex systems, respectively. Lipid peroxidation was evaluated by means of a fluorescence-based assay. Results. Our AKU model allowed us to prove the efficacy of ascorbic acid combined with N-acetylcysteine, taurine, phytic acid and lipoic acid in significantly inhibiting SAA production, pro-inflammatory cytokine release and membrane lipid peroxidation. Conclusion. All the tested antioxidant compounds were able to reduce the production of amyloid and may be the basis for establishing new therapies for AKU amyloidosis. PMID:23704321

  19. Induction of human spermine oxidase SMO(PAOh1) is regulated at the levels of new mRNA synthesis, mRNA stabilization and newly synthesized protein

    PubMed Central

    2004-01-01

    The oxidation of polyamines induced by antitumour polyamine analogues has been associated with tumour response to specific agents. The human spermine oxidase, SMO(PAOh1), is one enzyme that may play a direct role in the cellular response to the antitumour polyamine analogues. In the present study, the induction of SMO(PAOh1) enzyme activity by CPENSpm [N1-ethyl-N11-(cyclopropyl)methyl-4,8,diazaundecane] is demonstrated to be a result of newly synthesized mRNA and protein. Inhibition of new RNA synthesis by actinomycin D inhibits both the appearance of SMO(PAOh1) mRNA and enzyme activity. Similarly, inhibition of newly synthesized protein with cycloheximide prevents analogue-induced enzyme activity. Half-life determinations indicate that stabilization of SMO(PAOh1) protein does not play a significant role in analogue-induced activity. However, half-life experiments using actinomycin D indicate that CPENSpm treatment not only increases mRNA expression, but also leads to a significant increase in mRNA half-life (17.1 and 8.8 h for CPENSpm-treated cells and control respectively). Using reporter constructs encompassing the SMO(PAOh1) promoter region, a 30–90% increase in transcription is observed after exposure to CPENSpm. The present results are consistent with the hypothesis that analogue-induced expression of SMO(PAOh1) is a result of increased transcription and stabilization of SMO(PAOh1) mRNA, leading to increased protein production and enzyme activity. These data indicate that the major level of control of SMO(PAOh1) expression in response to polyamine analogues exposure is at the level of mRNA. PMID:15496143

  20. Codon influence on protein expression in E. coli correlates with mRNA levels

    PubMed Central

    Boël, Grégory; Wong, Kam-Ho; Su, Min; Luff, Jon; Valecha, Mayank; Everett, John K.; Acton, Thomas B.; Xiao, Rong; Montelione, Gaetano T.; Aalberts, Daniel P.; Hunt, John F.

    2016-01-01

    Degeneracy in the genetic code, which enables a single protein to be encoded by a multitude of synonymous gene sequences, has an important role in regulating protein expression, but substantial uncertainty exists concerning the details of this phenomenon. Here we analyze the sequence features influencing protein expression levels in 6,348 experiments using bacteriophage T7 polymerase to synthesize messenger RNA in Escherichia coli. Logistic regression yields a new codon-influence metric that correlates only weakly with genomic codon-usage frequency, but strongly with global physiological protein concentrations and also mRNA concentrations and lifetimes in vivo. Overall, the codon content influences protein expression more strongly than mRNA-folding parameters, although the latter dominate in the initial ~16 codons. Genes redesigned based on our analyses are transcribed with unaltered efficiency but translated with higher efficiency in vitro. The less efficiently translated native sequences show greatly reduced mRNA levels in vivo. Our results suggest that codon content modulates a kinetic competition between protein elongation and mRNA degradation that is a central feature of the physiology and also possibly the regulation of translation in E. coli. PMID:26760206

  1. Salivary Alpha-Amylase Activity Levels in Catatonic Schizophrenia Decrease after Electroconvulsive Therapy.

    PubMed

    Kanayama, Misako; Miyaoka, Tsuyoshi; Araki, Tomoko; Hayashida, Maiko; Hashioka, Sadayuki; Horiguchi, Jun

    2018-01-01

    Dysfunction of the autonomic nervous system (ANS) in schizophrenia has been detected by electrophysiological methods, but the underlying mechanisms remain unknown. Several studies have suggested that measuring salivary alpha-amylase activity levels is useful for evaluating the ANS activity and that sAA levels increase in schizophrenia and correlate with Brief Psychiatric Rating Scale (BPRS) scores. However, no study has examined the relationship between sAA activity levels and symptoms of schizophrenia with catatonic state. We present the case of a 59-year-old female with persistent catatonic schizophrenia treated by electroconvulsive therapy. We evaluated the ANS activity by measuring sAA activity levels before and after ECT, and we evaluated her symptoms using the BPRS and Bush-Francis Catatonia Rating Scale (BFCRS). ECT was highly effective and BPRS and BFCRS scores substantially decreased. sAA activity levels decreased from 125 kU/l to 33 kU/l. sAA activity levels could be a potential biomarker of schizophrenia with catatonic state.

  2. Synthesis and conformational analysis of linear homo- and heterooligomers from novel 2-C-branched sugar amino acids (SAAs).

    PubMed

    Tian, Guang-Zong; Hu, Jing; Zhang, Heng-Xi; Rademacher, Christoph; Zou, Xiao-Peng; Zheng, Hong-Ning; Xu, Fei; Wang, Xiao-Li; Linker, Torsten; Yin, Jian

    2018-04-26

    Sugar amino acids (SAAs), as biologically interesting structures bearing both amino and carboxylic acid functional groups represent an important class of multifunctional building blocks. In this study, we develop an easy access to novel SAAs in only three steps starting from nitro compounds in high yields in analytically pure form, easily available by ceric (IV) mediated radical additions. Such novel SAAs have been applied in the assembly of total nine carbopeptoids with the form of linear homo- and heterooligomers for the structural investigations employing circular dichroism (CD) spectroscopy, which suggest that the carbopeptoids emerge a well-extended, left (or right)-handed conformation similar to polyproline II (PPII) helices. NMR studies also clearly demonstrated the presence of ordered secondary structural elements. 2D-ROESY spectra were acquired to identify i+1 NH ↔  i C 1 H, i C 2 H correlations which support the conformational analysis of tetramers by CD spectroscopy. These findings provide interesting information of SAAs and their oligomers as potential scaffolds for discovering new drugs and materials.

  3. Changes in period mRNA levels in the brain and division of labor in honey bee colonies

    PubMed Central

    Toma, Dan P.; Bloch, Guy; Moore, Darrell; Robinson, Gene E.

    2000-01-01

    Previous research showed that age-related division of labor in honey bees is associated with changes in activity rhythms; young adult bees perform hive tasks with no daily rhythms, whereas older bees forage with strong daily rhythms. We report that this division of labor is also associated with differences in both circadian rhythms and mRNA levels of period, a gene well known for its role in circadian rhythms. The level of period mRNA in the brain oscillated in bees of all ages, but was significantly higher at all times in foragers. Elevated period mRNA levels cannot be attributed exclusively to aging, because bees induced to forage precociously because of a change in social environment had levels similar to normal age foragers. These results extend the regulation of a “clock gene” to a social context and suggest that there are connections at the molecular level between division of labor and chronobiology in social insects. PMID:10841583

  4. UAS-NAS Live Virtual Constructive Distributed Environment (LVC): LVC Gateway, Gateway Toolbox, Gateway Data Logger (GDL), SaaProc Software Design Description

    NASA Technical Reports Server (NTRS)

    Jovic, Srboljub

    2015-01-01

    This document provides the software design description for the two core software components, the LVC Gateway, the LVC Gateway Toolbox, and two participants, the LVC Gateway Data Logger and the SAA Processor (SaaProc).

  5. Minimal alteration in the ratio of circulatory fetal DNA to fetal corticotropin-releasing hormone mRNA level in preeclampsia.

    PubMed

    Zhong, Xiao Yan; Holzgreve, Wolfgang; Gebhardt, Stefan; Hillermann, Renate; Tofa, Kashefa Carelse; Gupta, Anurag Kumar; Huppertz, Berthold; Hahn, Sinuhe

    2006-01-01

    We have recently observed that fetal DNA and fetal corticotropin-releasing hormone (CRH) mRNA are associated with in vitro generated syncytiotrophoblast-derived microparticles, and that the ratio of fetal DNA to mRNA (CRH) varied according to whether the particles were derived by predominantly apoptotic, apo-necrotic or necrotic pathways. Hence, we examined whether these ratios varied in maternal plasma samples taken from normotensive and preeclamptic pregnancies in vivo. Maternal plasma samples were collected from 18 cases with preeclampsia and 29 normotensive term controls. Circulatory fetal CRH mRNA and DNA levels were quantified by real-time PCR and RT-PCR. Circulatory fetal mRNA and fetal DNA levels were significantly elevated in the preeclampsia study group when compared to normotensive controls. Alterations in the fetal mRNA to DNA ratio between the study and control groups were minimal, even when stratified into early (<34 weeks of gestation) and late (>34 weeks of gestation) onset preeclampsia. Our data suggest that although circulatory fetal DNA and mRNA levels are significantly elevated in preeclampsia, the ratios in maternal plasma are not dramatically altered. Copyright 2006 S. Karger AG, Basel.

  6. Postnatal handling does not normalize hypothalamic corticotropin-releasing factor mRNA levels in animals prenatally exposed to ethanol.

    PubMed

    Gabriel, Kara I; Glavas, Maria M; Ellis, Linda; Weinberg, Joanne

    2005-06-09

    Postnatal handling has been shown to attenuate some of the deficits in developmental outcome observed following prenatal ethanol exposure (E) although it appears to be ineffective at ameliorating the hypothalamic-pituitary-adrenal (HPA) hyperresponsiveness to stressors that has been observed in adult E animals. However, the effects of postnatal handling on central regulation of HPA activity in E animals, particularly with regard to alterations in steady-state hypothalamic corticotropin-releasing factor (CRF) activity, have not been examined. In the present study, offspring from E, pair-fed (PF), and ad-libitum-fed control (C) groups were exposed to daily handling during the first 2 weeks of life (H) or were left entirely undisturbed until weaning (NH). Basal CRF and arginine vasopressin (AVP) mRNA in the parvocellular portion of the paraventricular nucleus (pPVN) of the hypothalamus were assessed at 90-110 days of age. Prenatal ethanol exposure resulted in elevated basal pPVN CRF mRNA levels compared to those in ad-libitum-fed controls. Handling altered CRF mRNA levels in a sex-specific and prenatal treatment-specific manner. Females showed no significant effects of handling. In contrast, handling decreased CRF mRNA levels in PF and C but not E males compared to their NH counterparts. There were no effects of prenatal ethanol or postnatal handling on AVP mRNA levels. These findings indicate that prenatal ethanol exposure results in elevated basal CRF mRNA levels in adulthood and that handling appears to be ineffective in normalizing those elevations, supporting the suggestion that altered basal HPA regulation in E animals may, at least in part, underlie their HPA hyperresponsiveness to stressors.

  7. Induction of the SHARP-2 mRNA level by insulin is mediated by multiple signaling pathways.

    PubMed

    Kanai, Yukiko; Asano, Kosuke; Komatsu, Yoshiko; Takagi, Katsuhiro; Ono, Moe; Tanaka, Takashi; Tomita, Koji; Haneishi, Ayumi; Tsukada, Akiko; Yamada, Kazuya

    2017-02-01

    The rat enhancer of split- and hairy-related protein-2 (SHARP-2) is an insulin-inducible transcription factor which represses transcription of the rat phosphoenolpyruvate carboxykinase gene. In this study, a regulatory mechanism of the SHARP-2 mRNA level by insulin was analyzed. Insulin rapidly induced the level of SHARP-2 mRNA. This induction was blocked by inhibitors for phosphoinositide 3-kinase (PI 3-K), protein kinase C (PKC), and mammalian target of rapamycin (mTOR), actinomycin D, and cycloheximide. Whereas an adenovirus infection expressing a dominant negative form of atypical PKC lambda (aPKCλ) blocked the insulin-induction of the SHARP-2 mRNA level, insulin rapidly activated the mTOR. Insulin did not enhance transcriptional activity from a 3.7 kb upstream region of the rat SHARP-2 gene. Thus, we conclude that insulin induces the expression of the rat SHARP-2 gene at the transcription level via both a PI 3-K/aPKCλ- and a PI 3-K/mTOR- pathways and that protein synthesis is required for this induction.

  8. A hairpin within YAP mRNA 3′UTR functions in regulation at post-transcription level

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Gao, Yuen; Wang, Yuan; Feng, Jinyan

    2015-04-03

    The central dogma of gene expression is that DNA is transcribed into messenger RNAs, which in turn serve as the template for protein synthesis. Recently, it has been reported that mRNAs display regulatory roles that rely on their ability to compete for microRNA binding, independent of their protein-coding function. However, the regulatory mechanism of mRNAs remains poorly understood. Here, we report that a hairpin within YAP mRNA 3′untranslated region (3′UTR) functions in regulation at post-transcription level through generating endogenous siRNAs (esiRNAs). Bioinformatics analysis for secondary structure showed that YAP mRNA displayed a hairpin structure (termed standard hairpin, S-hairpin) within itsmore » 3′UTR. Surprisingly, we observed that the overexpression of S-hairpin derived from YAP 3′UTR (YAP-sh) increased the luciferase reporter activities of transcriptional factor NF-κB and AP-1 in 293T cells. Moreover, we identified that a fragment from YAP-sh, an esiRNA, was able to target mRNA 3′UTR of NF2 (a member of Hippo-signaling pathway) and YAP mRNA 3′UTR itself in hepatoma cells. Thus, we conclude that the YAP-sh within YAP mRNA 3′UTR may serve as a novel regulatory element, which functions in regulation at post-transcription level. Our finding provides new insights into the mechanism of mRNAs in regulatory function. - Highlights: • An S-hairpin within YAP mRNA 3′UTR possesses regulatory function. • YAP-sh acts as a regulatory element for YAP at post-transcription level. • YAP-sh-3p20, an esiRNA derived from YAP-sh, targets mRNAs of YAP and NF2. • YAP-sh-3p20 depresses the proliferation of HepG2 cells in vitro.« less

  9. EOP and SAA Undergraduates Who Left UC Davis without a Degree.

    ERIC Educational Resources Information Center

    Rasor, Marianne

    Undergraduate students enrolled in the Educational Opportunity Program (EOP) or the Student Affirmative Action (SAA) program at the University of California (UC), Davis, who withdrew before graduation were surveyed in 1981. Attention was directed to the respondents' educational experiences after leaving, their current employment, and their…

  10. The mRNA level of MLH1 in peripheral blood is a biomarker for the diagnosis of hereditary nonpolyposis colorectal cancer.

    PubMed

    Yu, Hong; Li, Hui; Cui, Yongan; Xiao, Wei; Dai, Guihong; Huang, Junxing; Wang, Chaofu

    2016-01-01

    Hereditary nonpolyposis colorectal cancer (HNPCC) is caused by functional defects in mismatch repair (MMR) genes, including mutL homolog 1 (MLH1) and mutS homolog 2 (MSH2). This study aimed to assess whether the mRNA expression of MLH1 in peripheral blood could be used as a biomarkers for the diagnosis of HNPCC. The mRNA level of MLH1 was determined in 19 HNPCC families (46 members) using real-time quantitative polymerase chain reaction (qPCR). The mRNA levels of MLH1 in HNPCC were significantly lower than controls (P < 0.001). Receiver operating characteristic (ROC) curve showed a high diagnostic value of the mRNA level of MLH1 for the diagnosis of HNPCC with the area under curve of 0.858. At an optimal cut-off value (0.511), the mRNA level of MLH1 had a sensitivity of 81.3% and a specificity of 86.7% for distinguishing HNPCC from controls. In conclusion, the mRNA expression of MLH1 in peripheral blood may serve as a biomarker for the diagnosis of HNPCC.

  11. High-level mRNA quantification of proliferation marker pKi-67 is correlated with favorable prognosis in colorectal carcinoma.

    PubMed

    Ihmann, Thomas; Liu, Jian; Schwabe, Wolfgang; Häusler, Peter; Behnke, Detlev; Bruch, Hans-Peter; Broll, Rainer; Windhövel, Ute; Duchrow, Michael

    2004-12-01

    The present study retrospectively examines the expression of pKi-67 mRNA and protein in colorectal carcinoma and their correlation to the outcome of patients. Immunohistochemistry and quantitative RT-PCR were used to analyze the expression of pKi-67 in 43 archival specimens of patients with curatively resected primary colorectal carcinoma, who were not treated with neo-adjuvant therapy. We determined a median pKi-67 (MIB-1) labeling index of 31.3% (range 10.3-66.4%), and a mean mRNA level of 0.1769 (DeltaC(T): range 0.01-0.69); indices and levels did not correlate. High pKi-67 mRNA DeltaC(T) values were associated with a significantly favorable prognosis, while pKi-67 labeling indices were not correlated to prognostic outcome. A multivariate analysis of clinical and biological factors indicated that tumor stage (UICC) and pKi-67 mRNA expression level were independent prognostic factors. Quantitatively determined pKi-67 mRNA can be a good and new prognostic indicator for primary resected colorectal carcinoma.

  12. Developmental changes in the hypothalamic mRNA expression levels of brain-derived neurotrophic factor and serum leptin levels: Their responses to fasting in male and female rats.

    PubMed

    Iwasa, Takeshi; Matsuzaki, Toshiya; Yano, Kiyohito; Munkhzaya, Munkhsaikhan; Tungalagsuvd, Altankhuu; Yiliyasi, Maira; Kuwahara, Akira; Irahara, Minoru

    2016-11-01

    The actions and responses of hypothalamic appetite regulatory factors change markedly during the neonatal to pre-pubertal period in order to maintain appropriate metabolic and nutritional conditions. In this study, we examined the developmental changes in the hypothalamic mRNA levels of brain-derived neurotrophic factor (BDNF), which is a potent anorectic factor and the changes in the sensitivity of the hypothalamic expression of this factor to fasting during the neonatal to pre-pubertal period. Under fed conditions, hypothalamic BDNF mRNA expression decreased during development in both male and female rats. Similarly, the serum levels of leptin, which is a positive regulator of hypothalamic BDNF expression, also tended to fall during the developmental period. The serum leptin level and the hypothalamic BDNF mRNA level were found to be positively correlated in both sexes under the fed conditions. Hypothalamic BDNF mRNA expression was decreased by 24h fasting (separating the rats from their mothers) in the early neonatal period (postnatal day 10) in both males and females, but no such changes were seen at postnatal day 20. Twenty-four hours' fasting (food deprivation) did not affect hypothalamic BDNF mRNA expression in the pre-pubertal period (postnatal day 30). On the other hand, the rats' serum leptin levels were decreased by 24h fasting (separating the rats from their mothers at postnatal day 10 and 20, and food deprivation at postnatal day 30) throughout the early neonatal to pre-pubertal period. The correlation between serum leptin and hypothalamic BDNF mRNA levels was not significant under the fasted conditions. It can be speculated that leptin partially regulates hypothalamic BDNF mRNA levels, but only in fed conditions. Such changes in hypothalamic BDNF expression might play a role in maintaining appropriate metabolic and nutritional conditions and promoting normal physical development. In addition, because maternal separation induces a negative energy

  13. [Effects of berberine on mRNA expression levels of PPARγ and adipocytokines in insulin-resistant adipocytes].

    PubMed

    Tu, Jun; Luo, Xin-Xin; Li, Bing-Tao; Li, Yu; Xu, Guo-Liang

    2016-06-01

    Adipocytokines are closely associated with insulin resistance (IR) in adipose tissues, and they are more and more seriously taken in the study of the development of diabetes. This experiment was mainly to study the effect of berberine on mRNA expression levels of PPARγ and adipocytokines in insulin resistant adipocytes, and investigate the molecular mechanism of berberine in enhancing insulin sensitization and application advantages of droplet digital PCR (ddPCR). ddPCR absolute quantification analysis was taken in this experiment to simply and intuitively determine the appropriate reference genes. ddPCR and quantitative Real-time PCR (qPCR) were used to compare the effect of different doses of berberine (10, 20, 50, 100 μmol•L⁻¹) on mRNA expression levels of PPARγ, adiponectin, resistin and leptin in IR 3T3-L1adipocytes. Antagonist GW9662 was added to study the inherent correlation between PPARγ and adiponectin mRNA expression levels. ddPCR results showed that the expression level of β-actin in adipocytes was stable, and suitable as reference gene for normalization of quantitative PCR data. Both of ddPCR and qPCR results showed that, as compared with IR models, the mRNA expression levels of adiponectin were decreased in the treatment with berberine (10, 20, 50, 100 μmol•L⁻¹) in a dose-dependent manner (P<0.01); the expression of PPARγ was decreased by 20, 50, 100 μmol•L⁻¹ berberine in a dose-dependent manner in qPCR assay (P<0.01) and decreased only by 50 and 100 μmol•L⁻¹ berberine in ddPCR assay (P<0.05). PPARγ specific antagonist GW9662 intervention experiment showed that adiponectin gene expression was directly relevant with PPARγ (P<0.05). ddPCR probe assay showed that various doses of berberine could significantly reduce mRNA expression levels of resistin and leptin (P<0.01) in a dose-dependent manner. In conclusion, berberine enhanced insulin sensitization effect not by up-regulating adiponect in expression of transcriptional

  14. Estradiol In Females May Negate Skeletal Muscle Myostatin Mrna Expression And Serum Myostatin Propeptide Levels After Eccentric Muscle Contractions

    PubMed Central

    Willoughby, Darryn S.; Wilborn, Colin D.

    2006-01-01

    Eccentric contractions produce a significant degree of inflammation and muscle injury that may increase the expression of myostatin. Due to its anti- oxidant and anti-flammatory effects, circulating 17-β estradiol (E2) may attenuate myostatin expression. Eight males and eight females performed 7 sets of 10 reps of eccentric contractions of the knee extensors at 150% 1-RM. Each female performed the eccentric exercise bout on a day that fell within her mid-luteal phase (d 21-23 of her 28-d cycle). Blood and muscle samples were obtained before and 6 and 24 h after exercise, while additional blood samples were obtained at 48 and 72 h after exercise. Serum E2 and myostatin LAP/propeptide (LAP/pro) levels were determined with ELISA, and myostatin mRNA expression determined using RT-PCR. Data were analyzed with two-way ANOVA and bivariate correlations (p < 0.05). Females had greater levels of serum E2 throughout the 72- h sampling period (p < 0.05). While males had greater body mass and fat-free mass, neither was correlated to the pre-exercise levels of myostatin mRNA and LAP/pro for either gender (p > 0.05). Compared to pre-exercise, males had significant increases (p < 0.05) in LAP/propetide and mRNA of 78% and 28%, respectively, at 24 h post-exercise, whereas females underwent respective decreases of 10% and 21%. E2 and LAP/propeptide were correlated at 6 h (r = -0.804, p = 0.016) and 24 h post- exercise (r = -0.841, p = 0.009) in males, whereas in females E2 levels were correlated to myostatin mRNA at 6 h (r =0.739, p = 0.036) and 24 h (r = 0.813, p = 0.014) post-exercise and LAP/propeptide at 6 h (r = 0.713, p = 0.047) and 24 h (r = 0.735, p = 0.038). In females, myostatin mRNA expression and serum LAP/propeptide levels do not appear to be significantly up-regulated following eccentric exercise, and may be due to higher levels of circulating E2. Key Points The pre-exercise levels of myostatin mRNA and propeptide were not significantly different between genders, and

  15. Serum vaspin levels and vaspin mRNA expression in subcutaneous adipose tissue in women with gestational diabetes mellitus.

    PubMed

    Mm, Wei Qian; Fan, Jianxia; Khor, Shuzin; Song, Mengfan; Hong, Wei; Dai, Xiaobei

    2014-11-01

    To compare serum vaspin level and mRNA and protein levels of vaspin in adipose tissue in women with gestational diabetes mellitus (GDM) and normal glucose tolerance (NGR), along with the correlation between serum vaspin level with fasting insulin (FINS), homeostasis model assessment of insulin resistance (HOMA-IR) and birth-weight. Thirty-seven women with GDM and 36 with NGR were enrolled. Total cholesterol (TC), triglyceride (TG), high density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), fasting plasma glucose (FPG), FINS and vaspin levels were measured. The mRNA and protein levels were detected using RT-PCR and Western blot. Pearson correlation analysis (PCA) was performed to reveal the correlation between serum vaspin level and FINS, HOMA-IR. Spearman correlation analysis (SCA) was conducted to examine the association between serum vaspin level and birth-weight. HDL-C level in GDM was lower than NGR group (P<0.05), and there were no statistical differences in TC, TG, LDL-C, FPG, FINS and HOMA-IR between the two groups. Serum vaspin level, mRNA and protein expression levels of vaspin in GDM were higher than NGR group (P<0.05). Serum vaspin level was not significantly correlated with FINS and HOMA-IR, but had a positive correlation with birth-weight (P=0.023). Serum vaspin level cannot serve as an independent predictor of IR. The increased serum vaspin level and increased vaspin mRNA and protein expression in adipose tissues in GDM women indicate that vaspin may be involved in the pathogenesis of GDM, but its exact mechanism needs further study. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  16. Serum amyloid A induction of cytokines in monocytes/macrophages and lymphocytes.

    PubMed

    Song, Changjie; Hsu, Kenneth; Yamen, Eric; Yan, Weixing; Fock, Jianyi; Witting, Paul K; Geczy, Carolyn L; Freedman, S Ben

    2009-12-01

    Serum amyloid A (SAA) is a biomarker of inflammation. Elevated blood levels in cardiovascular disease and local deposition in atheroma implies a role of SAA as a mediator rather than just a marker of inflammation. This study explored SAA-induced cytokine production and secretion by mononuclear cells. RT-PCR showed that SAA time-dependently induced cytokine mRNAs in peripheral blood mononuclear cells (PBMC) and THP-1 monocytoid cells, and dramatically increased IL-1beta, MCP-1, IL-6, IL-8, IL-10, GM-CSF, TNF, and MIP-1alpha secretion by PBMC to levels 28 to 25,000 fold above baseline, as measured with Bio-Plex kits; monocytes were the principle source. SAA induction of cytokines in monocyte-derived macrophages (MDM) was significantly higher than from monocytes from the same donors. SAA time-dependently induced transient and significant upregulation of NF-kappaB1 mRNA; inhibitor studies indicate that activation of NF-kappaB through the ERK1/2, p38 and JNK MAPKs and the PI3K pathway was involved. PBMC from 10 patients with coronary artery disease (CAD) spontaneously secreted higher levels of IL-6 and MIP-1alpha after 24h incubation than PBMC from normal controls, whereas SAA-induced levels of all cytokines were similar to controls. Aortic and coronary sinus sampling in 23 CAD patients indicated significant SAA release into the coronary circulation, not evident in 11 controls. SAA can increase monocyte and macrophage cytokine production, possibly at sites of atherosclerosis, thereby contributing to the pro-inflammatory state in coronary artery disease.

  17. Cytoplasmic delivery of ribozymes leads to efficient reduction in alpha-lactalbumin mRNA levels in C127I mouse cells.

    PubMed Central

    L'Huillier, P J; Davis, S R; Bellamy, A R

    1992-01-01

    Ribozymes targeted to five sites along the alpha-lactalbumin (alpha-lac) mRNA were delivered to the cytoplasm of mouse C127I mammary cells using the T7-vaccinia virus delivery system and the amount of alpha-lac mRNA was monitored 24-48 h post-transfection. Three target sites were selected in the alpha-lac coding region (nucleotides 15, 145 and 361) and two were located in the 3' non-coding region (nucleotides 442 and 694). Acting in trans and at a target:ribozyme ratio of 1:1000, ribozymes targeting sites 361 and 694 reduced alpha-lac mRNA by > 80%; another two ribozymes (targeting nucleotides 442 and 145) reduced mRNA levels by 80 and 60% respectively; the fifth ribozyme (targeting nucleotide 15, near the AUG) was largely ineffective. The kinetic activity (kcat) of each ribozyme in vitro was somewhat predictive of the activity of the two ribozymes that targeted nucleotides 361 and 694, but was not predictive of the in vivo activity of the other three ribozymes. Down-regulation of the intracellular levels of alpha-lac paralleled the ribozyme-dependent reduction achieved for mRNA. For site 442, the reduction in both mRNA and protein was attributed to the catalytic activity of the ribozyme rather than to the antisense effects of the flanking arms, because delivery of an engineered (catalytically-inactive) variant had no effect on mRNA levels and a minimal effect on the level of alpha-lac present in the cell. Images PMID:1425576

  18. Spaceflight has compartment- and gene-specific effects on mRNA levels for bone matrix proteins in rat femur

    NASA Technical Reports Server (NTRS)

    Evans, G. L.; Morey-Holton, E.; Turner, R. T.

    1998-01-01

    In the present study, we evaluated the possibility that the abnormal bone matrix produced during spaceflight may be associated with reduced expression of bone matrix protein genes. To test this possibility, we investigated the effects of a 14-day spaceflight (SLS-2 experiment) on steady-state mRNA levels for glyceraldehyde-3-phosphate dehydrogenase (GAPDH), osteocalcin, osteonectin, and prepro-alpha(1) subunit of type I collagen in the major bone compartments of rat femur. There were pronounced site-specific differences in the steady-state levels of expression of the mRNAs for the three bone matrix proteins and GAPDH in normal weight-bearing rats, and these relationships were altered after spaceflight. Specifically, spaceflight resulted in decreases in mRNA levels for GAPDH (decreased in proximal metaphysis), osteocalcin (decreased in proximal metaphysis), osteonectin (decreased in proximal and distal metaphysis), and collagen (decreased in proximal and distal metaphysis) compared with ground controls. There were no changes in mRNA levels for matrix proteins or GAPDH in the shaft and distal epiphysis. These results demonstrate that spaceflight leads to site- and gene-specific decreases in mRNA levels for bone matrix proteins. These findings are consistent with the hypothesis that spaceflight-induced decreases in bone formation are caused by concomitant decreases in expression of genes for bone matrix proteins.

  19. An eclogite-bearing continental tectonic slice in the Zermatt-Saas high-pressure ophiolites at Trockener Steg (Zermatt, Swiss Western Alps)

    NASA Astrophysics Data System (ADS)

    Weber, Sebastian; Bucher, Kurt

    2015-09-01

    The Theodul Glacier Unit (TGU) at "Trockener Steg" represents a continental slice, embedded within the ophiolitic Zermatt-Saas Zone. The Zermatt-Saas Zone is the remnant of the Piemonte-Liguria oceanic lithosphere, formed in the middle Jurassic and subducted up to eclogite facies conditions in the Early Tertiary. The close spatial association of the TGU to the Zermatt-Saas Zone permits a comparison of the metamorphic evolution of the units by detailed field mapping and a petrological investigation of eclogites. The eclogites from both tectono-metamorphic units can be clearly distinguished by their textures, mineral assemblages and by mineral and bulk-rock composition. Geothermobarometry and computed assemblage stability diagrams for the TGU eclogites indicate P-T conditions of 2.2 ± 0.1 GPa and 580 ± 50 °C. These derived P-T conditions must be considered as minimum peak metamorphic conditions the rocks achieved during subduction. The P-T data are different from those derived for eclogites of Zermatt-Saas Zone adjacent to the Theodul Glacier Unit, that reached maximal burial depths at 2.3-2.4 GPa and 500 ± 50 °C. While the estimates of the eclogites of Zermatt-Saas Zone are in good agreement with some of the previous studies, the contrasting P-T estimates for the TGU eclogites suggest that the Zermatt-Saas complex must be subdivided into several tectonic subunits. The non-uniform peak conditions over the "Trockener Steg" area and the maximum pressures conditions reported from ultra-high pressure localities within Zermatt-Saas Zone suggest, that individual tectonic slices have been assembled after detachment from the slab at the return-point, i.e. along the exhumation path. Detached packages of rocks may range from small tectonic slices up to several kilometer-sized fragments. The TGU is separated from the surrounding rocks of the ophiolite unit by two major tectonic contacts. In addition, the formation of biotite-rich crusts along the basal contact of the TGU

  20. Alternative Polyadenylation and Nonsense-Mediated Decay Coordinately Regulate the Human HFE mRNA Levels

    PubMed Central

    Martins, Rute; Proença, Daniela; Silva, Bruno; Barbosa, Cristina; Silva, Ana Luísa; Faustino, Paula; Romão, Luísa

    2012-01-01

    Nonsense-mediated decay (NMD) is an mRNA surveillance pathway that selectively recognizes and degrades defective mRNAs carrying premature translation-termination codons. However, several studies have shown that NMD also targets physiological transcripts that encode full-length proteins, modulating their expression. Indeed, some features of physiological mRNAs can render them NMD-sensitive. Human HFE is a MHC class I protein mainly expressed in the liver that, when mutated, can cause hereditary hemochromatosis, a common genetic disorder of iron metabolism. The HFE gene structure comprises seven exons; although the sixth exon is 1056 base pairs (bp) long, only the first 41 bp encode for amino acids. Thus, the remaining downstream 1015 bp sequence corresponds to the HFE 3′ untranslated region (UTR), along with exon seven. Therefore, this 3′ UTR encompasses an exon/exon junction, a feature that can make the corresponding physiological transcript NMD-sensitive. Here, we demonstrate that in UPF1-depleted or in cycloheximide-treated HeLa and HepG2 cells the HFE transcripts are clearly upregulated, meaning that the physiological HFE mRNA is in fact an NMD-target. This role of NMD in controlling the HFE expression levels was further confirmed in HeLa cells transiently expressing the HFE human gene. Besides, we show, by 3′-RACE analysis in several human tissues that HFE mRNA expression results from alternative cleavage and polyadenylation at four different sites – two were previously described and two are novel polyadenylation sites: one located at exon six, which confers NMD-resistance to the corresponding transcripts, and another located at exon seven. In addition, we show that the amount of HFE mRNA isoforms resulting from cleavage and polyadenylation at exon seven, although present in both cell lines, is higher in HepG2 cells. These results reveal that NMD and alternative polyadenylation may act coordinately to control HFE mRNA levels, possibly varying its

  1. Effects of seawater acclimation on mRNA levels of corticosteroid receptor genes in osmoregulatory and immune systems in trout

    USGS Publications Warehouse

    Yada, T.; Hyodo, S.; Schreck, C.B.

    2008-01-01

    Influence of environmental salinity on expression of distinct corticosteroid receptor (CR) genes, glucocorticoid receptor (GR)-1 and -2, and mineralcorticoid receptor (MR), was examined in osmoregulatory and hemopoietic organs and leucocytes of steelhead trout (Oncorhynchus mykiss). There was no significant difference in plasma cortisol levels between freshwater (FW)- or seawater (SW)-acclimated trout, whereas Na+, K+-ATPase was activated in gill of SW fish. Plasma lysozyme levels also showed a significant increase after acclimation to SW. In SW-acclimated fish, mRNA levels of GR-1, GR-2, and MR were significantly higher in gill and body kidney than those in FW. Head kidney and spleen showed no significant change in these CR mRNA levels after SW-acclimation. On the other hand, leucocytes isolated from head kidney and peripheral blood showed significant decreases in mRNA levels of CR in SW-acclimated fish. These results showed differential regulation of gene expression of CR between osmoregulatory and immune systems. ?? 2008 Elsevier Inc. All rights reserved.

  2. Nucleolin Mediates MicroRNA-directed CSF-1 mRNA Deadenylation but Increases Translation of CSF-1 mRNA*

    PubMed Central

    Woo, Ho-Hyung; Baker, Terri; Laszlo, Csaba; Chambers, Setsuko K.

    2013-01-01

    CSF-1 mRNA 3′UTR contains multiple unique motifs, including a common microRNA (miRNA) target in close proximity to a noncanonical G-quadruplex and AU-rich elements (AREs). Using a luciferase reporter system fused to CSF-1 mRNA 3′UTR, disruption of the miRNA target region, G-quadruplex, and AREs together dramatically increased reporter RNA levels, suggesting important roles for these cis-acting regulatory elements in the down-regulation of CSF-1 mRNA. We find that nucleolin, which binds both G-quadruplex and AREs, enhances deadenylation of CSF-1 mRNA, promoting CSF-1 mRNA decay, while having the capacity to increase translation of CSF-1 mRNA. Through interaction with the CSF-1 3′UTR miRNA common target, we find that miR-130a and miR-301a inhibit CSF-1 expression by enhancing mRNA decay. Silencing of nucleolin prevents the miRNA-directed mRNA decay, indicating a requirement for nucleolin in miRNA activity on CSF-1 mRNA. Downstream effects followed by miR-130a and miR-301a inhibition of directed cellular motility of ovarian cancer cells were found to be dependent on nucleolin. The paradoxical effects of nucleolin on miRNA-directed CSF-1 mRNA deadenylation and on translational activation were explored further. The nucleolin protein contains four acidic stretches, four RNA recognition motifs (RRMs), and nine RGG repeats. All three domains in nucleolin regulate CSF-1 mRNA and protein levels. RRMs increase CSF-1 mRNA, whereas the acidic and RGG domains decrease CSF-1 protein levels. This suggests that nucleolin has the capacity to differentially regulate both CSF-1 RNA and protein levels. Our finding that nucleolin interacts with Ago2 indirectly via RNA and with poly(A)-binding protein C (PABPC) directly suggests a nucleolin-Ago2-PABPC complex formation on mRNA. This complex is in keeping with our suggestion that nucleolin may work with PABPC as a double-edged sword on both mRNA deadenylation and translational activation. Our findings underscore the complexity of

  3. Sequences within the 5' untranslated region regulate the levels of a kinetoplast DNA topoisomerase mRNA during the cell cycle.

    PubMed Central

    Pasion, S G; Hines, J C; Ou, X; Mahmood, R; Ray, D S

    1996-01-01

    Gene expression in trypanosomatids appears to be regulated largely at the posttranscriptional level and involves maturation of mRNA precursors by trans splicing of a 39-nucleotide miniexon sequence to the 5' end of the mRNA and cleavage and polyadenylation at the 3' end of the mRNA. To initiate the identification of sequences involved in the periodic expression of DNA replication genes in trypanosomatids, we have mapped splice acceptor sites in the 5' flanking region of the TOP2 gene, which encodes the kinetoplast DNA topoisomerase, and have carried out deletion analysis of this region on a plasmid-encoded TOP2 gene. Block deletions within the 5' untranslated region (UTR) identified two regions (-608 to -388 and -387 to -186) responsible for periodic accumulation of the mRNA. Deletion of one or the other of these sequences had no effect on periodic expression of the mRNA, while deletion of both regions resulted in constitutive expression of the mRNA throughout the cell cycle. Subcloning of these sequences into the 5' UTR of a construct lacking both regions of the TOP2 5' UTR has shown that an octamer consensus sequence present in the 5' UTR of the TOP2, RPA1, and DHFR-TS mRNAs is required for normal cycling of the TOP2 mRNA. Mutation of the consensus octamer sequence in the TOP2 5' UTR in a plasmid construct containing only a single consensus octamer and that shows normal cycling of the plasmid-encoded TOP2 mRNA resulted in substantial reduction of the cycling of the mRNA level. These results imply a negative regulation of TOP2 mRNA during the cell cycle by a mechanism involving redundant elements containing one or more copies of a conserved octamer sequence within the 5' UTR of TOP2 mRNA. PMID:8943327

  4. Sequences within the 5' untranslated region regulate the levels of a kinetoplast DNA topoisomerase mRNA during the cell cycle.

    PubMed

    Pasion, S G; Hines, J C; Ou, X; Mahmood, R; Ray, D S

    1996-12-01

    Gene expression in trypanosomatids appears to be regulated largely at the posttranscriptional level and involves maturation of mRNA precursors by trans splicing of a 39-nucleotide miniexon sequence to the 5' end of the mRNA and cleavage and polyadenylation at the 3' end of the mRNA. To initiate the identification of sequences involved in the periodic expression of DNA replication genes in trypanosomatids, we have mapped splice acceptor sites in the 5' flanking region of the TOP2 gene, which encodes the kinetoplast DNA topoisomerase, and have carried out deletion analysis of this region on a plasmid-encoded TOP2 gene. Block deletions within the 5' untranslated region (UTR) identified two regions (-608 to -388 and -387 to -186) responsible for periodic accumulation of the mRNA. Deletion of one or the other of these sequences had no effect on periodic expression of the mRNA, while deletion of both regions resulted in constitutive expression of the mRNA throughout the cell cycle. Subcloning of these sequences into the 5' UTR of a construct lacking both regions of the TOP2 5' UTR has shown that an octamer consensus sequence present in the 5' UTR of the TOP2, RPA1, and DHFR-TS mRNAs is required for normal cycling of the TOP2 mRNA. Mutation of the consensus octamer sequence in the TOP2 5' UTR in a plasmid construct containing only a single consensus octamer and that shows normal cycling of the plasmid-encoded TOP2 mRNA resulted in substantial reduction of the cycling of the mRNA level. These results imply a negative regulation of TOP2 mRNA during the cell cycle by a mechanism involving redundant elements containing one or more copies of a conserved octamer sequence within the 5' UTR of TOP2 mRNA.

  5. Chronic periodontitis can affect the levels of potential oral cancer salivary mRNA biomarkers.

    PubMed

    Cheng, Y-S L; Jordan, L; Chen, H-S; Kang, D; Oxford, L; Plemons, J; Parks, H; Rees, T

    2017-06-01

    More than 100 salivary constituents have been found to show levels significantly different in patients with oral squamous cell carcinoma (OSCC) from those found in healthy controls, and therefore have been suggested to be potential salivary biomarkers for OSCC detection. However, many of these potential OSCC salivary biomarkers are also involved in chronic inflammation, and whether the levels of these biomarkers could be affected by the presence of chronic periodontitis was not known. The objective of this pilot study was therefore to measure the levels of seven previously reported potential OSCC salivary mRNA biomarkers in patients with chronic periodontitis and compare them to levels found in patients with OSCC and healthy controls. The seven salivary mRNAs were interleukin (IL)-8, IL-1β, dual specificity phosphatase 1, H3 histone family 3A, ornithine decarboxylase antizyme 1, S100 calcium-binding protein P (S100P) and spermidine/spermine N1-acetyltransferase 1. Unstimulated whole saliva samples were collected from a total of 105 human subjects from the following four study groups: OSCC; CPNS (chronic periodontitis, moderate to severe degree, non-smokers); CPS (chronic periodontitis, moderate to severe degree, smokers); and healthy controls. Levels of each mRNA in patient groups (OSCC or chronic periodontitis) relative to the healthy controls were determined by a pre-amplification reverse transcription-quantitative polymerase chain reaction approach with nested gene-specific primers. Results were recorded and analyzed by the Bio-Rad CFX96 Real-Time System. Mean fold changes between each pair of patient vs. control groups were analyzed by the Mann-Whitney U-test with Bonferroni corrections. Only S100P showed significantly higher levels in patients with OSCC compared to both patients with CPNS (p = 0.003) and CPS (p = 0.007). The difference in S100P levels between patients with OSCC and healthy controls was also marginally significant (p = 0.009). There was no

  6. Hepatic Expression of Serum Amyloid A1 Is Induced by Traumatic Brain Injury and Modulated by Telmisartan

    PubMed Central

    Villapol, Sonia; Kryndushkin, Dmitry; Balarezo, Maria G.; Campbell, Ashley M.; Saavedra, Juan M.; Shewmaker, Frank P.; Symes, Aviva J.

    2016-01-01

    Traumatic brain injury affects the whole body in addition to the direct impact on the brain. The systemic response to trauma is associated with the hepatic acute-phase response. To further characterize this response, we performed controlled cortical impact injury on male mice and determined the expression of serum amyloid A1 (SAA1), an apolipoprotein, induced at the early stages of the acute-phase response in liver and plasma. After cortical impact injury, induction of SAA1 was detectable in plasma at 6 hours post-injury and in liver at 1 day post-injury, followed by gradual diminution over time. In the liver, cortical impact injury increased neutrophil and macrophage infiltration, apoptosis, and expression of mRNA encoding the chemokines CXCL1 and CXCL10. An increase in angiotensin II AT1 receptor mRNA at 3 days post-injury was also observed. Administration of the AT1 receptor antagonist telmisartan 1 hour post-injury significantly decreased liver SAA1 levels and CXCL10 mRNA expression, but did not affect CXCL1 expression or the number of apoptotic cells or infiltrating leukocytes. To our knowledge, this is the first study to demonstrate that SAA1 is induced in the liver after traumatic brain injury and that telmisartan prevents this response. Elucidating the molecular pathogenesis of the liver after brain injury will assist in understanding the efficacy of therapeutic approaches to brain injury. PMID:26435412

  7. Comparison of mRNA levels of three ethylene receptors in senescing flowers of carnation (Dianthus caryophyllus L.).

    PubMed

    Shibuya, Kenichi; Nagata, Masayasu; Tanikawa, Natsu; Yoshioka, Toshihito; Hashiba, Teruyoshi; Satoh, Shigeru

    2002-03-01

    Three ethylene receptor genes, DC-ERS1, DC-ERS2 and DC-ETR1, were previously identified in carnation (Dianthus caryophyllus L.). Here, the presence of mRNAs for respective genes in flower tissues and their changes during flower senescence are investigated by Northern blot analysis. DC-ERS2 and DC-ETR1 mRNAs were present in considerable amounts in petals, ovaries and styles of the flower at the full-opening stage. In the petals the level of DC-ERS2 mRNA showed a decreasing trend toward the late stage of flower senescence, whereas it increased slightly in ovaries and was unchanged in styles throughout the senescence period. However, DC-ETR1 mRNA showed no or little changes in any of the tissues during senescence. Exogenously applied ethylene did not affect the levels of DC-ERS2 and DC-ETR1 mRNAs in petals. Ethylene production in the flowers was blocked by treatment with 1,1-dimethyl-4-(phenylsulphonyl)semicarbazide (DPSS), but the mRNA levels for DC-ERS2 and DC-ETR1 decreased in the petals. DC-ERS1 mRNA was not detected in any cases. These results indicate that DC-ERS2 and DC-ETR1 are ethylene receptor genes responsible for ethylene perception and that their expression is regulated in a tissue-specific manner and independently of ethylene in carnation flowers during senescence.

  8. Wastewater treatment plant effluent alters pituitary gland gonadotropin mRNA levels in juvenile coho salmon (Oncorhynchus kisutch).

    PubMed

    Harding, Louisa B; Schultz, Irvin R; da Silva, Denis A M; Ylitalo, Gina M; Ragsdale, Dave; Harris, Stephanie I; Bailey, Stephanie; Pepich, Barry V; Swanson, Penny

    2016-09-01

    It is well known that endocrine disrupting compounds (EDCs) present in wastewater treatment plant (WWTP) effluents interfere with reproduction in fish, including altered gonad development and induction of vitellogenin (Vtg), a female-specific egg yolk protein precursor produced in the liver. As a result, studies have focused on the effects of EDC exposure on the gonad and liver. However, impacts of environmental EDC exposure at higher levels of the hypothalamic-pituitary-gonad axis are less well understood. The pituitary gonadotropins, follicle-stimulating hormone (Fsh) and luteinizing hormone (Lh) are involved in all aspects of gonad development and are subject to feedback from gonadal steroids making them a likely target of endocrine disruption. In this study, the effects of WWTP effluent exposure on pituitary gonadotropin mRNA expression were investigated to assess the utility of Lh beta-subunit (lhb) as a biomarker of estrogen exposure in juvenile coho salmon (Oncorhynchus kisutch). First, a controlled 72-h exposure to 17α-ethynylestradiol (EE2) and 17β-trenbolone (TREN) was performed to evaluate the response of juvenile coho salmon to EDC exposure. Second, juvenile coho salmon were exposed to 0, 20 or 100% effluent from eight WWTPs from the Puget Sound, WA region for 72h. Juvenile coho salmon exposed to 2 and 10ng EE2L(-1) had 17-fold and 215-fold higher lhb mRNA levels relative to control fish. Hepatic vtg mRNA levels were dramatically increased 6670-fold, but only in response to 10ng EE2L(-1) and Fsh beta-subunit (fshb) mRNA levels were not altered by any of the treatments. In the WWTP effluent exposures, lhb mRNA levels were significantly elevated in fish exposed to five of the WWTP effluents. In contrast, transcript levels of vtg were not affected by any of the WWTP effluent exposures. Mean levels of natural and synthetic estrogens in fish bile were consistent with pituitary lhb expression, suggesting that the observed lhb induction may be due to

  9. Dietary fatty acids regulate hepatic low density lipoprotein (LDL) transport by altering LDL receptor protein and mRNA levels.

    PubMed Central

    Horton, J D; Cuthbert, J A; Spady, D K

    1993-01-01

    The concentration of LDL in plasma is strongly influenced by the amount and the type of lipid in the diet. Recent studies in the hamster have shown that dietary fatty acids differentially affect circulating LDL levels primarily by altering receptor-dependent LDL uptake in the liver. To investigate the mechanistic basis of this effect, rates of receptor-dependent LDL transport in the liver were correlated with LDL receptor protein and mRNA levels in hamsters fed safflower oil or coconut oil and varying amounts of cholesterol. Hepatic LDL receptor activity was significantly lower in animals fed coconut oil than in animals fed safflower oil at all levels of cholesterol intake (26, 53, and 61% lower at cholesterol intakes of 0, 0.06, and 0.12%, respectively). These fatty acid-induced changes in hepatic LDL receptor activity were accompanied by parallel changes in hepatic LDL receptor protein and mRNA levels, suggesting that dietary fatty acids regulate the LDL receptor pathway largely at the mRNA level. Images PMID:8349814

  10. An Investigation of Alerting and Prioritization Criteria for Sense and Avoid (SAA)

    DTIC Science & Technology

    2013-10-01

    TECHNICAL REPORT RDMR-TM-13-01 AN INVESTIGATION OF ALERTING AND PRIORITIZATION CRITERIA FOR SENSE AND AVOID ( SAA ...OFFICIAL ENDORSEMENT OR APPROVAL OF THE USE OF SUCH COMMERCIAL HARDWARE OR SOFTWARE . i/ii (Blank) REPORT DOCUMENTATION PAGE Form Approved...burden to Washington Headquarters Services , Directorate for Information Operations and Reports, 1215 Jefferson Davis Highway, Suite 1204, Arlington

  11. Maternal plasma levels of cell-free β-HCG mRNA as a prenatal diagnostic indicator of placenta accrete.

    PubMed

    Zhou, J; Li, J; Yan, P; Ye, Y H; Peng, W; Wang, S; Wang, X Tong

    2014-09-01

    Several biomarkers, including maternal serum creatinine kinase and α-fetoprotein, have been described as potential tools for the diagnosis of placental abnormalities. This study aimed to determine whether maternal plasma mRNA levels of the β subunit of human chorionic gonadotropin (β-HCG) could predict placenta accreta prenatally. Sixty-eight singleton pregnant women with prior cesarean deliveries (CDs) were classified into three groups: normal placentation (35 women, control group); placenta previa alone (21 women, placenta previa group); and both placenta previa and placenta accreta (12 women, placenta previa/accreta group). Maternal plasma concentrations of cell-free β-HCG mRNA were measured by real-time reverse-transcription polymerase chain reaction and were expressed as multiples of the median (MoM). Cell-free β-HCG mRNA concentrations (MoM, range) were significantly higher in women with placenta accreta (3.65, 2.78-7.19) than in women with placenta previa (0.94, 0.00-2.97) or normal placentation (1.00, 0.00-2.69) (Steel-Dwass test, P < 0.01 and P < 0.01, respectively). In the placenta previa/accreta group, the concentration of cell-free β-HCG mRNA was significantly higher among women who underwent CDs with hysterectomy (4.41, 3.49-7.19) than among women whose CDs did not result in hysterectomy (3.20, 2.78-3.70) (Mann-Whitney U test, P = 0.012). An increased level of cell-free β-HCG mRNA in the maternal plasma of women with placenta accreta may arise from direct uteroplacental transfer of cell-free placental mRNA molecules. The concentration of cell-free β-HCG mRNA in maternal plasma may be applicable to the prenatal diagnosis of placenta accreta, especially to identify women with placenta accreta likely to require hysterectomy. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Lower glutamic acid decarboxylase 65kD mRNA and protein levels in the prefrontal cortex in schizoaffective disorder but not schizophrenia

    PubMed Central

    Glausier, JR; Kimoto, S; Fish, KN; Lewis, DA

    2014-01-01

    Background Altered GABA signaling in the prefrontal cortex (PFC) has been associated with cognitive dysfunction in schizophrenia and schizoaffective disorder. PFC levels of the GABA-synthesizing enzyme glutamic acid decarboxylase 67kD (GAD67) has been consistently reported to be lower in these disorders, but the status of the second GABA-synthesizing enzyme, GAD65, remains unclear. Methods GAD65 mRNA levels were quantified in PFC area 9 by quantitative polymerase chain reaction from 62 subjects with schizophrenia or schizoaffective disorder and 62 matched healthy comparison subjects. GAD65 relative protein levels were quantified in a subset of subject pairs by confocal immunofluorescence microscopy. Results Mean GAD65 mRNA levels were 13.6% lower in schizoaffective disorder subjects, but did not differ in schizophrenia subjects, relative to their matched healthy comparison subjects. In the subjects with schizoaffective disorder, mean GAD65 protein levels were 19.4% lower and were correlated with GAD65 mRNA levels. Lower GAD65 mRNA and protein measures within schizoaffective disorder subjects was not attributable to factors commonly comorbid with the diagnosis. Conclusions In concert with previous studies, these findings suggest that schizoaffective disorder is associated with lower levels of both GAD65 and GAD67 mRNA and protein in the PFC, whereas subjects with schizophrenia have lower mean levels of only GAD67 mRNA and protein. Because cognitive function is generally better preserved in subjects with schizoaffective disorder relative to subjects with schizophrenia, these findings may support an interpretation that GAD65 down-regulation provides a homeostatic response complementary to GAD67 down-regulation expression that serves to reduce inhibition in the face of lower PFC network activity. PMID:24993056

  13. Neurotrophins and their receptors in the rat pituitary gland: regulation of BDNF and trkB mRNA levels by adrenal hormones.

    PubMed

    Kononen, J; Soinila, S; Persson, H; Honkaniemi, J; Hökfelt, T; Pelto-Huikko, M

    1994-12-01

    We studied the expression of messenger ribonucleic acids (mRNAs) for neurotrophins and neurotrophin receptors in the rat pituitary gland and examined the influence of adrenal hormones on their mRNA levels, using in situ hybridization and Northern blot analysis. The only neurotrophin present at detectable levels in the pituitary was brain-derived neurotrophic factor (BDNF), which was observed in the anterior and intermediate lobes. Several transcripts of the putative receptor for BDNF, trkB, were present in the anterior and posterior lobes of the pituitary. A low amount of trkC mRNA was found in both the anterior and the intermediate lobe. Dexamethasone treatment decreased both BDNF and trkB mRNA levels in the anterior lobe of the pituitary. Adrenalectomy had no effect on trkB expression, but it decreased BDNF mRNA levels in comparison to the control animals. This effect could not be reversed by dexamethasone substitution, suggesting that BDNF, mRNA levels may be regulated not only by glucocorticoids but also by other adrenal hormones. These results demonstrate that BDNF, trkB and trkC are expressed in the pituitary gland and that glucocorticoids and possibly other adrenal hormones may modulate pituitary functions by regulating the expression of neurotrophic factors and their receptors. Whether BDNF acts as a secreted hormone, a trophic factor, or has autocrine/paracrine functions within the pituitary through its receptor, trkB, remains to be studied.

  14. Effects of Age on Bone mRNA Levels of Sclerostin and Other Genes Relevant to Bone Metabolism in Humans

    PubMed Central

    Roforth, Matthew M.; Fujita, Koji; McGregor, Ulrike I.; Kirmani, Salman; McCready, Louise K.; Peterson, James M.; Drake, Matthew T.; Monroe, David G.; Khosla, Sundeep

    2013-01-01

    Although aging is associated with a decline in bone formation in humans, the molecular pathways contributing to this decline remain unclear. Several previous clinical studies have shown that circulating sclerostin levels increase with age, raising the possibility that increased production of sclerostin by osteocytes leads to the age-related impairment in bone formation. Thus, in the present study, we examined circulating sclerostin levels as well as bone mRNA levels of sclerostin using quantitative polymerase chain reaction (QPCR) analyses in needle bone biopsies from young (mean age, 30.0 years) versus old (mean age, 72.9 years) women. In addition, we analyzed the expression of genes in a number of pathways known to be altered with skeletal aging, based largely on studies in mice. While serum sclerostin levels were 46% higher (p < 0.01) in the old as compared to the young women, bone sclerostin mRNA levels were no different between the two groups (p = 0.845). However, genes related to notch signaling were significantly upregulated (p = 0.003 when analyzed as a group) in the biopsies from the old women. In an additional analysis of 118 genes including those from genome-wide association studies related to bone density and/or fracture, BMP/TGFβ family genes, selected growth factors and nuclear receptors, and Wnt/Wnt-related genes, we found that mRNA levels of the Wnt inhibitor, SFRP1, were significantly increased (by 1.6-fold, p = 0.0004, false discovery rate [q] = 0.04) in the biopsies from the old as compared to the young women. Our findings thus indicate that despite increases in circulating sclerostin levels, bone sclerostin mRNA levels do not increase in elderly women. However, aging is associated with alterations in several key pathways and genes in humans that may contribute to the observed impairment in bone formation. These include notch signaling, which represents a potential therapeutic target for increasing bone formation in humans. Our studies further

  15. Coupled Evolution of Transcription and mRNA Degradation

    PubMed Central

    Dori-Bachash, Mally; Shema, Efrat; Tirosh, Itay

    2011-01-01

    mRNA levels are determined by the balance between transcription and mRNA degradation, and while transcription has been extensively studied, very little is known regarding the regulation of mRNA degradation and its coordination with transcription. Here we examine the evolution of mRNA degradation rates between two closely related yeast species. Surprisingly, we find that around half of the evolutionary changes in mRNA degradation were coupled to transcriptional changes that exert opposite effects on mRNA levels. Analysis of mRNA degradation rates in an interspecific hybrid further suggests that opposite evolutionary changes in transcription and in mRNA degradation are mechanistically coupled and were generated by the same individual mutations. Coupled changes are associated with divergence of two complexes that were previously implicated both in transcription and in mRNA degradation (Rpb4/7 and Ccr4-Not), as well as with sequence divergence of transcription factor binding motifs. These results suggest that an opposite coupling between the regulation of transcription and that of mRNA degradation has shaped the evolution of gene regulation in yeast. PMID:21811398

  16. Maternal serum amyloid A level as a novel marker of primary unexplained recurrent early pregnancy loss.

    PubMed

    Ibrahim, Moustafa I; Ramy, Ahmed R; Abdelhamid, Ahmed S; Ellaithy, Mohamed I; Omar, Amna; Harara, Rany M; Fathy, Hayam; Abolouz, Ashraf S

    2017-03-01

    To assess maternal serum amyloid A (SAA) levels among women with primary unexplained recurrent early pregnancy loss (REPL). A prospective study was conducted among women with missed spontaneous abortion in the first trimester at Ain Shams University Maternity Hospital, Cairo, Egypt, between January 21 and December 25, 2014. Women with at least two consecutive primary unexplained REPLs and no previous live births were enrolled. A control group was formed of women with no history of REPL who had at least one previous uneventful pregnancy with no adverse outcomes. Serum samples were collected to measure SAA levels. The main outcome was the association between SAA and primary unexplained REPL. Each group contained 96 participants. Median SAA level was significantly higher among women with REPL (50.0 μg/mL, interquartile range 26.0-69.0) than among women in the control group (11.6 μg/mL, interquartile range 6.2-15.5; P<0.001). The SAA level was an independent indicator of primary unexplained REPL, after adjusting for maternal age and gestational age (odds ratio 1.12, 95% confidence interval 1.06-1.19; P<0.001). Elevated SAA levels found among women with primary unexplained REPL could represent a novel biomarker for this complication of pregnancy. © 2016 International Federation of Gynecology and Obstetrics.

  17. Serum Amyloid A Induces Toll-Like Receptor 2-Dependent Inflammatory Cytokine Expression and Atrophy in C2C12 Skeletal Muscle Myotubes.

    PubMed

    Passey, Samantha L; Bozinovski, Steven; Vlahos, Ross; Anderson, Gary P; Hansen, Michelle J

    2016-01-01

    Skeletal muscle wasting is an important comorbidity of Chronic Obstructive Pulmonary Disease (COPD) and is strongly correlated with morbidity and mortality. Patients who experience frequent acute exacerbations of COPD (AECOPD) have more severe muscle wasting and reduced recovery of muscle mass and function after each exacerbation. Serum levels of the pro-inflammatory acute phase protein Serum Amyloid A (SAA) can rise more than 1000-fold in AECOPD and are predictively correlated with exacerbation severity. The direct effects of SAA on skeletal muscle are poorly understood. Here we have examined SAA effects on pro-inflammatory cachectic cytokine expression (IL-6 and TNFα) and atrophy in C2C12 myotubes. SAA increased IL-6 (31-fold) and TNFα (6.5-fold) mRNA levels compared to control untreated cells after 3h of SAA treatment, and increased secreted IL-6 protein at 24h. OxPAPC, a dual TLR2 and TLR4 inhibitor, reduced the response to SAA by approximately 84% compared to SAA alone, and the TLR2 neutralising antibody T2.5 abolished SAA-induced expression of IL-6, indicating that SAA signalling in C2C12 myotubes is primarily via TLR2. SAA also reduced myotube width by 10-13% and induced a 2.5-fold increase in the expression of the muscle atrophy gene Atrogin-1, suggesting direct effects of SAA on muscle wasting. Blocking of TLR2 inhibited the SAA-induced decrease in myotube width and Atrogin-1 gene expression, indicating that SAA induces atrophy through TLR2. These data demonstrate that SAA stimulates a robust pro-inflammatory response in skeletal muscle myotubes via the TLR2-dependent release of IL-6 and TNFα. Furthermore, the observed atrophy effects indicate that SAA could also be directly contributing to the wasting and poor recovery of muscle mass. Therapeutic strategies targeting this SAA-TLR2 axis may therefore ameliorate muscle wasting in AECOPD and a range of other inflammatory conditions associated with loss of muscle mass.

  18. Decreased salivary alpha-amylase levels are associated with performance deficits during sleep loss.

    PubMed

    Pajcin, Maja; Banks, Siobhan; White, Jason M; Dorrian, Jill; Paech, Gemma M; Grant, Crystal; Johnson, Kayla; Tooley, Katie; Fidock, Justin; Kamimori, Gary H; Della Vedova, Chris B

    2017-04-01

    During sleep deprivation, neurobehavioral functions requiring sustained levels of attention and alertness are significantly impaired. Discrepancies between subjective measures of sleepiness and objective performance during sustained operations have led to interest in physiological monitoring of operator performance. Alertness, vigilance, and arousal are modulated by the wake-promoting actions of the central noradrenergic system. Salivary alpha-amylase (sAA) has been proposed as a sensitive peripheral measure of noradrenergic activity, but limited research has investigated the relationship between sAA and performance. In a laboratory-controlled environment, we investigated the relationship between sAA levels, subjective sleepiness, and performance during two days (50h) of total sleep deprivation. Beginning at 09:00, twelve healthy participants (5 females) aged 22.5±2.5years (mean±SD) provided saliva samples, recorded ratings of subjective sleepiness, completed a brief 3-min psychomotor vigilance task (PVT-B) and performed a 40-min simulated driving task, at regular 3h intervals during wakefulness. Ratings of subjective sleepiness exhibited a constant linear increase (p<0.001) during sleep deprivation. In contrast, sAA levels showed a marked diurnal profile, with levels increasing during the day (p<0.001) and steadily declining in the evening and early-morning (p<0.001). PVT-B (mean reaction time and mean slowest 10% reaction time) and simulated driving performance (speed deviation and lane deviation) also exhibited diurnal profiles across the two days of sleep deprivation. Performance peaked in the afternoon (p<0.001) and then steadily worsened as wakefulness continued into the evening and early-morning (p<0.001). Further analysis revealed that higher sAA levels in the hour preceding each performance assessment were associated with better PVT-B and driving performance (p<0.001). These findings suggest that sAA measures may be suitable indicators of performance

  19. Prolonged treatment with imatinib mesylate in patients with advanced chronic myeloid leukemia causes a reduction of bcr/abl mRNA levels independent of cytogenetic response.

    PubMed

    Cariani, E; Capucci, M; Micheletti, M; Spalenza, F; Zanella, I; Albertini, A; Rossi, G

    2003-06-01

    Bcr/abl mRNA levels were monitored in 13 patients with chronic myeloid leukemia receiving imatinib mesylate over a period of 78 weeks. During treatment median bcr/abl mRNA levels progressively declined from 77.2 normalized dose (nD) at baseline to 11.28 nD after 13 weeks ( P<0.05) and to 1.28 nD after 78 weeks ( P<0.05). After 13 weeks, bcr/abl mRNA levels were significantly lower in cytogenetic responders compared to nonresponders ( P<0.05), but subsequent decrease in the transcript levels caused the loss of any correlation to the cytogenetic status. These results suggest that bcr/abl mRNA levels may reflect cytogenetic response only during the early phases of imatinib therapy.

  20. RNase MRP cleaves the CLB2 mRNA to promote cell cycle progression: novel method of mRNA degradation.

    PubMed

    Gill, Tina; Cai, Ti; Aulds, Jason; Wierzbicki, Sara; Schmitt, Mark E

    2004-02-01

    RNase mitochondrial RNA processing (RNase MRP) mutants have been shown to have an exit-from-mitosis defect that is caused by an increase in CLB2 mRNA levels, leading to increased Clb2p (B-cyclin) levels and a resulting late anaphase delay. Here we describe the molecular defect behind this delay. CLB2 mRNA normally disappears rapidly as cells complete mitosis, but the level remains high in RNase MRP mutants. This is in direct contrast to other exit-from-mitosis mutants and is the result of an increase in CLB2 mRNA stability. We found that highly purified RNase MRP cleaved the 5' untranslated region (UTR) of the CLB2 mRNA in several places in an in vitro assay. In vivo, we identified RNase MRP-dependent cleavage products on the CLB2 mRNA that closely matched in vitro products. Disposal of these products was dependent on the 5'-->3' exoribonuclease Xrn1 and not the exosome. Our results demonstrate that the endoribonuclease RNase MRP specifically cleaves the CLB2 mRNA in its 5'-UTR to allow rapid 5' to 3' degradation by the Xrn1 nuclease. Degradation of the CLB2 mRNA by the RNase MRP endonuclease provides a novel way to regulate the cell cycle that complements the protein degradation machinery. In addition, these results denote a new mechanism of mRNA degradation not seen before in the yeast Saccharomyces cerevisiae.

  1. The internal structure of eclogite-facies ophiolite complexes: Implications from the Austroalpine outliers within the Zermatt-Saas Zone, Western Alps

    NASA Astrophysics Data System (ADS)

    Weber, Sebastian; Martinez, Raul

    2016-04-01

    The Western Alpine Penninic domain is a classical accretionary prism that formed after the closure of the Penninic oceans in the Paleogene. Continental and oceanic nappes were telescoped into the Western Alpine stack associated with continent-continent collision. Within the Western Alpine geologic framework, the ophiolite nappes of the Zermatt-Saas Zone and the Tsate Unit are the remnants of the southern branch of the Piemonte-Liguria ocean basin. In addition, a series of continental basement slices reported as lower Austroalpine outliers have preserved an eclogitic high-pressure imprint, and are tectonically sandwiched between these oceanic nappes. Since the outliers occur at an unusual intra-ophiolitic setting and show a polymetamorphic character, this group of continental slices is of special importance for understanding the tectono-metamorphic evolution of Western Alps. Recently, more geochronological data from the Austroalpine outliers have become available that make it possible to establish a more complete picture of their complex geological history. The Lu-Hf garnet-whole rock ages for prograde growth of garnet fall into the time interval of 52 to 62 Ma (Weber et al., 2015, Fassmer et al. 2015), but are consistently higher than the Lu-Hf garnet-whole rock ages from several other locations throughout the Zermatt-Saas zone that range from 52 to 38 Ma (Skora et al., 2015). This discrepancy suggests that the Austroalpine outliers may have been subducted earlier than the ophiolites of the Zermatt-Saas Zone and therefore have been tectonically emplaced into their present intra-ophiolite position. This points to the possibility that the Zermatt-Saas Zone consists of tectonic subunits, which reached their respective pressure peaks over a prolonged time period, approximately 10-20 Ma. The pressure-temperature estimates from several members of the Austroalpine outliers indicate a complex distribution of metamorphic peak conditions, without ultrahigh

  2. Lysophosphatidic acid receptor mRNA levels in heart and white adipose tissue are associated with obesity in mice and humans.

    PubMed

    Brown, Amy; Hossain, Intekhab; Perez, Lester J; Nzirorera, Carine; Tozer, Kathleen; D'Souza, Kenneth; Trivedi, Purvi C; Aguiar, Christie; Yip, Alexandra M; Shea, Jennifer; Brunt, Keith R; Legare, Jean-Francois; Hassan, Ansar; Pulinilkunnil, Thomas; Kienesberger, Petra C

    2017-01-01

    Lysophosphatidic acid (LPA) receptor signaling has been implicated in cardiovascular and obesity-related metabolic disease. However, the distribution and regulation of LPA receptors in the myocardium and adipose tissue remain unclear. This study aimed to characterize the mRNA expression of LPA receptors (LPA1-6) in the murine and human myocardium and adipose tissue, and its regulation in response to obesity. LPA receptor mRNA levels were determined by qPCR in i) heart ventricles, isolated cardiomyocytes, and perigonadal adipose tissue from chow or high fat-high sucrose (HFHS)-fed male C57BL/6 mice, ii) 3T3-L1 adipocytes and HL-1 cardiomyocytes under conditions mimicking gluco/lipotoxicity, and iii) human atrial and subcutaneous adipose tissue from non-obese, pre-obese, and obese cardiac surgery patients. LPA1-6 were expressed in myocardium and white adipose tissue from mice and humans, except for LPA3, which was undetectable in murine adipocytes and human adipose tissue. Obesity was associated with increased LPA4, LPA5 and/or LPA6 levels in mice ventricles and cardiomyocytes, HL-1 cells exposed to high palmitate, and human atrial tissue. LPA4 and LPA5 mRNA levels in human atrial tissue correlated with measures of obesity. LPA5 mRNA levels were increased in HFHS-fed mice and insulin resistant adipocytes, yet were reduced in adipose tissue from obese patients. LPA4, LPA5, and LPA6 mRNA levels in human adipose tissue were negatively associated with measures of obesity and cardiac surgery outcomes. This study suggests that obesity leads to marked changes in LPA receptor expression in the murine and human heart and white adipose tissue that may alter LPA receptor signaling during obesity.

  3. Lysophosphatidic acid receptor mRNA levels in heart and white adipose tissue are associated with obesity in mice and humans

    PubMed Central

    Perez, Lester J.; Nzirorera, Carine; Tozer, Kathleen; D’Souza, Kenneth; Trivedi, Purvi C.; Aguiar, Christie; Yip, Alexandra M.; Shea, Jennifer; Brunt, Keith R.; Legare, Jean-Francois; Hassan, Ansar; Pulinilkunnil, Thomas

    2017-01-01

    Background Lysophosphatidic acid (LPA) receptor signaling has been implicated in cardiovascular and obesity-related metabolic disease. However, the distribution and regulation of LPA receptors in the myocardium and adipose tissue remain unclear. Objectives This study aimed to characterize the mRNA expression of LPA receptors (LPA1-6) in the murine and human myocardium and adipose tissue, and its regulation in response to obesity. Methods LPA receptor mRNA levels were determined by qPCR in i) heart ventricles, isolated cardiomyocytes, and perigonadal adipose tissue from chow or high fat-high sucrose (HFHS)-fed male C57BL/6 mice, ii) 3T3-L1 adipocytes and HL-1 cardiomyocytes under conditions mimicking gluco/lipotoxicity, and iii) human atrial and subcutaneous adipose tissue from non-obese, pre-obese, and obese cardiac surgery patients. Results LPA1-6 were expressed in myocardium and white adipose tissue from mice and humans, except for LPA3, which was undetectable in murine adipocytes and human adipose tissue. Obesity was associated with increased LPA4, LPA5 and/or LPA6 levels in mice ventricles and cardiomyocytes, HL-1 cells exposed to high palmitate, and human atrial tissue. LPA4 and LPA5 mRNA levels in human atrial tissue correlated with measures of obesity. LPA5 mRNA levels were increased in HFHS-fed mice and insulin resistant adipocytes, yet were reduced in adipose tissue from obese patients. LPA4, LPA5, and LPA6 mRNA levels in human adipose tissue were negatively associated with measures of obesity and cardiac surgery outcomes. This study suggests that obesity leads to marked changes in LPA receptor expression in the murine and human heart and white adipose tissue that may alter LPA receptor signaling during obesity. PMID:29236751

  4. Structure of Serum Amyloid A Suggests a Mechanism for Selective Lipoprotein Binding and Functions: SAA as a Hub in Macromolecular Interaction Networks

    PubMed Central

    Frame, Nicholas M.; Gursky, Olga

    2016-01-01

    Serum amyloid A is a major acute-phase plasma protein that modulates innate immunity and cholesterol homeostasis. We combine sequence analysis with x-ray crystal structures to postulate that SAA acts as an intrinsically disordered hub mediating interactions among proteins, lipids and proteoglycans. A structural model of lipoprotein-bound SAA monomer is proposed wherein two α-helices from the N-domain form a concave hydrophobic surface that binds lipoproteins. A C-domain, connected to the N-domain via a flexible linker, binds polar/charged ligands including cell receptors, bridging them with lipoproteins and re-routing cholesterol transport. Our model is supported by the SAA cleavage in the inter-domain linker to generate the 1–76 fragment deposited in reactive amyloidosis. This model sheds new light on functions of this enigmatic protein. PMID:26918388

  5. Interactions between the HIV-1 Unspliced mRNA and Host mRNA Decay Machineries

    PubMed Central

    Toro-Ascuy, Daniela; Rojas-Araya, Bárbara; Valiente-Echeverría, Fernando; Soto-Rifo, Ricardo

    2016-01-01

    The human immunodeficiency virus type-1 (HIV-1) unspliced transcript is used both as mRNA for the synthesis of structural proteins and as the packaged genome. Given the presence of retained introns and instability AU-rich sequences, this viral transcript is normally retained and degraded in the nucleus of host cells unless the viral protein REV is present. As such, the stability of the HIV-1 unspliced mRNA must be particularly controlled in the nucleus and the cytoplasm in order to ensure proper levels of this viral mRNA for translation and viral particle formation. During its journey, the HIV-1 unspliced mRNA assembles into highly specific messenger ribonucleoproteins (mRNPs) containing many different host proteins, amongst which are well-known regulators of cytoplasmic mRNA decay pathways such as up-frameshift suppressor 1 homolog (UPF1), Staufen double-stranded RNA binding protein 1/2 (STAU1/2), or components of miRNA-induced silencing complex (miRISC) and processing bodies (PBs). More recently, the HIV-1 unspliced mRNA was shown to contain N6-methyladenosine (m6A), allowing the recruitment of YTH N6-methyladenosine RNA binding protein 2 (YTHDF2), an m6A reader host protein involved in mRNA decay. Interestingly, these host proteins involved in mRNA decay were shown to play positive roles in viral gene expression and viral particle assembly, suggesting that HIV-1 interacts with mRNA decay components to successfully accomplish viral replication. This review summarizes the state of the art in terms of the interactions between HIV-1 unspliced mRNA and components of different host mRNA decay machineries. PMID:27886048

  6. Hepatic expression of serum amyloid A1 is induced by traumatic brain injury and modulated by telmisartan.

    PubMed

    Villapol, Sonia; Kryndushkin, Dmitry; Balarezo, Maria G; Campbell, Ashley M; Saavedra, Juan M; Shewmaker, Frank P; Symes, Aviva J

    2015-10-01

    Traumatic brain injury affects the whole body in addition to the direct impact on the brain. The systemic response to trauma is associated with the hepatic acute-phase response. To further characterize this response, we performed controlled cortical impact injury on male mice and determined the expression of serum amyloid A1 (SAA1), an apolipoprotein, induced at the early stages of the acute-phase response in liver and plasma. After cortical impact injury, induction of SAA1 was detectable in plasma at 6 hours post-injury and in liver at 1 day post-injury, followed by gradual diminution over time. In the liver, cortical impact injury increased neutrophil and macrophage infiltration, apoptosis, and expression of mRNA encoding the chemokines CXCL1 and CXCL10. An increase in angiotensin II AT1 receptor mRNA at 3 days post-injury was also observed. Administration of the AT1 receptor antagonist telmisartan 1 hour post-injury significantly decreased liver SAA1 levels and CXCL10 mRNA expression, but did not affect CXCL1 expression or the number of apoptotic cells or infiltrating leukocytes. To our knowledge, this is the first study to demonstrate that SAA1 is induced in the liver after traumatic brain injury and that telmisartan prevents this response. Elucidating the molecular pathogenesis of the liver after brain injury will assist in understanding the efficacy of therapeutic approaches to brain injury. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  7. Creatine kinase and alpha-actin mRNA levels decrease in diabetic rat hearts

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Popovich, B.; Barrieux, A.; Dillmann, W.H.

    1987-05-01

    Diabetic cardiomyopathy is associated with cardiac atrophy and isoenzyme redistribution. To determine if tissue specific changes occur in mRNAs coding for ..cap alpha..-actin and creatine kinase (CK), they performed RNA blot analysis. Total ventricular RNA from control (C) and 4 wk old diabetic (D) rats were hybridized with /sup 32/P cDNA probes for ..cap alpha..-actin and CK. A tissue independent cDNA probe, CHOA was also used. Signal intensity was quantified by photodensitometry. D CK mRNA was 47 +/- 16% lower in D vs C. Insulin increases CK mRNA by 20% at 1.5 hs, and completely reverses the deficit after 4more » wks. D ..cap alpha..-actin mRNA is 66 +/- 18% lower in D vs C. Insulin normalized ..cap alpha..-actin mRNA by 5 hs. CHOA mRNA is unchanged in D vs C, but D + insulin CHOA mRNA is 30 +/- 2% lower than C. In rats with diabetic cardiomyopathy, muscle specific CK and ..cap alpha..-actin mRNAs are decreased. Insulin treatment reverses these changes.« less

  8. Analysis of acute-phase proteins, AHSG, C3, CLI, HP and SAA, reveals distinctive expression patterns associated with breast, colorectal and lung cancer.

    PubMed

    Dowling, Paul; Clarke, Colin; Hennessy, Kim; Torralbo-Lopez, Beatriz; Ballot, Jo; Crown, John; Kiernan, Ingrid; O'Byrne, Kenneth J; Kennedy, M John; Lynch, Vincent; Clynes, Martin

    2012-08-15

    Early detection, clinical management and disease recurrence monitoring are critical areas in cancer treatment in which specific biomarker panels are likely to be very important in each of these key areas. We have previously demonstrated that levels of alpha-2-heremans-schmid-glycoprotein (AHSG), complement component C3 (C3), clusterin (CLI), haptoglobin (HP) and serum amyloid A (SAA) are significantly altered in serum from patients with squamous cell carcinoma of the lung. Here, we report the abundance levels for these proteins in serum samples from patients with advanced breast cancer, colorectal cancer (CRC) and lung cancer compared to healthy controls (age and gender matched) using commercially available enzyme-linked immunosorbent assay kits. Logistic regression (LR) models were fitted to the resulting data, and the classification ability of the proteins was evaluated using receiver-operating characteristic curve and leave-one-out cross-validation (LOOCV). The most accurate individual candidate biomarkers were C3 for breast cancer [area under the curve (AUC) = 0.89, LOOCV = 73%], CLI for CRC (AUC = 0.98, LOOCV = 90%), HP for small cell lung carcinoma (AUC = 0.97, LOOCV = 88%), C3 for lung adenocarcinoma (AUC = 0.94, LOOCV = 89%) and HP for squamous cell carcinoma of the lung (AUC = 0.94, LOOCV = 87%). The best dual combination of biomarkers using LR analysis were found to be AHSG + C3 (AUC = 0.91, LOOCV = 83%) for breast cancer, CLI + HP (AUC = 0.98, LOOCV = 92%) for CRC, C3 + SAA (AUC = 0.97, LOOCV = 91%) for small cell lung carcinoma and HP + SAA for both adenocarcinoma (AUC = 0.98, LOOCV = 96%) and squamous cell carcinoma of the lung (AUC = 0.98, LOOCV = 84%). The high AUC values reported here indicated that these candidate biomarkers have the potential to discriminate accurately between control and cancer groups both individually and in combination with other proteins. Copyright © 2011 UICC.

  9. [The expressions of HSP 70 mRNA and c-fos mRNA in the skeletal muscle and cardiac muscle of rabbits by electrocuted].

    PubMed

    Wang, Ye; Liu, Min; Cheng, Wei-bo; He, Gui-qiong; Li, Fan; Liao, Zhi-gang

    2008-08-01

    To study the changes of HSP 70 mRNA and c-fos mRNA expression and to find a method to differentiate antemortem from postmortem electrocution. Fifteen New Zealand rabbits were randomly divided into three groups, the antemortem electrocution group, the postmortem electrocution group, and the control group. Each group consists of five rabbits. The levels of HSP 70 mRNA and c-fos mRNA in skeletal muscle and cardiac muscle were examined with quantitative fluorescent RT-PCR. The levels of HSP 70 mRNA and c-fos mRNA in the antemortem electrocution group increased significantly (P<0.05), compared with that of the postmortem electrocution group. The changes of HSP 70 mRNA and c-fos mRNA expression in skeletal muscle and cardiac muscle can be used as an indicator to distinguish antemortem from postmortem electrocution.

  10. Sex hormone-binding globulin and corticosteroid-binding globulin mRNA levels in infertile women with luteal phase deficiency.

    PubMed

    Misao, R; Nakanishi, Y; Fujimoto, J; Tamaya, T

    1995-09-01

    This study was designed to investigate the biological significance in intracellular expression of sex hormone-binding globulin (SHBG) and corticosteroid-binding globulin (CBG) mRNA in uterine endometrium with luteal phase deficiency (designated as out-of-phase endometrium or low serum progesterone level). The levels of such mRNAs were measured by the quantitative reverse transcription-polymerase chain reaction. Under the normal serum 17 beta-estradiol and progesterone levels in the mid-luteal phase, the levels of SHBG and CBG mRNAs in the out-of-phase endometria were not significantly different from those in the normal endometria. On the other hand, SHBG and CBG mRNA levels in the endometria of low serum midluteal progesterone level were significantly (p < 0.05) reduced and raised, respectively, compared with normal levels. These findings suggest that the synthesis of endometrial steroid-binding proteins in the out-of-phase endometrium is conserved, as that in the in-phase endometrium, whereas the decreased progesterone level might up-regulate CBG expression with down-regulation of SHBG expression.

  11. Low-level laser irradiation modulates brain-derived neurotrophic factor mRNA transcription through calcium-dependent activation of the ERK/CREB pathway.

    PubMed

    Yan, Xiaodong; Liu, Juanfang; Zhang, Zhengping; Li, Wenhao; Sun, Siguo; Zhao, Jian; Dong, Xin; Qian, Jixian; Sun, Honghui

    2017-01-01

    Low-level laser (LLL) irradiation has been reported to promote neuronal differentiation, but the mechanism remains unclear. Brain-derived neurotrophic factor (BDNF) has been confirmed to be one of the most important neurotrophic factors because it is critical for the differentiation and survival of neurons during development. Thus, this study aimed to investigate the effects of LLL irradiation on Bdnf messenger RNA (mRNA) transcription and the molecular pathway involved in LLL-induced Bdnf mRNA transcription in cultured dorsal root ganglion neurons (DRGNs) using Ca 2+ imaging, pharmacological detections, RNA interference, immunocytochemistry assay, Western blot, and qPCR analysis. We show here that LLL induced increases in the [Ca 2+ ] i level, Bdnf mRNA transcription, cAMP-response element-binding protein (CREB) phosphorylation, and extracellular signal-regulated kinase (ERK) phosphorylation, mediated by Ca 2+ release via inositol triphosphate receptor (IP3R)-sensitive calcium (Ca 2+ ) stores. Blockade of Ca 2+ increase suppressed Bdnf mRNA transcription, CREB phosphorylation, and ERK phosphorylation. Downregulation of phosphorylated (p)-CREB reduced Bdnf mRNA transcription triggered by LLL. Furthermore, blockade of ERK using PD98059 inhibitor reduced p-CREB and Bdnf mRNA transcription induced by LLL. Taken together, these findings establish the Ca 2+ -ERK-CREB cascade as a potential signaling pathway involved in LLL-induced Bdnf mRNA transcription. To our knowledge, this is the first report of the mechanisms of Ca 2+ -dependent Bdnf mRNA transcription triggered by LLL. These findings may help further explore the complex molecular signaling networks in LLL-triggered nerve regeneration in vivo and may also provide experimental evidence for the development of LLL for clinical applications.

  12. Post-transcriptional regulation tends to attenuate the mRNA noise and to increase the mRNA gain

    NASA Astrophysics Data System (ADS)

    Shi, Changhong; Wang, Shuqiang; Zhou, Tianshou; Jiang, Yiguo

    2015-10-01

    Post-transcriptional regulation is ubiquitous in prokaryotic and eukaryotic cells, but how it impacts gene expression remains to be fully explored. Here, we analyze a simple gene model in which we assume that mRNAs are produced in a constitutive manner but are regulated post-transcriptionally by a decapping enzyme that switches between the active state and the inactive state. We derive the analytical mRNA distribution governed by a chemical master equation, which can be well used to analyze the mechanism of how post-transcription regulation influences the mRNA expression level including the mRNA noise. We demonstrate that the mean mRNA level in the stochastic case is always higher than that in the deterministic case due to the stochastic effect of the enzyme, but the size of the increased part depends mainly on the switching rates between two enzyme states. More interesting is that we find that in contrast to transcriptional regulation, post-transcriptional regulation tends to attenuate noise in mRNA. Our results provide insight into the role of post-transcriptional regulation in controlling the transcriptional noise.

  13. PAI-1 mRNA expression and plasma level in rheumatoid arthritis: relationship with 4G/5G PAI-1 polymorphism.

    PubMed

    Muñoz-Valle, José Francisco; Ruiz-Quezada, Sandra Luz; Oregón-Romero, Edith; Navarro-Hernández, Rosa Elena; Castañeda-Saucedo, Eduardo; De la Cruz-Mosso, Ulises; Illades-Aguiar, Berenice; Leyva-Vázquez, Marco Antonio; Castro-Alarcón, Natividad; Parra-Rojas, Isela

    2012-12-01

    Rheumatoid arthritis (RA) is a chronic inflammatory disease affecting the synovial membrane, cartilage and bone. PAI-1 is a key regulator of the fibrinolytic system through which plasminogen is converted to plasmin. The plasmin activates the matrix metalloproteinase system, which is closely related with the joint damage and bone destruction in RA. The aim of this study was to investigate the relationship between 4G/5G PAI-1 polymorphism with mRNA expression and PAI-1 plasma protein levels in RA patients. 113 RA patients and 123 healthy subjects (HS) were included in the study. The 4G/5G PAI-1 polymorphism was determined by polymerase chain reaction-restriction fragment length polymorphism method; the PAI-1 mRNA expression was determined by real-time PCR; and the soluble PAI-1 (sPAI-1) levels were quantified using an ELISA kit. No significant differences in the genotype and allele frequencies of 4G/5G PAI-1 polymorphism were found between RA patients and HS. However, the 5G/5G genotype was the most frequent in both studied groups: RA (42%) and HS (44%). PAI-1 mRNA expression was slightly increased (0.67 fold) in RA patients with respect to HS (P = 0.0001). In addition, in RA patients, the 4G/4G genotype carriers showed increased PAI-1 mRNA expression (3.82 fold) versus 4G/5G and 5G/5G genotypes (P = 0.0001), whereas the sPAI-1 plasma levels did not show significant differences. Our results indicate that the 4G/5G PAI-1 polymorphism is not a marker of susceptibility in the Western Mexico. However, the 4G/4G genotype is associated with high PAI-1 mRNA expression but not with the sPAI-1 levels in RA patients.

  14. GenSAA: A tool for advancing satellite monitoring with graphical expert systems

    NASA Technical Reports Server (NTRS)

    Hughes, Peter M.; Luczak, Edward C.

    1993-01-01

    During numerous contacts with a satellite each day, spacecraft analysts must closely monitor real time data for combinations of telemetry parameter values, trends, and other indications that may signify a problem or failure. As satellites become more complex and the number of data items increases, this task is becoming increasingly difficult for humans to perform at acceptable performance levels. At the NASA Goddard Space Flight Center, fault-isolation expert systems have been developed to support data monitoring and fault detection tasks in satellite control centers. Based on the lessons learned during these initial efforts in expert system automation, a new domain-specific expert system development tool named the Generic Spacecraft Analyst Assistant (GenSAA) is being developed to facilitate the rapid development and reuse of real-time expert systems to serve as fault-isolation assistants for spacecraft analysts. Although initially domain-specific in nature, this powerful tool will support the development of highly graphical expert systems for data monitoring purposes throughout the space and commercial industry.

  15. Apolipoprotein A-II induces acute-phase response associated AA amyloidosis in mice through conformational changes of plasma lipoprotein structure.

    PubMed

    Yang, Mu; Liu, Yingye; Dai, Jian; Li, Lin; Ding, Xin; Xu, Zhe; Mori, Masayuki; Miyahara, Hiroki; Sawashita, Jinko; Higuchi, Keiichi

    2018-04-04

    During acute-phase response (APR), there is a dramatic increase in serum amyloid A (SAA) in plasma high density lipoproteins (HDL). Elevated SAA leads to reactive AA amyloidosis in animals and humans. Herein, we employed apolipoprotein A-II (ApoA-II) deficient (Apoa2 -/- ) and transgenic (Apoa2Tg) mice to investigate the potential roles of ApoA-II in lipoprotein particle formation and progression of AA amyloidosis during APR. AA amyloid deposition was suppressed in Apoa2 -/- mice compared with wild type (WT) mice. During APR, Apoa2 -/- mice exhibited significant suppression of serum SAA levels and hepatic Saa1 and Saa2 mRNA levels. Pathological investigation showed Apoa2 -/- mice had less tissue damage and less inflammatory cell infiltration during APR. Total lipoproteins were markedly decreased in Apoa2 -/- mice, while the ratio of HDL to low density lipoprotein (LDL) was also decreased. Both WT and Apoa2 -/- mice showed increases in LDL and very large HDL during APR. SAA was distributed more widely in lipoprotein particles ranging from chylomicrons to very small HDL in Apoa2 -/- mice. Our observations uncovered the critical roles of ApoA-II in inflammation, serum lipoprotein stability and AA amyloidosis morbidity, and prompt consideration of therapies for AA and other amyloidoses, whose precursor proteins are associated with circulating HDL particles.

  16. Quantification of Chitinase mRNA Levels in Human and Mouse Tissues by Real-Time PCR: Species-Specific Expression of Acidic Mammalian Chitinase in Stomach Tissues

    PubMed Central

    Ohno, Misa; Togashi, Yuto; Tsuda, Kyoko; Okawa, Kazuaki; Kamaya, Minori; Sakaguchi, Masayoshi; Sugahara, Yasusato; Oyama, Fumitaka

    2013-01-01

    Chitinase hydrolyzes chitin, which is an N-acetyl-D-glucosamine polymer that is present in a wide range of organisms, including insects, parasites and fungi. Although mammals do not contain any endogenous chitin, humans and mice express two active chitinases, chitotriosidase (Chit1) and acidic mammalian chitinase (AMCase). Because the level of expression of these chitinases is increased in many inflammatory conditions, including Gaucher disease and mouse models of asthma, both chitinases may play important roles in the pathophysiologies of these and other diseases. We recently established a quantitative PCR system using a single standard DNA and showed that AMCase mRNA is synthesized at extraordinarily high levels in mouse stomach tissues. In this study, we applied this methodology to the quantification of chitinase mRNAs in human tissues and found that both chitinase mRNAs were widely expressed in normal human tissues. Chit1 mRNA was highly expressed in the human lung, whereas AMCase mRNA was not overexpressed in normal human stomach tissues. The levels of these mRNAs in human tissues were significantly lower than the levels of housekeeping genes. Because the AMCase expression levels were quite different between the human and mouse stomach tissues, we developed a quantitative PCR system to compare the mRNA levels between human and mouse tissues using a human-mouse hybrid standard DNA. Our analysis showed that Chit1 mRNA is expressed at similar levels in normal human and mouse lung. In contrast, the AMCase expression level in human stomach was significantly lower than that expression level observed in mouse stomach. These mRNA differences between human and mouse stomach tissues were reflecting differences in the chitinolytic activities and levels of protein expression. Thus, the expression level of the AMCase in the stomach is species-specific. PMID:23826286

  17. Maximum production of fermentable sugars from barley straw using optimized soaking in aqueous ammonia (SAA) pretreatment

    USDA-ARS?s Scientific Manuscript database

    Soaking in aqueous ammonia (SAA) pretreatment was investigated to improve enzymatic digestibility and consequently to increase total fermentable sugar production from barley straw. Various effects of pretreatment process parameters, such as reaction temperature, reaction time, solid:liquid ratio, an...

  18. Assessement of serum amyloid A levels in the rehabilitation setting in the Florida manatee (Trichechus manatus latirostris).

    PubMed

    Cray, Carolyn; Dickey, Meranda; Brewer, Leah Brinson; Arheart, Kristopher L

    2013-12-01

    The acute phase protein serum amyloid A (SAA) has been previously shown to have value as a biomarker of inflammation and infection in many species, including manatees (Trichechus manatus latirostris). In the current study, results from an automated assay for SAA were used in a rehabilitation setting. Reference intervals were established from clinically normal manatees using the robust method: 0-46 mg/L. More than 30-fold higher mean SAA levels were observed in manatees suffering from cold stress and boat-related trauma. Poor correlations were observed between SAA and total white blood count, percentage of neutrophils, albumin, and albumin/globulin ratio. A moderate correlation was observed between SAA and the presence of nucleated red blood cells. The sensitivity of SAA testing was 93% and the specificity was 98%, representing the highest combined values of all the analytes. The results indicate that the automated method for SAA quantitation can provide important clinical data for manatees in a rehabilitation setting.

  19. Green tea polyphenols improve cardiac muscle mRNA and protein levels of signal pathways related to insulin and lipid metabolism and inflammation in insulin-resistant rats.

    PubMed

    Qin, Bolin; Polansky, Marilyn M; Harry, Dawson; Anderson, Richard A

    2010-05-01

    Epidemiological studies indicate that the consumption of green tea polyphenols (GTP) may reduce the risk of coronary artery disease. To explore the underlying mechanisms of action at the molecular level, we examined the effects of GTP on the cardiac mRNA and protein levels of genes involved in insulin and lipid metabolism and inflammation. In rats fed a high-fructose diet, supplementation with GTP (200 mg/kg BW daily dissolved in distilled water) for 6 wk, reduced systemic blood glucose, plasma insulin, retinol-binding protein 4, soluble CD36, cholesterol, triglycerides, free fatty acids and LDL-C levels, as well as the pro-inflammatory cytokines, tumor necrosis factor-alpha (TNF-alpha) and IL-6. GTP did not affect food intake, bodyweight and heart weight. In the myocardium, GTP also increased the insulin receptor (Ir), insulin receptor substrate 1 and 2 (Irs1 and Irs2), phosphoinositide-3-kinase (Pi3k), v-akt murine thymoma viral oncogene homolog 1 (Akt1), glucose transporter 1 and 4 (Glut1 and Glut4) and glycogen synthase 1 (Gys1) expression but inhibited phosphatase and tensin homolog deleted on chromosome ten (Pten) expression and decreased glycogen synthase kinase 3beta (Gsk3beta) mRNA expression. The sterol regulatory element-binding protein-1c (Srebp1c) mRNA, microsomal triglyceride transfer protein (Mttp) mRNA and protein, Cd36 mRNA and cluster of differentiation 36 protein levels were decreased and peroxisome proliferator-activated receptor (Ppar)gamma mRNA levels were increased. GTP also decreased the inflammatory factors: Tnf, Il1b and Il6 mRNA levels, and enhanced the anti-inflammatory protein, zinc-finger protein, protein and mRNA expression. In summary, consumption of GTP ameliorated the detrimental effects of high-fructose diet on insulin signaling, lipid metabolism and inflammation in the cardiac muscle of rats.

  20. [The effects of methionine and choline on the expression levels of CaMKII and CREB mRNA and proteins in rats exposed to lead].

    PubMed

    Feng, Chang; Fan, Guang-qin; Wu, Feng-yun; Lin, Fen; Li, Yan-shu; Chen, Ying

    2012-07-01

    To study the effects of methionine and choline on the expression levels of CaMKII and CREB mRNA and proteins in hippocampus of rats exposed to lead. Male SD rats were divided into five groups. (1) control group, (2) group exposed to lead+2 by drinking water with 0.40 g/L lead acetate, (3) group exposed to methionine and choline (1:1, 400 mg/kg), (4) group exposed to 0.40 g/L lead acetate plus methionine and choline (1:1, 100 mg/kg), (5) group exposed to 0.40 g/L lead acetate plus methionine and choline (1:1, 400 mg/kg). In 8 weeks after exposure, all rats were killed. Then CREB mRNA and CaMK II mRNA expression levels in hippocampus were detected by real-time PCR, CREB and CaMK II protein expression levels in hippocampus were measured by western blot assay. The expression levels (0.743 ± 0.185 and 0.729 ± 0.199) of CaMKII mRNA and CREB mRNA in the hippocampus of lead group were significantly lower than those (0.950 ± 0.238 and 0.901 ± 0.232) of control group (P < 0.05), also the expression levels (0.271 ± 0.045 and 0.212 ± 0.058) of CREB protein and pCREB protein in the hippocampus of lead group were significantly lower than those (0.319 ± 0.058 and 0.506 ± 0.125) of control group (P < 0.05). The expression levels (1.014 ± 0.210 and 1.126 ± 0.379) of CaMKII mRNA and the expression levels (1.029 ± 0.335 and 0.932 ± 0.251) of CREB mRNA in the hippocampus of 2 groups exposed to lead acetate plus methionine and choline were significantly higher than those of lead group (P < 0.05). The expression levels (0.407 ± 0.951 and 0.563 ± 0.178) of CREB protein and pCREB protein in the hippocampus of group exposed to lead acetate plus 400 mg/kg methionine and choline were significantly higher than those of lead group (P < 0.05). Methionine and choline could decrease the inhibition effects of lead on the expression of CaMKII and CREB mRNA or CREB and pCREB proteins in the hippocampus of rats.

  1. Regulation of the steady state level of Fc gamma RI mRNA by IFN-gamma and dexamethasone in human monocytes, neutrophils, and U-937 cells.

    PubMed

    Pan, L Y; Mendel, D B; Zurlo, J; Guyre, P M

    1990-07-01

    The high affinity IgG FcR Fc gamma RI, CD64, plays important roles in the immune response. Fc gamma RI is predominantly expressed on monocytes and macrophages, and barely detectable on neutrophils. rIFN-gamma markedly increases the expression of Fc gamma RI on neutrophils, monocytes, macrophages and myeloid cell lines such as U-937, HL-60, and THP-1. Glucocorticoids inhibit the augmentation of Fc gamma RI expression by rIFN-gamma on neutrophils and myeloid cell lines, but enhance the augmentation of Fc gamma RI expression by rIFN-gamma on monocytes. In this study, we examined the effect of rIFN-gamma and dexamethasone (Dex) on the steady state level of Fc gamma RI mRNA in U-937 cells, neutrophils, and monocytes by hybridizing total RNA with the Fc gamma RI cDNA probe, p135. We found that the amount of Fc gamma RI mRNA increased within 1 h of treatment with rIFN-gamma in all three cell types. This initial induction of Fc gamma RI mRNA by rIFN-gamma was completely blocked by an inhibitor of RNA synthesis, actinomycin D, suggesting that the rIFN-gamma-mediated induction of Fc gamma RI mRNA is dependent on gene transcription. Dex, used in combination with rIFN-gamma, partially blocked the induction of Fc gamma RI mRNA by rIFN-gamma in U-937 cells and neutrophils, but caused a synergistic increase in Fc gamma RI mRNA levels in monocytes. The inhibitory effect of Dex on the steady state level of Fc gamma RI mRNA in U-937 cells was blocked by an inhibitor of protein synthesis, cycloheximide, suggesting that Dex-induced proteins were involved in the regulation of Fc gamma RI expression. This study indicates that the regulation of Fc gamma RI expression on U-937 cells, neutrophils, and monocytes by rIFN-gamma and Dex occurs, at least in part, at the mRNA level. rIFN-gamma increases the steady state level of Fc gamma RI mRNA through a common pathway among U-937 cells, neutrophils, and monocytes, whereas the effect of Dex on rIFN-gamma-induced Fc gamma RI mRNA is cell

  2. Salivary alpha amylase and cortisol levels in children with global developmental delay and their relation with the expectation of dental care and behavior during the intervention.

    PubMed

    dos Santos, Márcio José Possari; Bernabé, Daniel Galera; Nakamune, Ana Cláudia de Melo Stevanato; Perri, Silvia Helena Venturoli; de Aguiar, Sandra Maria Herondina Coelho Ávila; de Oliveira, Sandra Helena Penha

    2012-01-01

    The purpose of this study was to analyze the alpha-amylase (sAA) and cortisol levels in children with Global developmental delay (GDD) before and after dental treatment and its association with the children's behavior during treatment. The morning salivary cortisol levels and activity of sAA of 33 children with GDD were evaluated before and after dental treatment and were compared to 19 healthy children. The behavior of children with GDD during dental care was assessed by the Frankl scale. Children with GDD showed lower levels of sAA activity than healthy children, but this result was not significant. The salivary cortisol levels were similar between GDD and healthy children. GDD children showed increased levels of sAA (but not cortisol) prior to the dental treatment as compared to the post-treatment phase. GDD children who showed less favorable behavior during dental care had higher levels of sAA and salivary cortisol than GDD children with more favorable behavior, but only the sAA results were significant. In conclusion, GDD children show hyperactivity of the SNS-axis in anticipation of dental treatment which indicates the need for strategies to reduce their anxiety levels before and during dental care. Copyright © 2011 Elsevier Ltd. All rights reserved.

  3. Evaluation of the Trade Space Between UAS Maneuver Performance and SAA System Performance Requirements

    NASA Technical Reports Server (NTRS)

    Jack, Devin P.; Hoffler, Keith D.; Johnson, Sally C.

    2014-01-01

    A need exists to safely integrate Unmanned Aircraft Systems (UAS) into the National Airspace System. Replacing manned aircraft's see-and-avoid capability in the absence of an onboard pilot is one of the key challenges associated with safe integration. Sense-and-avoid (SAA) systems will have to achieve yet-to-be-determined required separation distances for a wide range of encounters. They will also need to account for the maneuver performance of the UAS they are paired with. The work described in this paper is aimed at developing an understanding of the trade space between UAS maneuver performance and SAA system performance requirements. An assessment of current manned and unmanned aircraft performance was used to establish potential UAS performance test matrix bounds. Then, nearterm UAS integration work was used to narrow down the scope. A simulator was developed with sufficient fidelity to assess SAA system performance requirements for a wide range of encounters. The simulator generates closest-point-of-approach (CPA) data from the wide range of UAS performance models maneuvering against a single intruder with various encounter geometries. The simulator is described herein and has both a graphical user interface and batch interface to support detailed analysis of individual UAS encounters and macro analysis of a very large set of UAS and encounter models, respectively. Results from the simulator using approximate performance data from a well-known manned aircraft is presented to provide insight into the problem and as verification and validation of the simulator. Analysis of climb, descent, and level turn maneuvers to avoid a collision is presented. Noting the diversity of backgrounds in the UAS community, a description of the UAS aerodynamic and propulsive design and performance parameters is included. Initial attempts to model the results made it clear that developing maneuver performance groups is required. Discussion of the performance groups developed and how

  4. High concentrate-induced subacute ruminal acidosis (SARA) increases plasma acute phase proteins (APPs) and cortisol in goats.

    PubMed

    Jia, Y Y; Wang, S Q; Ni, Y D; Zhang, Y S; Zhuang, S; Shen, X Z

    2014-09-01

    The aim of this study was to investigate changes of stress status in dairy goats induced to subacute ruminal acidosis (SARA). The level of acute phase proteins (APPs) including haptoglobin (HP) and serum amyloid A (SAA) in plasma and their mRNA expression in liver, as well as plasma cortisol and genes expression of key factors controlling cortisol synthesis in adrenal cortex were compared between SARA and control goats. SARA was induced by feeding high concentrate diet (60% concentrate of dry matter) for 3 weeks (SARA, n=6), while control goats (Con, n=6) received a low concentrate diet (40% concentrate of dry matter) during the experimental time. SARA goats showed ruminal pH below 5.8 for more than 3 h per day, which was significantly lower than control goats (pH>6.0). SARA goats demonstrated a significant increase of hepatic HP and SAA mRNA expression (P<0.05), and the level of HP but not SAA in plasma was markedly increased compared with control (P<0.05). The level of cortisol in plasma showed a trend to increase in SARA goats (0.05mRNA expression of 17α-hydroxylase cytochrome (P45017α ) (P<0.01) and 3β-hydroxysteroid dehydrogenase (3β-HSD) (P<0.05) was significantly increased in SARA goats. The contents of 3β-HSD and P450 side-chain cleavage protein were increased by 58.6% and 39.4%, respectively, but did not reach the statistical significance (P>0.05). These results suggested that SARA goats experienced a certain stress status, exhibiting an increase in HP production and cortisol secretion.

  5. The effects of running exercise on oxidative capacity and PGC-1α mRNA levels in the soleus muscle of rats with metabolic syndrome.

    PubMed

    Nagatomo, Fumiko; Fujino, Hidemi; Kondo, Hiroyo; Kouzaki, Motoki; Gu, Ning; Takeda, Isao; Tsuda, Kinsuke; Ishihara, Akihiko

    2012-03-01

    Skeletal muscles in animals with metabolic syndrome exhibit reduced oxidative capacity. We investigated the effects of running exercise on fiber characteristics, oxidative capacity, and mRNA levels in the soleus muscles of rats with metabolic syndrome [SHR/NDmcr-cp (cp/cp); CP]. We divided 5-week-old CP rats into non-exercise (CP) and exercise (CP-Ex) groups. Wistar-Kyoto rats (WKY) were used as the control group. CP-Ex rats were permitted voluntary exercise on running wheels for 10 weeks. Triglyceride levels were higher and adiponectin levels lower in the CP and CP-Ex groups than in the WKY group. However, triglyceride levels were lower and adiponectin levels higher in the CP-Ex group than in the CP group. The soleus muscles in CP-Ex rats contained only high-oxidative type I fibers, whereas those in WKY and CP rats contained type I, IIA, and IIC fibers. Muscle succinate dehydrogenase (SDH) activity was higher in the CP-Ex group than in the CP group; there was no difference in SDH activity between the WKY and CP-Ex groups. Muscle proliferator-activated receptor γ coactivator-1α (PGC-1α) mRNA levels were higher in the CP-Ex group than in the CP group; there was no difference in PGC-1α mRNA levels between the WKY and CP-Ex groups. In CP-Ex rats, longer running distance was associated with increased muscle SDH activity and PGC-1α mRNA levels. We concluded that running exercise restored decreased muscle oxidative capacity and PGC-1α mRNA levels and improved hypertriglyceridemia in rats with metabolic syndrome.

  6. Effect of lipoic acid on paraoxonase-1 and paraoxonase-3 protein levels, mRNA expression and arylesterase activity in liver hepatoma cells.

    PubMed

    Ozgun, Eray; Sayilan Ozgun, Gulben; Tabakcioglu, Kiymet; Suer Gokmen, Selma; Sut, Necdet; Eskiocak, Sevgi

    2017-10-01

    Paraoxonase-1 (PON1) and PON3 (PON3) are anti-atherosclerotic enzymes, synthesized primarily in liver and bound to HDL in circulation. The aim of the present study was to investigate the effects of therapeutic doses of lipoic acid on PON1 and PON3 protein levels, mRNA expression and arylesterase activity in liver. We treated HepG2 cells with 10, 40 and 200 μM lipoic acid for 72 h. Cell viability was evaluated by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide assay. PON1 and PON3 protein levels were measured by Western blotting, their mRNA expression was measured by quantitative PCR and arylesterase activity was measured spectrophotometrically. 200 µM lipoic acid caused a significant increase on PON1 and PON3 protein levels and arylesterase activity as compared with control, 10 µM and 40 µM lipoic acid-treated cells. 200 µM lipoic acid also caused a significant decrease on PON1 mRNA expression whereas on a significant increase PON3 mRNA expression as compared with control, 10 µM and 40 µM lipoic acid-treated cells. Our study showed that although lipoic acid up-regulates PON3 but down-regulates PON1 mRNA expression, it increases both PON1 and PON3 protein levels and arylesterase activity in HepG2 cells. We can report that lipoic acid may be useful for preventing atherosclerosis at therapeutic doses.

  7. TREM-1 SNP rs2234246 regulates TREM-1 protein and mRNA levels and is associated with plasma levels of L-selectin

    PubMed Central

    Aldasoro Arguinano, Alex-Ander; Dadé, Sébastien; Stathopoulou, Maria; Derive, Marc; Coumba Ndiaye, Ndeye; Xie, Ting; Masson, Christine; Gibot, Sébastien

    2017-01-01

    High levels of TREM-1 are associated with cardiovascular and inflammatory diseases risks and the most recent studies have showed that TREM-1 deletion or blockade is associated with up to 60% reduction of the development of atherosclerosis. So far, it is unknown whether the levels of TREM-1 protein are genetically regulated. Moreover, TREM family receptors have been suggested to regulate the cellular adhesion process. The goal of this study was to investigate whether polymorphisms within TREM-1 are regulating the variants of serum TREM-1 levels and the expression levels of their mRNA. Furthermore, we aimed to point out associations between polymorphisms on TREM-1 and blood levels of selectins. Among the 10 SNPs studied, the minor allele T of rs2234246, was associated with increased sTREM-1 in the discovery population (p-value = 0.003), explaining 33% of its variance, and with increased levels of mRNA (p-value = 0.007). The same allele was associated with increased soluble L-selectin levels (p-value = 0.011). The higher levels of sTREM-1 and L-selectin were confirmed in the replication population (p-value = 0.0007 and p-value = 0.018 respectively). We demonstrated for the first time one SNP on TREM-1, affecting its expression levels. These novel results, support the hypothesis that TREM-1 affects monocytes extravasation and accumulation processes leading to atherogenesis and atherosclerotic plaque progression, possibly through increased inflammation and subsequent higher expression of sL-selectin. PMID:28771614

  8. Prolonged exposure of chromaffin cells to nitric oxide down-regulates the activity of soluble guanylyl cyclase and corresponding mRNA and protein levels

    PubMed Central

    Ferrero, Rut; Torres, Magdalena

    2002-01-01

    Background Soluble guanylyl cyclase (sGC) is the main receptor for nitric oxide (NO) when the latter is produced at low concentrations. This enzyme exists mainly as a heterodimer consisting of one α and one β subunit and converts GTP to the second intracellular messenger cGMP. In turn, cGMP plays a key role in regulating several physiological processes in the nervous system. The aim of the present study was to explore the effects of a NO donor on sGC activity and its protein and subunit mRNA levels in a neural cell model. Results Continuous exposure of bovine adrenal chromaffin cells in culture to the nitric oxide donor, diethylenetriamine NONOate (DETA/NO), resulted in a lower capacity of the cells to synthesize cGMP in response to a subsequent NO stimulus. This effect was not prevented by an increase of intracellular reduced glutathione level. DETA/NO treatment decreased sGC subunit mRNA and β1 subunit protein levels. Both sGC activity and β1 subunit levels decreased more rapidly in chromaffin cells exposed to NO than in cells exposed to the protein synthesis inhibitor, cycloheximide, suggesting that NO decreases β1 subunit stability. The presence of cGMP-dependent protein kinase (PKG) inhibitors effectively prevented the DETA/NO-induced down regulation of sGC subunit mRNA and partially inhibited the reduction in β1 subunits. Conclusions These results suggest that activation of PKG mediates the drop in sGC subunit mRNA levels, and that NO down-regulates sGC activity by decreasing subunit mRNA levels through a cGMP-dependent mechanism, and by reducing β1 subunit stability. PMID:12350235

  9. Principles of mRNA transport in yeast.

    PubMed

    Heym, Roland Gerhard; Niessing, Dierk

    2012-06-01

    mRNA localization and localized translation is a common mechanism by which cellular asymmetry is achieved. In higher eukaryotes the mRNA transport machinery is required for such diverse processes as stem cell division and neuronal plasticity. Because mRNA localization in metazoans is highly complex, studies at the molecular level have proven to be cumbersome. However, active mRNA transport has also been reported in fungi including Saccharomyces cerevisiae, Ustilago maydis and Candida albicans, in which these events are less difficult to study. Amongst them, budding yeast S. cerevisiae has yielded mechanistic insights that exceed our understanding of other mRNA localization events to date. In contrast to most reviews, we refrain here from summarizing mRNA localization events from different organisms. Instead we give an in-depth account of ASH1 mRNA localization in budding yeast. This approach is particularly suited to providing a more holistic view of the interconnection between the individual steps of mRNA localization, from transcriptional events to cytoplasmic mRNA transport and localized translation. Because of our advanced mechanistic understanding of mRNA localization in yeast, the present review may also be informative for scientists working, for example, on mRNA localization in embryogenesis or in neurons.

  10. Complex mutual regulation of facilitates chromatin transcription (FACT) subunits on both mRNA and protein levels in human cells.

    PubMed

    Safina, Alfiya; Garcia, Henry; Commane, Mairead; Guryanova, Olga; Degan, Seamus; Kolesnikova, Kateryna; Gurova, Katerina V

    2013-08-01

    Facilitates chromatin transcription (FACT) is a chromatin remodeling complex with two subunits: SSRP1 and SPT16. Mechanisms controlling FACT levels are of interest, since the complex is not expressed in most differentiated cells, but is frequently upregulated in cancer, particularly in poorly differentiated, aggressive tumors. Moreover, inhibition of FACT expression or function in tumor cells interferes with their survival. Here we demonstrate that SSRP1 and SPT16 protein levels decline upon induction of cellular differentiation or senescence in vitro and that similar declines in protein levels for both SSRP1 and SPT16 occur upon RNAi-mediated knockdown of either SSRP1 or SPT16. The interdependence of SSRP1 and SPT16 protein levels was found to be due to their association with SSRP1 and SPT16 mRNAs, which stabilizes the proteins. In particular, presence of SSRP1 mRNA is critical for SPT16 protein stability. In addition, binding of SSRP1 and SPT16 mRNAs to the FACT complex increases the stability and efficiency of translation of the mRNAs. These data support a model in which the FACT complex is stable when SSRP1 mRNA is present, but quickly degrades when SSRP1 mRNA levels drop. In the absence of FACT complex, SSRP1 and SPT16 mRNAs are unstable and inefficiently translated, making reactivation of FACT function unlikely in normal cells. Thus, we have described a complex and unusual mode of regulation controlling cellular FACT levels that results in amplified and stringent control of FACT activity. The FACT dependence of tumor cells suggests that mechanisms controlling FACT levels could be targeted for anticancer therapy.

  11. Association between ERCC1 and TS mRNA levels and disease free survival in colorectal cancer patients receiving oxaliplatin and fluorouracil (5-FU) adjuvant chemotherapy.

    PubMed

    Li, Sheng; Zhu, Liangjun; Yao, Li; Xia, Lei; Pan, Liangxi

    2014-08-29

    Aim was to explore the association of ERCC1 and TS mRNA levels with the disease free survival (DFS) in Chinese colorectal cancer (CRC) patients receiving oxaliplatin and 5-FU based adjuvant chemotherapy. Total 112 Chinese stage II-III CRC patients were respectively treated by four different chemotherapy regimens after curative operation. The TS and ERCC1 mRNA levels in primary tumor were measured by real-time RT-PCR. Kaplan-Meier curves and log-rank tests were used for DFS analysis. The Cox proportional hazards model was used for prognostic analysis. In univariate analysis, the hazard ratio (HR) for the mRNA expression levels of TS and ERCC1 (logTS: HR = 0.820, 95% CI = 0.600 - 1.117, P = 0.210; logERCC1: HR = 1.054, 95% CI = 0.852 - 1.304, P = 0.638) indicated no significant association of DFS with the TS and ERCC1 mRNA levels. In multivariate analyses, tumor stage (IIIc: reference, P = 0.083; IIb: HR = 0.240, 95% CI = 0.080 - 0.724, P = 0.011; IIc: HR < 0.0001, P = 0.977; IIIa: HR = 0.179, 95% CI = 0.012 - 2.593, P = 0.207) was confirmed to be the independent prognostic factor for DFS. Moreover, the Kaplan-Meier DFS curves showed that TS and ERCC1 mRNA levels were not significantly associated with the DFS (TS: P = 0.264; ERCC1: P = 0.484). The mRNA expression of ERCC1 and TS were not applicable to predict the DFS of Chinese stage II-III CRC patients receiving 5-FU and oxaliplatin based adjuvant chemotherapy.

  12. GreenIT Service Level Agreements

    NASA Astrophysics Data System (ADS)

    von Laszewski, Gregor; Wang, Lizhe

    In this paper we are introducing a framework towards the inclusion of Green IT metrics as part of service level agreements for future Grids and Clouds. As part of this effort we need to revisit Green IT metrics and proxies that we consider optimizing against in order to develop GreenIT as a Services (GaaS) that can be reused as part of a Software as a Service (SaaS) and Infrastructure Infrastructureas a service (IaaS) framework. We report on some of our ongoing efforts and demonstrate how we already achieve impact on the environment with our services.

  13. Macrophages from Behcet's Disease Patients Express Decreased Level of Aryl Hydrocarbon Receptor (AHR) mRNA.

    PubMed

    Palizgir, Mohammad Taghi; Akhtari, Maryam; Mahmoudi, Mahdi; Mostafaei, Shayan; Rezaeimanesh, Alireza; Akhlaghi, Massoomeh; Shahram, Farhad

    2017-10-01

    Aryl hydrocarbon receptor (AHR) is a ligand-activated transcription factor, connecting environmental stimulators with the immune system. M1 macrophages are a part of immune system that contribute to the inflammatory events in the pathogenesis of Behcet's disease (BD). The effect of AHR on the macrophages in BD patients is still unclear. In this study, we investigated the mRNA expression of AHR in the monocyte-derived and M1 macrophages in active BD patients in comparison to healthy controls. Isolated monocytes from 10 healthy controls and 10 active BD patients were differentiated to macrophages by macrophage-colony stimulating factor (M-CSF) for 7 days. Cells were then polarized to M1 macrophages by lipopolysaccharide (LPS) and interferon-γ (IFNγ) for 24h. Monocyte purity and macrophage markers expression were analyzed by flow cytometry. Analysis of AHR mRNA expression was performed by SYBR Green real-time PCR. Our results showed that AHR expression is significantly down-regulated in M1 macrophages compare to monocyte-derived macrophages. It was shown that both monocyte-derived macrophages and M1 macrophages from BD patients significantly express lower level of AHR mRNA compared to healthy individuals. Our results demonstrate an anti-inflammatory role for AHR in macrophages, which suggest that decreased AHR expression is associated with pro-inflammatory M1 macrophage and BD susceptibility.

  14. Fibroblast growth factor-21 and omentin-1 hepatic mRNA expression and serum levels in morbidly obese women with non-alcoholic fatty liver disease.

    PubMed

    Waluga, M; Kukla, M; Zorniak, M; Kajor, M; Liszka, L; Dyaczynski, M; Kowalski, G; Zadlo, D; Waluga, E; Olczyk, P; Buldak, R J; Berdowska, A; Hartleb, M

    2017-06-01

    Fibroblast growth factor-21 (FGF21) and omentin-1 have been recognized as potent antidiabetic agents with potential hepatoprotective activity. The aim of this study was to evaluate hepatic FGF21 and omentin-1 mRNA expression as well as their serum levels as predictive markers of liver injury and insulin resistance in morbidly obese women with non-alcoholic fatty liver disease (NAFLD). This study included 56 severely obese women who underwent intraoperative wedge liver biopsy during the bariatric surgery. Hepatic FGF21 and omentin-1 mRNA were assessed by quantitative real-time PCR, while their serum concentrations were measured with commercially available enzyme-linked immunosorbent assays. The FGF21 serum level was significantly higher in patients with a greater extent of steatosis (grade 2 and 3) compared to those without or with mild steatosis (grade 0 and 1) (P = 0.049). Receiver Operating Characteristic analysis, however, showed poor discriminant power for the FGF21 serum levels in differentiating between more and less extensive steatosis with an AUC = 0.666. There was a tendency towards higher levels of hepatic FGF21 mRNA in patients with lobular inflammation and fibrosis and towards lower levels in the case of hepatocyte ballooning and steatosis. There was a positive mutual correlation between hepatic FGF21 and omentin-1 mRNA levels (r = 0.78; P < 0.001). Fibrosis stage was associated with serum glucose and homeostatic model assessment for insulin resistance (HOMA-IR) (P = 0.03 and P = 0.02, respectively). Serum omentin-1 was not associated with histopathological features. The hepatic omentin-1 mRNA levels showed a tendency to be lower in patients with advanced steatosis and hepatocyte ballooning. In conclusion, our study, which focused on hepatic FGF21 and omentin-1 mRNA expression, confirmed marked expression of both molecules in the liver of morbidly obese patients with NAFLD. More extensive steatosis was associated with evident changes in the serum FGF21

  15. Macrophage colony-stimulating factor accelerates wound healing and upregulates TGF-beta1 mRNA levels through tissue macrophages.

    PubMed

    Wu, L; Yu, Y L; Galiano, R D; Roth, S I; Mustoe, T A

    1997-10-01

    Macrophage colony-stimulating factor (M-CSF) is produced by many cell types involved in wound repair, yet it acts specifically on monocytes and macrophages. The monocyte-derived cell is thought to be important in wound healing, but the importance of the role of tissue macrophages in wound healing has not been well defined. Dermal ulcers were created in normal and ischemic ears of young rabbits. Either rhM-CSF (17 microg/wound) or buffer was applied to each wound. Wounds were bisected and analyzed histologically at Days 7 and 10 postwounding. The amounts of epithelial growth and granulation tissue deposition were measured in all wounds. The level of increase of TGF-beta1 mRNA level in M-CSF-treated wounds was examined using competitive RT-PCR. M-CSF increased new granulation tissue formation by 37% (N = 21, P < 0.01) and 50% (P < 0.01) after single and multiple treatments, respectively, in nonischemic wounds. TGF-beta1 mRNA levels in rhM-CSF-treated wounds increased 5.01-fold (N = 8) over vehicle-treated wounds under nonischemic conditions. In contrast, no effect could be detected in ischemic wounds treated with rhM-CSF, and these wounds only showed a 1.66-fold increase in TGF-beta1 mRNA levels when compared to ischemic wounds treated with vehicle alone. GAPDH, a housekeeping gene, showed no change. As mesenchymal cells lack receptors for M-CSF, the improved healing of wounds treated with topical rhM-CSF must reflect a generalized enhancement of activation and function of tissue macrophages, as demonstrated by upregulation of TGF-beta. The lack of effect under ischemic conditions suggests that either macrophage activity and/or response to M-CSF is adversely affected under those conditions; this may suggest the pathogenesis of impaired wound healing at the cellular level. Copyright 1997 Academic Press.

  16. Comparison of Tight Junction Protein-Related Gene mRNA Expression Levels between Male and Female Gastroesophageal Reflux Disease Patients.

    PubMed

    Kim, Jin Joo; Kim, Nayoung; Park, Ji Hyun; Kim, Young Sun; Lee, Sun Min; Lee, Dong Ho; Jung, Hyun Chae

    2018-03-21

    Male predominance has been observed in the erosive reflux disease (ERD), but reverse finding in nonerosive reflux disease (NERD). This suggests sex-specific medicine approach is needed but its mechanism is remained to be elucidated. We aimed to compare clinical characteristics and mRNA expression levels of tight junction-related proteins between male and female gastroesophageal reflux disease (GERD) patients. Sixteen healthy controls, 45 ERD, and 14 NERD patients received upper endoscopies and completed questionnaires. Quantitative real-time polymerase chain reactions (qPCR) of occludin (OCLN), zonal occludens (ZO) 1, claudin1 (CDLN1) and claudin4 (CDLN4), and neurokinin 1 receptor (NK1R) were performed in the distal esophageal mucosal specimen. These results were analyzed by sex. Female GERD patients were affected more by reflux symptoms than males. The impairment of overall QoL was more prominent in female patients with reflux symptoms than male patients (5.6±0.2 vs. 4.9±0.6, p=0.009). The levels of OCLN mRNA expression were significantly lower in the male ERD group. On the other hand, those of CLDN1, CLDN4, and NK1R except ZO-1 were significantly higher in the male ERD group. We demonstrated that female ERD/NERD patients were affected more by GERD and male ERD patients showed significant changes of tight junction protein mRNA expression levels.

  17. Changes in mRNA levels for brain-derived neurotrophic factor after wheel running in rats selectively bred for high- and low-aerobic capacity

    PubMed Central

    Groves-Chapman, Jessica L.; Murray, Patrick S.; Stevens, Kristin L.; Monroe, Derek; Koch, Lauren G.; Britton, Steven L.; Holmes, Philip V.

    2012-01-01

    We evaluated levels of exercise-induced brain-derived neurotrophic factor (BDNF) messenger RNA (mRNA) within the hippocampal formation in rats selectively bred for 1) high intrinsic (i.e., untrained) aerobic capacity (High Capacity Runners, HCR), 2) low intrinsic aerobic capacity (Low Capacity Runners, LCR), and 3) unselected Sprague-Dawley (SD) rats with or without free access to running wheels for three weeks. The specific aim of the study was to determine whether a dose-response relationship exists between cumulative running distance and levels of BDNF mRNA. No additional treatments or behavioral manipulations were used. HCR, LCR, and SD rats were grouped by strain and randomly assigned to sedentary or activity (voluntary access to activity wheel) conditions. Animals were killed after 21 days of exposure to the assigned conditions. Daily running distances (mean ± standard deviation meters/d) during week three were: HCR (4726 ± 3220), SD (2293 ± 3461), LCR (672 ± 323). Regardless of strain, levels of BDNF mRNA in CA1 were elevated in wheel runners compared to sedentary rats and this difference persisted after adjustment for age (p=0.040). BDNF mRNA was not affected by intrinsic aerobic capacity and was not related to total running distance. The results support that BDNF mRNA expression is increased by unlimited access to activity wheel running for 3 weeks but is not dependent upon accumulated running distance. PMID:22024546

  18. Efficacy of Omega Fatty Acid Supplementation on mRNA Expression Level of Tumor Necrosis Factor Alpha in Patients with Gastric Adenocarcinoma.

    PubMed

    Hosseinzadeh, Asghar; Ardebili, Seyed Mojtaba Mohaddes

    2016-09-01

    Tumor necrosis factor alpha (TNF-α), a multifunctional cytokine, is involved in apoptosis, cell proliferation, cell survival, and inflammation. It plays a dual role in cancer development and progression. It has been revealed that polyunsaturated fatty acids (PUFAs) modulate the production and activity of TNF family cytokines. The objective of the present study was to evaluate the effect of PUFAs on messenger RNA expression levels of TNF-α in patients with gastric adenocarcinoma. Thirty-four chemotherapy-naive patients diagnosed with gastric adenocarcinoma were randomly divided into two groups. The first group (17 individuals) received cisplatin without supplements and the second group (17 individuals) received cisplatin plus orally administered PUFA supplements for 3 weeks, based on treatment strategies. The gastric biopsy samples were obtained from all participants before and after treatment, and TNF-α mRNA expression levels were evaluated by quantitative real-time PCR procedure. Our findings revealed that TNF-α mRNA expression is downregulated in group II, after receiving cisplatin and omega fatty acid supplement for 3 weeks. However, this difference is not statistically significant (p > 0.05). TNF-α mRNA expression did not show significant alteration in group I, after receiving cisplatin alone. Taken together, we concluded that omega fatty acids reduce TNF-α expression at the mRNA level in patients with gastric adenocarcinoma. These data suggest that TNF-α may act as a potential target for the therapy of human gastric adenocarcinoma.

  19. High Intensity High Volume Interval Training Improves Endurance Performance and Induces a Nearly Complete Slow-to-Fast Fiber Transformation on the mRNA Level.

    PubMed

    Eigendorf, Julian; May, Marcus; Friedrich, Jan; Engeli, Stefan; Maassen, Norbert; Gros, Gerolf; Meissner, Joachim D

    2018-01-01

    We present here a longitudinal study determining the effects of two 3 week-periods of high intensity high volume interval training (HIHVT) (90 intervals of 6 s cycling at 250% maximum power, P max /24 s) on a cycle ergometer. HIHVT was evaluated by comparing performance tests before and after the entire training (baseline, BSL, and endpoint, END) and between the two training sets (intermediate, INT). The mRNA expression levels of myosin heavy chain (MHC) isoforms and markers of energy metabolism were analyzed in M. vastus lateralis biopsies by quantitative real-time PCR. In incremental tests peak power (P peak ) was increased, whereas V ˙ O 2peak was unaltered. Prolonged time-to-exhaustion was found in endurance tests with 65 and 80% P max at INT and END. No changes in blood levels of lipid metabolites were detected. Training-induced decreases of hematocrit indicate hypervolemia. A shift from slow MHCI/β to fast MHCIIa mRNA expression occurred after the first and second training set. The mRNA expression of peroxisome proliferator-activated receptor gamma coactivator 1α (PGC-1α), a master regulator of oxidative energy metabolism, decreased after the second training set. In agreement, a significant decrease was also found for citrate synthase mRNA after the second training set, indicating reduced oxidative capacity. However, mRNA expression levels of glycolytic marker enzyme glyceraldehyde-3-phosphate dehydrogenase did not change after the first and second training set. HIHVT induced a nearly complete slow-to-fast fiber type transformation on the mRNA level, which, however, cannot account for the improvements of performance parameters. The latter might be explained by the well-known effects of hypervolemia on exercise performance.

  20. The Nucleotide Sequence and Spliced pol mRNA Levels of the Nonprimate Spumavirus Bovine Foamy Virus

    PubMed Central

    Holzschu, Donald L.; Delaney, Mari A.; Renshaw, Randall W.; Casey, James W.

    1998-01-01

    We have determined the complete nucleotide sequence of a replication-competent clone of bovine foamy virus (BFV) and have quantitated the amount of splice pol mRNA processed early in infection. The 544-amino-acid Gag protein precursor has little sequence similarity with its primate foamy virus homologs, but the putative nucleocapsid (NC) protein, like the primate NCs, contains the three glycine-arginine-rich regions that are postulated to bind genomic RNA during virion assembly. The BFV gag and pol open reading frames overlap, with pro and pol in the same translational frame. As with the human foamy virus (HFV) and feline foamy virus, we have detected a spliced pol mRNA by PCR. Quantitatively, this mRNA approximates the level of full-length genomic RNA early in infection. The integrase (IN) domain of reverse transcriptase does not contain the canonical HH-CC zinc finger motif present in all characterized retroviral INs, but it does contain a nearby histidine residue that could conceivably participate as a member of the zinc finger. The env gene encodes a protein that is over 40% identical in sequence to the HFV Env. By comparison, the Gag precursor of BFV is predicted to be only 28% identical to the HFV protein. PMID:9499074

  1. Low BMI is correlated with increased TGF-β and IL-10 mRNA levels in the peripheral blood of breast cancer patients.

    PubMed

    Liu, Chao; Wang, Qian; Sun, Bing; Meng, Xiangying; Li, Lan; Yang, Liuchun; Cong, Yang; Liu, Jiannan; Xuan, Liang; Huang, Yan; Wu, Shikai

    2018-03-01

    Transforming growth factor-β (TGF-β), interleukin-10 (IL-10), and forkhead box P3 (Foxp3) have important roles in breast cancer development. Previous studies confirmed a correlation between these immune molecules and tumor characteristics, but their association with nutritional status in breast cancer is largely unknown. We aimed to investigate the association between body mass index (BMI), hemoglobin, total protein, albumin, globulin (GLB), albumin/GLB ratio (AGR), pre-albumin, prognostic nutritional index, and TGF-β, IL-10, and Foxp3 mRNA expression in patients with breast cancer. Quantitative real-time PCR was used to detect the mRNA expression of TGF-β, IL-10, and Foxp3 in the peripheral blood of 107 patients with breast cancer and 21 healthy controls. We found that TGF-β mRNA levels were 2.6-fold, 3.2-fold, and 2.3-fold higher in patients with low BMI (<23), low AGR, and high GLB, respectively, than in their counterparts (P < 0.05). In addition, IL-10 mRNA expression levels in patients with normal BMI (<23) were 2.8-fold and 3.5-fold higher than in those who were overweight (23≤ BMI <25) and obese (BMI ≥ 25), respectively (P < 0.05). In addition, TGF-β, IL-10, and Foxp3 mRNA levels were significantly higher in patients with breast cancer than in healthy controls (P < 0.05). In summary, our results suggest that nutritional status, especially BMI, may strongly affect systematic immune function in patients with breast cancer. © 2018 IUBMB Life, 70(3):237-245, 2018. © 2018 International Union of Biochemistry and Molecular Biology.

  2. CYP3A5 mRNA degradation by nonsense-mediated mRNA decay.

    PubMed

    Busi, Florent; Cresteil, Thierry

    2005-09-01

    The total CYP3A5 mRNA level is significantly greater in carriers of the CYP3A5*1 allele than in CYP3A5*3 homozygotes. Most of the CYP3A5*3 mRNA includes an intronic sequence (exon 3B) containing premature termination codons (PTCs) between exons 3 and 4. Two models were used to investigate the degradation of CYP3A5 mRNA: a CYP3A5 minigene consisting of CYP3A5 exons and introns 3 to 6 transfected into MCF7 cells, and the endogenous CYP3A5 gene expressed in HepG2 cells. The 3'-untranslated region g.31611C>T mutation has no effect on CYP3A5 mRNA decay. Splice variants containing exon 3B were more unstable than wild-type (wt) CYP3A5 mRNA. Cycloheximide prevents the recognition of PTCs by ribosomes: in transfected MCF7 and HepG2 cells, cycloheximide slowed down the degradation of exon 3B-containing splice variants, suggesting the participation of nonsense-mediated decay (NMD). When PTCs were removed from pseudoexon 3B or when UPF1 small interfering RNA was used to impair the NMD mechanism, the decay of the splice variant was reduced, confirming the involvement of NMD in the degradation of CYP3A5 splice variants. Induction could represent a source of variability for CYP3A5 expression and could modify the proportion of splice variants. The extent of CYP3A5 induction was investigated after exposure to barbiturates or steroids: CYP3A4 was markedly induced in a pediatric population compared with untreated neonates. However, no effect could be detected in either the total CYP3A5 RNA, the proportion of splice variant RNA, or the protein level. Therefore, in these carriers, induction is unlikely to switch on the phenotypic CYP3A5 expression in carriers of CYP3A5*3/*3.

  3. Gravitational loading of a simulated launch alters mRNA expression in osteoblasts

    NASA Technical Reports Server (NTRS)

    Fitzgerald, J.; Hughes-Fulford, M.

    1996-01-01

    Serum-deprived mouse osteoblastic cells (MC3T3-E1a) were centrifuged under a regime designed to simulate a space shuttle launch (maximum of 3g). Messenger RNA levels for eight genes involved in bone growth and maintenance were determined using RT-PCR. Following 30 min of centrifugation, mRNA level for early response gene c-fos was significantly increased 89% (P < 0.05). The c-fos induction was transient and returned to control levels after 3 h. The mRNA level for the mineralization marker gene osteocalcin was significantly decreased to 44% of control level (P < 0.005) 3 h after centrifugation. No changes in mRNA levels were detected for c-myc, TGFbeta1, TGFbeta2, cyclophilin A, or actin. No basal mRNA level for TGFbeta3 was detected. In addition, no change in the steady-state synthesis of prostaglandin E2 was detected, possibly due to lack of lipid substrates in serum-deprived cells, suggesting that the increase in c-fos mRNA in response to gravitational loading is a result of mechanical stimulation. These results indicate that a small magnitude mechanical loading, such as that experienced during a shuttle launch, can alter mRNA levels in quiescent osteoblastic cells.

  4. Primary induction of vitellogenin mRNA in the rooster by 17beta-estradiol.

    PubMed Central

    Burns, A T; Deeley, R G; Gordon, J I; Udell, D S; Mullinix, K P; Goldberger, R F

    1978-01-01

    We have studied the kinetics of vitellogenin mRNA accumulation in rooster liver after a primary injection of 17beta-estradiol. The levels of vitellogenin mRNA have been determined both by hybridization of total cellular RNA to vitellogenin cDNA and by translation of vitellogenin mRNA in a wheat germ cell-free system. The results obtained by both methods of analysis are in good agreement and indicate that vitellogenin mRNA is present in the liver of normal roosters at a level of 0-5 molecules per liver cell and increases in amount during the 3 days following injection of estrogen, reaching a level of almost 6000 molecules per cell at the peak of the response. The level of vitellogenin mRNA declined exponentially during the next 14 days with a half-life of 29 hr, reaching a level of less than 10 molecules per cell at 17 days after injection of the hormone. The levels of vitellogenin mRNA after stimulation with estrogen have been correlated with the in vivo rate of synthesis of the vitellogenin polypeptide. The results indicate that the rate of vitellogenin synthesis is closely correlated with the level of vitellogenin mRNA. On the basis of these findings, we conclude that vitellogenin mRNA does not exist in the liver in an untranslated form after withdrawal from estrogen. PMID:273910

  5. RNA/DNA ratio and LPL and MyoD mRNA expressions in muscle of Oreochromis niloticus fed with elevated levels of palm oil

    NASA Astrophysics Data System (ADS)

    Ayisi, Christian Larbi; Zhao, Jinliang

    2016-02-01

    Palm oil is of great potential as one of the sustainable alternatives to fish oil (FO) in aquafeeds. In this present study, five isonitrogenous diets (32% crude protein) with elevated palm oil levels of 0%, 2%, 4%, 6% and 8% were used during an 8-week feeding trial to evaluate its effects on RNA/DNA ratio and lipoprotein lipase (LPL) and MyoD mRNA expressions in muscle of Oreochromis niloticus. The results showed that RNA, DNA content as well as ratio of RNA to DNA were significantly affected ( P < 0.05), in each case the highest was recorded in fish group subjected to 6% palm oil level. There was a strong positive correlation between nucleic acid concentration (RNA concentration and RNA: DNA ratio) and specific growth rate (SGR), protein efficiency ratio (PER), while a negative correlation existed between nucleic acid concentration (RNA concentration and RNA: DNA ratio) and feed conversion ratio (FCR). The mRNA expressions of LPL and MyoD in muscle were not significantly affected by the different palm oil levels, although the highest expression was observed in fish fed with 6% palm oil level. There also existed a strong positive correlation between the mRNA expression of LPL, MyoD and SGR, PER, while their correlation with FCR was negative. In conclusion, elevated palm oil affected the RNA, DNA concentration as well as RNA/DNA ratio significantly, although the mRNA expression of LPL and MyoD were not affected significantly by elevated palm oil levels.

  6. Expression of beta 3-adrenoceptor mRNA in rat tissues.

    PubMed

    Evans, B A; Papaioannou, M; Bonazzi, V R; Summers, R J

    1996-01-01

    1. This study examines the expression of beta 3-adrenoceptor messenger RNA (beta 3-AR mRNA) in rat tissues to allow comparison with atypical beta-adrenoceptors determined by functional and radioligand binding techniques. 2. A reverse transcription/polymerase chain reaction protocol has been developed for determining the relative amounts of beta 3-AR mRNA in rat tissues. 3. Measurement of adipsin and uncoupling protein (UCP) mRNA was used to examine all tissues for the presence of white and brown adipose tissue which may contribute beta 3-AR mRNA. 4. The beta 3-AR mRNA is expressed at high levels in brown and white adipose tissue, stomach fundus, the longitudinal/circular smooth muscle of both colon and ileum, and colon submucosa. There was substantial expression of adipsin in colon submucosa and moderate expression in fundus, suggesting that in these regions at least some of the beta 3-AR signal may be contributed by fat. Pylorus and colon mucosa showed moderate levels of beta 3-AR mRNA with lower levels of adipsin. Ileum mucosa and submucosa showed low but readily detectable levels of beta 3-AR. 5. Expression of adipsin in rat skeletal muscles coupled to very low levels of beta 3-AR mRNA indicates that the observed beta 3-AR may be due to the presence of intrinsic fat. beta 3-AR mRNA was virtually undetectable in heart, lung and liver. These results raise the possibility that the atypical beta-AR demonstrated by functional and/or binding studies in muscle and in heart is not the beta 3-AR. 6. By use of two different sets of primers for amplification of beta 3-AR cDNA, no evidence was found for differential splicing of the mRNA in any of the tissues examined. 7. The detection of beta 3-AR mRNA in the gut mucosa and submucosa suggests that in addition to its established roles in lipolysis, thermogenesis and regulation of gut motility beta 3-AR may subserve other functions in the gastrointestinal tract. The absence of beta 3-AR mRNA in rat heart or its presence with

  7. Geomagnetic South Atlantic Anomaly and global sea level rise: A direct connection?

    NASA Astrophysics Data System (ADS)

    de Santis, A.; Qamili, E.; Spada, G.; Gasperini, P.

    2012-01-01

    We highlight the existence of an intriguing and to date unreported relationship between the surface area of the South Atlantic Anomaly (SAA) of the geomagnetic field and the current trend in global sea level rise. These two geophysical variables have been growing coherently during the last three centuries, thus strongly suggesting a causal relationship supported by some statistical tests. The monotonic increase of the SAA surface area since 1600 may have been associated with an increased inflow of radiation energy through the inner Van Allen belt with a consequent warming of the Earth's atmosphere and finally global sea level rise. An alternative suggestive and original explanation is also offered, in which pressure changes at the core-mantle boundary cause surface deformations and relative sea level variations. Although we cannot establish a clear connection between SAA dynamics and global warming, the strong correlation between the former and global sea level supports the idea that global warming may be at least partly controlled by deep Earth processes triggering geomagnetic phenomena, such as the South Atlantic Anomaly, on a century time scale.

  8. Correlation of Cyfra 21-1 levels in saliva and serum with CK19 mRNA expression in oral squamous cell carcinoma.

    PubMed

    Malhotra, Rewa; Urs, Aadithya B; Chakravarti, Anita; Kumar, Suman; Gupta, V K; Mahajan, Bhawna

    2016-07-01

    Oral squamous cell carcinoma (OSCC) accounts for 90 % of malignant lesions of oral cavity. The study assessed the potential of Cyfra 21-1 as a tumor marker in OSCC. The study included 50 patients of OSCC to evaluate levels of Cyfra 21-1 in serum and saliva by electrochemiluminescent immunoassay (ECLIA) and CK19 messenger RNA (mRNA) expression in tissue by florescent quantitative real-time reverse transcriptase polymerase chain reaction (RT-PCR) along with healthy individuals as control. The salivary and serum Cyfra 21-1 levels in patients of OSCC were significantly higher compared to controls (p value < 0.01). There was a 2.75-fold increase in CK19 mRNA expression in OSCC cases compared to controls. A significant positive correlation was found between serum and salivary Cyfra 21-1, serum Cyfra 21-1, and CK19 mRNA expression and between salivary Cyfra 21-1 and CK19 mRNA expression. Among these, correlation between serum and salivary Cyfra 21-1 was highly significant. Salivary and serum Cyfra 21-1 showed significantly elevated levels in grade II OSCC compared to grade I histopathologically. Elevated levels of salivary Cyfra 21-1 were associated with recurrence in OSCC patients. Reverse operating curve constructed using 3 ng/ml as a cutoff for serum Cyfra 21-1 revealed the sensitivity and specificity to be 88 and 78.2 %, respectively. Using a cutoff value of 8.5 ng/ml for salivary Cyfra 21-1, the sensitivity was found to be 93.8 % and specificity 84.3 %. We advocate salivary Cyfra 21-1 as a better diagnostic marker over serum Cyfra 21-1 as well as a potential marker in the prognosis of OSCC.

  9. Serum amyloid A: an ozone-induced circulating factor with potentially important functions in the lung-brain axis.

    PubMed

    Erickson, Michelle A; Jude, Joseph; Zhao, Hengjiang; Rhea, Elizabeth M; Salameh, Therese S; Jester, William; Pu, Shelley; Harrowitz, Jenna; Nguyen, Ngan; Banks, William A; Panettieri, Reynold A; Jordan-Sciutto, Kelly L

    2017-09-01

    Accumulating evidence suggests that O 3 exposure may contribute to CNS dysfunction. Here, we posit that inflammatory and acute-phase proteins in the circulation increase after O 3 exposure and systemically convey signals of O 3 exposure to the CNS. To model acute O 3 exposure, female Balb/c mice were exposed to 3 ppm O 3 or forced air for 2 h and were studied after 6 or 24 h. Of 23 cytokines and chemokines, only KC/CXCL1 was increased in blood 6 h after O 3 exposure. The acute-phase protein serum amyloid A (A-SAA) was significantly increased by 24 h, whereas C-reactive protein was unchanged. A-SAA in blood correlated with total leukocytes, macrophages, and neutrophils in bronchoalveolar lavage from O 3 -exposed mice. A-SAA mRNA and protein were increased in the liver. We found that both isoforms of A-SAA completely crossed the intact blood-brain barrier, although the rate of SAA2.1 influx was approximately 5 times faster than that of SAA1.1. Finally, A-SAA protein, but not mRNA, was increased in the CNS 24 h post-O 3 exposure. Our findings suggest that A-SAA is functionally linked to pulmonary inflammation in our O 3 exposure model and that A-SAA could be an important systemic signal of O 3 exposure to the CNS.-Erickson, M. A., Jude, J., Zhao, H., Rhea, E. M., Salameh, T. S., Jester, W., Pu, S., Harrowitz, J., Nguyen, N., Banks, W. A., Panettieri, R. A., Jr., Jordan-Sciutto, K. L. Serum amyloid A: an ozone-induced circulating factor with potentially important functions in the lung-brain axis. © FASEB.

  10. Gestational Protein Restriction Increases Cardiac Connexin 43 mRNA levels in male adult rat offspring

    PubMed Central

    Rossini, Kamila Fernanda; de Oliveira, Camila Andrea; Rebelato, Hércules Jonas; Esquisatto, Marcelo Augusto Marreto; Catisti, Rosana

    2017-01-01

    Background The dietary limitation during pregnancy influences the growth and development of the fetus and offspring and their health into adult life. The mechanisms underlying the adverse effects of gestational protein restriction (GPR) in the development of the offspring hearts are not well understood. Objectives The aim of this study was to evaluate the effects of GPR on cardiac structure in male rat offspring at day 60 after birth (d60). Methods Pregnant Wistar rats were fed a normal-protein (NP, 17% casein) or low-protein (LP, 6% casein) diet. Blood pressure (BP) values from 60-day-old male offspring were measured by an indirect tail-cuff method using an electro sphygmomanometer. Hearts (d60) were collected for assessment of connexin 43 (Cx43) mRNA expression and morphological and morphometric analysis. Results LP offspring showed no difference in body weight, although they were born lighter than NP offspring. BP levels were significantly higher in the LP group. We observed a significant increase in the area occupied by collagen fibers, a decrease in the number of cardiomyocytes by 104 µm2, and an increase in cardiomyocyte area associated with an increased Cx43 expression. Conclusion GPR changes myocardial levels of Cx43 mRNA in male young adult rats, suggesting that this mechanism aims to compensate the fibrotic process by the accumulation of collagen fibers in the heart interstitium. PMID:28678925

  11. Thyroid hormone deiodinase type 2 mRNA levels in sea lamprey (Petromyzon marinus) are regulated during metamorphosis and in response to a thyroid challenge.

    PubMed

    Stilborn, S Salina M; Manzon, Lori A; Schauenberg, Jennifer D; Manzon, Richard G

    2013-03-01

    Thyroid hormones (THs) are crucial for normal vertebrate development and are the one obligate morphogen that drives amphibian metamorphosis. However, contrary to other metamorphosing vertebrates, lampreys exhibit a sharp drop in serum TH early in metamorphosis, and anti-thyroid agents such as potassium perchlorate (KClO(4)) induce metamorphosis. The type 2 deiodinase (D2) enzyme is a key regulator of TH availability during amphibian metamorphosis. We set out to determine how D2 may be involved in the regulation of lamprey metamorphosis and thyroid homeostasis. We cloned a 1.8Kb Petromyzon marinus D2 cDNA that includes the entire protein coding region and a selenocysteine (Sec) codon. Northern blotting indicated that the lamprey D2 mRNA is the longest reported to date (>9Kb). Using real-time PCR, we showed that intestinal and hepatic D2 mRNA levels were elevated prior to and during the early stages of metamorphosis and then declined dramatically to low levels that were sustained for the remainder of metamorphosis. These data are consistent with previously reported changes in serum TH levels and deiodinase activity. Treatment of larvae with either TH or KClO(4) significantly affected D2 mRNA levels in the intestine and liver. These D2 mRNA levels during metamorphosis and in response to thyroid challenges suggest that D2 may function in the regulation of TH levels during lamprey metamorphosis and the maintenance of TH homeostasis. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. MHC2TA mRNA levels and human herpesvirus 6 in multiple sclerosis patients treated with interferon beta along two-year follow-up

    PubMed Central

    2012-01-01

    Background In previous studies we found that MHC2TA +1614 genotype frequency was very different when MS patients with and without human herpesvirus 6 (HHV-6) in serum samples were compared; a different clinical behavior was also described. The purpose of the study was: 1. To evaluate if MHC2TA expression in MS patients was influenced by interferon beta (IFN-beta) treatment. 2. To study MHC2TA expression in MS patients with and without minor allele C. 3. To analyze the relation between MHC2TA mRNA levels and HHV-6 active infection in MS patients. Methods Blood and serum samples of 154 MS patients were collected in five programmed visits: basal (prior to beginning IFN-beta treatment), six, twelve, eighteen and twenty-four months later. HHV-6 in serum and MHC2TA mRNA levels were evaluated by PCR and RT-PCR, respectively. Neutralizing antibodies (NAbs) against IFN-beta were analyzed by the cytopathic effect assay. Results We found that MHC2TA mRNA levels were significantly lower among MS patients with HHV-6 active infection at the basal visit (without treatment) than in those MS patients without HHV-6 active infection at the basal visit (p = 0.012); in all the positive samples we only found variant A. Furthermore, 58/99 (58.6%) MS patients without HHV-6 along the five programmed visits and an increase of MHC2TA expression after two-years of IFN-beta treatment were clinical responders vs. 5/21 (23.8%) among those MS patients with HHV-6 and a decrease of MHC2TA mRNA levels along the two-years with IFN-beta treatment (p = 0.004); no differences were found between patients with and without NAbs. Conclusions MHC2TA mRNA levels could be decreased by the active replication of HHV-6; the absence of HHV-6 in serum and the increase of MHC2TA expression could be further studied as markers of good clinical response to IFN-beta treatment. PMID:23009575

  13. The mRNA expression levels of uncoupling proteins 1 and 2 in mononuclear cells from patients with metabolic disorders: obesity and type 2 diabetes mellitus.

    PubMed

    Margaryan, Sona; Witkowicz, Agata; Partyka, Anna; Yepiskoposyan, Levon; Manukyan, Gayane; Karabon, Lidia

    2017-10-19

    Type 2 diabetes mellitus (T2DM) and obesity are metabolic disorders whose major hallmark is insulin resistance. Impaired mitochondrial activity, such as reduced ratio of energy production to respiration, has been implicated in the development of insulin resistance. Uncoupling proteins (UCPs) are proton carriers, expressed in the mitochondrial inner membrane, that uncouple oxygen consumption by the respiratory chain from ATP synthesis. The aim of the study was to determine transcriptional levels of UCP1 and UCP2 in peripheral blood mononuclear cells (PBMCs) from patients with metabolic disorders: T2DM, obesity and from healthy individuals. The mRNA levels of UCP1, UCP2 were determined by Real-Time PCR method using Applied Biosystems assays. The UCP1 mRNA expression level was not detectable in the majority of studied samples, while very low expression was found in PBMCs from 3 obese persons. UCP2 mRNA expression level was detectable in all samples. The median mRNA expression of UCP2 was lower in all patients with metabolic disorders as compared to the controls (0.20+0.14 vs. 0.010+0.009, p=0.05). When compared separately, the differences of medians UCP2 mRNA expression level between the obese individuals and the controls as well as between the T2DM patients and the controls did not reach statistical significance. Decreased UCP2 gene expression in mononuclear cells from obese and diabetic patients might contribute to the immunological abnormalities in these metabolic disorders and suggests its role as a candidate gene in future studies of obesity and diabetes.

  14. Influence of developmental stage and genotype on liver mRNA levels among wild, domesticated, and hybrid rainbow trout (Oncorhynchus mykiss).

    PubMed

    White, Samantha L; Sakhrani, Dionne; Danzmann, Roy G; Devlin, Robert H

    2013-10-02

    Release of domesticated strains of fish into nature may pose a threat to wild populations with respect to their evolved genetic structure and fitness. Understanding alterations that have occurred in both physiology and genetics as a consequence of domestication can assist in evaluating the risks posed by introgression of domesticated genomes into wild genetic backgrounds, however the molecular causes of these consequences are currently poorly defined. The present study has examined levels of mRNA in fast-growing pure domesticated (D), slow-growing age-matched pure wild (Wa), slow-growing size-matched pure wild (Ws), and first generation hybrid cross (W/D) rainbow trout (Oncorhynchus mykiss) to investigate the influence of genotype (domesticated vs. wild, and their interactions in hybrids) and developmental stage (age- or size-matched animals) on genetic responses (i.e. dominant vs. recessive) and specific physiological pathways. Highly significant differences in mRNA levels were found between domesticated and wild-type rainbow trout genotypes (321 mRNAs), with many mRNAs in the wild-domesticated hybrid progeny showing intermediate levels. Differences were also found between age-matched and size-matched wild-type trout groups (64 mRNAs), with unique mRNA differences for each of the wild-type groups when compared to domesticated trout (Wa: 114 mRNAs, Ws: 88 mRNAs), illustrating an influence of fish developmental stage affecting findings when used as comparator groups to other genotypes. Analysis of differentially expressed mRNAs (found for both wild-type trout to domesticated comparisons) among the genotypes indicates that 34.8% are regulated consistent with an additive genetic model, whereas 39.1% and 26.1% show a recessive or dominant mode of regulation, respectively. These molecular data are largely consistent with phenotypic data (growth and behavioural assessments) assessed in domesticated and wild trout strains. The present molecular data are concordant with

  15. Different Relationship between hsp70 mRNA and hsp70 Levels in the Heat Shock Response of Two Salmonids with Dissimilar Temperature Preference

    PubMed Central

    Lewis, Mario; Götting, Miriam; Anttila, Katja; Kanerva, Mirella; Prokkola, Jenni M.; Seppänen, Eila; Kolari, Irma; Nikinmaa, Mikko

    2016-01-01

    The heat shock response (HSR) refers to the rapid production of heat shock proteins (hsps) in response to a sudden increase in temperature. Its regulation by heat shock factors is a good example of how gene expression is transcriptionally regulated by environmental stresses. In contrast, little is known about post-transcriptional regulation of the response. The heat shock response is often used to characterize the temperature tolerance of species with the rationale that whenever the response sets on, a species is approaching its lethal temperature. It has commonly been considered that an increase in hsp mRNA gives an accurate indication that the same happens to the protein level, but this need not be the case. With climate change, understanding the effects of temperature on gene expression of especially polar organisms has become imperative to evaluate how both biodiversity and commercially important species respond, since temperature increases are expected to be largest in polar areas. Here we studied the HSR of two phylogenetically related Arctic species, which differ in their temperature tolerance with Arctic charr having lower maximally tolerated temperature than Atlantic salmon. Arctic charr acclimated to 15°C and exposed to 7°C temperature increase for 30 min showed both an increase in hsp70 mRNA and hsp70 whereas in salmon only hsp70 mRNA increased. Our results indicate that the temperature for transcriptional induction of hsp can be different from the one required for a measurable change in inducible hsp level. The species with lower temperature tolerance, Arctic charr, are experiencing temperature stress already at the higher acclimation temperature, 15°C, as their hsp70 mRNA and hsp70 levels were higher, and they grow less than fish at 8°C (whereas for salmon the opposite is true). Consequently, charr experience more drastic heat shock than salmon. Although further studies are needed to establish the temperature range and length of exposure where hsp

  16. [Effects of lipopolysaccharides extracted from Porphyromonas endodontalis on the expression of IL-1beta mRNA and IL-6 mRNA in osteoblasts].

    PubMed

    Yang, Di; Li, Ren; Qiu, Li-Hong; Li, Chen

    2009-04-01

    To quantify the IL-1 beta mRNA and IL-6 mRNA expression induced by lipopolysaccharides (LPS)extracted from Porphyromonas endodontalis(P.e) in osteoblasts, and to relate P.e-LPS to bone absorption pathogenesis in lesions of chronical apical periodontitis. MG63 was treated with different concentrations of P.e-LPS(0-50 microg/mL) for different hours(0-24h). The expression of IL-1 beta mRNA and IL-6 mRNA was detected by reverse transcription polymerase chain reaction (RT-PCR).Statistical analysis was performed using one- way ANOVA and Dunnett t test with SPSS11.0 software package. The level of IL-1 beta mRNA and IL-6 mRNA increased significantly after treatment with P.e-LPS at more than 5 microg/mL (P<0.01)and for more than 1 hour (P<0.01), which indicated that P.e-LPS induced osteoblasts to express IL-1 beta mRNA and IL-6 mRNA in dose and time dependent manners. P.e-LPS may promote bone resorption in lesions of chronical apical periodontitis by inducing IL-1 beta mRNA and IL-6 mRNA expression in osteoblasts.

  17. Effects of different dietary intake on mRNA levels of MSTN, IGF-I, and IGF-II in the skeletal muscle of Dorper and Hu sheep hybrid F1 rams.

    PubMed

    Xing, H J; Wang, Z Y; Zhong, B S; Ying, S J; Nie, H T; Zhou, Z R; Fan, Y X; Wang, F

    2014-07-24

    MSTN, IGF-І(insulin-like growth factor-І) and IGF-II (insulin-like growth factor-II) regulate skeletal muscle growth. This study investigated the effects of different dietary intake levels on skeletal muscles. Sheep was randomly assigned to 3 feeding groups: 1) the maintenance diet (M), 2) 1.4 x the maintenance diet (1.4M), and 3) 2.15 x the maintenance diet (2.15M). Before slaughtering the animals, blood samples were collected to measure plasma urea, growth hormone, and insulin concentrations. After slaughtering, the longissimus dorsi, semitendinosus, semimembranosus, gastrocnemius, soleus, and chest muscle were removed to record various parameters, including the mRNA expression levels of MSTN and IGFs, in addition to skeletal muscle fiber diameter and cross-sectional area. The result showed that as dietary intake improved, the mRNA expression levels of MSTN and IGF-II decreased, whereas IGF-Іexpression increased. The mRNA expression levels of MSTN and IGFs were significantly different in the same skeletal muscle under different dietary intake. The skeletal muscle fiber diameter and cross-sectional area increased with greater dietary intake, as observed for the mRNA expression of IGF-І; however, it contrasted to that observed for the mRNA expression of MSTN and IGF-II. In conclusion, dietary intake levels have a certain influence on MSTN and IGFs mRNA expression levels, in addition to skeletal muscle fiber diameter and cross-sectional area. This study contributes valuable information for enhancing the molecular-based breeding of sheep.

  18. Immune-stimulatory effects of a bacteria-based probiotic on peripheral leukocyte subpopulations and cytokine mRNA expression levels in scouring holstein calves.

    PubMed

    Qadis, Abdul Qadir; Goya, Satoru; Yatsu, Minoru; Kimura, Atsushi; Ichijo, Toshihiro; Sato, Shigeru

    2014-05-01

    Subpopulations of peripheral leukocytes and cytokine mRNA expression levels were evaluated in scouring and healthy Holstein calves (age 10 ± 5 days; n=42) treated with a probiotic consisting of Lactobacillus plantarum, Enterococcus faecium and Clostridium butyricum. The calves were assigned to the scouring or healthy group and then subdivided into pathogen-positive treated (n=8), pathogen-positive control (n=8), pathogen-negative treated (n=6), pathogen-negative control (n=6), healthy treated (n=6) and healthy control (n=8) groups. A single dose of the probiotic (3.0 g/100 kg body weight) was given to each calf in the treatment groups for 5 days. Blood samples were collected on the first day of scour occurrence (day 0) and on day 7. In the scouring calves, smaller peripheral leukocyte subpopulations and cytokine mRNA expression levels were noted on day 0. The numbers of CD3(+) T cells; CD4(+), CD8(+) and WC1(+) γδ T cell subsets; and CD14(+), CD21(+) and CD282(+) (TLR2) cells were significantly increased in the scouring and healthy treated calves on day 7. Furthermore, interleukin-6, tumor necrosis factor-alpha and interferon-gamma mRNA expression was elevated in the peripheral leukocytes of the scouring and healthy treated calves on day 7. The scouring calves given the probiotic recovered on day 7. A significantly smaller number of peripheral leukocytes and lower cytokine mRNA expression level might be induced by scouring in calves. Repeated probiotic administration might stimulate cellular immunity and encourage recovery from scouring in pre-weaning Holstein calves.

  19. Effect of sulfide, osmotic, and thermal stresses on taurine transporter mRNA levels in the gills of the hydrothermal vent-specific mussel Bathymodiolus septemdierum.

    PubMed

    Nakamura-Kusakabe, Ikumi; Nagasaki, Toshihiro; Kinjo, Azusa; Sassa, Mieko; Koito, Tomoko; Okamura, Kei; Yamagami, Shosei; Yamanaka, Toshiro; Tsuchida, Shinji; Inoue, Koji

    2016-01-01

    Hydrothermal vent environmental conditions are characterized by high sulfide concentrations, fluctuating osmolality, and irregular temperature elevations caused by vent effluents. These parameters represent potential stressors for organisms that inhabit the area around hydrothermal vents. Here, we aimed to obtain a better understanding of the adaptation mechanisms of marine species to hydrothermal vent environments. Specifically, we examined the effect of sulfide, osmolality, and thermal stress on the expression of taurine transporter (TAUT) mRNA in the gill of the deep-sea mussel Bathymodiolus septemdierum, which is a dominant species around hydrothermal vent sites. We analyzed TAUT mRNA levels by quantitative real-time polymerase chain reaction (PCR) in the gill of mussels exposed to sulfide (0.1 or 1mg/L Na2S·9H2O), hyper- (115% seawater) and hypo- (97.5%, 95.5%, and 85% seawater) osmotic conditions, and thermal stresses (12°C and 20°C) for 24 and 48h. The results showed that mussels exposed to relatively low levels of sulfide (0.1mg/L) and moderate heat stress (12°C) exhibited higher TAUT mRNA levels than the control. Although hyper- and hypo-osmotic stress did not significantly change TAUT mRNA levels, slight induction was observed in mussels exposed to low osmolality. Our results indicate that TAUT is involved in the coping mechanism of mussels to various hydrothermal vent stresses. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Distributed Energy Resource Optimization Using a Software as Service (SaaS) Approach at the University of California, Davis Campus

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Stadler, Michael; Marnay, Chris; Donadee, Jon

    2011-02-06

    Together with OSIsoft LLC as its private sector partner and matching sponsor, the Lawrence Berkeley National Laboratory (Berkeley Lab) won an FY09 Technology Commercialization Fund (TCF) grant from the U.S. Department of Energy. The goal of the project is to commercialize Berkeley Lab's optimizing program, the Distributed Energy Resources Customer Adoption Model (DER-CAM) using a software as a service (SaaS) model with OSIsoft as its first non-scientific user. OSIsoft could in turn provide optimization capability to its software clients. In this way, energy efficiency and/or carbon minimizing strategies could be made readily available to commercial and industrial facilities. Specialized versionsmore » of DER-CAM dedicated to solving OSIsoft's customer problems have been set up on a server at Berkeley Lab. The objective of DER-CAM is to minimize the cost of technology adoption and operation or carbon emissions, or combinations thereof. DER-CAM determines which technologies should be installed and operated based on specific site load, price information, and performance data for available equipment options. An established user of OSIsoft's PI software suite, the University of California, Davis (UCD), was selected as a demonstration site for this project. UCD's participation in the project is driven by its motivation to reduce its carbon emissions. The campus currently buys electricity economically through the Western Area Power Administration (WAPA). The campus does not therefore face compelling cost incentives to improve the efficiency of its operations, but is nonetheless motivated to lower the carbon footprint of its buildings. Berkeley Lab attempted to demonstrate a scenario wherein UCD is forced to purchase electricity on a standard time-of-use tariff from Pacific Gas and Electric (PG&E), which is a concern to Facilities staff. Additionally, DER-CAM has been set up to consider the variability of carbon emissions throughout the day and seasons. Two distinct

  1. Lack of acute phase response in the livers of mice exposed to diesel exhaust particles or carbon black by inhalation

    PubMed Central

    Saber, Anne T; Halappanavar, Sabina; Folkmann, Janne K; Bornholdt, Jette; Boisen, Anne Mette Z; Møller, Peter; Williams, Andrew; Yauk, Carole; Vogel, Ulla; Loft, Steffen; Wallin, Håkan

    2009-01-01

    Background Epidemiologic and animal studies have shown that particulate air pollution is associated with increased risk of lung and cardiovascular diseases. Although the exact mechanisms by which particles induce cardiovascular diseases are not known, studies suggest involvement of systemic acute phase responses, including C-reactive protein (CRP) and serum amyloid A (SAA) in humans. In this study we test the hypothesis that diesel exhaust particles (DEP) – or carbon black (CB)-induced lung inflammation initiates an acute phase response in the liver. Results Mice were exposed to filtered air, 20 mg/m3 DEP or CB by inhalation for 90 minutes/day for four consecutive days; we have previously shown that these mice exhibit pulmonary inflammation (Saber AT, Bornholdt J, Dybdahl M, Sharma AK, Loft S, Vogel U, Wallin H. Tumor necrosis factor is not required for particle-induced genotoxicity and pulmonary inflammation., Arch. Toxicol. 79 (2005) 177–182). As a positive control for the induction of an acute phase response, mice were exposed to 12.5 mg/kg of lipopolysaccharide (LPS) intraperitoneally. Quantitative real time RT-PCR was used to examine the hepatic mRNA expression of acute phase proteins, serum amyloid P (Sap) (the murine homologue of Crp) and Saa1 and Saa3. While significant increases in the hepatic expression of Sap, Saa1 and Saa3 were observed in response to LPS, their levels did not change in response to DEP or CB. In a comprehensive search for markers of an acute phase response, we analyzed liver tissue from these mice using high density DNA microarrays. Globally, 28 genes were found to be significantly differentially expressed in response to DEP or CB. The mRNA expression of three of the genes (serine (or cysteine) proteinase inhibitor, clade A, member 3C, apolipoprotein E and transmembrane emp24 domain containing 3) responded to both exposures. However, these changes were very subtle and were not confirmed by real time RT-PCR. Conclusion Our findings

  2. Low levels of PRB3 mRNA are associated with dopamine-agonist resistance and tumor recurrence in prolactinomas.

    PubMed

    Wang, Fei; Gao, Hua; Li, Chuzhong; Bai, Jiwei; Lu, Runchun; Cao, Lei; Wu, Yongtu; Hong, Lichuan; Wu, Yonggang; Lan, Xiaolei; Zhang, Yazhuo

    2014-01-01

    Prolactinomas, or prolactin-secreting adenomas, constitute the most common type of hyperfunctioning pituitary adenoma. Dopamine agonists are used as first-line medication for prolactinomas, but the tumors are resistant to the therapy in 5-18 % of patients. To explore potential mechanisms of resistance to bromocriptine (a dopamine agonist), we analyzed six responsive prolactinomas and six resistant prolactinomas by whole-exome sequencing. We identified ten genes with sequence variants that were differentially found in the two groups of tumors. The expression of these genes was then quantified by real-time reverse-transcription PCR (RT-qPCR) in the 12 prolactinomas and in six normal pituitary glands. The mRNA levels of one of the genes, PRB3, were about fourfold lower in resistant prolactinomas than in the responsive tumors (p = 0.02). Furthermore, low PRB3 expression was also associated with tumor recurrence. Our results suggest that low levels of PRB3 mRNA may have a role in dopamine-agonist resistance and tumor recurrence of prolactinomas.

  3. Light-regulated protein and mRNA synthesis in root caps of maize

    NASA Technical Reports Server (NTRS)

    Feldman, L. J.; Piechulla, B.; Sun, P. S.

    1988-01-01

    Illumination of maize roots initiates changes in mRNA levels and in the activities of proteins within the root cap. Using Northern analysis we showed a 5-6 fold increase in the levels of three specific mRNAs and a 14-fold increase in plastid mRNA. This increase is rapid, occurring within 30 minutes of illumination. With prolonged periods of darkness following illumination, messages return to levels observed in dark, control caps. For two species of mRNA illumination results in a reduction in message levels. Light-stimulated increases in the levels of specific mRNAs are proportionally greater than are increases in the activities of corresponding proteins. We suggest that the light-stimulated increase in protein activity in root caps may be preceded by and occur as a consequence of enhanced levels of mRNA. Our work suggests that photomorphogenesis in roots could involve changes in the levels of a wide variety of mRNAs within the root cap.

  4. Regulation of bone sialoprotein mRNA by steroid hormones

    PubMed Central

    1989-01-01

    In this report we demonstrate an increase in the steady-state level of bone sialoprotein (BSP) mRNA in rat calvaria and a rat osteosarcoma cell line (ROS 17/2.8) after treatment with the synthetic glucocorticoid, dexamethasone. In contrast, 1.25-dihydroxyvitamin D3 reduced the amount of BSP mRNA in calvaria and inhibited the dexamethasone induction in ROS 17/2.8 cells. The increase in BSP mRNA is most likely due to an increase in the transcriptional rate. The stability of mRNA was unchanged after dexamethasone treatment with a half-life of approximately 5 h. Nuclear transcription experiments with nuclei isolated from ROS 17/2.8 cells showed an increased BSP mRNA synthesis in cells treated with dexamethasone. PMID:2592421

  5. Conjunctival mucin mRNA expression in contact lens wear.

    PubMed

    Corrales, Rosa M; Galarreta, David; Herreras, Jose M; Saez, Victoria; Arranz, Isabel; González, Maria J; Mayo, Agustin; Calonge, Margarita; Chaves, Felipe J

    2009-09-01

    To investigate the influence of the water content in non-ionic hydrogel contact lenses (HCL) on the mRNA levels of human conjunctival mucin genes (MUCs). Sixteen healthy subjects with no history of contact lenses wear were selected and randomized into two equal groups. Group 1 subjects wore low water content (38%, Soflens 38) non-ionic HCLs. Group 2 wore high water content (66%, Soflens 66) non-ionic HCLs. Conjunctival impression cytology was applied to the superior bulbar conjunctiva of both eyes before, 6 months, and 1 year after HCL fitting, and 15 days after discontinuation of wearing. Total RNA was isolated, retrotranscribed, and amplified by conventional polymerase chain reaction (PCR) and by quantitative real time PCR to study the mRNA levels of MUCs and to analyze variations during the study period. Time- and HCL-dependent variations in mRNA expression were analyzed using Student's test. From the known MUCs, transcripts from MUC1, MUC2, MUC4, MUC5AC, MUC7, MUC13, MUC15, MUC16, and MUC17 genes were detected in all subjects before HCL fitting. Except for MUC2, the expression of some MUC genes significantly increased whereas others significantly decreased at either the 6- and 12-month period. Statistically significant differences between both HCL groups (p < 0.001) were found in the MUC4, MUC13, and MUC15 mRNA expression after 1 year of wear and after the 15 days without HCL wear. However, these differences were not clearly related to the water content of the lenses. Low and high water content non-ionic HCLs induced different changes in the mRNA levels of several MUCs, but the water content was not related to the changes. Recovery to basal levels of conjunctival MUC mRNA expression after wearing HCL lenses for a year takes longer than 15 days for some MUCs.

  6. flexCloud: Deployment of the FLEXPART Atmospheric Transport Model as a Cloud SaaS Environment

    NASA Astrophysics Data System (ADS)

    Morton, Don; Arnold, Dèlia

    2014-05-01

    FLEXPART (FLEXible PARTicle dispersion model) is a Lagrangian transport and dispersion model used by a growing international community. We have used it to simulate and forecast the atmospheric transport of wildfire smoke, volcanic ash and radionuclides. Additionally, FLEXPART may be run in backwards mode to provide information for the determination of emission sources such as nuclear emissions and greenhouse gases. This open source software is distributed in source code form, and has several compiler and library dependencies that users need to address. Although well-documented, getting it compiled, set up, running, and post-processed is often tedious, making it difficult for the inexperienced user. Our interest is in moving scientific modeling and simulation activities from site-specific clusters and supercomputers to a cloud model as a service paradigm. Choosing FLEXPART for our prototyping, our vision is to construct customised IaaS images containing fully-compiled and configured FLEXPART codes, including pre-processing, execution and postprocessing components. In addition, with the inclusion of a small web server in the image, we introduce a web-accessible graphical user interface that drives the system. A further initiative being pursued is the deployment of multiple, simultaneous FLEXPART ensembles in the cloud. A single front-end web interface is used to define the ensemble members, and separate cloud instances are launched, on-demand, to run the individual models and to conglomerate the outputs into a unified display. The outcome of this work is a Software as a Service (Saas) deployment whereby the details of the underlying modeling systems are hidden, allowing modelers to perform their science activities without the burden of considering implementation details.

  7. Fine-tuning of ULK1 mRNA and protein levels is required for autophagy oscillation

    PubMed Central

    Ciccosanti, Fabiola

    2016-01-01

    Autophagy is an intracellular degradation pathway whose levels are tightly controlled to secure cell homeostasis. Unc-51–like kinase 1 (ULK1) is a conserved serine–threonine kinase that plays a central role in the initiation of autophagy. Here, we report that upon autophagy progression, ULK1 protein levels are specifically down-regulated by the E3 ligase NEDD4L, which ubiquitylates ULK1 for degradation by the proteasome. However, whereas ULK1 protein is degraded, ULK1 mRNA is actively transcribed. Upon reactivation of mTOR-dependent protein synthesis, basal levels of ULK1 are promptly restored, but the activity of newly synthesized ULK1 is inhibited by mTOR. This prepares the cell for a new possible round of autophagy stimulation. Our results thus place NEDD4L and ULK1 in a key position to control oscillatory activation of autophagy during prolonged stress to keep the levels of this process under a safe and physiological threshold. PMID:27932573

  8. Salt Stress Increases the Level of Translatable mRNA for Phosphoenolpyruvate Carboxylase in Mesembryanthemum crystallinum1

    PubMed Central

    Ostrem, James A.; Olson, Steve W.; Schmitt, Jürgen M.; Bohnert, Hans J.

    1987-01-01

    Mesembryanthemum crystallinum responds to salt stress by switching from C3 photosynthesis to Crassulacean acid metabolism (CAM). During this transition the activity of phosphoenolpyruvate carboxylase (PEPCase) increases in soluble protein extracts from leaf tissue. We monitored CAM induction in plants irrigated with 0.5 molar NaCl for 5 days during the fourth, fifth, and sixth week after germination. Our results indicate that the age of the plant influenced the response to salt stress. There was no increase in PEPCase protein or PEPCase enzyme activity when plants were irrigated with 0.5 molar NaCl during the fourth and fifth week after germination. However, PEPCase activity increased within 2 to 3 days when plants were salt stressed during the sixth week after germination. Immunoblot analysis with anti-PEPCase antibodies showed that PEPCase synthesis was induced in both expanded leaves and in newly developing axillary shoot tissue. The increase in PEPCase protein was paralleled by an increase in PEPCase mRNA as assayed by immunoprecipitation of PEPCase from the in vitro translation products of RNA from salt-stressed plants. These results demonstrate that salinity increased the level of PEPCase in leaf and shoot tissue via a stress-induced increase in the steady-state level of translatable mRNA for this enzyme. Images Fig. 2 Fig. 3 Fig. 4 PMID:16665596

  9. Immune-Stimulatory Effects of a Bacteria-Based Probiotic on Peripheral Leukocyte Subpopulations and Cytokine mRNA Expression Levels in Scouring Holstein Calves

    PubMed Central

    QADIS, Abdul Qadir; GOYA, Satoru; YATSU, Minoru; KIMURA, Atsushi; ICHIJO, Toshihiro; SATO, Shigeru

    2014-01-01

    ABSTRACT Subpopulations of peripheral leukocytes and cytokine mRNA expression levels were evaluated in scouring and healthy Holstein calves (age 10 ± 5 days; n=42) treated with a probiotic consisting of Lactobacillus plantarum, Enterococcus faecium and Clostridium butyricum. The calves were assigned to the scouring or healthy group and then subdivided into pathogen-positive treated (n=8), pathogen-positive control (n=8), pathogen-negative treated (n=6), pathogen-negative control (n=6), healthy treated (n=6) and healthy control (n=8) groups. A single dose of the probiotic (3.0 g/100 kg body weight) was given to each calf in the treatment groups for 5 days. Blood samples were collected on the first day of scour occurrence (day 0) and on day 7. In the scouring calves, smaller peripheral leukocyte subpopulations and cytokine mRNA expression levels were noted on day 0. The numbers of CD3+ T cells; CD4+, CD8+ and WC1+ γδ T cell subsets; and CD14+, CD21+ and CD282+ (TLR2) cells were significantly increased in the scouring and healthy treated calves on day 7. Furthermore, interleukin-6, tumor necrosis factor-alpha and interferon-gamma mRNA expression was elevated in the peripheral leukocytes of the scouring and healthy treated calves on day 7. The scouring calves given the probiotic recovered on day 7. A significantly smaller number of peripheral leukocytes and lower cytokine mRNA expression level might be induced by scouring in calves. Repeated probiotic administration might stimulate cellular immunity and encourage recovery from scouring in pre-weaning Holstein calves. PMID:24451928

  10. Differential utilization of decapping enzymes in mammalian mRNA decay pathways

    PubMed Central

    Li, You; Song, Mangen; Kiledjian, Megerditch

    2011-01-01

    mRNA decapping is a crucial step in the regulation of mRNA stability and gene expression. Dcp2 is an mRNA decapping enzyme that has been widely studied. We recently reported the presence of a second mammalian cytoplasmic decapping enzyme, Nudt16. Here we address the differential utilization of the two decapping enzymes in specified mRNA decay processes. Using mouse embryonic fibroblast (MEF) cell lines derived from a hypomorphic knockout of the Dcp2 gene with undetectable levels of Dcp2 or MEF cell lines harboring a Nudt16-directed shRNA to generate reduced levels of Nudt16, we demonstrate the distinct roles for Dcp2 and Nudt16 in nonsense-mediated mRNA decay (NMD), decay of ARE-containing mRNA and miRNA-mediated silencing. Our results indicated that NMD preferentially utilizes Dcp2 rather than Nudt16; Dcp2 and Nudt16 are redundant in miRNA-mediated silencing; and Dcp2 and Nudt16 are differentially utilized for ARE-mRNA decay. These data demonstrate that the two distinct decapping enzymes can uniquely function in specific mRNA decay processes in mammalian cells. PMID:21224379

  11. Stress Biomarkers, Mood States, and Sleep during a Major Competition: “Success” and “Failure” Athlete's Profile of High-Level Swimmers

    PubMed Central

    Chennaoui, Mounir; Bougard, Clément; Drogou, Catherine; Langrume, Christophe; Miller, Christian; Gomez-Merino, Danielle; Vergnoux, Frédéric

    2016-01-01

    The aim of this study was to evaluate stress markers, mood states, and sleep indicators in high-level swimmers during a major 7-days competition according to the outcomes. Nine swimmers [six men and three women (age: 22 ± 2 and 22 ± 4 years, respectively)] were examined. Before (PRE) and after (POST) each race (series, semi-finals, and finals), salivary concentrations of cortisol, α-amylase (sAA), and chromogranin-A (CgA) were determined. Mood states were assessed by the profile of mood state (POMS) questionnaire completed before and after the 7-days, and self-reported sleep diaries were completed daily. In the “failure” group, cortisol and sAA significantly increased between PRE-POST measurements (p < 0.05), while sCgA was not changed. Significant overall decrease of cortisol (-52.6%) and increase of sAA (+68.7%) was shown in the “failure group.” In this group, fatigue, confusion and depression scores, and sleep duration before the finals increased. The results in the “success” group show tendencies for increased cortisol and sCgA concentrations in response to competition, while sAA was not changed. Cortisol levels before the semi-finals and finals and sCgA levels before the finals were positively correlated to the fatigue score in the “failure” group only (r = 0.89). sAA levels before and after the semi-finals were negatively correlated to sleep duration measured in the subsequent night (r = −0.90). In conclusion, the stress of the competition could trigger a negative mood profile and sleep disturbance which correspond to different responses of biomarkers related to the hypothalamo-pituitary-adrenal axis and the sympathetic nervous system (SNS) activity, cortisol, sAA, and CgA. PMID:27014092

  12. Effectiveness of memantine on depression-like behavior, memory deficits and brain mRNA levels of BDNF and TrkB in rats subjected to repeated unpredictable stress.

    PubMed

    Amidfar, Meysam; Kim, Yong-Ku; Wiborg, Ove

    2018-06-01

    Previous clinical and preclinical studies have indicated that the N-methyl-d-aspartate (NMDA) receptor antagonist, memantine, has neuroprotective properties as well as antidepressant effects. The present study was designed to examine behavioral and molecular effects of memantine administration in rats subjected to the repeated unpredictable stress (RUS) paradigm. Rats were split into four groups at random including control+saline, control+memantine, stressed+saline and stressed+memantine. After 10days of exposure to the RUS paradigm, rats were administered memantine (20mg/kg) intraperitoneally (ip) for 14days. Depression-like behavior and memory performance were assessed by measuring immobility time in the forced swim test and passive avoidance test, respectively. The mRNA levels of BDNF and TrkB in the prefrontal cortex and hippocampus were measured by real-time quantitative PCR. Our results demonstrated that the RUS paradigm caused depression-like behavior and impairment of memory retrieval in rats. We did not find significant changes in BDNF or TrkB mRNA levels in hippocampus, but mRNA levels of TrkB in the prefrontal cortex showed a significant downregulation. Administration of memantine reversed depression-like behavior and memory impairment and significantly increased BDNF and TrkB mRNA levels in both prefrontal cortex and hippocampus of stress exposed rats. Our study supports the hypothesis that drugs with antagonistic properties on the NMDA receptor, such as memantine, might be efficient in treatment of major depression. Our results also suggest that upregulated mRNA levels of BDNF and TrkB in the brain might be essential for antidepressant-like activity of memantine in stress exposed rats. Copyright © 2017 Institute of Pharmacology, Polish Academy of Sciences. Published by Elsevier B.V. All rights reserved.

  13. Long-term treatment with haloperidol affects neuropeptide S and NPSR mRNA levels in the rat brain.

    PubMed

    Palasz, Artur; Rojczyk, Ewa; Golyszny, Milosz; Filipczyk, Lukasz; Worthington, John J; Wiaderkiewicz, Ryszard

    2016-04-01

    The brainstem-derived neuropeptide S (NPS) has a multidirectional regulatory activity, especially as a potent anxiolytic factor. Accumulating data suggests that neuroleptics affect peptidergic signalling in various brain structures. However, there is no information regarding the influence of haloperidol on NPS and NPS receptor (NPSR) expression. We assessed NPS and NPSR mRNA levels in brains of rats treated with haloperidol using quantitative real-time polymerase chain reaction. Chronic haloperidol treatment (4 weeks) led to a striking upregulation of NPS and NPSR expression in the rat brainstem. Conversely, the NPSR mRNA expression was decreased in the hippocampus and striatum. This stark increase of NPS in response to haloperidol treatment supports the hypothesis that this neuropeptide is involved in the dopamine-dependent anxiolytic actions of neuroleptics and possibly also in the pathophysiology of mental disorders. Furthermore, our findings underline the complex nature of potential interactions between dopamine receptors and brain peptidergic pathways, which has potential clinical applications.

  14. The Antagonistic Effect of Selenium on Cadmium-Induced Damage and mRNA Levels of Selenoprotein Genes and Inflammatory Factors in Chicken Kidney Tissue.

    PubMed

    Wang, Xinyue; Bao, Rongkun; Fu, Jing

    2018-02-01

    Selenium (Se) is a necessary trace mineral in the diet of humans and animals. Cadmium (Cd) is a toxic heavy metal that can damage animal organs, especially the kidneys. Antagonistic interactions between Se and Cd have been reported in previous studies. However, little is known about the effects of Se against Cd toxicity and on the mRNA levels of 25 selenoprotein genes and inflammatory factors in chicken kidneys. In the current study, we fed chickens with a Se-treated, Cd-treated, or Se/Cd treated diet for 90 days. We then analyzed the mRNA expression of inflammatory factors (including prostaglandin E synthase (PTGES), nuclear factor-kappa B (NF-κB), tumor necrosis factor-α (TNF-α), and cyclooxygenase-2 (COX-2)) and 25 selenoprotein genes (Gpx1, Gpx2, Gpx3, Gpx4, Txnrd1, Txnrd2, Txnrd3, Dio1, Dio2, Dio3, SPS2, Sepp1, SelPb, Sep15, Selh, Seli, Selm, Selo, Sels, Sepx1, Selu, Selk, Selw, Seln, Selt). The results demonstrated that Cd exposure increased the Cd content in the chicken kidneys, renal tubular epithelial cells underwent denaturation and necrosis, and the tubules became narrow or disappeared. However, Se supplementation reduced the Cd content in chicken kidneys and induced normal development of renal tubular epithelial cells. In addition, we also observed that Se alleviated the Cd-induced increase in the mRNA levels of inflammatory factors and ameliorated the Cd-induced downtrend in the mRNA levels of 25 selenoprotein genes in chicken kidneys.

  15. Adipocyte resistin mRNA levels are down-regulated by laparoscopic ovarian electrocautery in both obese and lean women with polycystic ovary syndrome.

    PubMed

    Seow, Kok-Min; Juan, Chi-Chang; Ho, Low-Tone; Hsu, Yung-Pei; Lin, Yu-Hung; Huang, Lee-Wen; Hwang, Jiann-Loung

    2007-04-01

    The aim of this study was to investigate serum and adipocyte mRNA expression of resistin in lean and obese women with polycystic ovary syndrome (PCOS) before and 3 months after laparoscopic ovarian electrocauterization (LOE). Adipose tissue obtained from 12 women with PCOS (six obese and six lean, body mass index > 27 kg m(-1) as threshold point) before and after LOE was analysed. Gene expression of resistin was measured by semi-quantitative RT-PCR. Ten lean, age-matched healthy women served as controls. Both lean and obese women with PCOS had significantly higher fasting and 2 h insulin and homeostasis model insulin resistance index (HOMA(IR)) values and lower fasting glucose-to-insulin ratios (G(0)/I(0)) than did the controls. The serum levels of glucose and insulin and HOMA(IR) were significantly decreased, and the G(0)/I(0) ratio was significantly increased 3 months after LOE. No difference was found in serum resistin levels between controls and either obese or lean women with PCOS before LOE, nor between PCOS patients before and after LOE. However, resistin mRNA expression levels in both lean and obese women with PCOS before LOE were significantly higher than that in controls and were decreased significantly after LOE back to control levels. Local resistin activity may be actively involved in the pathogenesis of PCOS. LOE reduces insulin resistance and down-regulates resistin mRNA expression in lean and obese women with PCOS.

  16. Kinematics of syn- and post-exhumational shear zones at Lago di Cignana (Western Alps, Italy): constraints on the exhumation of Zermatt-Saas (ultra)high-pressure rocks and deformation along the Combin Fault and Dent Blanche Basal Thrust

    NASA Astrophysics Data System (ADS)

    Kirst, Frederik; Leiss, Bernd

    2017-01-01

    Kinematic analyses of shear zones at Lago di Cignana in the Italian Western Alps were used to constrain the structural evolution of units from the Piemont-Ligurian oceanic realm (Zermatt-Saas and Combin zones) and the Adriatic continental margin (Dent Blanche nappe) during Palaeogene syn- and post-exhumational deformation. Exhumation of Zermatt-Saas (U)HP rocks to approximately lower crustal levels at ca. 39 Ma occurred during normal-sense top-(S)E shearing under epidote-amphibolite-facies conditions. Juxtaposition with the overlying Combin zone along the Combin Fault at mid-crustal levels occurred during greenschist-facies normal-sense top-SE shearing at ca. 38 Ma. The scarcity of top-SE kinematic indicators in the hanging wall of the Combin Fault probably resulted from strain localization along the uppermost Zermatt-Saas zone and obliteration by subsequent deformation. A phase of dominant pure shear deformation around 35 Ma affected units in the direct footwall and hanging wall of the Combin Fault. It is interpreted to reflect NW-SE crustal elongation during updoming of the nappe stack as a result of underthrusting of European continental margin units and the onset of continental collision. This phase was partly accompanied and followed by ductile bulk top-NW shearing, especially at higher structural levels, which transitioned into semi-ductile to brittle normal-sense top-NW deformation due to Vanzone phase folding from ca. 32 Ma onwards. Our structural observations suggest that syn-exhumational deformation is partly preserved within units and shear zones exposed at Lago di Cignana but also that the Combin Fault and Dent Blanche Basal Thrust experienced significant post-exhumational deformation reworking and overprinting earlier structures.

  17. Reduction in salivary α-amylase levels following a mind-body intervention in cancer survivors--an exploratory study.

    PubMed

    Lipschitz, David L; Kuhn, Renee; Kinney, Anita Y; Donaldson, Gary W; Nakamura, Yoshio

    2013-09-01

    The main aim of this exploratory study was to assess whether salivary α-amylase (sAA) and salivary cortisol levels would be positively modulated by sleep-focused mind-body interventions in female and male cancer survivors. We conducted a randomized controlled trial in which 57 cancer survivors with self-reported sleep disturbance received either a Sleep Hygiene Education (SHE; n=18) control, or one of two experimental mind-body interventions, namely, Mind-Body Bridging (MBB; n=19) or Mindfulness Meditation (MM; n=20). Interventions were three sessions each conducted once per week for three consecutive weeks. Saliva cortisol and sAA were measured at baseline and 1 week after the last session. Participants also completed a sleep scale at the same time points when saliva was collected for biomarker measurement. Our study revealed that at post-intervention assessment, mean sAA levels upon awakening ("Waking" sample) declined in MBB compared with that of SHE. Mean Waking cortisol levels did not differ among treatment groups but declined slightly in SHE. Self-reported sleep improved across the three interventions at Post-assessment, with largest improvements in the MBB intervention. In this exploratory study, sleep focused mind-body intervention (MBB) attenuated Waking sAA levels, suggesting positive influences of a mind-body intervention on sympathetic activity in cancer survivors with sleep disturbance. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Modulation of DNA repair capacity and mRNA expression levels of XRCC1, hOGG1 and XPC genes in styrene-exposed workers.

    PubMed

    Hanova, Monika; Stetina, Rudolf; Vodickova, Ludmila; Vaclavikova, Radka; Hlavac, Pavel; Smerhovsky, Zdenek; Naccarati, Alessio; Polakova, Veronika; Soucek, Pavel; Kuricova, Miroslava; Manini, Paola; Kumar, Rajiv; Hemminki, Kari; Vodicka, Pavel

    2010-11-01

    Decreased levels of single-strand breaks in DNA (SSBs), reflecting DNA damage, have previously been observed with increased styrene exposure in contrast to a dose-dependent increase in the base-excision repair capacity. To clarify further the above aspects, we have investigated the associations between SSBs, micronuclei, DNA repair capacity and mRNA expression in XRCC1, hOGG1 and XPC genes on 71 styrene-exposed and 51 control individuals. Styrene concentrations at workplace and in blood characterized occupational exposure. The workers were divided into low (below 50 mg/m³) and high (above 50 mg/m³)) styrene exposure groups. DNA damage and DNA repair capacity were analyzed in peripheral blood lymphocytes by Comet assay. The mRNA expression levels were determined by qPCR. A significant negative correlation was observed between SSBs and styrene concentration at workplace (R=-0.38, p=0.001); SSBs were also significantly higher in men (p=0.001). The capacity to repair irradiation-induced DNA damage was the highest in the low exposure group (1.34±1.00 SSB/10⁹ Da), followed by high exposure group (0.72±0.81 SSB/10⁹ Da) and controls (0.65±0.82 SSB/10⁹ Da). The mRNA expression levels of XRCC1, hOGG1 and XPC negatively correlated with styrene concentrations in blood and at workplace (p<0.001) and positively with SSBs (p<0.001). Micronuclei were not affected by styrene exposure, but were higher in older persons and in women (p<0.001). In this study, we did not confirm previous findings on an increased DNA repair response to styrene-induced genotoxicity. However, negative correlations of SSBs and mRNA expression levels of XRCC1, hOGG1 and XPC with styrene exposure warrant further highly-targeted study. Copyright © 2010 Elsevier Inc. All rights reserved.

  19. Chitinase mRNA Levels Determined by QPCR in Crab-Eating Monkey (Macaca fascicularis) Tissues: Species-Specific Expression of Acidic Mammalian Chitinase and Chitotriosidase.

    PubMed

    Uehara, Maiko; Tabata, Eri; Ishii, Kazuhiro; Sawa, Akira; Ohno, Misa; Sakaguchi, Masayoshi; Matoska, Vaclav; Bauer, Peter O; Oyama, Fumitaka

    2018-05-09

    Mice and humans express two active chitinases: acidic mammalian chitinase (AMCase) and chitotriosidase (CHIT1). Both chitinases are thought to play important roles in specific pathophysiological conditions. The crab-eating monkey ( Macaca fascicularis ) is one of the most frequently used nonhuman primate models in basic and applied biomedical research. Here, we performed gene expression analysis of two chitinases in normal crab-eating monkey tissues by way of quantitative real-time polymerase chain reaction (qPCR) using a single standard DNA molecule. Levels of AMCase and CHIT1 messenger RNAs (mRNAs) were highest in the stomach and the lung, respectively, when compared to other tissues. Comparative gene expression analysis of mouse, monkey, and human using monkey⁻mouse⁻human hybrid standard DNA showed that the AMCase mRNA levels were exceptionally high in mouse and monkey stomachs while very low in the human stomach. As for the CHIT1 mRNA, we detected higher levels in the monkey lung when compared with those of mouse and human. The differences of mRNA expression between the species in the stomach tissues were basically reflecting the levels of the chitinolytic activities. These results indicate that gene expression of AMCase and CHIT1 differs between mammalian species and requiring special attention in handling data in chitinase-related studies in particular organisms.

  20. Gene expression in scrapie. Cloning of a new scrapie-responsive gene and the identification of increased levels of seven other mRNA transcripts.

    PubMed

    Dandoy-Dron, F; Guillo, F; Benboudjema, L; Deslys, J P; Lasmézas, C; Dormont, D; Tovey, M G; Dron, M

    1998-03-27

    To define genes associated with or responsible for the neurodegenerative changes observed in transmissible spongiform encephalopathies, we analyzed gene expression in scrapie-infected mouse brain using "mRNA differential display." The RNA transcripts of eight genes were increased 3-8-fold in the brains of scrapie-infected animals. Five of these genes have not previously been reported to exhibit increased expression in this disease: cathepsin S, the C1q B-chain of complement, apolipoprotein D, and two previously unidentified genes denominated scrapie-responsive gene (ScRG)-1 and ScRG-2, which are preferentially expressed in brain tissue. Increased expression of the three remaining genes, beta2 microglobulin, F4/80, and metallothionein II, has previously been reported to occur in experimental scrapie. Kinetic analysis revealed a concomitant increase in the levels of ScRG-1, cathepsin S, the C1q B-chain of complement, and beta2 microglobulin mRNA as well as glial fibrillary acidic protein and F4/80 transcripts, markers of astrocytosis and microglial activation, respectively. In contrast, the level of ScRG-2, apolipoprotein D, and metallothionein II mRNA was only increased at the terminal stage of the disease. ScRG-1 mRNA was found to be preferentially expressed in glial cells and to code for a short protein of 47 amino acids with a strong hydrophobic N-terminal region.

  1. Increased duodenal DMT-1 expression and unchanged HFE mRNA levels in HFE-associated hereditary hemochromatosis and iron deficiency.

    PubMed

    Byrnes, V; Barrett, S; Ryan, E; Kelleher, T; O'Keane, C; Coughlan, B; Crowe, J

    2002-01-01

    HFE-associated hereditary hemochromatosis is characterized by imbalances of iron homeostasis and alterations in intestinal iron absorption. The identification of the HFE gene and the apical iron transporter divalent metal transporter-1, DMT-1, provide a direct method to address the mechanisms of iron overload in this disease. The aim of this study was to evaluate the regulation of duodenal HFE and DMT-1 gene expression in HFE-associated hereditary hemochromatosis. Small bowel biopsies and serum iron indices were obtained from a total of 33 patients. The study population comprised 13 patients with hereditary hemochromatosis (C282Y homozygous), 10 patients with iron deficiency anemia, and 10 apparently healthy controls, all of whom were genotyped for the two common mutations in the HFE gene (C282Y and H63D). Total RNA was isolated from tissue and amplified via RT-PCR for HFE, DMT-1, and the internal control GAPDH. DMT-1 protein expression was additionally assessed by immunohistochemistry. Levels of HFE mRNA did not differ significantly between patient groups (P = 0.09), specifically between C282Y homozygotes and iron deficiency anemic patients, when compared to controls (P = 0.09, P = 0.9, respectively). In contrast, DMT-1 mRNA levels were at least twofold greater in patients with hereditary hemochromatosis and iron deficiency anemia when compared to controls (P = 0.02, P = 0.01, respectively). Heightened DMT-1 protein expression correlated with mRNA levels in all patients. Loss of HFE function in hereditary hemochromatosis is not derived from inhibition of its gene expression. DMT-1 expression in C282Y homozygote subjects is consistent with the hypothesis of a "paradoxical" duodenal iron deficiency in hereditary hemochromatosis. The observed twofold upregulation of the DMT-1 is consistent with the slow but steady increase in body iron stores observed in those presenting with clinical features of hereditary hemochromatosis.

  2. Detection of functional polymorphisms influencing the promoter activity of the SAA2 gene and their association with milk production traits in Chinese Holstein cows.

    PubMed

    Yang, S; Li, C; Xie, Y; Cui, X; Li, X; Wei, J; Zhang, Y; Yu, Y; Wang, Y; Zhang, S; Zhang, Q; Sun, D

    2015-12-01

    Our previous RNA sequencing experiment showed that the serum amyloid A2 (SAA2) gene was one of the most promising candidates for milk protein and fat traits in dairy cattle. The SAA2 gene encodes an apolipoprotein related to high-density lipoproteins. To further validate its genetic effects, genotype-phenotype associations were performed in this study. Through resequencing of the entire coding region and the 5'-regulatory region of the SAA2 gene using pooled DNA of 12 unrelated sires, one novel 3-bp insertion-deletion and five previously reported SNPs were detected. These identified SNPs were genotyped and tested for association with five milk production-related traits in 717 Chinese Holstein cows. After Bonferroni correction for multiple t-tests, five of them were found to be statistically significant for milk yield, fat yield and protein yield (P < 0.0001~0.0053). Haplotype-based association analysis revealed a similar effect on fat yield and protein yield (P = 0.0005, P = 0.0032 respectively). Then, using luciferase report assay, the regulatory effect of the three SNPs located in the promoter region (c.-22G>A; c.17G>C; c.114G>A) was evaluated on transcriptional activity. In HEK-293 cell lines, we found that constructs GCG and AGG showed higher luciferase activity compared with GCA (P < 0.01, P < 0.01 respectively). Meanwhile, the prediction of the putative differential transcription factor binding site revealed that c.17G>C and c.114G>A caused the alteration in the transcription factor. Overall, the findings presented here provide the first evidence for associations of the SAA2 gene with milk fat and protein traits, which appears to be a key candidate for milk production traits in dairy cattle. © 2015 Stichting International Foundation for Animal Genetics.

  3. Salivary Alpha Amylase and Cortisol Levels in Children with Global Developmental Delay and Their Relation with the Expectation of Dental Care and Behavior during the Intervention

    ERIC Educational Resources Information Center

    dos Santos, Marcio Jose Possari; Bernabe, Daniel Galera; Nakamune, Ana Claudia de Melo Stevanato; Perri, Silvia Helena Venturoli; de Aguiar, Sandra Maria Herondina Coelho Avila; de Oliveira, Sandra Helena Penha

    2012-01-01

    The purpose of this study was to analyze the alpha-amylase (sAA) and cortisol levels in children with Global developmental delay (GDD) before and after dental treatment and its association with the children's behavior during treatment. The morning salivary cortisol levels and activity of sAA of 33 children with GDD were evaluated before and after…

  4. Comparison of plate-cage construct and stand-alone anchored spacer in the surgical treatment of three-level cervical spondylotic myelopathy: a preliminary clinical study.

    PubMed

    Shi, Sheng; Liu, Zu-De; Li, Xin-Feng; Qian, Lie; Zhong, Gui-Bin; Chen, Fang-Jing

    2015-09-01

    Although stand-alone cages were advocated to be superior to plate-cage construct (PCC) because of comparable clinical outcomes and fewer plate-related complications, cage dislocation and subsidence were frequently mentioned in multilevel fusion. There are some concerns about whether these issues can be effectively prevented in multilevel anterior cervical discectomy and fusion (ACDF) by stand-alone anchored spacer (SAAS). The aim was to compare clinical outcomes, radiologic parameters, and complications of PCC and SAAS in the treatment of three-level cervical spondylotic myelopathy (CSM). This was a retrospective comparative study. A total of 38 consecutive patients with three-level CSM (ACDF with PCC, 20 patients; ACDF with SAAS, 18 patients) were reviewed. Clinical outcomes were assessed using Japanese Orthopaedic Association and Neck Disability Index. The radiologic evaluations included cervical alignment (CA), segmental angle (SA), postoperative curvature loss (PCL), and incidence of subsidence. All the aforementioned parameters were compared before and after surgery between two groups. Besides, the aforementioned results were also compared between the two groups. The complications were also recorded. The mean follow-up period was 30.3 months. No significant differences were observed in clinical outcomes between the two groups (p>.05). Additionally, no significant differences existed in fusion rate between the two groups. There were significant differences in PCL of SA and CA and correction of SA between the two groups (p<.05). Besides, the incidence of subsidence (9 of 54 levels, 16.7%) was recorded in the SAAS group, and the potential of SAAS to reduce the incidence of postoperative dysphagia was not proven. No other complications were observed in this study. In the surgical treatment of three-level CSM, PCC is superior to SAAS in correction and maintenance of SA and avoiding cage subsidence, although the technique of ACDF with SAAS yielded encouraging

  5. Enzymatic fractionation of SAA-pretreated barley straw for production of fuel ethanol and astaxanthin as a value-added co-product

    USDA-ARS?s Scientific Manuscript database

    Barley straw was used to demonstrate a process for production of ethanol and astaxanthin as a value-added co-product. Barley straw was pretreated by soaking in aqueous ammonia (SAA) using the previously determined optimum conditions. The pretreated barley straw was first hydrolyzed with Accellerase®...

  6. [Transcription factors NF-kB, HIF-1, HIF-2, growth factor VEGF, VEGFR2 and carboanhydrase IX mRNA and protein level in the development of kidney cancer metastasis].

    PubMed

    Spirina, L V; Usynin, Y A; Yurmazov, Z A; Slonimskaya, E M; Kolegova, E S; Kondakova, I V

    2017-01-01

    Here, we have investigated the participation of nuclear factors NF-kB, HIF-1 and HIF-2, VEGF, VEGFR2, and carboanhydrase IX in clear-cell renal cancer. We have determined the expression and protein level of transcription factors, VEGF, VEGFR2, and carboanhydrase IX in tumor and normal tissues of 30 patients with kidney cancer. The Real-Time PCR and ELISA were used in the study. The low levels of HIF-1 mRNA expression associated with high levels of HIF-1 protein were also associated with metastasis. The expression levels of VEGF, VEGFR2, and their protein levels are increased in primary tumors of patients with disseminated kidney cancer compared to nonmetastatic cancer. No correlation was revealed between the content of mRNA and encoded proteins in the kidney cancer tissues. The changes in the ratios of mRNA levels and the respective proteins (HIF-1α, HIF-2, NF-kB, VEGF, VEGFR2, and carboanhydrase IX) may contribute to kidney-cancer metastasis.

  7. Global survey of mRNA levels and decay rates of Chlamydia trachomatis trachoma and lymphogranuloma venereum biovars.

    PubMed

    Ferreira, Rita; Borges, Vítor; Borrego, Maria José; Gomes, João Paulo

    2017-07-01

    Interpreting the intricate bacterial transcriptomics implies understanding the dynamic relationship established between de novo transcription and the degradation of transcripts. Here, we performed a comparative overview of gene expression levels and mRNA decay rates for different-biovar (trachoma and lymphogranuloma venereum) strains of the obligate intracellular bacterium Chlamydia trachomatis . By using RNA-sequencing to measure gene expression levels at mid developmental stage and mRNA decay rates upon rifampicin-based transcription blockage, we observed that: i ) 60-70% of the top-50 expressed genes encode proteins with unknown function and proteins involved in "Translation, ribosomal structure and biogenesis" for all strains; ii ) the expression ranking by genes' functional categories was in general concordant among different-biovar strains; iii ) the median of the half-life time (t 1/2 ) values of transcripts were 15-17 min, indicating that the degree of transcripts' stability seems to correlate with the bacterial intracellular life-style, as these values are considerably higher than the ones observed in other studies for facultative intracellular and free-living bacteria; iv ) transcript decay rates were highly heterogeneous within each C. trachomatis strain and did not correlate with steady-state expression levels; v ) only at very few instances (essentially at gene functional category level) was possible to unveil dissimilarities potentially underlying phenotypic differences between biovars. In summary, the unveiled transcriptomic scenario, marked by a general lack of correlation between transcript production and degradation and a huge inter-transcript heterogeneity in decay rates, likely reflects the challenges underlying the unique biphasic developmental cycle of C. trachomatis and its intricate interactions with the human host, which probably exacerbate the complexity of the bacterial transcription regulation.

  8. Correlation of mRNA and protein levels: Cell type-specific gene expression of cluster designation antigens in the prostate

    PubMed Central

    Pascal, Laura E; True, Lawrence D; Campbell, David S; Deutsch, Eric W; Risk, Michael; Coleman, Ilsa M; Eichner, Lillian J; Nelson, Peter S; Liu, Alvin Y

    2008-01-01

    Background: Expression levels of mRNA and protein by cell types exhibit a range of correlations for different genes. In this study, we compared levels of mRNA abundance for several cluster designation (CD) genes determined by gene arrays using magnetic sorted and laser-capture microdissected human prostate cells with levels of expression of the respective CD proteins determined by immunohistochemical staining in the major cell types of the prostate – basal epithelial, luminal epithelial, stromal fibromuscular, and endothelial – and for prostate precursor/stem cells and prostate carcinoma cells. Immunohistochemical stains of prostate tissues from more than 50 patients were scored for informative CD antigen expression and compared with cell-type specific transcriptomes. Results: Concordance between gene and protein expression findings based on 'present' vs. 'absent' calls ranged from 46 to 68%. Correlation of expression levels was poor to moderate (Pearson correlations ranged from 0 to 0.63). Divergence between the two data types was most frequently seen for genes whose array signals exceeded background (> 50) but lacked immunoreactivity by immunostaining. This could be due to multiple factors, e.g. low levels of protein expression, technological sensitivities, sample processing, probe set definition or anatomical origin of tissue and actual biological differences between transcript and protein abundance. Conclusion: Agreement between these two very different methodologies has great implications for their respective use in both molecular studies and clinical trials employing molecular biomarkers. PMID:18501003

  9. mRNA levels of enzymes and receptors implicated in arachidonic acid metabolism in gliomas.

    PubMed

    De Armas, Rafael; Durand, Karine; Guillaudeau, Angélique; Weinbreck, Nicolas; Robert, Sandrine; Moreau, Jean-Jacques; Caire, François; Acosta, Gisela; Pebet, Matias; Chaunavel, Alain; Marin, Benoît; Labrousse, François; Denizot, Yves

    2010-07-01

    Gliomas are tumors of the central nervous system derived from glial cells. They show cellular heterogeneity and lack specific diagnostic markers. Although a possible role for the eicosanoid cascade has been suggested in glioma tumorigenesis, the relationship between enzymes and receptors implicated in arachidonic acid metabolism, with histological tumor type has not yet been determined. Quantitative real-time reverse transcription-polymerase chain reaction was performed to measure and compare transcript levels of enzymes and receptors implicated in both lipoxygenase and cyclooxygenase pathways between oligodendrogliomas, astrocytomas, glioblastomas and mixed oligoastrocytomas. Arachidonic acid metabolism-related enzymes and receptor transcripts (i) were underexpressed in classical oligodendrogliomas compared to astrocytomas and/or glioblastomas, (ii) differed between astrocytomas and glioblastomas and (iii) had an intermediate expression in mixed oligoastrocytomas. mRNA levels of enzymes and receptors implicated both in lipoxygenase and cyclooxygenase pathways differed significantly in gliomas according to the histological type. Copyright 2010 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.

  10. TSHB mRNA is linked to cholesterol metabolism in adipose tissue.

    PubMed

    Moreno-Navarrete, José María; Moreno, María; Ortega, Francisco; Xifra, Gemma; Hong, Shangyu; Asara, John M; Serrano, José C E; Jové, Mariona; Pissios, Pavlos; Blüher, Matthias; Ricart, Wifredo; Portero-Otin, Manuel; Fernández-Real, José Manuel

    2017-10-01

    Subclinical hypothyroidism is known to be associated with increased serum cholesterol. Since thyroid-stimulating hormone (TSH) exerts an inductor effect on cholesterol biosynthesis, we aimed to investigate the relationship between TSH mRNA and cholesterol metabolism in human adipose tissue (AT). Cross-sectionally, AT TSH-β ( TSHB ) mRNA was evaluated in 4 independent cohorts in association with serum total and LDL cholesterol, and AT lipidomics. Longitudinally, the effects of statins and of diet and exercise on AT TSHB mRNA were also examined. The bidirectional relationship between cholesterol and TSHB were studied in isolated human adipocytes. TSHB mRNA was consistently detected in AT from euthyroid subjects, and positively associated with serum total- and LDL-cholesterol, and with AT-specific cholesterol metabolism-associated lipids [arachidonoyl cholesteryl ester, C8-dihydroceramide, N -stearoyl-d-sphingosine, and GlcCer(18:0, 24:1)]. Reduction of cholesterol with statins and with diet and exercise interventions led to decreased TSHB mRNA in human AT, whereas excess cholesterol up-regulated TSHB mRNA in human adipocytes. In addition, recombinant human TSH α/β administration resulted in increased HMGCR mRNA levels in human adipocytes. In mice, subcutaneous AT Tshb expression levels correlated directly with circulating cholesterol levels. In summary, current results provide novel evidence of TSHB as a paracrine factor that is modulated in parallel with cholesterol metabolism in human AT.-Moreno-Navarrete, J. M., Moreno, M., Ortega, F., Xifra, G., Hong, S., Asara, J. M., Serrano, J. C. E., Jové, M., Pissios, P., Blüher, M., Ricart, W., Portero-Otin, M., Fernández-Real, J. M. TSHB mRNA is linked to cholesterol metabolism in adipose tissue. © FASEB.

  11. Preprotachykinin A mRNA expression in the rat brain during development.

    PubMed

    Brené, S; Lindefors, N; Friedman, W J; Persson, H

    1990-12-15

    Expression of preprotachykinin A (PPT-A) mRNA was analyzed by northern blots using mRNA prepared from rat brain at 12 different developmental stages ranging from embryonic day 15 (E15) to adult. A single PPT-A mRNA of 1.3 kb was detected throughout development. PPT-A mRNA was detected as early as E15 and an approximately 3-fold increase occurred at birth. This amount remained until 3 weeks of age when the level increased, reaching a peak at 5 weeks of age. Adult amounts were approximately 3-fold higher than the levels at birth. The distribution of PPT-A mRNA-expressing cells in rat brain was studied by in situ hybridization on sections from embryonic day 20, postnatal days 4 and 7 as well as adult. Cells expressing PPT-A mRNA were detected in the forebrain at all 4 ages analyzed. However, the hybridization pattern and the labeling intensity varied in different brain regions during development. In cingulate cortex, intense labeling was seen in numerous cells at embryonic day 20 and postnatal days 4 and 7, whereas in the adult cingulate cortex only a few scattered labeled cells were observed. In frontoparietal cortex labeled cells were found from postnatal day 4 to adult, with the highest density of labeled cells at P7. Developmental differences in both the distribution of PPT-A mRNA-expressing cells and the level of PPT-A mRNA expression were also found in caudate-putamen, lateral hypothalamus and amygdala. Thus, our results show several changes in PPT-A mRNA expression during ontogeny, indicating a region and time-specific regulation of PPT-A mRNA expression during brain maturation.

  12. A Nascent Peptide Signal Responsive to Endogenous Levels of Polyamines Acts to Stimulate Regulatory Frameshifting on Antizyme mRNA.

    PubMed

    Yordanova, Martina M; Wu, Cheng; Andreev, Dmitry E; Sachs, Matthew S; Atkins, John F

    2015-07-17

    The protein antizyme is a negative regulator of cellular polyamine concentrations from yeast to mammals. Synthesis of functional antizyme requires programmed +1 ribosomal frameshifting at the 3' end of the first of two partially overlapping ORFs. The frameshift is the sensor and effector in an autoregulatory circuit. Except for Saccharomyces cerevisiae antizyme mRNA, the frameshift site alone only supports low levels of frameshifting. The high levels usually observed depend on the presence of cis-acting stimulatory elements located 5' and 3' of the frameshift site. Antizyme genes from different evolutionary branches have evolved different stimulatory elements. Prior and new multiple alignments of fungal antizyme mRNA sequences from the Agaricomycetes class of Basidiomycota show a distinct pattern of conservation 5' of the frameshift site consistent with a function at the amino acid level. As shown here when tested in Schizosaccharomyces pombe and mammalian HEK293T cells, the 5' part of this conserved sequence acts at the nascent peptide level to stimulate the frameshifting, without involving stalling detectable by toe-printing. However, the peptide is only part of the signal. The 3' part of the stimulator functions largely independently and acts at least mostly at the nucleotide level. When polyamine levels were varied, the stimulatory effect was seen to be especially responsive in the endogenous polyamine concentration range, and this effect may be more general. A conserved RNA secondary structure 3' of the frameshift site has weaker stimulatory and polyamine sensitizing effects on frameshifting. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  13. Immediate-early gene response to repeated immobilization: Fos protein and arc mRNA levels appear to be less sensitive than c-fos mRNA to adaptation.

    PubMed

    Ons, Sheila; Rotllant, David; Marín-Blasco, Ignacio J; Armario, Antonio

    2010-06-01

    Stress exposure resulted in brain induction of immediate-early genes (IEGs), considered as markers of neuronal activation. Upon repeated exposure to the same stressor, reduction of IEG response (adaptation) has been often observed, but there are important discrepancies in literature that may be in part related to the particular IEG and methodology used. We studied the differential pattern of adaptation of the IEGs c-fos and arc (activity-regulated cytoskeleton-associated protein) after repeated exposure to a severe stressor: immobilization on wooden boards (IMO). Rats repeatedly exposed to IMO showed reduced c-fos mRNA levels in response to acute IMO in most brain areas studied: the medial prefrontal cortex (mPFC), lateral septum (LS), medial amygdala (MeA), paraventricular nucleus of the hypothalamus (PVN) and locus coeruleus. In contrast, the number of neurons showing Fos-like immunoreactivity was only reduced in the MeA and the various subregions of the PVN. IMO-induced increases in arc gene expression were restricted to telencephalic regions and reduced by repeated IMO only in the mPFC. Double-labelling in the LS of IMO-exposed rats revealed that arc was expressed in only one-third of Fos+ neurons, suggesting two populations of Fos+ neurons. These data suggest that c-fos mRNA levels are more affected by repeated IMO than corresponding protein, and that arc gene expression does not reflect adaptation in most brain regions, which may be related to its constitutive expression. Therefore, the choice of a particular IEG and the method of measurement are important for proper interpretation of the impact of chronic repeated stress on brain activation.

  14. Quantitative assessment of hTERT mRNA expression in dysplastic nodules of HBV-related hepatocarcinogenesis.

    PubMed

    Oh, Bong-Kyeong; Kim, Young-Joo; Park, Young Nyun; Choi, Jinsub; Kim, Kyung Sik; Park, Chanil

    2006-04-01

    Telomerase reverse transcriptase (hTERT) is the rate-limiting determinant of telomerase, which is critical for carcinogenesis. Dysplastic nodules (DNs) appear to be preneoplastic lesions of hepatocellular carcinomas (HCCs). In this study, in order to characterize DNs, hTERT mRNA, hTERT gene dosage, and mRNA for c-myc, a transcriptional activator of hTERT were studied in human multi-step hepatocarcinogenesis. Fifty four hepatic nodules including 5 large regenerative nodules, 14 low-grade DNs, 7 high-grade DNs, 11 DNs with HCC foci and 17 HCCs, 23 livers with chronic hepatitis/cirrhosis, and 6 normal livers were examined. Transcript levels were measured by real-time quantitative RT-PCR and gene dosages by real-time PCR and Southern blotting. The hTERT mRNA levels increased with the progression of hepatocarcinogenesis, and a significant induction in the transition between low- and high-grade DNs was seen. Most high-grade DNs strongly expressed hTERT mRNA at levels similar to those of HCCs. Twenty-one percent of low-grade DNs had high levels of hTERT mRNA, up to those of high-grade DNs and there was no difference in the pathological features between low-grade DNs with and without increased hTERT mRNA levels. No correlation was found between hTERT mRNA levels, hTERT gene dosage, and c-myc mRNA levels. These results suggest that the induction of hTERT mRNA is an important early event and that its measurement by real-time quantitative RT-PCR is a useful tool to detect premalignant/malignant tendencies in hepatic nodules. However, hTERT gene dosage and c-myc expression are not the main mechanisms regulating hTERT expression in hepatocarcinogenesis.

  15. Effect of conventional and intensified milk replacer feeding programs on performance, vaccination response, and neutrophil mRNA levels of Holstein calves.

    PubMed

    Hengst, B A; Nemec, L M; Rastani, R R; Gressley, T F

    2012-09-01

    This study compared conventional and intensified milk replacer feeding regimens on growth, intake, respiratory and fecal scores, vaccination response, and neutrophil mRNA levels. Holstein calves were randomly assigned to a 10-wk study on d 2 of life. Treatments were conventional (CON; n=8) and intensified (INT; n=7) milk replacer feeding programs. Conventional calves were fed a 20.8% crude protein and 21.0% fat milk replacer at 1.25% of birth body weight (BW) from wk 1 to 6 of life and 0.625% of birth BW during wk 7. A 29.3% crude protein and 16.2% fat milk replacer was fed to INT calves at 1.5% of birth BW during wk 1, 2% of current BW from wk 2 to 6, and 1% of current BW during wk 7. All calves were given milk replacer twice daily during wk 1 to 6, once daily during wk 7, and were weaned completely during wk 8. Calf starter intake was measured daily through wk 8. Body weight and withers height were measured weekly. Fecal and respiratory scores were recorded twice daily at feeding. Calves were vaccinated against ovalbumin at the end of wk 1, 3, and 5. Blood samples were collected at the end of wk 1, 3, 5, and 8 for analysis of serum anti-ovalbumin IgG concentration and for isolation of neutrophils. Quantitative PCR was used to measure neutrophil mRNA levels of 7 functionality genes. Treatment did not affect total DMI or anti-ovalbumin IgG response. Intensified milk replacer feeding increased average daily gain, protein intake, fat intake, and feed efficiency compared with the CON feeding program. Compared with CON calves, INT calves had greater fecal scores, indicating looser feces and greater respiratory scores, indicating more respiratory problems. Calves assigned to the INT treatment had increased neutrophil mRNA levels of L-selectin, and at wk 8, neutrophil cytosolic factor 1 was increased and toll-like receptor 4 tended to be increased compared with CON calves. This suggests greater activation of neutrophils in INT calves postweaning, but differences were

  16. The rapid destabilization of p53 mRNA in immortal chicken embryo fibroblast cells.

    PubMed

    Kim, H; You, S; Foster, L K; Farris, J; Foster, D N

    2001-08-23

    The steady-state levels of p53 mRNA were dramatically lower in immortal chicken embryo fibroblast (CEF) cell lines compared to primary CEF cells. In the presence of cycloheximide (CHX), the steady-state levels of p53 mRNA markedly increased in immortal CEF cell lines, similar to levels found in primary cells. The de novo synthetic rates of p53 mRNA were relatively similar in primary and immortal cells grown in the presence or absence of CHX. Destabilization of p53 mRNA was observed in the nuclei of immortal, but not primary, CEF cells. The half-life of p53 mRNA in primary cells was found to be a relatively long 23 h compared to only 3 h in immortal cells. The expression of transfected p53 cDNA was inhibited in immortal cells, but restored upon CHX treatment. The 5'-region of the p53 mRNA was shown to be involved in the rapid p53 mRNA destabilization in immortal cells by expression analysis of 5'- and 3'-deleted p53 cDNAs as well as fusion mRNA constructs of N-terminal p53 and N-terminal deleted LacZ genes. Together, it is suggestive that the downregulation of p53 mRNA in immortal CEF cells occurs through a post-transcriptional destabilizing mechanism.

  17. Intake of branched-chain amino acids influences the levels of MAFbx mRNA and MuRF-1 total protein in resting and exercising human muscle.

    PubMed

    Borgenvik, Marcus; Apró, William; Blomstrand, Eva

    2012-03-01

    Resistance exercise and amino acids are two major factors that influence muscle protein turnover. Here, we examined the effects of resistance exercise and branched-chain amino acids (BCAA), individually and in combination, on the expression of anabolic and catabolic genes in human skeletal muscle. Seven subjects performed two sessions of unilateral leg press exercise with randomized supplementation with BCAA or flavored water. Biopsies were collected from the vastus lateralis muscle of both the resting and exercising legs before and repeatedly after exercise to determine levels of mRNA, protein phosphorylation, and amino acid concentrations. Intake of BCAA reduced (P < 0.05) MAFbx mRNA by 30 and 50% in the resting and exercising legs, respectively. The level of MuRF-1 mRNA was elevated (P < 0.05) in the exercising leg two- and threefold under the placebo and BCAA conditions, respectively, whereas MuRF-1 total protein increased by 20% (P < 0.05) only in the placebo condition. Phosphorylation of p70(S6k) increased to a larger extent (∼2-fold; P < 0.05) in the early recovery period with BCAA supplementation, whereas the expression of genes regulating mTOR activity was not influenced by BCAA. Muscle levels of phenylalanine and tyrosine were reduced (13-17%) throughout recovery (P < 0.05) in the placebo condition and to a greater extent (32-43%; P < 0.05) following BCAA supplementation in both resting and exercising muscle. In conclusion, BCAA ingestion reduced MAFbx mRNA and prevented the exercise-induced increase in MuRF-1 total protein in both resting and exercising leg. Further-more, resistance exercise differently influenced MAFbx and MuRF-1 mRNA expression, suggesting both common and divergent regulation of these two ubiquitin ligases.

  18. Is Having Low Motivation the Same as Not Having High Motivation? Comparing the CSAS-R and the SAAS-R

    ERIC Educational Resources Information Center

    McCoach, Betsy; Newton, Sarah D.; Siegle, Del; Baslanti, Ugur; Picho, Katherine

    2016-01-01

    The purpose of the current study was to examine the relationships among the Challenges to Scholastic Achievement Scale-Revised (CSAS-R) and The School Attitude Assessment Survey-Revised (SAAS-R) items and factors to determine whether the negative scales on the CSAS-R seemed to be measuring the same constructs as those that are measured on the…

  19. Variations in endothelin receptor B subtype 2 (EDNRB2) coding sequences and mRNA expression levels in 4 Muscovy duck plumage colour phenotypes.

    PubMed

    Wu, N; Qin, H; Wang, M; Bian, Y; Dong, B; Sun, G; Zhao, W; Chang, G; Xu, Q; Chen, G

    2017-04-01

    1. Endothelin receptor B subtype 2 (EDNRB2) is a paralog of EDNRB, which encodes a 7-transmembrane G-protein coupled receptor. Previous studies reported that EDNRB was essential for melanoblast migration in mammals and ducks. 2. Muscovy ducks have different plumage colour phenotypes. Variations in EDNRB2 coding sequences (CDSs) and mRNA expression levels were investigated in 4 different Muscovy duck plumage colour phenotypes, including black, black mutant, silver and white head. 3. The EDNRB2 gene from Muscovy duck was cloned; it had a length of 6435 bp and encoded 437 amino acids. The coding region was screened and potential single nucleotide polymorphisms were identified. Eight mutations were obtained, including one missense variant (c.64C > T) and 7 synonymous substitutions. The substitutions were associated with plumage colour phenotypes. 4. The EDNRB2 mRNA expression levels were compared between feather pulp from black birds and black mutant birds. The results indicated that EDNRB2 transcripts in feather pulp were significantly higher in black feathers than in white feathers. 5. The results determined the variation of EDNRB2 CDS and mRNA expression in Muscovy ducks of various plumage colours.

  20. Effects of environmental stress on mRNA expression levels of seven genes related to oxidative stress and growth in Atlantic salmon Salmo salar L. of farmed, hybrid and wild origin.

    PubMed

    Solberg, Monica F; Kvamme, Bjørn Olav; Nilsen, Frank; Glover, Kevin A

    2012-12-05

    Ten generations of domestication selection has caused farmed Atlantic salmon Salmo salar L. to deviate from wild salmon in a range of traits. Each year hundreds of thousands of farmed salmon escape into the wild. Thus, interbreeding between farmed escapees and wild conspecifics represents a significant threat to the genetic integrity of wild salmon populations. In a previous study we demonstrated how domestication has inadvertently selected for reduced responsiveness to stress in farmed salmon. To complement that study, we have evaluated the expression of seven stress-related genes in head kidney of salmon of farmed, hybrid and wild origin exposed to environmentally induced stress. In general, the crowding stressor used to induce environmental stress did not have a strong impact on mRNA expression levels of the seven genes, except for insulin-like growth factor-1 (IGF-1) that was downregulated in the stress treatment relative to the control treatment. mRNA expression levels of glutathione reductase (GR), Cu/Zn superoxide dismutase (Cu/Zn SOD), Mn superoxide dismutase (Mn SOD), glutathione peroxidase (GP) and IGF-1 were affected by genetic origin, thus expressed significantly different between the salmon of farmed, hybrid or wild origin. A positive relationship was detected between body size of wild salmon and mRNA expression level of the IGF-1 gene, in both environments. No such relationship was observed for the hybrid or farmed salmon. Farmed salmon in this study displayed significantly elevated mRNA levels of the IGF-1 gene relative to the wild salmon, in both treatments, while hybrids displayed a non additive pattern of inheritance. As IGF-1 mRNA levels are positively correlated to growth rate, the observed positive relationship between body size and IGF-1 mRNA levels detected in the wild but neither in the farmed nor the hybrid salmon, could indicate that growth selection has increased IGF-1 levels in farmed salmon to the extent that they may not be limiting

  1. Effects of environmental stress on mRNA expression levels of seven genes related to oxidative stress and growth in Atlantic salmon Salmo salar L. of farmed, hybrid and wild origin

    PubMed Central

    2012-01-01

    Background Ten generations of domestication selection has caused farmed Atlantic salmon Salmo salar L. to deviate from wild salmon in a range of traits. Each year hundreds of thousands of farmed salmon escape into the wild. Thus, interbreeding between farmed escapees and wild conspecifics represents a significant threat to the genetic integrity of wild salmon populations. In a previous study we demonstrated how domestication has inadvertently selected for reduced responsiveness to stress in farmed salmon. To complement that study, we have evaluated the expression of seven stress-related genes in head kidney of salmon of farmed, hybrid and wild origin exposed to environmentally induced stress. Results In general, the crowding stressor used to induce environmental stress did not have a strong impact on mRNA expression levels of the seven genes, except for insulin-like growth factor-1 (IGF-1) that was downregulated in the stress treatment relative to the control treatment. mRNA expression levels of glutathione reductase (GR), Cu/Zn superoxide dismutase (Cu/Zn SOD), Mn superoxide dismutase (Mn SOD), glutathione peroxidase (GP) and IGF-1 were affected by genetic origin, thus expressed significantly different between the salmon of farmed, hybrid or wild origin. A positive relationship was detected between body size of wild salmon and mRNA expression level of the IGF-1 gene, in both environments. No such relationship was observed for the hybrid or farmed salmon. Conclusion Farmed salmon in this study displayed significantly elevated mRNA levels of the IGF-1 gene relative to the wild salmon, in both treatments, while hybrids displayed a non additive pattern of inheritance. As IGF-1 mRNA levels are positively correlated to growth rate, the observed positive relationship between body size and IGF-1 mRNA levels detected in the wild but neither in the farmed nor the hybrid salmon, could indicate that growth selection has increased IGF-1 levels in farmed salmon to the extent

  2. Attenuation of Proinflammatory Responses by S-[6]-Gingerol via Inhibition of ROS/NF-Kappa B/COX2 Activation in HuH7 Cells.

    PubMed

    Li, Xiao-Hong; McGrath, Kristine C Y; Tran, Van H; Li, Yi-Ming; Duke, Colin C; Roufogalis, Basil D; Heather, Alison K

    2013-01-01

    Introduction. Hepatic inflammation underlies the pathogenesis of chronic diseases such as insulin resistance and type 2 diabetes mellitus. S-[6]-Gingerol has been shown to have anti-inflammatory properties. Important inflammatory mediators of interleukins include nuclear factor κ B (NF κ B) and cyclooxygenase 2 (COX2). We now explore the mechanism of anti-inflammatory effects of S-[6]-gingerol in liver cells. Methods. HuH7 cells were stimulated with IL1β to establish an in vitro hepatic inflammatory model. Results. S-[6]-Gingerol attenuated IL1β-induced inflammation and oxidative stress in HuH7 cells, as evidenced by decreasing mRNA levels of inflammatory factor IL6, IL8, and SAA1, suppression of ROS generation, and increasing mRNA levels of DHCR24. In addition, S-[6]-gingerol reduced IL1β-induced COX2 upregulation as well as NF κ B activity. Similar to the protective effects of S-[6]-gingerol, both NS-398 (a selective COX2 inhibitor) and PDTC (a selective NF κ B inhibitor) suppressed mRNA levels of IL6, IL8, and SAA1. Importantly, PDTC attenuated IL1β-induced overexpression of COX2. Of particular note, the protective effect of S-[6]-gingerol against the IL1β-induced inflammatory response was similar to that of BHT, an ROS scavenger. Conclusions. The findings of this study demonstrate that S-[6]-gingerol protects HuH7 cells against IL1β-induced inflammatory insults through inhibition of the ROS/NF κ B/COX2 pathway.

  3. Changes in liver PPARalpha mRNA expression in response to two levels of high-safflower-oil diets correlate with changes in adiposity and serum leptin in rats and mice.

    PubMed

    Hsu, Shan-Ching; Huang, Ching-jang

    2007-02-01

    The ligand-dependent transcription factor peroxisome proliferator-activated receptor alpha (PPARalpha) is known to be activated by common fatty acids and to regulate the expression of genes of various lipid oxidation pathways and transport. High-fat diets provide more fatty acids, which presumably could enhance lipid catabolism through up-regulation of PPARalpha signaling. However, high intake of fat could also lead to obesity. To examine PPARalpha signaling in high-fat feeding and obesity, this study examined the hepatic mRNA expression of PPARalpha and some of its target genes in Wistar rats and C57BL/6J mice fed two levels (20% or 30% wt/wt) of high-safflower-oil (SFO; oleic-acid-rich) diets until animals showed significantly higher body weight (13 weeks for rats and 22 weeks for mice) than those of control groups fed a 5% SFO diet. At the end of these respective feeding periods, only the rats fed 30% SFO and the mice fed 20% SFO among the two groups fed high-fat diets showed significantly higher body weight, white adipose tissue weight, serum leptin and mRNA expression of PPARalpha (P<.05) compared to the respective control groups. Despite elevated acyl-CoA (a PPARalpha target gene) protein and activity in both groups fed high-fat diets, the mRNA expression level of most PPARalpha target genes examined correlated mainly to PPARalpha mRNA levels and not to fat intake or liver lipid levels. The observation that the liver PPARalpha mRNA expression in groups fed high-fat diets was significantly higher only in obese animals with elevated serum leptin implied that obesity and associated hyperleptinemia might have a stronger impact than dietary SFO intake per se on PPARalpha-regulated mRNA expression in the liver.

  4. Role of mRNA Methylation in Prostate Cancer

    DTIC Science & Technology

    2015-02-01

    position of adenosine (m6A) is a post-transcriptional modification of mRNA. However, little is known regarding the biological meanings of this epigenetic ...its level is altered in various cancer cell lines. FTO, the fat mass and obesity associated gene, was recently shown as an m6A demethylase. FTO gene...mRNA. However, little is known regarding the biological meanings of this epigenetic regulation of mRNA. Recent technological advances have made it

  5. Region specific regulation of glutamic acid decarboxylase mRNA expression by dopamine neurons in rat brain.

    PubMed

    Lindefors, N; Brene, S; Herrera-Marschitz, M; Persson, H

    1989-01-01

    In situ hybridization histochemistry and RNA blots were used to study the expression of glutamic acid decarboxylase (GAD) mRNA in rats with or without a unilateral lesion of midbrain dopamine neurons. Two populations of GAD mRNA positive neurons were found in the intact caudate-putamen, substantia nigra and fronto-parietal cortex. In caudate-putamen, only one out of ten of the GAD mRNA positive neurons expressed high levels, while in substantia nigra every second of the positive neurons expressed high levels of GAD mRNA. Relatively few, but intensively labelled neurons were found in the intact fronto-parietal cerebral cortex. In addition, one out of six of the GAD mRNA positive neurons in the fronto-parietal cortex showed a low labeling. On the ipsilateral side, the forebrain dopamine deafferentation induced an increase in the number of neurons expressing high levels of GAD mRNA in caudate-putamen, and a decrease in fronto-parietal cortex. A smaller decrease was also seen in substantia nigra. However, the total number of GAD mRNA positive neurons were not significantly changed in any of these brain regions. The changes in the levels of GAD mRNA after the dopamine lesion were confirmed by RNA blot analysis. Hence, midbrain dopamine neurons appear to control neuronal expression of GAD mRNA by a tonic down-regulation in a fraction of GAD mRNA positive neurons in caudate-putamen, and a tonic up-regulation in a fraction of GAD mRNA positive neurons in fronto-parietal cortex and substantia nigra.

  6. Increased Bone Marrow (BM) Plasma Level of Soluble CD30 and Correlations with BM Plasma Level of Interferon (IFN)-γ, CD4/CD8 T-Cell Ratio and Disease Severity in Aplastic Anemia

    PubMed Central

    Shi, Jun; Ge, Meili; Li, Xingxin; Shao, Yingqi; Yao, Jianfeng; Zheng, Yizhou

    2014-01-01

    Idiopathic aplastic anemia (AA) is an immune-mediated bone marrow failure syndrome. Immune abnormalities such as decreased lymphocyte counts, inverted CD4/CD8 T-cell ratio and increased IFN-γ-producing T cells have been found in AA. CD30, a surface protein belonging to the tumor necrosis factor receptor family and releasing from cell surface as a soluble form (sCD30) after activation, marks a subset of activated T cells secreting IFN-γ when exposed to allogeneic antigens. Our study found elevated BM plasma levels of sCD30 in patients with SAA, which were closely correlated with disease severity, including absolute lymphocyte count (ALC) and absolute netrophil count (ANC). We also noted that sCD30 levels were positively correlated with plasma IFN-γ levels and CD4/CD8 T-cell ratio in patients with SAA. In order to explain these phenomena, we stimulated T cells with alloantigen in vitro and found that CD30+ T cells were the major source of IFN-γ, and induced CD30+ T cells from patients with SAA produced significantly more IFN-γ than that from healthy individuals. In addition, increased proportion of CD8+ T cells in AA showed enhanced allogeneic response by the fact that they expressed more CD30 during allogeneic stimulation. sCD30 levels decreased in patients responded to immunosuppressive therapy. In conclusion, elevated BM plasma levels of sCD30 reflected the enhanced CD30+ T cell-mediated immune response in SAA. CD30 as a molecular marker that transiently expresses on IFN-γ-producing T cells, may participate in mediating bone marrow failure in AA, which also can facilitate our understanding of AA pathogenesis to identify new therapeutic targets. PMID:25383872

  7. Selective decline of Nogo mRNA in the aging brain.

    PubMed

    Trifunovski, Alexandra; Josephson, Anna; Bickford, Paula C; Olson, Lars; Brené, Stefan

    2006-06-26

    The Nogo system has recently been implicated not only in regeneration but also in modulating plasticity. One reason for declining memory functions in aging may be altered plasticity in the aged hippocampus and cortex cerebri. Therefore, we have examined the levels of mRNA encoding Nogo, OMgp and MAG, as well as the receptor components NgR, Lingo-1 and Troy in cortex and hippocampus of young (4 months), middle aged (16 months) and old (24 months) Fisher 344 rats. No significant changes of receptor components or the ligands OMgp or MAG were observed. Nogo mRNA, however, was significantly decreased in hippocampal subregions of aged animals. The specific decrease of Nogo mRNA levels in hippocampus and possibly cortex cerebri may relate to age-dependent decline of brain plasticity.

  8. UCP2 mRNA expression is dependent on glucose metabolism in pancreatic islets

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Dalgaard, Louise T., E-mail: ltd@ruc.dk; Department of Science, Systems and Models, Roskilde University

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer UCP2 mRNA levels are decreased in islets of Langerhans from glucokinase deficient mice. Black-Right-Pointing-Pointer UCP2 mRNA up-regulation by glucose is dependent on glucokinase. Black-Right-Pointing-Pointer Absence of UCP2 increases GSIS of glucokinase heterozygous pancreatic islets. Black-Right-Pointing-Pointer This may protect glucokinase deficient mice from hyperglycemic damages. -- Abstract: Uncoupling Protein 2 (UCP2) is expressed in the pancreatic {beta}-cell, where it partially uncouples the mitochondrial proton gradient, decreasing both ATP-production and glucose-stimulated insulin secretion (GSIS). Increased glucose levels up-regulate UCP2 mRNA and protein levels, but the mechanism for UCP2 up-regulation in response to increased glucose is unknown. The aim was tomore » examine the effects of glucokinase (GK) deficiency on UCP2 mRNA levels and to characterize the interaction between UCP2 and GK with regard to glucose-stimulated insulin secretion in pancreatic islets. UCP2 mRNA expression was reduced in GK+/- islets and GK heterozygosity prevented glucose-induced up-regulation of islet UCP2 mRNA. In contrast to UCP2 protein function UCP2 mRNA regulation was not dependent on superoxide generation, but rather on products of glucose metabolism, because MnTBAP, a superoxide dismutase mimetic, did not prevent the glucose-induced up-regulation of UCP2. Glucose-stimulated insulin secretion was increased in UCP2-/- and GK+/- islets compared with GK+/- islets and UCP2 deficiency improved glucose tolerance of GK+/- mice. Accordingly, UCP2 deficiency increased ATP-levels of GK+/- mice. Thus, the compensatory down-regulation of UCP2 is involved in preserving the insulin secretory capacity of GK mutant mice and might also be implicated in limiting disease progression in MODY2 patients.« less

  9. Induction of vitellogenin synthesis by estrogen in avian liver: relationship between level of vitellogenin mRNA and vitellogenin synthesis.

    PubMed Central

    Mullinix, K P; Wetekam, W; Deeley, R G; Gordon, J I; Meyers, M; Kent, K A; Goldberger, R F

    1976-01-01

    We have investigated the estrogen-mediated induction of vitellogenin synthesis in rooster liver. We compared the concentrations of vitellogenin messenger RNA (mRNA) in the liver with the concentrations of vitellogenin in the sera of roosters that had recieved various treatments with estrogen. We found no vitellogenin mRNA in the livers of the unstimulated roosters. An initial injection of estrogen was attended by de novo synthesis of vitellogenin mRNA in the liver and accumulation of vitellogenin in the serum. When vitellogenin was no longer present in the serum or liver (the "post-estrogen-serum-negative" state), the liver was found to contain appreciable amounts of vitellogenin mRNA. This mRNA was of the same size as that found in the liver of the rooster actively synthesizing vitellogenin in response to estrogen. Whereas vitellogenin mRNA was in large polysomes in the livers of the roosters actively synthesizing vitellogenin, the vitellogenin mRNA in the liver of the post-estrogen-serum-negative rooster was not associated with polysomes. The possible relevance of these findings to the fact that the rooster responds differently to a primary stimulation with estrogen than to subsequent stimulations is discussed. PMID:1064017

  10. mRNA Levels of Placental Iron and Zinc Transporter Genes Are Upregulated in Gambian Women with Low Iron and Zinc Status.

    PubMed

    Jobarteh, Modou Lamin; McArdle, Harry J; Holtrop, Grietje; Sise, Ebrima A; Prentice, Andrew M; Moore, Sophie E

    2017-07-01

    Background: The role of the placenta in regulating micronutrient transport in response to maternal status is poorly understood. Objective: We investigated the effect of prenatal nutritional supplementation on the regulation of placental iron and zinc transport. Methods: In a randomized trial in rural Gambia [ENID (Early Nutrition and Immune Development)], pregnant women were allocated to 1 of 4 nutritional intervention arms: 1 ) iron and folic acid (FeFol) tablets (FeFol group); 2 ) multiple micronutrient (MMN) tablets (MMN group); 3 ) protein energy (PE) as a lipid-based nutrient supplement (LNS; PE group); and 4 ) PE and MMN (PE+MMN group) as LNS. All arms included iron (60 mg/d) and folic acid (400 μg/d). The MMN and PE+MMN arms included 30 mg supplemental Zn/d. In a subgroup of ∼300 mother-infant pairs, we measured maternal iron status, mRNA levels of genes encoding for placental iron and zinc transport proteins, and cord blood iron levels. Results: Maternal plasma iron concentration in late pregnancy was 45% and 78% lower in the PE and PE+MMN groups compared to the FeFol and MMN groups, respectively ( P < 0.001). The mRNA levels of the placental iron uptake protein transferrin receptor 1 were 30-49% higher in the PE and PE+MMN arms than in the FeFol arm ( P < 0.031), and also higher in the PE+MMN arm (29%; P = 0.042) than in the MMN arm. Ferritin in infant cord blood was 18-22% lower in the LNS groups ( P < 0.024). Zinc supplementation in the MMN arm was associated with higher maternal plasma zinc concentrations (10% increase; P < 0.001) than in other intervention arms. mRNA levels for intracellular zinc-uptake proteins, in this case zrt, irt-like protein (ZIP) 4 and ZIP8, were 96-205% lower in the PE+MMN arm than in the intervention arms without added zinc ( P < 0.025). Furthermore, mRNA expression of ZIP1 was 85% lower in the PE+MMN group than in the PE group ( P = 0.003). Conclusion: In conditions of low maternal iron and in the absence of supplemental

  11. Maintenance of CCL5 mRNA stores by post-effector and memory CD8 T cells is dependent on transcription and is coupled to increased mRNA stability.

    PubMed

    Marçais, Antoine; Tomkowiak, Martine; Walzer, Thierry; Coupet, Charles-Antoine; Ravel-Chapuis, Aymeric; Marvel, Jacqueline

    2006-10-01

    Immunological memory is associated with the display of improved effector functions by cells of the adaptive immune system. The storage of untranslated mRNA coding for the CCL5 chemokine by CD8 memory cells is a new process supporting the immediate display of an effector function. Here, we show that, after induction during the primary response, high CCL5 mRNA levels are specifically preserved in CD8 T cells. We have investigated the mechanisms involved in the long-term maintenance of CCL5 mRNA levels by memory CD8 T cells. We demonstrate that the CCL5 mRNA half-life is increased in memory CD8 T cells and that these cells constitutively transcribe ccl5 gene. By inhibiting ccl5 transcription using IL-4, we demonstrate the essential role of transcription in the maintenance of CCL5 mRNA stores. Finally, we show that these stores are spontaneously reconstituted when the inhibitory signal is removed, indicating that the transcription of ccl5 is a default feature of memory CD8 T cells imprinted in their genetic program.

  12. L-DOPA decarboxylase mRNA levels provide high diagnostic accuracy and discrimination between clear cell and non-clear cell subtypes in renal cell carcinoma.

    PubMed

    Papadopoulos, Emmanuel I; Petraki, Constantina; Gregorakis, Alkiviadis; Chra, Eleni; Fragoulis, Emmanuel G; Scorilas, Andreas

    2015-06-01

    Renal cell carcinoma (RCC) is the most frequent type of kidney cancer. RCC patients frequently present with arterial hypertension due to various causes, including intrarenal dopamine deficiency. L-DOPA decarboxylase (DDC) is the gene encoding the enzyme that catalyzes the biosynthesis of dopamine in humans. Several studies have shown that the expression levels of DDC are significantly deregulated in cancer. Thus, we herein sought to analyze the mRNA levels of DDC and evaluate their clinical significance in RCC. DDC levels were analyzed in 58 surgically resected RCC tumors and 44 adjacent non-cancerous renal tissue specimens via real-time PCR. Relative levels of DDC were estimated by applying the 2(-ΔΔC)T method, while their diagnostic accuracy and correlation with the clinicopathological features of RCC tumors were assessed by comprehensive statistical analysis. DDC mRNA levels were found to be dramatically downregulated (p<0.001) in RCC tumors, exhibiting remarkable diagnostic accuracy as assessed by ROC curve analysis (AUC: 0.910; p<0.001) and logistic regression (OR: 0.678; p=0.001). Likewise, DDC was found to be differentially expressed between clear cell RCC and the group of non-clear cell subtypes (p=0.001) consisted of papillary and chromophobe RCC specimens. Furthermore, a statistically significant inverse correlation was also observed when the mRNA levels of DDC were analyzed in relation to tumor grade (p=0.049). Our data showed that DDC constitutes a highly promising molecular marker for RCC, exhibiting remarkable diagnostic accuracy and potential to discriminate between clear cell and non-clear cell histological subtypes of RCC. Copyright © 2015. Published by Elsevier Inc.

  13. Biomaterials for mRNA Delivery

    PubMed Central

    Islam, Mohammad Ariful; Reesor, Emma K. G.; Xu, Yingjie; Zope, Harshal R.; Zetter, Bruce R.; Shi, Jinjun

    2015-01-01

    Messenger RNA (mRNA) has recently emerged with remarkable potential as an effective alternative to DNA-based therapies because of several unique advantages. mRNA does not require nuclear entry for transfection activity and has a negligible chance of integrating into the host genome which excludes the possibility of potentially detrimental genomic alternations. Chemical modification of mRNA has further enhanced its stability and decreased its activation of innate immune responses. Additionally, mRNA has been found to have rapid expression and predictable kinetics. Nevertheless, the ubiquitous application of mRNA remains challenging given its unfavorable attributes, such as large size, negative charge and susceptibility to enzymatic degradation. Further refinement of mRNA delivery modalities is therefore essential for its development as a therapeutic tool. This review provides an exclusive overview of current state-of-the-art biomaterials and nanotechnology platforms for mRNA delivery, and discusses future prospects to bring these exciting technologies into clinical practice. PMID:26280625

  14. Tau mRNA 3'UTR-to-CDS ratio is increased in Alzheimer disease.

    PubMed

    García-Escudero, Vega; Gargini, Ricardo; Martín-Maestro, Patricia; García, Esther; García-Escudero, Ramón; Avila, Jesús

    2017-08-10

    Neurons frequently show an imbalance in expression of the 3' untranslated region (3'UTR) relative to the coding DNA sequence (CDS) region of mature messenger RNAs (mRNA). The ratio varies among different cells or parts of the brain. The Map2 protein levels per cell depend on the 3'UTR-to-CDS ratio rather than the total mRNA amount, which suggests powerful regulation of protein expression by 3'UTR sequences. Here we found that MAPT (the microtubule-associated protein tau gene) 3'UTR levels are particularly high with respect to other genes; indeed, the 3'UTR-to-CDS ratio of MAPT is balanced in healthy brain in mouse and human. The tau protein accumulates in Alzheimer diseased brain. We nonetheless observed that the levels of RNA encoding MAPT/tau were diminished in these patients' brains. To explain this apparently contradictory result, we studied MAPT mRNA stoichiometry in coding and non-coding regions, and found that the 3'UTR-to-CDS ratio was higher in the hippocampus of Alzheimer disease patients, with higher tau protein but lower total mRNA levels. Our data indicate that changes in the 3'UTR-to-CDS ratio have a regulatory role in the disease. Future research should thus consider not only mRNA levels, but also the ratios between coding and non-coding regions. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Interconnections between mRNA degradation and RDR-dependent siRNA production in mRNA turnover in plants.

    PubMed

    Tsuzuki, Masayuki; Motomura, Kazuki; Kumakura, Naoyoshi; Takeda, Atsushi

    2017-03-01

    Accumulation of an mRNA species is determined by the balance between the synthesis and the degradation of the mRNA. Individual mRNA molecules are selectively and actively degraded through RNA degradation pathways, which include 5'-3' mRNA degradation pathway, 3'-5' mRNA degradation pathway, and RNA-dependent RNA polymerase-mediated mRNA degradation pathway. Recent studies have revealed that these RNA degradation pathways compete with each other in mRNA turnover in plants and that plants have a hidden layer of non-coding small-interfering RNA production from a set of mRNAs. In this review, we summarize the current information about plant mRNA degradation pathways in mRNA turnover and discuss the potential roles of a novel class of the endogenous siRNAs derived from plant mRNAs.

  16. Gibberellin (GA3) enhances cell wall invertase activity and mRNA levels in elongating dwarf pea (Pisum sativum) shoots

    NASA Technical Reports Server (NTRS)

    Wu, L. L.; Mitchell, J. P.; Cohn, N. S.; Kaufman, P. B.

    1993-01-01

    The invertase (EC 3.2.1.26) purified from cell walls of dwarf pea stems to homogeneity has a molecular mass of 64 kilodaltons (kD). Poly(A)+RNA was isolated from shoots of dwarf pea plants, and a cDNA library was constructed using lambda gt11 as an expression vector. The expression cDNA library was screened with polyclonal antibodies against pea cell wall invertase. One invertase cDNA clone was characterized as a full-length cDNA with 1,863 base pairs. Compared with other known invertases, one homologous region in the amino acid sequence was found. The conserved motif, Asn-Asp-Pro-Asn-Gly, is located near the N-terminal end of invertase. Northern blot analysis showed that the amount of invertase mRNA (1.86 kb) was rapidly induced to a maximal level 4 h after GA3 treatment, then gradually decreased to the control level. The mRNA level at 4 h in GA3-treated peas was fivefold higher than that of the control group. The maximal increase in activity of pea cell wall invertase elicited by GA3 occcured at 8 h after GA3 treatment. This invertase isoform was shown immunocytochemically to be localized in the cell walls, where a 10-fold higher accumulation occurred in GA3-treated tissue compared with control tissue. This study indicates that the expression of the pea shoot cell-wall invertase gene could be regulated by GA3 at transcriptional and/or translational levels.

  17. Whole-genome analysis of mRNA decay in Plasmodium falciparum reveals a global lengthening of mRNA half-life during the intra-erythrocytic development cycle.

    PubMed

    Shock, Jennifer L; Fischer, Kael F; DeRisi, Joseph L

    2007-01-01

    The rate of mRNA decay is an essential element of post-transcriptional regulation in all organisms. Previously, studies in several organisms found that the specific half-life of each mRNA is precisely related to its physiologic role, and plays an important role in determining levels of gene expression. We used a genome-wide approach to characterize mRNA decay in Plasmodium falciparum. We found that, globally, rates of mRNA decay increase dramatically during the asexual intra-erythrocytic developmental cycle. During the ring stage of the cycle, the average mRNA half-life was 9.5 min, but this was extended to an average of 65 min during the late schizont stage of development. Thus, a major determinant of mRNA decay rate appears to be linked to the stage of intra-erythrocytic development. Furthermore, we found specific variations in decay patterns superimposed upon the dominant trend of progressive half-life lengthening. These variations in decay pattern were frequently enriched for genes with specific cellular functions or processes. Elucidation of Plasmodium mRNA decay rates provides a key element for deciphering mechanisms of genetic control in this parasite, by complementing and extending previous mRNA abundance studies. Our results indicate that progressive stage-dependent decreases in mRNA decay rate function are a major determinant of mRNA accumulation during the schizont stage of intra-erythrocytic development. This type of genome-wide change in mRNA decay rate has not been observed in any other organism to date, and indicates that post-transcriptional regulation may be the dominant mechanism of gene regulation in P. falciparum.

  18. Effect of 50 Hz electric field in diacylglycerol acyltransferase mRNA expression level and plasma concentration of triacylglycerol, free fatty acid, phospholipid and total cholesterol

    PubMed Central

    2012-01-01

    Background The effects of exposure to a 50 Hz electric field (EF) on plasma level of triacylglycerol, free fatty acids, total cholesterol and phospholipid and mRNA expression level of diacylglycerol acyltransferase (DGAT) 1 and 2 in liver and intestines from C57BL/6 J mice were studied. Methods The test was based on comparison between mice post treated with 50 Hz EF of 45 kV/m intensity for 30 min per day for 11 days or without EF. DGATs mRNA expression was analyzed by real-time quantitative polymerase chain reaction. Results There was no difference in the gene expression level of DGAT1 in liver and intestines. The DGAT2 gene expression level in liver derived from mice treated with EF was significantly lower than those in the control (P < 0.001). Both plasma total cholesterol (P < 0.01) and phospholipid (P < 0.05) in the group exposed to EF were lower than those in the control, but there was no difference in triacylglycerol or free fatty acid levels. Conclusion Exposure to 50 Hz EF decrease the plasma levels of total cholesterol and phospholipids, and downregulated DGAT2 mRNA expression in liver. The mechanisms for the effects of EF on lipid metabolism are not well understand yet, but altered DGAT2 activity may be involved. PMID:22676350

  19. Oestradiol reduces Liver Receptor Homolog-1 mRNA transcript stability in breast cancer cell lines

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Lazarus, Kyren A.; Environmental and Biotechnology Centre, Swinburne University, Hawthorn, Victoria 3122; Zhao, Zhe

    2013-08-30

    Highlights: •LRH-1 is an orphan nuclear receptor that regulates tumor proliferation. •In breast cancer, high mRNA expression is associated with ER+ status. •In ER−ve cells, despite very low mRNA, we found abundant LRH-1 protein. •Our data show distinctly different LRH-1 protein isoforms in ER− and ER+ breast cancer cells. •This is due to differences in LRH-1 mRNA and protein stability rates. -- Abstract: The expression of orphan nuclear receptor Liver Receptor Homolog-1 (LRH-1) is elevated in breast cancer and promotes proliferation, migration and invasion in vitro. LRH-1 expression is regulated by oestrogen (E{sub 2}), with LRH-1 mRNA transcript levels highermore » in oestrogen receptor α (ERα) positive (ER+) breast cancer cells compared to ER− cells. However, the presence of LRH-1 protein in ER− cells suggests discordance between mRNA transcript levels and protein expression. To understand this, we investigated the impact of mRNA and protein stability in determining LRH-1 protein levels in breast cancer cells. LRH-1 transcript levels were significantly higher in ER+ versus ER− breast cancer cells lines; however LRH-1 protein was expressed at similar levels. We found LRH-1 mRNA and protein was more stable in ER− compared to ER+ cell lines. The tumor-specific LRH-1 variant isoform, LRH-1v4, which is highly responsive to E{sub 2}, showed increased mRNA stability in ER− versus ER+ cells. In addition, in MCF-7 and T47-D cell lines, LRH-1 total mRNA stability was reduced with E{sub 2} treatment, this effect mediated by ERα. Our data demonstrates that in ER− cells, increased mRNA and protein stability contribute to the abundant protein expression levels. Expression and immunolocalisation of LRH-1 in ER− cells as well as ER− tumors suggests a possible role in the development of ER− tumors. The modulation of LRH-1 bioactivity may therefore be beneficial as a treatment option in both ER− and ER+ breast cancer.« less

  20. The Co-Induced Effects of Molybdenum and Cadmium on the Trace Elements and the mRNA Expression Levels of CP and MT in Duck Testicles.

    PubMed

    Xia, Bing; Chen, Hua; Hu, Guoliang; Wang, Liqi; Cao, Huabin; Zhang, Caiying

    2016-02-01

    To investigate the chronic toxicity of molybdenum (Mo) and cadmium (Cd) on the trace elements and the mRNA expression levels of ceruloplasmin (CP) and metallothionein (MT) in duck testicles, 120 healthy 11-day-old male ducks were randomly divided into six groups with 20 ducks in each group. Ducks were treated with the diet containing different dosages of Mo or Cd. The source of Mo and Cd was hexaammonium molybdate ([(NH4)6Mo7O24·4H2O]) and cadmium sulfate (3CdSO4·8H2O), respectively, in this study. After being treated for 60 and 120 days, ten male birds in each group were randomly selected and euthanized and then testicles were aseptically collected for determining the mRNA expression levels of MT and CP, antioxidant indexes, and contents of trace elements in the testicle. In addition, testicle tissues at 120 days were subjected to histopathological analysis with the optical microscope. The results showed that co-exposure to Mo and Cd resulted in an increase in malondialdehyde (MDA) level while decrease in xanthine oxidase (XOD) and catalase (CAT) activities. The mRNA expression level of MT gene was upregulated while CP was decreased in combination groups. Contents of Mo, copper (Cu), iron (Fe), and zinc (Zn) decreased in combined groups while Cd increased in Cd and combined groups at 120 days. Furthermore, severe congestion, low sperm count, and malformation were observed in low dietary of Mo combined with Cd group and high dietary of Mo combined with Cd group. Our results suggested that Mo and Cd might aggravate testicular degeneration synergistically through altering the mRNA expression levels of MT and CP, increasing lipid peroxidation through inhibiting related enzyme activities and disturbing homeostasis of trace elements in testicles. Interaction of Mo and Cd may have a synergistic effect on the testicular toxicity.

  1. Pregnancy amelioration of arthritis in SKG mice corresponds with alterations in serum amyloid A3 levels.

    PubMed

    Shaw, Laura A; Stefanski, Adrianne L; Peterson, Lisa K; Rumer, Kristen K; Vondracek, Andrea; Phang, Tzu L; Sakaguchi, Shimon; Winn, Virginia D; Dragone, Leonard L

    2012-06-30

    OBJECTIVES: Pregnancy leads to rheumatoid arthritis remission in humans. The objective of this study was to determine if the SKG mouse could serve as a model for pregnancy-associated inflammatory arthritis amelioration. In addition, the maternal peripheral blood mononuclear cell (PBMC) transcriptome was assessed to define a biomarker associated with remission. METHODS: Cohorts of zymosan-treated pregnant SKG mice and controls were monitored for arthritis progression. Microarray analysis evaluated alterations in gene expression in maternal PBMCs at embryonic day 14.5 (E14.5) between arthritic and pregnancy-remitted mice. A selected target, serum amyloid A3 (SAA3), was further investigated using quantitative reverse transcriptase PCR (qRT-PCR) and an enzyme-linked immunosorbent assay (ELISA). RESULTS: Pregnancy resulted in complete or partial remission in the majority of the zymosan-treated SKG mice. Twenty-seven transcripts were differentially expressed in the PBMCs between arthritic and pregnancy-remitted mice. Expression and plasma SAA3 levels decreased with pregnancy-induced arthritis amelioration and plasma SAA3 levels correlated with arthritis severity. CONCLUSIONS: These results establish the SKG mouse as a model system to study pregnancy-induced amelioration of arthritis. These studies also establish SAA3 as a biomarker of arthritis amelioration in SKG mice. This model can be used to elucidate the molecular and cellular mechanisms underlying the impact of pregnancy on the maternal immune system that results in arthritis amelioration.

  2. Single-cell mRNA cytometry via sequence-specific nanoparticle clustering and trapping

    NASA Astrophysics Data System (ADS)

    Labib, Mahmoud; Mohamadi, Reza M.; Poudineh, Mahla; Ahmed, Sharif U.; Ivanov, Ivaylo; Huang, Ching-Lung; Moosavi, Maral; Sargent, Edward H.; Kelley, Shana O.

    2018-05-01

    Cell-to-cell variation in gene expression creates a need for techniques that can characterize expression at the level of individual cells. This is particularly true for rare circulating tumour cells, in which subtyping and drug resistance are of intense interest. Here we describe a method for cell analysis—single-cell mRNA cytometry—that enables the isolation of rare cells from whole blood as a function of target mRNA sequences. This approach uses two classes of magnetic particles that are labelled to selectively hybridize with different regions of the target mRNA. Hybridization leads to the formation of large magnetic clusters that remain localized within the cells of interest, thereby enabling the cells to be magnetically separated. Targeting specific intracellular mRNAs enablescirculating tumour cells to be distinguished from normal haematopoietic cells. No polymerase chain reaction amplification is required to determine RNA expression levels and genotype at the single-cell level, and minimal cell manipulation is required. To demonstrate this approach we use single-cell mRNA cytometry to detect clinically important sequences in prostate cancer specimens.

  3. mRNA stability in mammalian cells.

    PubMed Central

    Ross, J

    1995-01-01

    This review concerns how cytoplasmic mRNA half-lives are regulated and how mRNA decay rates influence gene expression. mRNA stability influences gene expression in virtually all organisms, from bacteria to mammals, and the abundance of a particular mRNA can fluctuate manyfold following a change in the mRNA half-life, without any change in transcription. The processes that regulate mRNA half-lives can, in turn, affect how cells grow, differentiate, and respond to their environment. Three major questions are addressed. Which sequences in mRNAs determine their half-lives? Which enzymes degrade mRNAs? Which (trans-acting) factors regulate mRNA stability, and how do they function? The following specific topics are discussed: techniques for measuring eukaryotic mRNA stability and for calculating decay constants, mRNA decay pathways, mRNases, proteins that bind to sequences shared among many mRNAs [like poly(A)- and AU-rich-binding proteins] and proteins that bind to specific mRNAs (like the c-myc coding-region determinant-binding protein), how environmental factors like hormones and growth factors affect mRNA stability, and how translation and mRNA stability are linked. Some perspectives and predictions for future research directions are summarized at the end. PMID:7565413

  4. Increased Levels of Cell-Free Human Placental Lactogen mRNA at 28-32 Gestational Weeks in Plasma of Pregnant Women With Placenta Previa and Invasive Placenta

    PubMed Central

    Sekizawa, Akihiko; Ventura, Walter; Koide, Keiko; Hori, Kyouko; Okai, Takashi; Masashi, Yoshida; Furuya, Kenichi; Mizumoto, Yoshifumi

    2014-01-01

    We compared the levels of cell-free human placental lactogen (hPL) messenger RNA (mRNA) in maternal plasma at 28 to 32 weeks of gestation between women with diagnosis of placenta previa or invasive placenta and women with an uneventful pregnancy. Sensitivity and specificity of hPL mRNA for the prediction of invasive placenta were further explored. Plasma hPL mRNA were quantified by real-time reverse-transcriptase polymerase chain reaction in women with placenta previa (n = 13), invasive placenta (n = 5), and normal pregnancies (n = 92). Median (range) hPL mRNA was significantly higher in women with placenta previa, 782 (10-2301) copies/mL of plasma, and in those with invasive placenta, 615 (522-2102) copies/mL of plasma, when compared to normal pregnancies, 90 (4-4407) copies/mL of plasma, P < .01 and P < .05, respectively. We found a sensitivity of 100% and a specificity of 61.5% for the prediction of invasive placenta among women with placenta previa. In conclusion, expression of hPL mRNA is increased in plasma of women with placenta previa and invasive placenta at 28 to 32 weeks of gestation. PMID:23744883

  5. Increased levels of cell-free human placental lactogen mRNA at 28-32 gestational weeks in plasma of pregnant women with placenta previa and invasive placenta.

    PubMed

    Kawashima, Akihiro; Sekizawa, Akihiko; Ventura, Walter; Koide, Keiko; Hori, Kyouko; Okai, Takashi; Masashi, Yoshida; Furuya, Kenichi; Mizumoto, Yoshifumi

    2014-02-01

    We compared the levels of cell-free human placental lactogen (hPL) messenger RNA (mRNA) in maternal plasma at 28 to 32 weeks of gestation between women with diagnosis of placenta previa or invasive placenta and women with an uneventful pregnancy. Sensitivity and specificity of hPL mRNA for the prediction of invasive placenta were further explored. Plasma hPL mRNA were quantified by real-time reverse-transcriptase polymerase chain reaction in women with placenta previa (n = 13), invasive placenta (n = 5), and normal pregnancies (n = 92). Median (range) hPL mRNA was significantly higher in women with placenta previa, 782 (10-2301) copies/mL of plasma, and in those with invasive placenta, 615 (522-2102) copies/mL of plasma, when compared to normal pregnancies, 90 (4-4407) copies/mL of plasma, P < .01 and P < .05, respectively. We found a sensitivity of 100% and a specificity of 61.5% for the prediction of invasive placenta among women with placenta previa. In conclusion, expression of hPL mRNA is increased in plasma of women with placenta previa and invasive placenta at 28 to 32 weeks of gestation.

  6. Immediate Effects of Traditional Thai Massage on Psychological Stress as Indicated by Salivary Alpha-Amylase Levels in Healthy Persons.

    PubMed

    Sripongngam, Thanarat; Eungpinichpong, Wichai; Sirivongs, Dhavee; Kanpittaya, Jaturat; Tangvoraphonkchai, Kamonwan; Chanaboon, Sutin

    2015-10-05

    BACKGROUND Stress can cause psychological and physiological changes. Many studies revealed that massage can decrease stress. However, traditional Thai massage has not been well researched in this regard. The purpose of this study was to investigate the immediate effects of traditional Thai massage (TTM) on salivary alpha-amylase levels (sAA), heart rate variability (HRV), autonomic nervous system (ANS) function, and plasma renin activity (PRA). MATERIAL AND METHODS Twenty-nine healthy participants were randomly allocated into either a traditional Thai massage (TTM) group or Control (C) group, after which they were switched to the other group with a 2-week wash-out period. Each of them was given a 10-minute mental arithmetic test to induce psychological stress before a 1-hour session of TTM or rest. RESULTS Within-groups comparison revealed that sAA was significantly decreased (p<0.05) in the TTM group but not in the C group. HRV and ANS function were significantly increased (p<0.05) and PRA was significantly decreased (p<0.05) in both groups. However, low frequency per high frequency ratio (LF/HF ratio) and ANS balance status were not changed. Only sAA was found to be significantly different between groups (p<0.05). CONCLUSIONS We conclude that both TTM and rest can reduce psychological stress, as indicated by decreased sAA levels, increased parasympathetic activity, decreased sympathetic activity, and decreased PRA. However, TTM may have a modest effect on stress reduction as indicated by a reduced sAA.

  7. Immediate Effects of Traditional Thai Massage on Psychological Stress as Indicated by Salivary Alpha-Amylase Levels in Healthy Persons

    PubMed Central

    Sripongngam, Thanarat; Eungpinichpong, Wichai; Sirivongs, Dhavee; Kanpittaya, Jaturat; Tangvoraphonkchai, Kamonwan; Chanaboon, Sutin

    2015-01-01

    Background Stress can cause psychological and physiological changes. Many studies revealed that massage can decrease stress. However, traditional Thai massage has not been well researched in this regard. The purpose of this study was to investigate the immediate effects of traditional Thai massage (TTM) on salivary alpha-amylase levels (sAA), heart rate variability (HRV), autonomic nervous system (ANS) function, and plasma renin activity (PRA). Material/Methods Twenty-nine healthy participants were randomly allocated into either a traditional Thai massage (TTM) group or Control (C) group, after which they were switched to the other group with a 2-week wash-out period. Each of them was given a 10-minute mental arithmetic test to induce psychological stress before a 1-hour session of TTM or rest. Results Within-groups comparison revealed that sAA was significantly decreased (p<0.05) in the TTM group but not in the C group. HRV and ANS function were significantly increased (p<0.05) and PRA was significantly decreased (p<0.05) in both groups. However, low frequency per high frequency ratio (LF/HF ratio) and ANS balance status were not changed. Only sAA was found to be significantly different between groups (p<0.05). Conclusions We conclude that both TTM and rest can reduce psychological stress, as indicated by decreased sAA levels, increased parasympathetic activity, decreased sympathetic activity, and decreased PRA. However, TTM may have a modest effect on stress reduction as indicated by a reduced sAA. PMID:26436433

  8. PTB and TIAR binding to insulin mRNA 3'- and 5'UTRs; implications for insulin biosynthesis and messenger stability.

    PubMed

    Fred, Rikard G; Mehrabi, Syrina; Adams, Christopher M; Welsh, Nils

    2016-09-01

    Insulin expression is highly controlled on the posttranscriptional level. The RNA binding proteins (RBPs) responsible for this result are still largely unknown. To identify RBPs that bind to insulin mRNA we performed mass spectrometry analysis on proteins that bound synthetic oligonucloetides mimicing the 5'- and the 3'-untranslated regions (UTRs) of rat and human insulin mRNA in vitro . We observed that the RBPs heterogeneous nuclear ribonucleoprotein (hnRNP) U, polypyrimidine tract binding protein (PTB), hnRNP L and T-cell restricted intracellular antigen 1-related protein (TIA-1-related protein; TIAR) bind to insulin mRNA sequences, and that the in vitro binding affinity of these RBPs changed when INS-1 cells were exposed to glucose, 3-isobutyl-1-methylxanthine (IBMX) or nitric oxide. High glucose exposure resulted in a modest increase in PTB and TIAR binding to an insulin mRNA sequence. The inducer of nitrosative stress DETAnonoate increased markedly hnRNP U and TIAR mRNA binding. An increased PTB to TIAR binding ratio in vitro correlated with higher insulin mRNA levels and insulin biosynthesis rates in INS-1 cells. To further investigate the importance of RNA-binding proteins for insulin mRNA stability, we decreased INS-1 and EndoC-βH1 cell levels of PTB and TIAR by RNAi. In both cell lines, decreased levels of PTB resulted in lowered insulin mRNA levels while decreased levels of TIAR resulted in increased insulin mRNA levels. Thapsigargin-induced stress granule formation was associated with a redistribution of TIAR from the cytosol to stress granules. These experiments indicate that alterations in insulin mRNA stability and translation correlate with differential RBP binding. We propose that the balance between PTB on one hand and TIAR on the other participates in the control of insulin mRNA stability and utilization for insulin biosynthesis.

  9. Overexpression of nucleolin in chronic lymphocytic leukemia cells induces stabilization of bcl2 mRNA

    PubMed Central

    Otake, Yoko; Soundararajan, Sridharan; Sengupta, Tapas K.; Kio, Ebenezer A.; Smith, James C.; Pineda-Roman, Mauricio; Stuart, Robert K.; Spicer, Eleanor K.

    2007-01-01

    B-cell chronic lymphocytic leukemia (CLL) is characterized by the accumulation of clonal B cells that are resistant to apoptosis as a result of bcl2 oncogene overexpression. Studies were done to determine the mechanism for the up-regulation of bcl-2 protein observed in CD19+ CLL cells compared with CD19+ B cells from healthy volunteers. The 11-fold higher level of bcl-2 protein in CLL cells was positively correlated with a 26-fold elevation in the cytosolic level of nucleolin, a bcl2 mRNA–stabilizing protein. Measurements of the bcl2 heterogeneous nuclear/bcl2 mRNA (hnRNA)/mRNA ratios and the rates of bcl2 mRNA decay in cell extracts indicated that the 3-fold higher steady-state level of bcl2 mRNA in CLL cells was the result of increased bcl2 mRNA stability. Nucleolin was present throughout the nucleus and cytoplasm of CLL cells, whereas in normal B cells nucleolin was only detected in the nucleus. The addition of recombinant human nucleolin to extracts of normal B cells markedly slowed the rate of bcl2 mRNA decay. SiRNA knockdown of nucleolin in MCF-7 cells resulted in decreased levels of bcl2 mRNA and protein but no change in β-actin. These results indicate that bcl-2 overexpression in CLL cells is related to stabilization of bcl2 mRNA by nucleolin. PMID:17179226

  10. Effects of long-term treatment with the luteinizing hormone-releasing hormone (LHRH) agonist Decapeptyl and the LHRH antagonist Cetrorelix on the levels of pituitary LHRH receptors and their mRNA expression in rats

    PubMed Central

    Horvath, Judit E.; Bajo, Ana M.; Schally, Andrew V.; Kovacs, Magdolna; Herbert, Francine; Groot, Kate

    2002-01-01

    The effects of depot formulations of the luteinizing hormone-releasing hormone (LHRH) agonist Decapeptyl (25 μg/day) for 30 days or LHRH antagonist Cetrorelix pamoate (100 μg/day) for 30 days and daily injections of 100 μg of Decapeptyl for 10 days on the expression of mRNA for pituitary LHRH receptor (LHRH-R) and the levels of LHRH-R protein were evaluated in rats. Serum sex steroid concentrations and the weights of the reproductive organs were greatly reduced in all groups treated with analogs, demonstrating an efficient blockade of the pituitary–gonadal axis. Decapeptyl microcapsules elevated serum LH in female rats, but decreased it in male rats. LHRH-R mRNA expression in female pituitaries was reduced to 41% and 56–65% on days 10 and 30, respectively, whereas LHRH-R protein was 64% of control on day 10 and returned to pretreatment levels on day 30. Decapeptyl microcapsules reduced LHRH-R mRNA expression in male pituitaries to 58% on day 30 but not LHRH-R protein. Daily injections of Decapeptyl caused a desensitization of LH responses in female rats, while raising LHRH-R mRNA expression in female rats by 23% and LHRH-R protein levels by 119%. Cetrorelix pamoate reduced serum LH in female rats and diminished LHRH-R mRNA to 30% and 26% and LHRH-R protein to 57% and 48% on days 10 and 30, respectively. Elevated LHRH-R protein levels of ovariectomized rats were reduced after 10-day treatment with Cetrorelix or 100 μg/day Decapeptyl. Thus, changes in the mRNA expression after treatment with Cetrorelix, but not always Decapeptyl, paralleled those of LHRH-R protein. The inhibitory effect of Cetrorelix on serum LH, pituitary LHRH-R mRNA, and LHRH-R protein was greater than that of Decapeptyl. PMID:12409615

  11. Low Incidence along with Low mRNA Levels of EGFRvIII in Prostate and Colorectal Cancers Compared to Glioblastoma

    PubMed Central

    Peciak, Joanna; Stec, Wojciech J; Treda, Cezary; Ksiazkiewicz, Magdalena; Janik, Karolina; Popeda, Marta; Smolarz, Maciej; Rosiak, Kamila; Hulas-Bigoszewska, Krystyna; Och, Waldemar; Rieske, Piotr; Stoczynska-Fidelus, Ewelina

    2017-01-01

    Background: The presence as well as the potential role of EGFRvIII in tumors other than glioblastoma still remains a controversial subject with many contradictory data published. Previous analyses, however, did not consider the level of EGFRvIII mRNA expression in different tumor types. Methods: Appropriately designed protocol for Real-time quantitative reverse-transcription PCR (Real-time qRT-PCR) was applied to analyze EGFRvIII and EGFRWT mRNA expression in 155 tumor specimens. Additionally, Western Blot (WB) analysis was performed for selected samples. Stable cell lines showing EGFRvIII expression (CAS-1 and DK-MG) were analyzed by means of WB, immunocytochemistry (ICC) and fluorescence in situ hybridization (FISH). Results: Our analyses revealed EGFRvIII expression in 27.59% of glioblastomas (8/29), 8.11% of colorectal cancers (3/37), 6.52% of prostate cancers (3/46) and none of breast cancers (0/43). Despite the average relative expression of EGFRvIII varying greatly among tumors of different tissues (approximately 800-fold) or even within the same tissue group (up to 8000-fold for GB), even the marginal expression of EGFRvIII mRNA can be detrimental to cancer progression, as determined by the analysis of stable cell lines endogenously expressing the oncogene. Conclusion: EGFRvIII plays an unquestionable role in glioblastomas with high expression of this oncogene. Our data suggests that EGFRvIII importance should not be underestimated even in tumors with relatively low expression of this oncogene. PMID:28123609

  12. Production of fermentable sugars from corn fiber using soaking in aqueous ammonia (saa) pretreatment and fermentation to succinic acid by Escherichia coli afp184

    USDA-ARS?s Scientific Manuscript database

    Conversion of corn fiber (CF), a by-product from the corn-to-ethanol conversion process, into fermentable sugar and succinic acid was investigated using soaking in aqueous ammonia (SAA) pretreatment followed by biological conversions including enzymatic hydrolysis and fermentation using genetically ...

  13. Distribution of cholecystokinin mRNA and peptides in the human brain.

    PubMed

    Lindefors, N; Brené, S; Kopp, J; Lindén, A; Brodin, E; Sedvall, G; Persson, H

    1991-01-01

    Expression of preprocholecystokinin mRNA was studied in regions of post mortem human brain using RNA blot analysis (Northern blot) and in situ hybridization. Northern blot analysis using a cDNA probe showed high levels of an approximately 0.8 kb preprocholecystokinin mRNA in all regions of neocortex examined. Lower levels of preprocholecystokinin mRNA were detected in amygdaloid body and thalamus. In situ hybridization analysis using the same cDNA probe revealed numerous weakly labelled neurons in different areas of human neocortex and less numerous neurons in hippocampus and amygdaloid body. High-performance liquid-chromatography and gel-chromatography combined with radioimmunoassay of cholecystokinin-like immunoreactivity from human cerebral cortex and caudate nucleus revealed two major forms, one coeluting with sulphated cholecystokinin-8 and the other coeluting with sulphated cholecystokinin-58. Two minor components coeluting with cholecystokinin-4 and cholecystokinin-5 were also detected. The finding of cholecystokinin-like immunoreactivity corresponding to cholecystokinin-8 and cholecystokinin-58 in caudate nucleus where no preprocholecystokinin mRNA was found, indicates the presence of these peptides in afferent nerve terminals.

  14. Vibrational force alters mRNA expression in osteoblasts

    NASA Technical Reports Server (NTRS)

    Tjandrawinata, R. R.; Vincent, V. L.; Hughes-Fulford, M.

    1997-01-01

    Serum-deprived mouse osteoblastic (MC3T3E1) cells were subjected to a vibrational force modeled by NASA to simulate a space shuttle launch (7.83 G rms). The mRNA levels for eight genes were investigated to determine the effect of vibrational force on mRNA expression. The mRNA levels of two growth-related protooncogenes, c-fos and c-myc, were up-regulated significantly within 30 min after vibration, whereas those of osteocalcin as well as transforming growth factor-beta1 were decreased significantly within 3 h after vibration. No changes were detected in the levels of beta-actin, histone H4, or cytoplasmic phospholipase A2 after vibration. No basal levels of cyclooxygenase-2 expression were detected. In addition, the extracellular concentrations of prostaglandin E2 (PGE2), a potent autocrine/paracrine growth factor in bone, were not significantly altered after vibration most likely due to the serum deprivation state of the osteoblasts. In comparison with the gravitational launch profile, vibrational-induced changes in gene expression were greater both in magnitude and number of genes activated. Taken together, these data suggest that the changes in mRNA expression are due to a direct mechanical effect of the vibrational force on the osteoblast cells and not to changes in the local PGE2 concentrations. The finding that launch forces induce gene expression is of utmost importance since many of the biological experiments do not dampen vibrational loads on experimental samples. This lack of dampening of vibrational forces may partially explain why 1-G onboard controls sometimes do not reflect 1-G ground controls. These data may also suggest that scientists use extra ground controls that are exposed to launch forces, have these forces dampened on launched samples, or use facilities such as Biorack that provide an onboard 1-G centrufuge in order to control for space shuttle launch forces.

  15. TNF-α messenger ribonucleic acid (mRNA) in patients with nonalcoholic steatohepatitis.

    PubMed

    Alaaeddine, Nada; Sidaoui, Joseph; Hilal, George; Serhal, Reem; Abedelrahman, Abir; Khoury, Salem

    2012-01-01

    tumor necrosis factor (TNF)-α plays a significant role in the pathogenesis of nonalcoholic steatohepatitis (NASH). A few studies have confirmed high TNF-α plasma protein levels in patients with NASH compared to healthy volunteers. We herein aimed to revisit these findings using other molecular techniques. a cross-sectional evaluation of patients newly diagnosed with NASH. A quantitative assay for the measurement of TNF-α messenger ribonucleic acid (mRNA) was performed for NASH patients and controls using real-time reverse transcription polymerase chain reaction (RT-PCR). in 39 patients with NASH (mean age 38.6 ± 9.4 years, range 28-60 years; 79% males), the mean TNF-α mRNA level was significantly higher than that found for controls (137.6 ± 102.3 ng/mL versus 83.5 ± 43.8 ng/mL, respectively; P = 0.012). A TNF-α mRNA cut-off of 100 ng/mL predicted NASH most optimally (AUC 0.685 ± 0.066, P = 0.01; with 66.7% sensitivity and 74.1% specificity). Serum TNF-α and soluble TNF-α receptor II (sTNFRII) levels were significantly higher in patients compared to controls using ELISA. high TNF-α mRNA levels, determined by RT-PCR, characterize patients with NASH.

  16. 5-hydroxytryptamine level and 5-HT2A receptor mRNA expression in the guinea pigs eyes with spectacle lens-induced myopia

    PubMed Central

    Yang, Ji-Wen; Xu, Yan-Chun; Sun, Lin; Tian, Xiao-Dan

    2010-01-01

    AIM To investigate 5-hydroxytryptamine (5-HT) function and 5-HT receptor 2A (5-HT2A) mRNA expression in the formation of lens-induced myopia (LIM). METHODS Lens-induced myopia construction method was applied to generate myopia on guinea pig right eye (LIM eye). RESULTS LIM eyes formed significant myopia with longer axial length. 5-HT level in retina, choroids and sclera from LIM eyes was significantly higher than that in control group. 5-HT2A mRNA expression was also significantly up-regulated. CONCLUSION Refraction lens could induce myopia in guinea pig and 5-HT may play an important role in the formation of myopia by binding with 5-HT2A receptor. PMID:22553578

  17. Exon 2-mediated c-myc mRNA decay in vivo is independent of its translation.

    PubMed Central

    Pistoi, S; Roland, J; Babinet, C; Morello, D

    1996-01-01

    We have previously shown that the steady-state level of c-myc mRNA in vivo is primarily controlled by posttranscriptional regulatory mechanisms. To identify the sequences involved in this process, we constructed a series of H-2/myc transgenic lines in which various regions of the human c-MYC gene were placed under the control of the quasi-ubiquitous H-2K class I regulatory sequences. We demonstrated that the presence of one of the two coding exons, exon 2 or exon 3, is sufficient to confer a level of expression of transgene mRNA similar to that of endogenous c-myc in various adult tissues as well as after partial hepatectomy or after protein synthesis inhibition. We now focus on the molecular mechanisms involved in modulation of expression of mRNAs containing c-myc exon 2 sequences, with special emphasis on the coupling between translation and c-myc mRNA turnover. We have undertaken an analysis of expression, both at the mRNA level and at the protein level, of new transgenic constructs in which the translation is impaired either by disruption of the initiation codon or by addition of stop codons upstream of exon 2. Our results show that the translation of c-myc exon 2 is not required for regulated expression of the transgene in the different situations analyzed, and therefore they indicate that the mRNA destabilizing function of exon 2 is independent of translation by ribosomes. Our investigations also reveal that, in the thymus, some H-2/myc transgenes express high levels of mRNA but low levels of protein. Besides the fact that these results suggest the existence of tissue-specific mechanisms that control c-myc translatability in vivo, they also bring another indication of the uncoupling of c-myc mRNA translation and degradation. PMID:8756668

  18. Conserved Non-Coding Sequences are Associated with Rates of mRNA Decay in Arabidopsis.

    PubMed

    Spangler, Jacob B; Feltus, Frank Alex

    2013-01-01

    Steady-state mRNA levels are tightly regulated through a combination of transcriptional and post-transcriptional control mechanisms. The discovery of cis-acting DNA elements that encode these control mechanisms is of high importance. We have investigated the influence of conserved non-coding sequences (CNSs), DNA patterns retained after an ancient whole genome duplication event, on the breadth of gene expression and the rates of mRNA decay in Arabidopsis thaliana. The absence of CNSs near α duplicate genes was associated with a decrease in breadth of gene expression and slower mRNA decay rates while the presence CNSs near α duplicates was associated with an increase in breadth of gene expression and faster mRNA decay rates. The observed difference in mRNA decay rate was fastest in genes with CNSs in both non-transcribed and transcribed regions, albeit through an unknown mechanism. This study supports the notion that some Arabidopsis CNSs regulate the steady-state mRNA levels through post-transcriptional control mechanisms and that CNSs also play a role in controlling the breadth of gene expression.

  19. Conserved Non-Coding Sequences are Associated with Rates of mRNA Decay in Arabidopsis

    PubMed Central

    Spangler, Jacob B.; Feltus, Frank Alex

    2013-01-01

    Steady-state mRNA levels are tightly regulated through a combination of transcriptional and post-transcriptional control mechanisms. The discovery of cis-acting DNA elements that encode these control mechanisms is of high importance. We have investigated the influence of conserved non-coding sequences (CNSs), DNA patterns retained after an ancient whole genome duplication event, on the breadth of gene expression and the rates of mRNA decay in Arabidopsis thaliana. The absence of CNSs near α duplicate genes was associated with a decrease in breadth of gene expression and slower mRNA decay rates while the presence CNSs near α duplicates was associated with an increase in breadth of gene expression and faster mRNA decay rates. The observed difference in mRNA decay rate was fastest in genes with CNSs in both non-transcribed and transcribed regions, albeit through an unknown mechanism. This study supports the notion that some Arabidopsis CNSs regulate the steady-state mRNA levels through post-transcriptional control mechanisms and that CNSs also play a role in controlling the breadth of gene expression. PMID:23675377

  20. Genome wide assessment of mRNA in astrocyte protrusions by direct RNA sequencing reveals mRNA localization for the intermediate filament protein nestin.

    PubMed

    Thomsen, Rune; Pallesen, Jonatan; Daugaard, Tina F; Børglum, Anders D; Nielsen, Anders L

    2013-11-01

    Subcellular RNA localization plays an important role in development, cell differentiation, and cell migration. For a comprehensive description of the population of protrusion localized mRNAs in astrocytes we separated protrusions from cell bodies in a Boyden chamber and performed high-throughput direct RNA sequencing. The mRNAs with localization in astrocyte protrusions encode proteins belonging to a variety of functional groups indicating involvement of RNA localization for a palette of cellular functions. The mRNA encoding the intermediate filament protein Nestin was among the identified mRNAs. By RT-qPCR and RNA FISH analysis we confirmed Nestin mRNA localization in cell protrusions and also protrusion localization of Nestin protein. Nestin mRNA localization was dependent of Fragile X mental retardation syndrome proteins Fmrp and Fxr1, and the Nestin 3'-UTR was sufficient to mediate protrusion mRNA localization. The mRNAs for two other intermediate filament proteins in astrocytes, Gfap and Vimentin, have moderate and no protrusion localization, respectively, showing that individual intermediate filament components have different localization mechanisms. The correlated localization of Nestin mRNA with Nestin protein in cell protrusions indicates the presence of a regulatory mechanism at the mRNA localization level for the Nestin intermediate filament protein with potential importance for astrocyte functions during brain development and maintenance. Copyright © 2013 Wiley Periodicals, Inc.

  1. Tudor-SN, a component of stress granules, regulates growth under salt stress by modulating GA20ox3 mRNA levels in Arabidopsis

    PubMed Central

    Yan, Chunxia; Yan, Zongyun; Wang, Yizheng; Yan, Xiaoyuan; Han, Yuzhen

    2014-01-01

    The Tudor-SN protein (TSN) is universally expressed and highly conserved in eukaryotes. In Arabidopsis, TSN is reportedly involved in stress adaptation, but the mechanism involved in this adaptation is not understood. Here, we provide evidence that TSN regulates the mRNA levels of GA20ox3, a key enzyme for gibberellin (GA) biosynthesis. The levels of GA20ox3 transcripts decreased in TSN1/TSN2 RNA interference (RNAi) transgenic lines and increased in TSN1 over-expression (OE) transgenic lines. The TSN1 OE lines displayed phenotypes that may be attributed to the overproduction of GA. No obvious defects were observed in the RNAi transgenic lines under normal conditions, but under salt stress conditions these lines displayed slower growth than wild-type (WT) plants. Two mutants of GA20ox3, ga20ox3-1 and -2, also showed slower growth under stress than WT plants. Moreover, a higher accumulation of GA20ox3 transcripts was observed under salt stress. The results of a western blot analysis indicated that higher levels of TSN1 accumulated after salt treatment than under normal conditions. Subcellular localization studies showed that TSN1 was uniformly distributed in the cytoplasm under normal conditions but accumulated in small granules and co-localized with RBP47, a marker protein for stress granules (SGs), in response to salt stress. The results of RNA immunoprecipitation experiments indicated that TSN1 bound GA20ox3 mRNA in vivo. On the basis of these findings, we conclude that TSN is a novel component of plant SGs that regulates growth under salt stress by modulating levels of GA20ox3 mRNA. PMID:25205572

  2. β-glucuronidase use as a single internal control gene may confound analysis in FMR1 mRNA toxicity studies.

    PubMed

    Kraan, Claudine M; Cornish, Kim M; Bui, Quang M; Li, Xin; Slater, Howard R; Godler, David E

    2018-01-01

    Relationships between Fragile X Mental Retardation 1 (FMR1) mRNA levels in blood and intragenic FMR1 CGG triplet expansions support the pathogenic role of RNA gain of function toxicity in premutation (PM: 55-199 CGGs) related disorders. Real-time PCR (RT-PCR) studies reporting these findings normalised FMR1 mRNA level to a single internal control gene called β-glucuronidase (GUS). This study evaluated FMR1 mRNA-CGG correlations in 33 PM and 33 age- and IQ-matched control females using three normalisation strategies in peripheral blood mononuclear cells (PBMCs): (i) GUS as a single internal control; (ii) the mean of GUS, Eukaryotic Translation Initiation Factor 4A2 (EIF4A2) and succinate dehydrogenase complex flavoprotein subunit A (SDHA); and (iii) the mean of EIF4A2 and SDHA (with no contribution from GUS). GUS mRNA levels normalised to the mean of EIF4A2 and SDHA mRNA levels and EIF4A2/SDHA ratio were also evaluated. FMR1mRNA level normalised to the mean of EIF4A2 and SDHA mRNA levels, with no contribution from GUS, showed the most significant correlation with CGG size and the greatest difference between PM and control groups (p = 10-11). Only 15% of FMR1 mRNA PM results exceeded the maximum control value when normalised to GUS, compared with over 42% when normalised to the mean of EIF4A2 and SDHA mRNA levels. Neither GUS mRNA level normalised to the mean RNA levels of EIF4A2 and SDHA, nor to the EIF4A2/SDHA ratio were correlated with CGG size. However, greater variability in GUS mRNA levels were observed for both PM and control females across the full range of CGG repeat as compared to the EIF4A2/SDHA ratio. In conclusion, normalisation with multiple control genes, excluding GUS, can improve assessment of the biological significance of FMR1 mRNA-CGG size relationships.

  3. Tris(2-butoxyethyl)phosphate and triethyl phosphate alter embryonic development, hepatic mRNA expression, thyroid hormone levels, and circulating bile acid concentrations in chicken embryos

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Egloff, Caroline; Crump, Doug, E-mail: doug.crump@ec.gc.ca; Porter, Emily

    The organophosphate flame retardants tris(2-butoxyethyl) phosphate (TBOEP) and triethyl phosphate (TEP) are used in a wide range of applications to suppress or delay the ignition and spread of fire. Both compounds have been detected in the environment and TBOEP was recently measured in free-living avian species. In this study, TBOEP and TEP were injected into the air cell of chicken embryos at concentrations ranging from 0 to 45,400 ng/g and 0 to 241,500 ng/g egg, respectively. Pipping success, development, hepatic mRNA expression of 9 target genes, thyroid hormone levels, and circulating bile acid concentrations were determined. Exposure to the highestmore » doses of TBOEP and TEP resulted in negligible detection of the parent compounds in embryonic contents at pipping indicating their complete metabolic degradation. TBOEP exposure had limited effects on chicken embryos, with the exception of hepatic CYP3A37 mRNA induction. TEP exposure decreased pipping success to 68%, altered growth, increased liver somatic index (LSI) and plasma bile acids, and modulated genes associated with xenobiotic and lipid metabolism and the thyroid hormone pathway. Plasma thyroxine levels were decreased at all TEP doses, including an environmentally-relevant concentration (8 ng/g), and gallbladder hypotrophy was evident at ≥ 43,200 ng/g. Tarsus length and circulating thyroxine concentration emerged as potential phenotypic anchors for the modulation of transthyretin mRNA. The increase in plasma bile acids and LSI, gallbladder hypotrophy, and discoloration of liver tissue represented potential phenotypic outcomes associated with modulation of hepatic genes involved with xenobiotic and lipid metabolism. - Highlights: • TBOEP is not embryolethal to chicken embryos. • TEP affected embryonic viability, morphometric endpoints, and thyroid hormone levels. • TEP altered mRNA levels of xenobiotic and lipid metabolism genes. • TEP increased plasma bile acids and caused gallbladder

  4. Unmasking Upstream Gene Expression Regulators with miRNA-corrected mRNA Data

    PubMed Central

    Bollmann, Stephanie; Bu, Dengpan; Wang, Jiaqi; Bionaz, Massimo

    2015-01-01

    Expressed micro-RNA (miRNA) affects messenger RNA (mRNA) abundance, hindering the accuracy of upstream regulator analysis. Our objective was to provide an algorithm to correct such bias. Large mRNA and miRNA analyses were performed on RNA extracted from bovine liver and mammary tissue. Using four levels of target scores from TargetScan (all miRNA:mRNA target gene pairs or only the top 25%, 50%, or 75%). Using four levels of target scores from TargetScan (all miRNA:mRNA target gene pairs or only the top 25%, 50%, or 75%) and four levels of the magnitude of miRNA effect (ME) on mRNA expression (30%, 50%, 75%, and 83% mRNA reduction), we generated 17 different datasets (including the original dataset). For each dataset, we performed upstream regulator analysis using two bioinformatics tools. We detected an increased effect on the upstream regulator analysis with larger miRNA:mRNA pair bins and higher ME. The miRNA correction allowed identification of several upstream regulators not present in the analysis of the original dataset. Thus, the proposed algorithm improved the prediction of upstream regulators. PMID:27279737

  5. Elevated levels of serum-soluble triggering receptor expressed on myeloid cells-1 in patients with IBD do not correlate with intestinal TREM-1 mRNA expression and endoscopic disease activity.

    PubMed

    Saurer, Leslie; Rihs, Silvia; Birrer, Michèle; Saxer-Seculic, Nikolina; Radsak, Markus; Mueller, Christoph

    2012-10-01

    Triggering receptor expressed on myeloid cells-1 (TREM-1) is a potent amplifier of pro-inflammatory responses. We have previously demonstrated a substantial increase in TREM-1-expressing macrophages in the inflamed intestinal mucosa of patients with inflammatory bowel diseases (IBD). TREM-1 is also produced as a soluble receptor (sTREM-1). Here, we aimed to determine whether serum sTREM-1 could be used as a surrogate marker of disease activity in patients with IBD. Intestinal biopsies and concurrently collected sera from patients with Crohn's disease (CD) and Ulcerative colitis (UC) enrolled in the Swiss IBD cohort study were analyzed for intestinal TREM-1 mRNA and serum sTREM-1 expression. TREM-1 mRNA and sTREM-1 were correlated with the endoscopically determined disease activity. Serum sTREM-1 and TREM-1 mRNA expression levels were further determined in sera and colonic tissues collected at various time-points post disease induction in an experimental mouse model of colitis and correlated with disease activity. Expression of TREM-1 mRNA was upregulated in intestinal biopsies from patients with active disease but not in patients with quiescent disease. Serum sTREM-1 was elevated in IBD patients compared to normal controls. No substantial differences in sTREM-1 expression levels were found in patients with active versus quiescent disease. In colitic mice, colonic TREM-1 mRNA and serum sTREM-1 were also upregulated. While colonic TREM-1 mRNA expression levels correlated with disease activity, augmented serum sTREM-1 in fact associated with a milder course of disease. Analysis of sTREM-1 as a surrogate marker of disease activity in patients with IBD warrants caution. Copyright © 2012 European Crohn's and Colitis Organisation. Published by Elsevier B.V. All rights reserved.

  6. Symptoms of prenatal depression are associated with raised salivary alpha-amylase levels.

    PubMed

    Braithwaite, Elizabeth C; Ramchandani, Paul G; Lane, Tracy A; Murphy, Susannah E

    2015-10-01

    Prenatal depression increases risk for a number of adverse offspring outcomes, however the biological mechanisms underlying this association remain unclear. It has been suggested that maternal glucocorticoids may mediate this link, though supporting evidence has been mixed. An alternative mechanism of effect may be via depression-induced changes in maternal sympathetic nervous system (SNS) function. We examined this hypothesis by determining the relationship between symptoms of maternal prenatal depression and diurnal salivary alpha-amylase (sAA) levels. 76 pregnant women were recruited during either the second or third trimester of pregnancy. Participants self-reported depressive symptoms using the Edinburgh postnatal depression scale. Saliva samples, to be assayed for alpha-amylase activity, were collected at home over two working days. Participants with depressive symptoms in later pregnancy had elevated awakening sAA levels compared with non-depressed controls (t(73) = -2.737, p = 0.008), and continued to have raised sAA throughout the day (F(1) = 10.924, p = 0.002). Our findings highlight that symptoms of depression during late pregnancy are associated with increased maternal SNS activity. Thus, changes in maternal SNS function, which may include increased vasoconstriction and reduced foetal blood flow, could, in part, mediate associations between prenatal depression and adverse offspring outcomes. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Cell type-specific regulation of beta2-adrenoceptor mRNA by agonists.

    PubMed

    Danner, S; Lohse, M J

    1997-07-16

    Prolonged agonist stimulation of beta2-adrenoceptors results in receptor down-regulation which is often paralleled by a reduction of the corresponding mRNA. In this study, we investigated the agonist-dependent regulation of beta2-adrenoceptor mRNA in DDT1-MF2 smooth muscle cells and C6 glioma cells. In DDT1-MF2 cells the half-life of the mRNA was 12 h in monolayer compared to 2 h in suspension cultures. Under both conditions, the agonist isoproterenol reduced this half-life by a factor of 2. In contrast, in C6 glioma cells isoproterenol had no effect on the mRNA stability, even though it reduced mRNA levels by approximately 50%. Isoproterenol-induced downregulation of beta2-adrenoceptor mRNA was completely blocked in C6 cells by the presence of a protein synthesis inhibitor, while this was not so in DDT1-MF2-cells. These data show that beta2-adrenoceptor downregulation occurs via cell-type specific mechanisms.

  8. Pregnancy amelioration of arthritis in SKG mice corresponds with alterations in serum amyloid A3 levels

    PubMed Central

    Shaw, Laura A; Stefanski, Adrianne L; Peterson, Lisa K; Rumer, Kristen K; Vondracek, Andrea; Phang, Tzu L; Sakaguchi, Shimon; Winn, Virginia D; Dragone, Leonard L

    2012-01-01

    Objectives: Pregnancy leads to rheumatoid arthritis remission in humans. The objective of this study was to determine if the SKG mouse could serve as a model for pregnancy-associated inflammatory arthritis amelioration. In addition, the maternal peripheral blood mononuclear cell (PBMC) transcriptome was assessed to define a biomarker associated with remission. Methods: Cohorts of zymosan-treated pregnant SKG mice and controls were monitored for arthritis progression. Microarray analysis evaluated alterations in gene expression in maternal PBMCs at embryonic day 14.5 (E14.5) between arthritic and pregnancy-remitted mice. A selected target, serum amyloid A3 (SAA3), was further investigated using quantitative reverse transcriptase PCR (qRT-PCR) and an enzyme-linked immunosorbent assay (ELISA). Results: Pregnancy resulted in complete or partial remission in the majority of the zymosan-treated SKG mice. Twenty-seven transcripts were differentially expressed in the PBMCs between arthritic and pregnancy-remitted mice. Expression and plasma SAA3 levels decreased with pregnancy-induced arthritis amelioration and plasma SAA3 levels correlated with arthritis severity. Conclusions: These results establish the SKG mouse as a model system to study pregnancy-induced amelioration of arthritis. These studies also establish SAA3 as a biomarker of arthritis amelioration in SKG mice. This model can be used to elucidate the molecular and cellular mechanisms underlying the impact of pregnancy on the maternal immune system that results in arthritis amelioration. PMID:23097751

  9. Reduced m6A mRNA methylation is correlated with the progression of human cervical cancer

    PubMed Central

    Kong, Beihua; Song, Chen; Cong, Jianglin; Hou, Jianqing; Wang, Shaoguang

    2017-01-01

    The m6A mRNA methylation involves in mRNA splicing, degradation and translation. Recent studies have revealed that reduced m6A mRNA methylation might promote cancer development. However, the role of m6A mRNA methylation in cervical cancer development remains unknown. Therefore, we investigated the role of m6A methylation in cervical cancer in the current study. We first evaluated the m6A mRNA methylation level in 286 pairs of cervical cancer samples and their adjacent normal tissues by dot blot assay. Then the role of m6A on patient survival rates and cervical cancer progression were assessed. The m6A level was significantly reduced in the cervical cancer when comparing with the adjacent normal tissue. The m6A level reduction was significantly correlated with the FIGO stage, tumor size, differentiation, lymph invasion and cancer recurrence. It was also shown to be an independent prognostic indicator of disease-free survival and overall survival for patients with cervical cancer. Reducing m6A level via manipulating the m6A regulators expression promoted cervical cancer cell proliferation. And increasing m6A level significantly suppressed tumor development both in vitro and in vivo. Our results showed that the reduced m6A level is tightly associated with cervical cancer development and m6A mRNA methylation might be a potential therapeutic target in cervical cancer. PMID:29228737

  10. Manganese exposure alters extracellular GABA, GABA receptor and transporter protein and mRNA levels in the developing rat brain

    PubMed Central

    Anderson, Joel G.; Fordahl, Steve C.; Cooney, Paula T.; Weaver, Tara L.; Colyer, Christa L.; Erikson, Keith M.

    2011-01-01

    Unlike other essential trace elements (e.g., zinc and iron) it is the toxicity of manganese (Mn) that is more common in human populations than its deficiency. Data suggest alterations in dopamine biology may drive the effects associated with Mn neurotoxicity, though recently γ-aminobutyric acid (GABA) has been implicated. In addition, iron deficiency (ID), a common nutritional problem, may cause disturbances in neurochemistry by facilitating accumulation of Mn in the brain. Previous data from our lab have shown decreased brain tissue levels of GABA as well as decreased 3H-GABA uptake in synaptosomes as a result of Mn exposure and ID. These results indicate a possible increase in the concentration of extracellular GABA due to alterations in expression of GABA transport and receptor proteins. In this study weanling-male Sprague-Dawley rats were randomly placed into one of four dietary treatment groups: control (CN; 35 mg Fe/kg diet), iron-deficient (ID; 6 mg Fe/kg diet), CN with Mn supplementation (via the drinking water; 1 g Mn/L) (CNMn), and ID with Mn supplementation (IDMn). Using in vivo microdialysis, an increase in extracellular GABA concentrations in the striatum was observed in response to Mn exposure and ID although correlational analysis reveals that extracellular GABA is related more to extracellular iron levels and not Mn. A diverse effect of Mn exposure and ID was observed in the regions examined via Western blot and RT-PCR analysis, with effects on mRNA and protein expression of GAT-1, GABAA, and GABAB differing between and within the regions examined. For example, Mn exposure reduced GAT-1 protein expression by approximately 50% in the substantia nigra, while increasing mRNA expression approximately four-fold, while in the caudate putamen mRNA expression was decreased with no effect on protein expression. These data suggest that Mn exposure results in an increase in extracellular GABA concentrations via altered expression of transport and receptor

  11. Effect of long term caffeine treatment on A1 and A2 adenosine receptor binding and on mRNA levels in rat brain.

    PubMed

    Johansson, B; Ahlberg, S; van der Ploeg, I; Brené, S; Lindefors, N; Persson, H; Fredholm, B B

    1993-04-01

    The effect of long-term oral treatment with caffeine on A1 and A2 receptors in the rat brain was studied. Caffeine was added to the drinking water and the animals were sacrificed after a 12 day treatment period. The plasma caffeine concentration was close to 100 microM. A1 receptors were studied using quantitative autoradiography with [3H]cyclohexyladenosine (CHA). Caffeine treatment increased the number of A1 receptors in the CA3 subfield of the hippocampus from 337 to 393 fmol/mg with no change in KD (0.692 vs. 0.675 nM). A1 mRNA was measured using Northern blots and quantitative in situ hybridization. There was no increase in A1 mRNA. A2a receptors, located in dopamine rich regions of the rat brain, were studied with quantitative autoradiography using [3H]CGS 21680 as the ligand, and the A2a mRNA was determined using quantitative in situ hybridization. Caffeine treatment produced no significant change in either receptor number or mRNA, even though the apparent Bmax tended to increase from 322 +/- 8 to 352 +/- 8 fmol/mg. The results show that treatment with caffeine in a dose that causes tolerance to several effects of caffeine and increases some effects of adenosine analogues increases the number of A1 receptors without any change in A1 mRNA, suggesting that the adaptive changes are at a post-translational level. There were no significant changes in A2 receptors indicating that the two types are regulated differently and/or that the amount of endogenous agonist is sufficient to regulate A1, but not A2 receptors.

  12. Evaluation of mRNA expression levels and electrophysiological function of neuron-like cells derived from canine bone marrow stromal cells.

    PubMed

    Nakano, Rei; Edamura, Kazuya; Sugiya, Hiroshi; Narita, Takanori; Okabayashi, Ken; Moritomo, Tadaaki; Teshima, Kenji; Asano, Kazushi; Nakayama, Tomohiro

    2013-10-01

    To investigate the in vitro differentiation of canine bone marrow stromal cells (BMSCs) into functional, mature neurons. Bone marrow from 6 adult dogs. BMSCs were isolated from bone marrow and chemically induced to develop into neurons. The morphology of the BMSCs during neuronal induction was monitored, and immunocytochemical analyses for neuron markers were performed after the induction. Real-time PCR methods were used to evaluate the mRNA expression levels of markers for neural stem or progenitor cells, neurons, and ion channels, and western blotting was used to assess the expression of neuronal proteins before and after neuronal induction. The electrophysiological properties of the neuron-like cells induced from canine BMSCs were evaluated with fluorescent dye to monitor Ca(2)+ influx. Canine BMSCs developed a neuron-like morphology after neuronal induction. Immunocytochemical analysis revealed that these neuron-like cells were positive for neuron markers. After induction, the cells' mRNA expression levels of almost all neuron and ion channel markers increased, and the protein expression levels of nestin and neurofilament-L increased significantly. However, the neuron-like cells derived from canine BMSCs did not have the Ca(2)+ influx characteristic of spiking neurons. Although canine BMSCs had neuron-like morphological and biochemical properties after induction, they did not develop the electrophysiological characteristics of neurons. Thus, these results have suggested that canine BMSCs could have the capacity to differentiate into a neuronal lineage, but the differentiation protocol used may have been insufficient to induce development into functional neurons.

  13. Cytochrome P450IA mRNA expression in feral Hudson River tomcod

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kreamer, G.L.; Squibb, K.; Gioeli, D.

    1991-06-01

    The authors sought to determine if levels of cytochrome P450IA gene expression are environmentally induced in feral populations of Hudson River tomcod, a cancer prone fish, and whether laboratory exposure of tomcod to artificially spiked and naturally contaminated Hudson sediments can elicit a significant response. Using Northern blot analysis, they found levels of P450IA mRNA in tomcod collected from two Hudson River sites higher than those in tomcod from a river in Maine. Depuration of environmentally induced Hudson tomcod P450IA mRNA was rapid, with an initial detectable decline in P450 gene expression by 8 hr and basal levels reached bymore » 5 days. Intraperitoneal injection of {beta}-napthoflavone in depurated Hudson tomcod resulted in a 15-fold induction of P450 gene expression within 26 hr. Exposure of depurated Hudson tomcod to natural sediment spiked with two PAHs resulted in a 7-fold induction of P450 gene expression. Exposure of depurated tomcod to sediment from a contaminated Hudson site also resulted in a 7- to 15-fold induction of P450IA mRNA expression. Northern blot analysis revealed a second polymorphic cytochrome P450IA mRNA band in some tomcod which was also detected by Southern blot analysis. Induction of cytochrome P450IA mRNA in Atlantic tomcod may provide a sensitive biomarker of environmentally relevant concentrations of some pollutants in the Hudson and other northeastern tidal rivers.« less

  14. Regulation of mRNA Trafficking by Nuclear Pore Complexes

    PubMed Central

    Bonnet, Amandine; Palancade, Benoit

    2014-01-01

    Over the last two decades, multiple studies have explored the mechanisms governing mRNA export out of the nucleus, a crucial step in eukaryotic gene expression. During transcription and processing, mRNAs are assembled into messenger ribonucleoparticles (mRNPs). mRNPs are then exported through nuclear pore complexes (NPCs), which are large multiprotein assemblies made of several copies of a limited number of nucleoporins. A considerable effort has been put into the dissection of mRNA export through NPCs at both cellular and molecular levels, revealing the conserved contributions of a subset of nucleoporins in this process, from yeast to vertebrates. Several reports have also demonstrated the ability of NPCs to sort out properly-processed mRNPs for entry into the nuclear export pathway. Importantly, changes in mRNA export have been associated with post-translational modifications of nucleoporins or changes in NPC composition, depending on cell cycle progression, development or exposure to stress. How NPC modifications also impact on cellular mRNA export in disease situations, notably upon viral infection, is discussed. PMID:25184662

  15. Emphysema induced by elastase alters the mRNA relative levels from DNA repair genes in acute lung injury in response to sepsis induced by lipopolysaccharide administration in Wistar rats.

    PubMed

    Sergio, Luiz Philippe S; Lucinda, Leda M F; Reboredo, Maycon M; de Paoli, Flavia; Fonseca, Lídia M C; Pinheiro, Bruno V; Mencalha, Andre L; Fonseca, Adenilson S

    2018-03-01

    Purpose/Aim of the study: Patients suffering from chronic obstructive pulmonary disease (COPD) in association with acute respiratory distress syndrome (ARDS) present oxidative stress in lung cells, with production of free radicals and DNA lesions in pulmonary and adjacent cells. Once the DNA molecule is damaged, a set of enzymatic mechanisms are trigged to preserve genetic code integrity and cellular homeostasis. These enzymatic mechanisms include the base and the nucleotide excision repair pathways, as well as telomere regulation. Thus, the aim of this work was to evaluate the mRNA levels from APEX1, ERCC2, TP53, and TRF2 genes in lung tissue from Wistar rats affected by acute lung injury in response to sepsis and emphysema. Adult male Wistar rats were randomized into 4 groups (n = 6, for each group): control, emphysema, sepsis, and emphysema with sepsis. Pulmonary emphysema was induced by intratracheal instillation of elastase (12 IU/animal) and sepsis induced by intraperitoneal Escherichia coli lipopolysaccharide (LPS) injection (10 mg/kg). Lungs were removed, and samples were withdrawn for histological analysis and total RNA extraction, cDNA synthesis, and mRNA level evaluation by real time quantitative polymerase chain reaction. Data show acute lung injury by LPS and emphysema by elastase and that APEX1, ERCC2, TP53, and TRF2 mRNA levels are increased significantly (p < 0.01) in emphysema with sepsis group. Our results suggest that alteration in mRNA levels from DNA repair and genomic stability could be part of cell response to acute lung injury in response to emphysema and sepsis.

  16. Increased GAD67 mRNA levels are correlated with in vivo GABA synthesis in the MPTP-treated catecholamine-depleted goldfish brain.

    PubMed

    Hibbert, Benjamin; Fung, Irene; McAuley, Rebecca; Larivière, Katherine; MacNeil, Brian; Bafi-Yeboa, Nana; Livesey, John; Trudeau, Vance

    2004-09-28

    The role of catecholamine neuronal input on GABAergic activity in the hypothalamus, telencephalon, optic tectum, and cerebellum was investigated in early recrudescent female goldfish (Carassius auratus). A new quantitative technique was developed and validated, permitting concomitant quantification and correlational analysis of glutamic acid decarboxylase 65 (GAD65), GAD67, and GAD3 mRNA levels and in vivo GABA synthesis. Catecholamine depletion was achieved by the administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP; 50 microg/g body weight) and dopamine (DA) depletion verified by HPLC. Endogenous GABA levels were increased by intraperitoneal administration of gamma-vinyl GABA (GVG; 300 microg/g body weight), an inhibitor of the GABA catabolic enzyme GABA transaminase. Treatment with MPTP resulted in a greater than twofold increase in GABA synthesis rate in the optic tectum and telencephalon. The increase in GABA synthesis rate was highly correlated with an increase in GAD67, but not GAD65 or GAD3 mRNA levels. These results suggest that catecholaminergic input exerts inhibitory effects on GABA synthesis rates through the modulation of GAD67 in the optic tectum and telencephalon. Together with previously published observations in rodents and primates, it is suggested that catecholaminergic control of GABA synthesis must have evolved more than 200 million years ago, before the emergence of the teleost fishes.

  17. Running and cocaine both upregulate dynorphin mRNA in medial caudate putamen.

    PubMed

    Werme, M; Thorén, P; Olson, L; Brené, S

    2000-08-01

    Physical activities such as long-distance running can be habit forming and associated with a sense of well-being to a degree that justifies comparison with drug-induced addictive behaviours. To understand molecular similarities and dissimilarities controlling these behaviours in humans we compared the effects of running in running wheels to the effects of chronic cocaine or morphine administration on mRNA levels in brain reward pathways in the inbred Fischer and Lewis rat strains. These strains are both inbred from the Sprague-Dawley strain; Lewis rats display a higher preference towards addictive drugs and running than do Fischer rats. After chronic cocaine or running a similar increase of dynorphin mRNA in medial caudate putamen was found in the Lewis rat, suggesting common neuronal adaptations in this brain region to both cocaine and running. Fischer and Lewis rats both responded to cocaine with increased dynorphin mRNA levels in medial caudate putamen. However, only Lewis rats increased dynorphin mRNA after running, possibly reflecting the much higher degree of running by the Lewis strain as compared to the Fischer strain. Moreover, the running-induced upregulation of dynorphin mRNA was blocked by the opioid receptor antagonist naloxone. We suggest that running increases dynorphin mRNA by a mechanism that involves endogenous opioids. The voluntary wheel-running model in rats might be used to study natural reward and compulsive behaviours and possibly also to screen candidate drugs for treatment of compulsive disorders.

  18. Local and systemic response to intramammary lipopolysaccharide challenge during long-term manipulated plasma glucose and insulin concentrations in dairy cows.

    PubMed

    Vernay, M C M B; Wellnitz, O; Kreipe, L; van Dorland, H A; Bruckmaier, R M

    2012-05-01

    The metabolic load during periods of high milk production in dairy cows causes a variety of changes of metabolite blood concentrations including dramatically decreased glucose levels. These changes supposedly impair the immune system. The goal of this study was, therefore, to evaluate adaptations of the cow's immune system in response to an intramammary lipopolysaccharide (LPS) stimulation during a 3-d modification of plasma glucose and insulin induced by different clamp infusions. Seventeen midlactating dairy cows received a hypoglycemic hyperinsulinemic clamp induced by insulin infusion (HypoG; n=5), a euglycemic hyperinsulinemic clamp induced by insulin and glucose infusion (EuG; n=6), or infusion of saline solution (NaCl; n=6) for 56 h. At 48 h of infusion, 2 udder quarters were challenged with 200 μg of Escherichia coli LPS. At 48 h of infusion (immediately before LPS challenge), tumor necrosis factor α, lactoferrin, and serum amyloid A (SAA) mRNA abundance was increased in HypoG and Il-1β mRNA abundance was decreased in EuG. After LPS challenge, plasma glucose concentration did not decrease, although plasma insulin increased simultaneously in all groups either due to enhanced endogenous release (NaCl) or due to increased insulin infusion rate (HypoG; EuG). Plasma cortisol, rectal temperatures, and milk somatic cell count of challenged quarters increased, whereas plasma nonesterified fatty acid concentrations were similarly decreased across treatments. In mammary biopsies, increased mRNA expression of tumor necrosis factor α, IL-1β, IL-8, and IL-10, and SAA were observed in LPS-treated quarters of all groups, with a more pronounced increase in IL-1β, IL-10, and SAA expression in EuG. Nuclear factor-κB mRNA expression was upregulated in NaCl and EuG but not in HypoG in response to LPS. Lactoferrin, toll-like receptor 4, and cyclooxygenase-2 mRNA expression was increased in LPS-treated quarters of EuG only, and 5-lipoxygenase mRNA expression was decreased

  19. mRNA Cancer Vaccines-Messages that Prevail.

    PubMed

    Grunwitz, Christian; Kranz, Lena M

    2017-01-01

    During the last decade, mRNA became increasingly recognized as a versatile tool for the development of new innovative therapeutics. Especially for vaccine development, mRNA is of outstanding interest and numerous clinical trials have been initiated. Strikingly, all of these studies have proven that large-scale GMP production of mRNA is feasible and concordantly report a favorable safety profile of mRNA vaccines. Induction of T-cell immunity is a multi-faceted process comprising antigen acquisition, antigen processing and presentation, as well as immune stimulation. The effectiveness of mRNA vaccines is critically dependent on making the antigen(s) of interest available to professional antigen-presenting cells, especially DCs. Efficient delivery of mRNA into DCs in vivo remains a major challenge in the mRNA vaccine field. This review summarizes the principles of mRNA vaccines and highlights the importance of in vivo mRNA delivery and recent advances in harnessing their therapeutic potential.

  20. mRNA transcripts as molecular biomarkers in medicine and nutrition

    PubMed Central

    Sunde, Roger A.

    2010-01-01

    In medicine, mRNA transcripts are being developed as molecular biomarkers for the diagnosis and treatment of a number of diseases. These biomarkers offer early and more accurate prediction and diagnosis of disease and disease progression, and ability to identify individuals at risk. Use of microarrays also offers opportunity to identify orthogonal (uncorrelated) biomarkers not known to be linked with conventional biomarkers. Investigators are increasingly using blood as a surrogate tissue for biopsy and analysis; total RNA isolated from whole blood is predominantly from erythroid cells, and whole blood mRNA share more than 80% of the transcriptome with major tissues. Thus blood mRNA biomarkers for individualized disease prediction and diagnosis are an exciting area in medicine; mRNA biomarkers in nutrition have potential application that parallel these opportunities. Assessment of selenium (Se) status and requirements is one area where tissue mRNA levels have been used successfully. Selenoprotein-H and selenoprotein-W as well as glutathione peroxidase-1 (Gpx1) mRNAs are highly down-regulated in Se deficiency in rat liver, and the minimum dietary Se requirement is 0.06–0.07 μg Se/g based on these biomarkers, similar to requirements determined using conventional biomarkers. Blood Gpx1 mRNA can also be used to determine Se requirements in rats, showing that blood mRNA has potential for assessment of nutrient status. Future research is needed to develop mRNA biomarker panels for all nutrients that will discriminate between deficient, marginal, adequate, and supernutritional individuals and populations, and differentiate between individuals that will benefit versus be adversely affected by nutrient supplementation. PMID:20303730

  1. Predictive value of PD-L1 based on mRNA level in the treatment of stage IV melanoma with ipilimumab.

    PubMed

    Brüggemann, C; Kirchberger, M C; Goldinger, S M; Weide, B; Konrad, A; Erdmann, M; Schadendorf, D; Croner, R S; Krähenbühl, L; Kähler, K C; Hafner, C; Leisgang, W; Kiesewetter, F; Dummer, R; Schuler, G; Stürzl, M; Heinzerling, L

    2017-10-01

    PD-L1 is established as a predictive marker for therapy of non-small cell lung cancer with pembrolizumab. Furthermore, PD-L1 positive melanoma has shown more favorable outcomes when treated with anti-PD1 antibodies and dacarbazine compared to PD-L1 negative melanoma. However, the role of PD-L1 expression with regard to response to checkpoint inhibition with anti-CTLA-4 is not clear, yet. In addition, the lack of standardization in the immunohistochemical assessment of PD-L1 makes the comparison of results difficult. In this study, we investigated the PD-L1 gene expression with a new fully automated technique via RT-PCR and correlated the findings with the response to the anti-CTLA-4 antibody ipilimumab. Within a retrospective multi-center trial, PD-L1 gene expression was evaluated in 78 melanoma patients in a total of 111 pre-treatment tumor samples from 6 skin cancer centers and analyzed with regard to response to ipilimumab. For meaningful statistical analysis, the cohort was enriched for responders with 30 responders and 48 non-responders. Gene expression was assessed by quantitative RT-PCR after extracting mRNA from formalin-fixed paraffin embedded tumor tissue and correlated with results from immunohistochemical (IHC) stainings. The evaluation of PD-L1 expression based on mRNA level is feasible. Correlation between PD-L1 expression as assessed by IHC and RT-PCR showed varying levels of concordance depending on the antibody employed. RT-PCR should be further investigated to measure PD-L1 expression, since it is a semi-quantitative method with observer-independent evaluation. With this approach, there was no statistical significant difference in the PD-L1 expression between responders and non-responders to the therapy with ipilimumab. The evaluation of PD-L1 expression based on mRNA level is feasible. Correlation between PD-L1 expression as assessed by IHC and RT-PCR showed varying levels of concordance depending on the antibody employed. RT-PCR should be

  2. Effect of Supplemental Trace Minerals on Hsp-70 mRNA Expression in Commercial Broiler Chicken.

    PubMed

    Rajkumar, U; Vinoth, A; Reddy, E Pradeep Kumar; Shanmugam, M; Rao, S V Rama

    2018-01-02

    The effects of supplementing the organic forms of selenium (Se), chromium (Cr), and zinc (Zn) on Hsp-70 mRNA expression and body weight in broiler chickens were evaluated. 200 chicks were equally distributed into stainless steel battery brooders at the rate of 5 birds per pen and reared under heat stress condition up to 42 nd day. The chicks were fed with three experimental diets supplemented with organic forms of Se (0.30 mg/kg), Cr (2 mg/kg), and Zn (40 mg/kg) during the starter and finisher phases and a control diet without any supplementation. On the 21st and 42nd day, 20 birds from each period were sacrificed and samples were collected for analysis. Organic Se, Cr, and Zn supplementation significantly (P < 0.05) reduced the expression of Hsp-70 mRNA levels. The Hsp-70 mRNA expression levels were significantly (P < 0.05) different between the tissues studied with spleen having the lowest expression level. Hsp-70 mRNA expression level was not affected by age of the birds. The study concluded that organic trace mineral (oTM) supplementation resulted in low Hsp-70 mRNA expression, indicating reduced heat stress in broilers.

  3. Multihormonal induction of hepatic α2u-globulin mRNA as measured by hybridization to complementary DNA

    PubMed Central

    Kurtz, David T.; Feigelson, Philip

    1977-01-01

    A procedure is presented for the preparation of a 3H-labeled complementary DNA (cDNA) specific for the mRNA coding for α2u-globulin, a male rat liver protein under multihormonal control that represents approximately 1% of hepatic protein synthesis. Rat liver polysomes are incubated with monospecific rabbit antiserum to α2u-globulin, which binds to the nascent α2u-globulin chains on the polysomes. These antibody-polysome complexes are then adsorbed to goat antiserum to rabbit IgG that is covalently linked to p-aminobenzylcellulose. mRNA preparations are thus obtained that contain 30-40% α2u-globulin mRNA. A labeled cDNA is made to this α2u-globulin-enriched mRNA preparation by using RNA-dependent DNA polymerase (reverse transcriptase). To remove the non-α2u-globulin sequences, this cDNA preparation is hybridized to an RNA concentration × incubation time (R0t) of 1000 mol of ribonucleotide per liter × sec with female rat liver mRNA, which, though it shares the vast majority of mRNA sequences with male liver, contains no α2u-globulin mRNA sequences. The cDNA remaining single-stranded is isolated by hydroxylapatite chromatography and is shown to be specific for α2u-globulin mRNA by several criteria. Good correlation was found in all endocrine states studied between the hepatic level of α2u-globulin, the level of functional α2u-globulin mRNA as assayed in a wheat germ cell-free translational system, and the level of α2u-globulin mRNA sequences as measured by hybridization to the α2u-globulin cDNA. Thus, the hormonal control of hepatic α2u-globulin synthesis by sex steroids and thyroid hormone occurs through modulation of the cellular level of α2u-globulin mRNA sequences, presumably by hormonal control of transcriptive synthesis. PMID:73184

  4. Impact of fasting followed by short-term exposure to interleukin-6 on cytochrome P450 mRNA in mice.

    PubMed

    Rasmussen, Martin Krøyer; Bertholdt, Lærke; Gudiksen, Anders; Pilegaard, Henriette; Knudsen, Jakob G

    2018-01-05

    The gene expression of the cytochrome P450 (CYP) enzyme family is regulated by numerous factors. Fasting has been shown to induce increased hepatic CYP mRNA in both humans and animals. However, the coordinated regulation of CYP, CYP-regulating transcription factors, and transcriptional co-factors in the liver linking energy metabolism to detoxification has never been investigated. Interleukin-6 (IL-6) has been suggested to be released during fasting and has been shown to regulate CYP expression. The present study investigated the hepatic mRNA content of selected CYP, AhR, CAR, PXR and PPARα in mice fasted for 18h and subsequently exposed to IL-6. Furthermore, the impact of fasting on PGC-1α, HNF-4α, SIRT1 and SIRT3 mRNA was examined. Fasting induced a marked increase in Cyp2b10, Cyp2e1 and Cyp4a10 mRNA, while CYP1a1, Cyp1a2, Cyp2a4 and Cyp3a11 mRNA levels remained unchanged. In accordance, the mRNA levels of CAR and PPARα were also increased with fasting. The PGC-1α, SIRT1 and SIRT3 mRNA levels were also increased after fasting, while the HNF-4α mRNA levels remained unchanged. In mice subjected to IL-6 injection, the fasting-induced PXR, PPARα and PGC-1α mRNA responses were lower than after saline injection. In conclusion, fasting was demonstrated to be a strong inducer of hepatic CYP mRNA as well as selected transcription factors controlling the expression of the investigated CYP. Moreover, the mRNA levels of transcriptional co-factors acting as energy sensors and co-factors for CYP regulation was also increased in the liver, suggesting crosstalk at the molecular level between regulation of energy metabolism and detoxification. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Comparative effects of low-level laser therapy pre- and post-injury on mRNA expression of MyoD, myogenin, and IL-6 during the skeletal muscle repair.

    PubMed

    Alves, Agnelo Neves; Ribeiro, Beatriz Guimarães; Fernandes, Kristianne Porta Santos; Souza, Nadhia Helena Costa; Rocha, Lília Alves; Nunes, Fabio Daumas; Bussadori, Sandra Kalil; Mesquita-Ferrari, Raquel Agnelli

    2016-05-01

    This study analyzed the effect of pre-injury and post-injury irradiation with low-level laser therapy (LLLT) on the mRNA expression of myogenic regulatory factors and interleukin 6 (IL-6) during the skeletal muscle repair. Male rats were divided into six groups: control group, sham group, LLLT group, injury group; pre-injury LLLT group, and post-injury LLLT group. LLLT was performed with a diode laser (wavelength 780 nm; output power 40 mW' and total energy 3.2 J). Cryoinjury was induced by two applications of a metal probe cooled in liquid nitrogen directly onto the belly of the tibialis anterior (TA) muscle. After euthanasia, the TA muscle was removed for the isolation of total RNA and analysis of MyoD, myogenin, and IL-6 using real-time quantitative PCR. Significant increases were found in the expression of MyoD mRNA at 3 and 7 days as well as the expression of myogenin mRNA at 14 days in the post-injury LLLT group in comparison to injury group. A significant reduction was found in the expression of IL-6 mRNA at 3 and 7 days in the pre-injury LLLT and post-injury LLLT groups. A significant increase in IL-6 mRNA was found at 14 days in the post-injury LLLT group in comparison to the injury group. LLLT administered following muscle injury modulates the mRNA expression of MyoD and myogenin. Moreover, the both forms of LLLT administration were able to modulate the mRNA expression of IL-6 during the muscle repair process.

  6. Protopine and allocryptopine increase mRNA levels of cytochromes P450 1A in human hepatocytes and HepG2 cells independently of AhR.

    PubMed

    Vrba, Jiri; Vrublova, Eva; Modriansky, Martin; Ulrichova, Jitka

    2011-06-10

    The isoquinoline alkaloids protopine and allocryptopine are present in phytopreparations from medicinal plants, such as Fumaria officinalis. Since nothing is known about effects of the alkaloids on the expression of xenobiotic-metabolizing enzymes, we examined whether protopine or allocryptopine affect the expression of cytochromes P450 (CYPs) 1A1 and 1A2 in primary cultures of human hepatocytes and human hepatoma HepG2 cells. In HepG2 cells, protopine and allocryptopine significantly increased CYP1A1 mRNA levels after 24h exposure at concentrations from 25 and 10 μM, respectively, as shown by real-time PCR. Both protopine and allocryptopine also dose-dependently increased CYP1A1 and CYP1A2 mRNA levels in human hepatocytes. However, the effects of the tested alkaloids on both cell models were much lower than the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), a prototypical CYP1A inducer. Using gene reporter assays performed in transiently transfected HepG2 cells, we demonstrated that the induction of CYP1A1 expression by either protopine or allocryptopine was associated with mild or negligible activation of the aryl hydrocarbon receptor. In contrast to TCDD, CYP1A mRNA levels induced by protopine or allocryptopine in both HepG2 cells and human hepatocytes did not result in elevated CYP1A protein or activity levels as shown by western blotting and EROD assays, respectively. We conclude that the use of products containing protopine and/or allocryptopine may be considered safe in terms of possible induction of CYP1A enzymes. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  7. Polycystic ovary syndrome influences the level of serum amyloid A and activity of phospholipid transfer protein in HDL₂ and HDL₃.

    PubMed

    Gidwani, S; Phelan, N; McGill, J; McGowan, A; O'Connor, A; Young, I S; Gibney, J; McEneny, J

    2014-07-01

    Is polycystic ovary syndrome (PCOS) associated with altered levels of pro-inflammatory high-density lipoproteins (HDL) and activity of HDL-associated enzymes? In PCOS, HDL contained increased levels of the inflammatory marker serum amyloid A (SAA) and altered functioning of HDL-associated phospholipid transfer protein (PLTP), with these changes being independent of BMI, body fat and insulin resistance (IR). PCOS is associated with adipocyte-derived inflammation, which potentially increases the risk of cardiovascular disease and diabetes. SAA is an inflammatory marker that is released from hypertrophic adipocytes and interacts with HDL, reducing their anti-atherogenic properties. No studies have previously investigated if SAA-associated HDL influences the HDL-associated enzymes namely, PLTP and cholesterol ester transfer protein (CETP) in women with PCOS. Obese women with PCOS were matched with controls for BMI and percentage body fat (n = 100/group; cohort-1); a subset of these women (n = 64/group; cohort-2) were further matched for IR. HDL in blood samples was subfractionated into HDL₂ and HDL₃ by rapid ultracentrifugation. SAA was measured in serum, HDL₂ and HDL₃ by an enzyme-linked immunosorbent assay and the activities of PLTP and CETP were measured in HDL₂ and HDL₃ by fluorimetric assays. In the PCOS women from cohort-1, SAA was increased in serum, HDL₂ and HDL₃ (P = 0.038, 0.008 and 0.001 versus control, respectively), as was the activity of PLTP in HDL₂ and HDL₃ (P = 0.006 and 0.009 versus controls, respectively). In the PCOS women from cohort-2, SAA was increased in serum, HDL₂ and HDL₃, although only significantly in HDL₃ (P = 0.083, 0.120 and 0.034 versus controls, respectively), as was the activity of PLTP in HDL₂ and HDL₃, although this was only significant in HDL₂ (P = 0.045 and 0.070 versus controls, respectively). First, insulin sensitivity was not determined by the euglycaemic-hyperinsulinaemic clamp. Secondly, the

  8. Fractionation of corn fiber treated by soaking in aqueous ammonia (SAA) for isolation of hemicellulose B and production of C5 sugars by enzyme hydrolysis

    USDA-ARS?s Scientific Manuscript database

    A process was developed to fractionate and isolate the hemicellulose B component of corn fiber generated by corn wet milling. The process consisted of pretreatment by soaking in aqueous ammonia (SAA) followed by enzymatic cellulose hydrolysis, during which the hemicellulose B was solubilized by cle...

  9. Effect of iron on accumulation of exotoxin A-specific mRNA in Pseudomonas aeruginosa.

    PubMed Central

    Lory, S

    1986-01-01

    A DNA probe from an internal fragment of the exotoxin A structural gene was used to study the effects of selected culture conditions on steady-state levels of exotoxin-specific mRNA in Pseudomonas aeruginosa. Cells grown under conditions of iron deprivation began to synthesize and excrete the exotoxin A polypeptide during the late exponential phase of growth and throughout the stationary phase of growth, concomitant with a sharp increase in exotoxin A mRNA pools in P. aeruginosa cells. The addition of iron to the medium resulted in the failure of these cells to synthesize exotoxin A mRNA, despite significantly enhanced growth. The inhibition of the production of exotoxin A and the accumulation of its mRNA by iron was dose dependent, with a half-maximal inhibitory concentration of FeSO4 of 5 to 10 microM. A blockade of the initiation of transcription by rifampin resulted in the decay of exotoxin A mRNA, with a half-life of approximately 8 to 10 min, depending on the media used for growth. The addition of iron to cells actively engaged in exotoxin A synthesis also resulted in a gradual decrease in the amount of this mRNA in bacteria. However, the rate of decline of mRNA induced by iron was relatively slow (half-life, 90 min), with a considerable lag time between the iron addition and the first detectable effect on mRNA. While iron clearly appears to influence the production of exotoxin A at the transcriptional level, the molecular basis of this effect may involve several interacting factors affecting the initiation of transcription and perhaps mRNA turnover. Images PMID:2430950

  10. Absolute Measurements of Macrophage Migration Inhibitory Factor and Interleukin-1-β mRNA Levels Accurately Predict Treatment Response in Depressed Patients.

    PubMed

    Cattaneo, Annamaria; Ferrari, Clarissa; Uher, Rudolf; Bocchio-Chiavetto, Luisella; Riva, Marco Andrea; Pariante, Carmine M

    2016-10-01

    Increased levels of inflammation have been associated with a poorer response to antidepressants in several clinical samples, but these findings have had been limited by low reproducibility of biomarker assays across laboratories, difficulty in predicting response probability on an individual basis, and unclear molecular mechanisms. Here we measured absolute mRNA values (a reliable quantitation of number of molecules) of Macrophage Migration Inhibitory Factor and interleukin-1β in a previously published sample from a randomized controlled trial comparing escitalopram vs nortriptyline (GENDEP) as well as in an independent, naturalistic replication sample. We then used linear discriminant analysis to calculate mRNA values cutoffs that best discriminated between responders and nonresponders after 12 weeks of antidepressants. As Macrophage Migration Inhibitory Factor and interleukin-1β might be involved in different pathways, we constructed a protein-protein interaction network by the Search Tool for the Retrieval of Interacting Genes/Proteins. We identified cutoff values for the absolute mRNA measures that accurately predicted response probability on an individual basis, with positive predictive values and specificity for nonresponders of 100% in both samples (negative predictive value=82% to 85%, sensitivity=52% to 61%). Using network analysis, we identified different clusters of targets for these 2 cytokines, with Macrophage Migration Inhibitory Factor interacting predominantly with pathways involved in neurogenesis, neuroplasticity, and cell proliferation, and interleukin-1β interacting predominantly with pathways involved in the inflammasome complex, oxidative stress, and neurodegeneration. We believe that these data provide a clinically suitable approach to the personalization of antidepressant therapy: patients who have absolute mRNA values above the suggested cutoffs could be directed toward earlier access to more assertive antidepressant strategies

  11. Regulation of Hippocampal α1d Adrenergic Receptor mRNA by Corticosterone in Adrenalectomized Rats

    PubMed Central

    Day, Heidi E.W.; Kryskow, Elisa M.; Watson, Stanley J.; Akil, Huda; Campeau, Serge

    2008-01-01

    The hippocampal formation receives extensive noradrenergic projections and expresses high levels of mineralocorticoid (MR) and glucocorticoid (GR) receptors. Considerable evidence suggests that the noradrenergic system influences hippocampal corticosteroid receptors. However, there is relatively little data describing the influence of glucocorticoids on noradrenergic receptors in the hippocampal formation. α1d adrenergic receptor (ADR) mRNA is expressed at high levels in the hippocampal formation, within cells that express MR or GR. In order to determine whether expression of α1d ADR mRNA is influenced by circulating glucocorticoids, male rats underwent bilateral adrenalectomy (ADX) or sham surgery, and were killed after 1, 3, 7 or 14 days. Levels of α1d ADR mRNA were profoundly decreased in hippocampal subfields CA1, CA2 and CA3 and the medial and lateral blades of the dentate gyrus, as early as 1 day after ADX, as determined by in situ hybridization. The effect was specific for the hippocampal formation, with levels of α1d mRNA unaltered by ADX in the lateral amygdala, reticular thalamic nucleus, retrosplenial cortex or primary somatosensory cortex. Additional rats underwent ADX or sham surgery and received a corticosterone pellet (10 or 50 mg) or placebo for 7 days. Corticosterone replacement prevented the ADX-induced decrease in hippocampal α1d ADR mRNA, with the magnitude of effect depending on corticosterone dose and hippocampal subregion. These data indicate that α1d ADR mRNA expression in the hippocampal formation is highly sensitive to circulating levels of corticosterone, and provides further evidence for a close interaction between glucocorticoids and the noradrenergic system in the hippocampus. PMID:18534559

  12. Consequences of metaphase II oocyte cryopreservation on mRNA content.

    PubMed

    Chamayou, S; Bonaventura, G; Alecci, C; Tibullo, D; Di Raimondo, F; Guglielmino, A; Barcellona, M L

    2011-04-01

    We studied the consequences of freezing/thawing processes on mRNA contents in MII oocytes after slow-freezing/rapid thawing (SF/RT) and vitrification/warming (V/W) protocols, and compared the results to fresh MII oocytes. We quantified the nuclear transcript mRNA responsible for the translation of proteins belonging either to trans-regulatory protein family or to functional structural proteins such as proteins involved in DNA structural organization (NAP1L1, TOP1, H1F0H1), chromosomal structure maintenance (SMC, SCC3, RAD21, SMC1A, SMC1B, STAG3, REC8), mitochondrial energetic pathways (ATP5GJ, SDHC), cell cycle regulation and processes (CLTA, MAPK6, CKS2) and staminal cell potency-development competence stage (DPPA3, OCT4, FOXJ2). Surplus MII oocytes were donated from patients in IVF cycles and divided in three groups of 15 oocytes. Group 1 was comprised of non-cryopreserved oocytes and Groups 2 and 3 underwent SF/RT and V/W procedures, respectively. There was an overall decrease of mRNA extracted from cryopreserved oocytes compared to control group. Only 39.4% of mRNA content were preserved after SF/RT while 63.3% of mRNA content were maintained after V/W. Oocyte cryopreservation is associated with molecular injury associated with the decrease of stored mRNA. However the V/W protocol is more conservative than SF/RT resulting in a level of mRNA sufficient to maintain biologic functions in the subsequent fertilized oocyte. Copyright © 2011 Elsevier Inc. All rights reserved.

  13. Midbrain dopamine neurons regulate preprotachykinin-A mRNA expression in the rat forebrain during development.

    PubMed

    Brené, S; Lindefors, N; Persson, H

    1992-06-01

    Intracerebroventricular 6-hydroxydopamine injections were performed at postnatal days 3 and 6 in animals pretreated with the norepinephrine uptakeblocker desimipramine in order to generate a selective lesion of dopamine neurons. In situ hybridization was then used to analyze preprotachykinin-A (PPT-A) mRNA expression in the lesioned as well as in saline-injected control animals. The midbrain dopaminergic lesion caused a 22-25% increase in the level of PPT-A mRNA in cingulate cortex and frontoparietal cortex when analysed at 2 weeks of age, compared to saline-injected control animals. In contrast, the lesion caused no change in PPT-A mRNA expression in the neonatal caudate-putamen. These results indicate that dopamine neurons downregulate the expression of PPT-A mRNA specifically in cingulate cortex and frontoparietal cortex during early postnatal brain development. In the adult rat forebrain, lesioned at P3 and P6, no change in the level of PPT-A mRNA was seen in cingulate cortex and frontoparietal cortex. However, a 29% decrease in PPT-A mRNA was seen in the lateral caudate-putamen with no significant change in neurons of medial caudate-putamen. Thus, dopamine neurons appears to exert a region specific influence on PPT-A mRNA expression during brain development.

  14. Polysome Fractionation to Analyze mRNA Distribution Profiles.

    PubMed

    Panda, Amaresh C; Martindale, Jennifer L; Gorospe, Myriam

    2017-02-05

    Eukaryotic cells adapt to changes in external or internal signals by precisely modulating the expression of specific gene products. The expression of protein-coding genes is controlled at the transcriptional and post-transcriptional levels. Among the latter steps, the regulation of translation is particularly important in cellular processes that require rapid changes in protein expression patterns. The translational efficiency of mRNAs is altered by RNA-binding proteins (RBPs) and noncoding (nc)RNAs such as microRNAs (Panda et al. , 2014a and 2014b; Abdelmohsen et al. , 2014). The impact of factors that regulate selective mRNA translation is a critical question in RNA biology. Polyribosome (polysome) fractionation analysis is a powerful method to assess the association of ribosomes with a given mRNA. It provides valuable information about the translational status of that mRNA, depending on the number of ribosomes with which they are associated, and identifies mRNAs that are not translated (Panda et al. , 2016). mRNAs associated with many ribosomes form large polysomes that are predicted to be actively translated, while mRNAs associated with few or no ribosomes are expected to be translated poorly if at all. In sum, polysome fractionation analysis allows the direct determination of translation efficiencies at the level of the whole transcriptome as well as individual mRNAs.

  15. Regulation of mouse hepatic CYP2D9 mRNA expression by growth and adrenal hormones.

    PubMed

    Jarukamjorn, Kanokwan; Sakuma, Tsutomu; Jaruchotikamol, Atika; Oguro, Miki; Nemoto, Nobuo

    2006-02-01

    The constitutive expression of CYP2D9 is sexually dimorphic, namely, strong in males, but diminutive in females. Repetition of mimic growth hormone (GH) secretion pattern impressively returned the mRNA expression level to that in intact mice: the GH secretion pattern's regulation of CYP2D9 mRNA expression has been predominantly disrupted by exogenous GH-administration. The extensive decline of CYP2D9 mRNA expression becoming a sexually non-specific P450 in 9-week-old male mice exposed as neonates to monosodium L-glutamate (MSG) suggested that the male GH secretion pattern is a key to the regulation of male-specific CYP2D9 mRNA expression in adult mice. Dexamethasone (Dex) showed possibility to induce CYP2D9 mRNA expression in adult MSG-neonatally treated mice of either sex. However, the antagonism was observed by co-administration of Dex and GH in the males. Dex-administration in adrenalectomized mice significantly elevated CYP2D9 mRNA expression levels. These findings suggest that an adrenal hormone participates in the regulatory mechanism of CYP2D9 mRNA expression in association with GH.

  16. Replenishment of RANTES mRNA expression in activated eosinophils fromatopic asthmatics

    PubMed Central

    Velazquez, J R; Lacy, P; Moqbel, R

    2000-01-01

    Eosinophils have been shown to express the gene encoding regulated upon activation, normal T‐cell expressed and secreted (RANTES), a potent eosinophilotactic chemokine. RANTES protein expression in eosinophils has previously been shown to be up‐regulated by a number of agonists, including complement‐dependent factors (C3b/iC3b) and interferon‐γ (IFN‐γ). We hypothesized that gene expression of RANTES is regulated in these cells by eosinophil‐specific agonists. We analysed RANTES mRNA expression by reverse‐transcription polymerase chain reaction (RT‐PCR) in human peripheral blood eosinophils obtained from mild atopic asthmatics following stimulation over time. In resting eosinophils, a low level of RANTES mRNA was found to be constitutively expressed in all the atopic donors tested in this study (n = 6). Following stimulation with C3b/iC3b (serum‐coated surfaces), eosinophils released measurable levels of RANTES, while sustained transcript expression was detected for up to 24 hr of stimulation. In contrast, IFN‐γ (5 ng/ml) transiently and significantly (P < 0·05, n = 3) depleted relative amounts of RANTES PCR product (compared with β2‐microglobulin) after 1–4 hr of stimulation. RANTES transcript was again detectable after 24 hr of IFN‐γ incubation, suggesting that the pool of RANTES mRNA had been replenished. Other eosinophil‐active cytokines, interleukin‐3 (IL‐3), IL‐4, IL‐5 and granulocyte–macrophage colony‐stimulating factor, did not appear to modulate RANTES mRNA expression after 1 hr of incubation. The effect of IFN‐γ on RANTES mRNA was reversed by cycloheximide, suggesting that IFN‐γ may act by increasing the rate of translation of RANTES mRNA. These findings indicate that IFN‐γ may induce a rapid and transient effect on the translation and replenishment of RANTES mRNA in eosinophils. This novel observation supports the notion that eosinophils have the potential to replenish their stored and released

  17. Advanced Running Performance by Genetic Predisposition in Male Dummerstorf Marathon Mice (DUhTP) Reveals Higher Sterol Regulatory Element-Binding Protein (SREBP) Related mRNA Expression in the Liver and Higher Serum Levels of Progesterone

    PubMed Central

    Brenmoehl, Julia; Walz, Christina; Ponsuksili, Siriluck; Schwerin, Manfred; Fuellen, Georg; Hoeflich, Andreas

    2016-01-01

    Long-term-selected DUhTP mice represent a non-inbred model for inborn physical high-performance without previous training. Abundance of hepatic mRNA in 70-day male DUhTP and control mice was analyzed using the Affymetrix mouse array 430A 2.0. Differential expression analysis with PLIER corrected data was performed using AltAnalyze. Searching for over-representation in biochemical pathways revealed cholesterol metabolism being most prominently affected in DUhTP compared to unselected control mice. Furthermore, pathway analysis by AltAnalyze plus PathVisio indicated significant induction of glycolysis, fatty acid synthesis and cholesterol biosynthesis in the liver of DUhTP mice versus unselected control mice. In contrast, gluconeogenesis was partially inactivated as judged from the analysis of hepatic mRNA transcript abundance in DUhTP mice. Analysis of mRNA transcripts related to steroid hormone metabolism inferred elevated synthesis of progesterone and reduced levels of sex steroids. Abundance of steroid delta isomerase-5 mRNA (Hsd3b5, FC 4.97) was increased and steroid 17-alpha-monooxygenase mRNA (Cyp17a1, FC -11.6) was massively diminished in the liver of DUhTP mice. Assessment of steroid profiles by LC-MS revealed increased levels of progesterone and decreased levels of sex steroids in serum from DUhTP mice versus controls. Analysis of hepatic mRNA transcript abundance indicates that sterol regulatory element-binding protein-1 (SREBP-1) may play a major role in metabolic pathway activation in the marathon mouse model DUhTP. Thus, results from bioinformatics modeling of hepatic mRNA transcript abundance correlated with direct steroid analysis by mass spectrometry and further indicated functions of SREBP-1 and steroid hormones for endurance performance in DUhTP mice. PMID:26799318

  18. Low-level laser irradiation alters mRNA expression from genes involved in DNA repair and genomic stabilization in myoblasts

    NASA Astrophysics Data System (ADS)

    Trajano, L. A. S. N.; Sergio, L. P. S.; Silva, C. L.; Carvalho, L.; Mencalha, A. L.; Stumbo, A. C.; Fonseca, A. S.

    2016-07-01

    Low-level lasers are used for the treatment of diseases in soft and bone tissues, but few data are available regarding their effects on genomic stability. In this study, we investigated mRNA expression from genes involved in DNA repair and genomic stabilization in myoblasts exposed to low-level infrared laser. C2C12 myoblast cultures in different fetal bovine serum concentrations were exposed to low-level infrared laser (10, 35 and 70 J cm-2), and collected for the evaluation of DNA repair gene expression. Laser exposure increased gene expression related to base excision repair (8-oxoguanine DNA glycosylase and apurinic/apyrimidinic endonuclease 1), nucleotide excision repair (excision repair cross-complementation group 1 and xeroderma pigmentosum C protein) and genomic stabilization (ATM serine/threonine kinase and tumor protein p53) in normal and low fetal bovine serum concentrations. Results suggest that genomic stability could be part of a biostimulation effect of low-level laser therapy in injured muscles.

  19. Modulation of transferrin receptor mRNA by transferrin-gallium in human myeloid HL60 and lymphoid CCRF-CEM leukaemic cells.

    PubMed Central

    Ul-Haq, R; Chitambar, C R

    1993-01-01

    Gallium binds to the iron transport protein transferrin (Tf), is incorporated into cells through transferrin receptors (TfR) and inhibits iron-dependent DNA synthesis. Since cellular TfR expression is tightly regulated by the availability of iron, we investigated the effects of transferrin-gallium (Tf-Ga) on TfR mRNA levels in myeloid HL60 and lymphoid CCRF-CEM cells. In HL60 cells, Tf-Ga increased TfR mRNA levels in a dose-dependent fashion. This increase in TfR mRNA was blocked by Tf-Fe and by cycloheximide. Analysis of the rate of mRNA decay in the presence of actinomycin D revealed that the half-life of TfR mRNA was increased in HL60 cells incubated with Tf-Ga. The rate of transcription of TfR mRNA was not increased by Tf-Ga. In contrast with HL60 cells, CCRF-CEM cells displayed a decrease in the level of TfR mRNA after incubation with Tf-Ga. Tf-Ga inhibited iron uptake in both HL60 and CCRF-CEM cells but increased the level of TfR mRNA only in HL60 cells, suggesting that the Tf-Ga induction of TfR mRNA was not solely due to inhibition of cellular iron uptake. At growth-inhibitory concentrations, Tf-Ga increased the TfR mRNA level in HL60 cells but decreased it in CCRF-CEM cells. Our studies suggest that in HL60 cells, gallium regulates TfR expression at the post-transcriptional level by mechanisms which require de novo protein synthesis and involve interaction with iron. The divergent effects of Tf-Ga on TfR mRNA in myeloid HL60 and lymphoid CCRF-CEM cells suggest that differences exist in the regulation of TfR expression between these two cell types. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:8379943

  20. Elevation of D4 dopamine receptor mRNA in postmortem schizophrenic brain.

    PubMed

    Stefanis, N C; Bresnick, J N; Kerwin, R W; Schofield, W N; McAllister, G

    1998-01-01

    The D4 dopamine (DA) receptor has been proposed to be a target for the development of a novel antipsychotic drug based on its pharmacological and distribution profile. There is much interest in whether D4 DA receptor levels are altered in schizophrenia, but the lack of an available receptor subtype-specific radioligand made this difficult to quantitate. In this study, we examined whether D4 mRNA levels are altered in different brain regions of schizophrenics compared to controls. Ribonuclease protection assays were carried out on total RNA samples isolated postmortem from frontal cortex and caudate brain regions of schizophrenics and matched controls. 32P-labelled RNA probes to the D4 DA receptor and to the housekeeping gene, glyceraldehyde-3-phosphate dehydrogenase (G3PDH), were hybridised with the RNA samples, digested with ribonucleases to remove unhybridised probe, and separated on 6% sequencing gels. Densitometer analysis on the subsequent autoradiogams was used to calculate the relative optical density of D4 mRNA compared to G3PDH mRNA. Statistical analysis of the data revealed a 3-fold higher level (P<0.011) of D4 mRNA in the frontal cortex of schizophrenics compared to controls. No increase was seen in caudate. D4 receptors could play a role in mediating dopaminergic activity in frontal cortex, an activity which may be malfunctioning in schizophrenia.

  1. RNA-Binding Proteins Revisited - The Emerging Arabidopsis mRNA Interactome.

    PubMed

    Köster, Tino; Marondedze, Claudius; Meyer, Katja; Staiger, Dorothee

    2017-06-01

    RNA-protein interaction is an important checkpoint to tune gene expression at the RNA level. Global identification of proteins binding in vivo to mRNA has been possible through interactome capture - where proteins are fixed to target RNAs by UV crosslinking and purified through affinity capture of polyadenylated RNA. In Arabidopsis over 500 RNA-binding proteins (RBPs) enriched in UV-crosslinked samples have been identified. As in mammals and yeast, the mRNA interactomes came with a few surprises. For example, a plethora of the proteins caught on RNA had not previously been linked to RNA-mediated processes, for example proteins of intermediary metabolism. Thus, the studies provide unprecedented insights into the composition of the mRNA interactome, highlighting the complexity of RNA-mediated processes. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Pulmonary Response to Surface-Coated Nanotitanium Dioxide Particles Includes Induction of Acute Phase Response Genes, Inflammatory Cascades, and Changes in MicroRNAs: A Toxicogenomic Study

    PubMed Central

    Halappanavar, Sabina; Jackson, Petra; Williams, Andrew; Jensen, Keld A; Hougaard, Karin S; Vogel, Ulla; Yauk, Carole L; Wallin, Håkan

    2011-01-01

    Titanium dioxide nanoparticles (nanoTiO2) are used in various applications including in paints. NanoTiO2 inhalation may induce pulmonary toxicity and systemic effects. However, the underlying molecular mechanisms are poorly understood. In this study, the effects of inhaled surface-coated nanoTiO2 on pulmonary global messenger RNA (mRNA) and microRNA (miRNA) expression in mouse were characterized to provide insight into the molecular response. Female C57BL/6BomTac mice were exposed for 1 hr daily to 42.4 ± 2.9 (SEM) mg surface-coated nanoTiO2/m3 for 11 consecutive days by inhalation and were sacrificed 5 days following the last exposure. Physicochemical properties of the particles were determined. Pulmonary response to nanoTiO2 was characterized using DNA microarrays and pathway-specific PCR arrays and related to data on pulmonary inflammation from bronchial lavages. NanoTiO2 exposure resulted in increased levels of mRNA for acute phase markers serum amyloid A-1 (Saa1) and serum amyloid A-3 (Saa3), several C-X-C and C-C motif chemokines, and cytokine tumor necrosis factor genes. Protein analysis of Saa1 and 3 showed selective upregulation of Saa3 in lung tissues. Sixteen miRNAs were induced by more than 1.2-fold (adjusted P-value < 0.05) following exposure. Real time polymerase chain reaction confirmed the upregulation of miR-1, miR-449a and revealed dramatic induction of miR-135b (60-fold). Thus, inhalation of surface-coated nanoTiO2 results in changes in the expression of genes associated with acute phase, inflammation and immune response 5 days post exposure with concomitant changes in several miRNAs. The role of these miRNAs in pulmonary response to inhaled particles is unknown and warrants further research. Environ. Mol. Mutagen., 2011. © 2011 Wiley-Liss, Inc.† PMID:21259345

  3. Attenuated acute salivary α-amylase responses to gustatory stimulation with citric acid in thin children.

    PubMed

    Chen, Long Hui; Yang, Ze Min; Chen, Wei Wen; Lin, Jing; Zhang, Min; Yang, Xiao Rong; Zhao, Ling Bo

    2015-04-14

    Salivary α-amylase (sAA) is responsible for the 'pre-digestion' of starch in the oral cavity and accounts for up to 50 % of salivary protein in human saliva. An accumulating body of literature suggests that sAA is of nutritional importance; however, it is still not clear how sAA is related to individual's nutritional status. Although copy number variations (CNV) of the salivary amylase gene (AMY1) are associated with variation in sAA levels, a significant amount of sAA variation is not explained by AMY1 CNV. To measure sAA responses to gustatory stimulation with citric acid, we used sAA ratio (the ratio of stimulated sAA levels to those of resting sAA) and investigated acute sAA responses to citric acid in children with normal (Normal-BMI, n 22) and low (Low-BMI, n 21) BMI. The AMY1 gene copy number was determined by quantitative PCR. We, for the first time, demonstrated attenuated acute sAA responses (decreased sAA ratio) to gustatory stimulation in Low-BMI (thinness grade 3) children compared with the Normal-BMI children, which suggest that sAA responses to gustatory stimulation may be of nutritional importance. However, child's nutritional status was not directly related to their resting or stimulated sAA levels, and it was not associated with AMY1 gene copy number. Finally, AMY1 CNV might influence, but did not eventually determine, sAA levels in children.

  4. The decapping activator Edc3 and the Q/N-rich domain of Lsm4 function together to enhance mRNA stability and alter mRNA decay pathway dependence in Saccharomyces cerevisiae.

    PubMed

    Huch, Susanne; Müller, Maren; Muppavarapu, Mridula; Gommlich, Jessie; Balagopal, Vidya; Nissan, Tracy

    2016-10-15

    The rate and regulation of mRNA decay are major elements in the proper control of gene expression. Edc3 and Lsm4 are two decapping activator proteins that have previously been shown to function in the assembly of RNA granules termed P bodies. Here, we show that deletion of edc3, when combined with a removal of the glutamine/asparagine rich region of Lsm4 (edc3Δ lsm4ΔC) reduces mRNA stability and alters pathways of mRNA degradation. Multiple tested mRNAs exhibited reduced stability in the edc3Δ lsm4ΔC mutant. The destabilization was linked to an increased dependence on Ccr4-mediated deadenylation and mRNA decapping. Unlike characterized mutations in decapping factors that either are neutral or are able to stabilize mRNA, the combined edc3Δ lsm4ΔC mutant reduced mRNA stability. We characterized the growth and activity of the major mRNA decay systems and translation in double mutant and wild-type yeast. In the edc3Δ lsm4ΔC mutant, we observed alterations in the levels of specific mRNA decay factors as well as nuclear accumulation of the catalytic subunit of the decapping enzyme Dcp2. Hence, we suggest that the effects on mRNA stability in the edc3Δ lsm4ΔC mutant may originate from mRNA decay protein abundance or changes in mRNPs, or alternatively may imply a role for P bodies in mRNA stabilization. © 2016. Published by The Company of Biologists Ltd.

  5. The decapping activator Edc3 and the Q/N-rich domain of Lsm4 function together to enhance mRNA stability and alter mRNA decay pathway dependence in Saccharomyces cerevisiae

    PubMed Central

    Huch, Susanne; Müller, Maren; Muppavarapu, Mridula; Gommlich, Jessie; Balagopal, Vidya; Nissan, Tracy

    2016-01-01

    ABSTRACT The rate and regulation of mRNA decay are major elements in the proper control of gene expression. Edc3 and Lsm4 are two decapping activator proteins that have previously been shown to function in the assembly of RNA granules termed P bodies. Here, we show that deletion of edc3, when combined with a removal of the glutamine/asparagine rich region of Lsm4 (edc3Δ lsm4ΔC) reduces mRNA stability and alters pathways of mRNA degradation. Multiple tested mRNAs exhibited reduced stability in the edc3Δ lsm4ΔC mutant. The destabilization was linked to an increased dependence on Ccr4-mediated deadenylation and mRNA decapping. Unlike characterized mutations in decapping factors that either are neutral or are able to stabilize mRNA, the combined edc3Δ lsm4ΔC mutant reduced mRNA stability. We characterized the growth and activity of the major mRNA decay systems and translation in double mutant and wild-type yeast. In the edc3Δ lsm4ΔC mutant, we observed alterations in the levels of specific mRNA decay factors as well as nuclear accumulation of the catalytic subunit of the decapping enzyme Dcp2. Hence, we suggest that the effects on mRNA stability in the edc3Δ lsm4ΔC mutant may originate from mRNA decay protein abundance or changes in mRNPs, or alternatively may imply a role for P bodies in mRNA stabilization. PMID:27543059

  6. Expression kinetics of nucleoside-modified mRNA delivered in lipid nanoparticles to mice by various routes.

    PubMed

    Pardi, Norbert; Tuyishime, Steven; Muramatsu, Hiromi; Kariko, Katalin; Mui, Barbara L; Tam, Ying K; Madden, Thomas D; Hope, Michael J; Weissman, Drew

    2015-11-10

    In recent years, in vitro transcribed messenger RNA (mRNA) has emerged as a potential therapeutic platform. To fulfill its promise, effective delivery of mRNA to specific cell types and tissues needs to be achieved. Lipid nanoparticles (LNPs) are efficient carriers for short-interfering RNAs and have entered clinical trials. However, little is known about the potential of LNPs to deliver mRNA. Here, we generated mRNA-LNPs by incorporating HPLC purified, 1-methylpseudouridine-containing mRNA comprising codon-optimized firefly luciferase into stable LNPs. Mice were injected with 0.005-0.250mg/kg doses of mRNA-LNPs by 6 different routes and high levels of protein translation could be measured using in vivo imaging. Subcutaneous, intramuscular and intradermal injection of the LNP-encapsulated mRNA translated locally at the site of injection for up to 10days. For several days, high levels of protein production could be achieved in the lung from the intratracheal administration of mRNA. Intravenous and intraperitoneal and to a lesser extent intramuscular and intratracheal deliveries led to trafficking of mRNA-LNPs systemically resulting in active translation of the mRNA in the liver for 1-4 days. Our results demonstrate that LNPs are appropriate carriers for mRNA in vivo and have the potential to become valuable tools for delivering mRNA encoding therapeutic proteins. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Lower FOXO3 mRNA expression in granulosa cells is involved in unexplained infertility.

    PubMed

    Yamamoto, Hikaru; Yamashita, Yoshiki; Saito, Natsuho; Hayashi, Atsushi; Hayashi, Masami; Terai, Yoshito; Ohmichi, Masahide

    2017-06-01

    The aim of this study was to investigate whether FOXO1 and FOXO3 mRNA expression in granulosa cells is the cause of unexplained infertility. Thirty-one patients aged <40 years (13 with unexplained infertility and 18 with male partner infertility as a control group) whose serum anti-Müllerian hormone level was >0.5 ng/μL were enrolled in the study. All patients underwent oocyte retrieval under a short protocol from June 2012 to October 2013. Real-time PCR was carried out using mRNA extracted from granulosa cells retrieved from mature follicles. We compared FOXO1 and FOXO3 mRNA expression ratios in granulosa cells between the unexplained infertility group and the male infertility group. The relation between FOXO1 and FOXO3 mRNA expression ratios in granulosa cells and assisted reproduction technology clinical outcome was also examined. FOXO3 mRNA expression ratio was significantly lower in the unexplained infertility group than in the male infertility group. Moreover, FOXO3 mRNA expression ratio showed a positive correlation with both the number of retrieved oocytes and serum anti-Müllerian hormone level. A positive correlation was also identified between FOXO1 mRNA expression and total dose of hMG. As well, the number of retrieved oocytes in the unexplained infertility group was statistically lower than that in the male infertility group. A lower FOXO3 mRNA expression in granulosa cells leads to poor oocyte development in patients with unexplained infertility undergoing controlled ovarian stimulation for in vitro fertilization-embryo transfer. © 2017 Japan Society of Obstetrics and Gynecology.

  8. NONOates regulate KCl cotransporter-1 and -3 mRNA expression in vascular smooth muscle cells.

    PubMed

    Di Fulvio, Mauricio; Lauf, Peter K; Shah, Shalin; Adragna, Norma C

    2003-05-01

    Nitric oxide (NO) donors regulate KCl cotransport (KCC) activity and cotransporter-1 and -3 (KCC1 and KCC3) mRNA expression in sheep erythrocytes and in primary cultures of rat vascular smooth muscle cells (VSMCs), respectively. In this study, we used NONOates as rapid and slow NO releasers to provide direct evidence implicating NO as a regulator of KCC3 gene expression at the mRNA level. In addition, we used the expression of KCC3 mRNA to further investigate the mechanism of action of these NO donors at the cellular level. Treatment of VSMCs with rapid NO releasers, like NOC-5 and NOC-9, as well as with the direct NO-independent soluble guanylyl cyclase (sGC) stimulator YC-1, acutely increased KCC3 mRNA expression in a concentration- and time-dependent manner. The slow NO releaser NOC-18 had no effect on KCC3 gene expression. A specific NO scavenger completely prevented the NONOate-induced KCC3 mRNA expression. Inhibition of sGC with LY-83583 blocked the NONOate- and YC-1-induced KCC3 mRNA expression. This study shows that in primary cultures of rat VSMCs, the fast NO releasers NOC-9 and NOC-5, but not the slow NO releaser NOC-18, acutely upregulate KCC3 mRNA expression in a NO/sGC-dependent manner.

  9. Altered Sex Hormone Concentrations and Gonadal mRNA Expression Levels of Activin Signaling Factors in Hatchling Alligators From a Contaminated Florida Lake

    PubMed Central

    MOORE, BRANDON C.; KOHNO, SATOMI; COOK, ROBERT W.; ALVERS, ASHLEY L.; HAMLIN, HEATHER J.; WOODRUFF, TERESA K.; GUILLETTE, LOUIS J.

    2014-01-01

    Activins and estrogens participate in regulating the breakdown of ovarian germ cell nests and follicle assembly in mammals. In 1994, our group reported elevated frequencies of abnormal, multioocytic ovarian follicles in 6 month old, environmental contaminant-exposed female alligators after gonadotropin challenge. Here, we investigated if maternal contribution of endocrine disrupting contaminants to the egg subsequently alters estrogen/inhibin/activin signaling in hatchling female offspring, putatively predisposing an increased frequency of multioocytic follicle formation. We quantified basal and exogenous gonadotropin-stimulated concentrations of circulating plasma steroid hormones and ovarian activin signaling factor mRNA abundance in hatchling alligators from the same contaminated (Lake Apopka) and reference (Lake Woodruff) Florida lakes, as examined in 1994. Basal circulating plasma estradiol and testosterone concentrations were greater in alligators from the contaminated environment, whereas activin/inhibin βA subunit and follistatin mRNA abundances were lower than values measured in ovaries from reference lake animals. Challenged, contaminant-exposed animals showed a more robust increase in plasma estradiol concentration following an acute follicle stimulating hormone (FSH) challenge compared with reference site alligators. Aromatase and follistatin mRNA levels increased in response to an extended FSH challenge in the reference site animals, but not in the contaminant-exposed animals. In hatchling alligators, ovarian follicles have not yet formed; therefore, these endocrine differences are likely to affect subsequent ovarian development, including ovarian follicle assembly. PMID:20166196

  10. Differential regulation of preprotachykinin-A mRNA expression in striatum by excitation of hippocampal neurons.

    PubMed

    Brené, S; Lindefors, N; Herrera-Marschitz, M; Persson, H

    1993-07-01

    In this report we have studied the influence of hippocampal neurons on neuropeptide mRNA expression in both dorsal and ventral striatum in the rat. Intrahippocampal unilateral kainic acid injections were performed in control animals and in animals with a unilateral 6-hydroxydopamine-induced dopamine deafferentation of the striatum. In situ hybridization combined with quantitative image analysis was used to study the expression of preprotachykinin A mRNA encoding the neuropeptides substance P and neurokinin A. The 6-hydroxydopamine-induced lesion caused a decrease of preprotachykinin A mRNA levels in the ipsilateral dorsal striatum and in both sides of the ventral striatum. In normal rats, the intrahippocampal kainic acid injection caused a twofold increase in preprotachykinin A mRNA in the limbic parts of the striatum, which are innervated by the hippocampus. No effect of the kainic acid injection was seen in the lateral parts of the dorsal striatum, a region which does not appear to be innervated by the hippocampus. Animals with a 6-hydroxydopamine lesion showed a similar kainic acid-mediated increase in preprotachykinin A mRNA in parts of the ventral striatum. In the dopamine-lesioned dorsal striatum and ventral striatum the decreased preprotachykinin A mRNA levels were normalized by the intrahippocampal kainic acid injection. These results show that kainic acid-mediated excitation of hippocampal neurons causes a dopamine-independent induction of preprotachykinin A mRNA expression in parts of the ventral striatum, and reverses the dopamine deafferentation-induced decrease of preprotachykinin A mRNA in both dorsal and ventral striatum. Combined, our results suggest that hippocampal neurons can regulate preprotachykinin A mRNA expression in both the ventral and the dorsal striatum.

  11. Messenger RNA (mRNA) nanoparticle tumour vaccination

    NASA Astrophysics Data System (ADS)

    Phua, Kyle K. L.; Nair, Smita K.; Leong, Kam W.

    2014-06-01

    Use of mRNA-based vaccines for tumour immunotherapy has gained increasing attention in recent years. A growing number of studies applying nanomedicine concepts to mRNA tumour vaccination show that the mRNA delivered in nanoparticle format can generate a more robust immune response. Advances in the past decade have deepened our understanding of gene delivery barriers, mRNA's biological stability and immunological properties, and support the notion for engineering innovations tailored towards a more efficient mRNA nanoparticle vaccine delivery system. In this review we will first examine the suitability of mRNA for engineering manipulations, followed by discussion of a model framework that highlights the barriers to a robust anti-tumour immunity mediated by mRNA encapsulated in nanoparticles. Finally, by consolidating existing literature on mRNA nanoparticle tumour vaccination within the context of this framework, we aim to identify bottlenecks that can be addressed by future nanoengineering research.

  12. Exaptive origins of regulated mRNA decay in eukaryotes

    PubMed Central

    Hamid, Fursham M.

    2016-01-01

    Eukaryotic gene expression is extensively controlled at the level of mRNA stability and the mechanisms underlying this regulation are markedly different from their archaeal and bacterial counterparts. We propose that two such mechanisms, nonsense‐mediated decay (NMD) and motif‐specific transcript destabilization by CCCH‐type zinc finger RNA‐binding proteins, originated as a part of cellular defense against RNA pathogens. These branches of the mRNA turnover pathway might have been used by primeval eukaryotes alongside RNA interference to distinguish their own messages from those of RNA viruses and retrotransposable elements. We further hypothesize that the subsequent advent of “professional” innate and adaptive immunity systems allowed NMD and the motif‐triggered mechanisms to be efficiently repurposed for regulation of endogenous cellular transcripts. This scenario explains the rapid emergence of archetypical mRNA destabilization pathways in eukaryotes and argues that other aspects of post‐transcriptional gene regulation in this lineage might have been derived through a similar exaptation route. PMID:27438915

  13. Regulation of Egr-1, VIP, and Shh mRNA and Egr-1 protein in the mouse retina by light and image quality.

    PubMed

    Brand, Christine; Burkhardt, Eva; Schaeffel, Frank; Choi, Jeong Won; Feldkaemper, Marita Pauline

    2005-04-28

    To analyze mRNA expression changes of Egr-1, VIP, and Shh under different light and treatment conditions in mice. The mRNA expression levels of the three genes and additionally the Egr-1 protein expression were compared in form deprived eyes and eyes with normal vision. Moreover, the influence of dark to light and light to dark transitions and of changes in retinal illumination on mRNA levels was investigated. Form deprivation of mice was induced by fitting frosted diffusers over one eye and an attentuation matched neutral density (ND) filter over the other eye. To measure the effects of retinal illumination changes on mRNA expression, animals were bilaterally fitted with different ND filters. Semiquantitative real-time RT-PCR was used to measure the mRNA levels and immunohistochemistry was applied to localize and detect Egr-1 protein. The expression levels of both Egr-1 mRNA and protein were reduced in form deprived eyes compared to their fellow eyes after 30 min and 1 h, respectively. Egr-1 mRNA was strikingly upregulated both after dark to light and light to dark transitions, whereas minor changes in retinal illumination by covering the eyes with neutral density filters did not alter Egr-1 mRNA expression. In mice, the mRNA levels of VIP and Shh were not affected by form deprivation, but they were found to be regulated depending on the time of day. Both Egr-1 mRNA and protein expression levels were strongly regulated by light, especially by transitions between light and darkness. Image contrast may exert an additional influence on mRNA and protein expression of Egr-1, particularly in the cells in the ganglion cell layer and in bipolar cells.

  14. Cis-Regulatory Variants Affect CHRNA5 mRNA Expression in Populations of African and European Ancestry

    PubMed Central

    Wang, Jen-Chyong; Spiegel, Noah; Bertelsen, Sarah; Le, Nhung; McKenna, Nicholas; Budde, John P.; Harari, Oscar; Kapoor, Manav; Brooks, Andrew; Hancock, Dana; Tischfield, Jay; Foroud, Tatiana; Bierut, Laura J.; Steinbach, Joe Henry; Edenberg, Howard J.; Traynor, Bryan J.; Goate, Alison M.

    2013-01-01

    Variants within the gene cluster encoding α3, α5, and β4 nicotinic receptor subunits are major risk factors for substance dependence. The strongest impact on risk is associated with variation in the CHRNA5 gene, where at least two mechanisms are at work: amino acid variation and altered mRNA expression levels. The risk allele of the non-synonymous variant (rs16969968; D398N) primarily occurs on the haplotype containing the low mRNA expression allele. In populations of European ancestry, there are approximately 50 highly correlated variants in the CHRNA5-CHRNA3-CHRNB4 gene cluster and the adjacent PSMA4 gene region that are associated with CHRNA5 mRNA levels. It is not clear which of these variants contribute to the changes in CHRNA5 transcript level. Because populations of African ancestry have reduced linkage disequilibrium among variants spanning this gene cluster, eQTL mapping in subjects of African ancestry could potentially aid in defining the functional variants that affect CHRNA5 mRNA levels. We performed quantitative allele specific gene expression using frontal cortices derived from 49 subjects of African ancestry and 111 subjects of European ancestry. This method measures allele-specific transcript levels in the same individual, which eliminates other biological variation that occurs when comparing expression levels between different samples. This analysis confirmed that substance dependence associated variants have a direct cis-regulatory effect on CHRNA5 transcript levels in human frontal cortices of African and European ancestry and identified 10 highly correlated variants, located in a 9 kb region, that are potential functional variants modifying CHRNA5 mRNA expression levels. PMID:24303001

  15. Quantification of cytokine mRNA in peripheral blood mononuclear cells using branched DNA (bDNA) technology.

    PubMed

    Shen, L P; Sheridan, P; Cao, W W; Dailey, P J; Salazar-Gonzalez, J F; Breen, E C; Fahey, J L; Urdea, M S; Kolberg, J A

    1998-06-01

    Changes in the patterns of cytokine expression are thought to be of central importance in human infectious and inflammatory diseases. As such, there is a need for precise, reproducible assays for quantification of cytokine mRNA that are amenable to routine use in a clinical setting. In this report, we describe the design and performance of a branched DNA (bDNA) assay for the direct quantification of multiple cytokine mRNA levels in peripheral blood mononuclear cells (PBMCs). Oligonucleotide target probe sets were designed for several human cytokines, including TNFalpha, IL-2, IL-4, IL-6, IL-10, and IFNgamma. The bDNA assay yielded highly reproducible quantification of cytokine mRNAs, exhibited a broad linear dynamic range of over 3-log10, and showed a sensitivity sufficient to measure at least 3000 molecules. The potential clinical utility of the bDNA assay was explored by measuring cytokine mRNA levels in PBMCs from healthy and immunocompromised individuals. Cytokine expression levels in PBMCs from healthy blood donors were found to remain relatively stable over a one-month period of time. Elevated levels of IFNgamma mRNA were detected in PBMCs from HIV-1 seropositive individuals, but no differences in mean levels of TNFalpha or IL-6 mRNA were detected between seropositive and seronegative individuals. By providing a reproducible method for quantification of low abundance transcripts in clinical specimens, the bDNA assay may be useful for studies addressing the role of cytokine expression in disease.

  16. Regulation of cytoplasmic mRNA decay

    PubMed Central

    Schoenberg, Daniel R.; Maquat, Lynne E.

    2012-01-01

    Discoveries made over the past 20 years highlight the importance of mRNA decay as a means to modulate gene expression and thereby protein production. Up until recently, studies focused largely on identifying cis-acting sequences that serve as mRNA stability or instability elements, the proteins that bind these elements, how the process of translation influences mRNA decay, and the ribonucleases that catalyze decay. Now, current studies have begun to elucidate how the decay process is regulated. This review examines our current understanding of how mammalian-cell mRNA decay is controlled by different signaling pathways and lays out a framework for future research. PMID:22392217

  17. Novel RNA-binding activity of NQO1 promotes SERPINA1 mRNA translation.

    PubMed

    Di Francesco, Andrea; Di Germanio, Clara; Panda, Amaresh C; Huynh, Phu; Peaden, Robert; Navas-Enamorado, Ignacio; Bastian, Paul; Lehrmann, Elin; Diaz-Ruiz, Alberto; Ross, David; Siegel, David; Martindale, Jennifer L; Bernier, Michel; Gorospe, Myriam; Abdelmohsen, Kotb; de Cabo, Rafael

    2016-10-01

    NAD(P)H: quinone oxidoreductase (NQO1) is essential for cell defense against reactive oxidative species, cancer, and metabolic stress. Recently, NQO1 was found in ribonucleoprotein (RNP) complexes, but NQO1-interacting mRNAs and the functional impact of such interactions are not known. Here, we used ribonucleoprotein immunoprecipitation (RIP) and microarray analysis to identify comprehensively the subset of NQO1 target mRNAs in human hepatoma HepG2 cells. One of its main targets, SERPINA1 mRNA, encodes the serine protease inhibitor α-1-antitrypsin, A1AT, which is associated with disorders including obesity-related metabolic inflammation, chronic obstructive pulmonary disease (COPD), liver cirrhosis and hepatocellular carcinoma. Biotin pulldown analysis indicated that NQO1 can bind the 3' untranslated region (UTR) and the coding region (CR) of SERPINA1 mRNA. NQO1 did not affect SERPINA1 mRNA levels; instead, it enhanced the translation of SERPINA1 mRNA, as NQO1 silencing decreased the size of polysomes forming on SERPINA1 mRNA and lowered the abundance of A1AT. Luciferase reporter analysis further indicated that NQO1 regulates SERPINA1 mRNA translation through the SERPINA1 3'UTR. Accordingly, NQO1-KO mice had reduced hepatic and serum levels of A1AT and increased activity of neutrophil elastase (NE), one of the main targets of A1AT. We propose that this novel mechanism of action of NQO1 as an RNA-binding protein may help to explain its pleiotropic biological effects. Published by Elsevier Inc.

  18. Low-level laser therapy (LLLT) attenuates RhoA mRNA expression in the rat bronchi smooth muscle exposed to tumor necrosis factor-alpha.

    PubMed

    de Lima, Flávia Mafra; Bjordal, Jan M; Albertini, Regiane; Santos, Fábio V; Aimbire, Flavio

    2010-09-01

    Low-level laser therapy (LLLT) has been found to produce anti-inflammatory effects in a variety of disorders. Bronchial smooth muscle (BSM) hyperreactivity is associated with increased Ca+2 sensitivity and increased RhoA mRNA expression. In the current study, we investigated if LLLT could reduce BSM contraction force and RhoA mRNA expression in tumor necrosis factor-alpha (TNF-alpha)-induced BSM hyperreactivity. In the study, 112 male Wistar rats were divided randomly into 16 groups, and BSM was harvested and suspended in TNF-alpha baths for 6 and 24 h, respectively. Irradiation with LLLT was performed with a wavelength of 660 nm for 42 s with a dose of 1.3 J/cm2. This LLLT dose was administered once in the 6-h group and twice in the 24-h group. LLLT significantly decreased contraction force in BSM at 6 h (TNF-alpha + LLLT: 11.65+/-1.10 g/100 mg of tissue) (F=3115) and at 24 h (TNF-alpha+ LLLT: 14.15+/-1.1 g/100 mg of tissue) (F=3245, p<0.05) after TNF-alpha, respectively, when compared to vehicle-bathed groups (control). LLLT also significantly decreased the expression of RhoA mRNA in BSM segments at 6 h (1.22+/-0.20) (F=2820, p<0.05) and 24 h (2.13+/-0.20) (F=3324, p<0.05) when compared to BSM segments incubated with TNF-alpha without LLLT irradiation. We conclude that LLLT administered with this protocol, reduces RhoA mRNA expression and BSM contraction force in TNF-alpha-induced BSM hyperreactivity.

  19. Differential mRNA expression of neuroinflammatory modulators in the spinal cord and thalamus of type 2 diabetic monkeys.

    PubMed

    Ding, Huiping; Kiguchi, Norikazu; Kishioka, Shiroh; Ma, Tao; Peters, Christopher M; Ko, Mei-Chuan

    2018-05-11

    Given that diabetes-associated complications are closely associated with neuroinflammation, it is imperative to study potential changes in neuroinflammatory modulators in the central nervous system of diabetic primates. The mRNA levels of pro- and anti-inflammatory cytokines, toll-like receptors (TLRs), growth factors, and cannabinoid receptors were compared in the spinal dorsal horn (SDH) and thalamus of naturally occurring type 2 diabetic monkeys and an age-matched control group using reverse transcription and quantitative real-time polymerase chain reaction. In the SDH of diabetic monkeys, mRNA levels of proinflammatory cytokines (i.e. interleukin [IL]-1β and tumor necrosis factor [TNF] α), TLR1, and TLR2 were increased, whereas mRNA levels of IL-10, an anti-inflammatory cytokine, were decreased. No changes were observed in the mRNA levels of growth factors and cannabinoid receptors. In line with the mRNA data, TNFα immunoreactivity was significantly increased in diabetic monkeys. Moreover, mRNA expression levels of IL-1β, TNFα, TLR1, and TLR2 in the SDH were positively correlated with plasma glucose concentrations in all monkeys. Several ligands and receptors involved in neuroinflammation are simultaneously dysregulated in the spinal cord of diabetic monkeys. This primate disease model will facilitate the design of novel treatment approaches to ameliorate neuroinflammation-driven adverse effects in diabetic patients. © 2018 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and John Wiley & Sons Australia, Ltd.

  20. Serum amyloid A protein in amyloidosis, rheumatic, and neoplastic diseases

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Benson, M.D.; Cohen, A.S.

    1979-01-01

    Serum levels of amyloid protein A (SAA) have been shown to be elevated in different types of amyloidosis and in rheumatic diseases by radioimmunoassay using 125 iodine labeled AA and anti-AA. SAA levels were elevated in both primary and secondary amyloidosis, but there were highly significant differences between these levels. In heredofamilial amyloid, SAA levels were within normal limits. While the mean SAA level was elevated in persons over 70 years, the fact that some persons in this age group had normal levels suggested that marked elevation after age 70 may be due to occult inflammatory or neoplastic disease. Highmore » SAA levels in patients with rheumatoid arthritis correlated, in most cases, with physician evaluation of disease activity and Westergren ESR. SAA levels in patients with systemic lupus erythematosus were lower than those in patients with rheumatoid arthritis, and most patients with degenerative joint disease had normal levels. Very high levels of SAA were found in patients with neoplastic diseases. Patients with carcinoma of the lung and bowel had much higher levels than patients with carcinoma of the breast. Determination of SAA levels may be of value in evaluating different forms of systemic amyloidosis, assessing the activity of rheumatic disease, and screening for occult inflammatory or neoplastic disease.« less

  1. Expression and regulation of Icer mRNA in the Syrian hamster pineal gland.

    PubMed

    Diaz, Elena; Garidou, Marie-Laure; Dardente, Hugues; Salingre, Anthony; Pévet, Paul; Simonneaux, Valérie

    2003-04-10

    Inducible-cAMP early repressor (ICER) is a potent inhibitor of CRE (cAMP-related element)-driven gene transcription. In the rat pineal gland, it has been proposed to be part of the mechanisms involved in the shutting down of the transcription of the gene coding for arylalkylamine N-acetyltransferase (AA-NAT, the melatonin rhythm-generating enzyme). In this study, we report that ICER is expressed in the pineal gland of the photoperiodic rodent Syrian hamster although with some difference compared to the rat. In the Syrian hamster pineal, Icer mRNA levels, low at daytime, displayed a 20-fold increase during the night. Nighttime administration of a beta-adrenergic antagonist, propranolol, significantly reduced Icer mRNA levels although daytime administration of a beta-adrenergic agonist, isoproterenol, was unable to raise the low amount of Icer mRNA. These observations indicate that Icer mRNA expression is induced by the clock-driven norepinephrine release and further suggest that this stimulation is restricted to nighttime, as already observed for Aa-nat gene transcription. Furthermore, we found that the daily profile of Icer mRNA displayed photoperiodic variation with a lengthening of the nocturnal peak in short versus long photoperiod. These data indicate that ICER may be involved in both daily and seasonal regulation of melatonin synthesis in the Syrian hamster.

  2. Detection of MMP-9 and TIMP-3 mRNA expression in the villi of patients undergoing early spontaneous abortion: A report of 30 cases.

    PubMed

    Jiang, Guangli; Qi, Yuxia

    2015-05-01

    The aim of the present study was to investigate the correlation of matrix metalloproteinase (MMP)-9 and tissue inhibitor of matrix metalloproteinase inhibitor (TIMP)-3 expression with spontaneous abortion (SA) during early pregnancy. The villus tissues of 30 SA cases and 20 requested abortion cases were collected during surgery and constituted the SA and normal abortion (NA) groups, respectively. The total villous RNA was extracted and the expression levels of MMP -9 and TIMP-3 mRNA were detected by reverse transcription-polymerase chain reaction (RT-PCR) assay to calculate the MMP-9/TIMP-3 mRNA ratio. The MMP-9 mRNA expression level and MMP-9/TIMP-3 mRNA ratio of the SA group were significantly higher than those of the NA group (P<0.01), while the TIMP-3 mRNA levels of the two groups were similar (P>0.05). The MMP-9 mRNA expression level of the SA group was higher than that of the NA group; thus, the MMP-9/TIMP-3 mRNA ratio was higher. These results suggest that the expression level of MMP-9 mRNA and the MMP-9/TIMP-3 mRNA ratio are associated with SA.

  3. The cellular growth rate controls overall mRNA turnover, and modulates either transcription or degradation rates of particular gene regulons.

    PubMed

    García-Martínez, José; Delgado-Ramos, Lidia; Ayala, Guillermo; Pelechano, Vicent; Medina, Daniel A; Carrasco, Fany; González, Ramón; Andrés-León, Eduardo; Steinmetz, Lars; Warringer, Jonas; Chávez, Sebastián; Pérez-Ortín, José E

    2016-05-05

    We analyzed 80 different genomic experiments, and found a positive correlation between both RNA polymerase II transcription and mRNA degradation with growth rates in yeast. Thus, in spite of the marked variation in mRNA turnover, the total mRNA concentration remained approximately constant. Some genes, however, regulated their mRNA concentration by uncoupling mRNA stability from the transcription rate. Ribosome-related genes modulated their transcription rates to increase mRNA levels under fast growth. In contrast, mitochondria-related and stress-induced genes lowered mRNA levels by reducing mRNA stability or the transcription rate, respectively. We also detected these regulations within the heterogeneity of a wild-type cell population growing in optimal conditions. The transcriptomic analysis of sorted microcolonies confirmed that the growth rate dictates alternative expression programs by modulating transcription and mRNA decay.The regulation of overall mRNA turnover keeps a constant ratio between mRNA decay and the dilution of [mRNA] caused by cellular growth. This regulation minimizes the indiscriminate transmission of mRNAs from mother to daughter cells, and favors the response capacity of the latter to physiological signals and environmental changes. We also conclude that, by uncoupling mRNA synthesis from decay, cells control the mRNA abundance of those gene regulons that characterize fast and slow growth. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  4. Alterations of type IV collagen alpha chains in patients with chronic acquired glomerulopathies: mRNA levels, protein expression and urinary loss.

    PubMed

    Sanna-Cherchi, Simone; Carnevali, Maria Luisa; Martorana, Davide; Cravedi, Paolo; Maggiore, Umberto; Alinovi, Rossella; Bovino, Achiropita; Mattei, Silvia; Orlandini, Guido; Gatti, Rita; Savi, Mario; Sado, Yoshikazu; Neri, Tauro M; Allegri, Landino

    2007-01-01

    Type IV collagen is a major structural component of the normal kidney glomerulus. However, its role in chronic acquired glomerulopathies has been only partially elucidated. Urinary levels of col(IV)alpha1, col(IV)alpha3 and col(IV)alpha5 collagen chains were analyzed in 107 patients with chronic acquired glomerulopathies. In a subgroup of 33 patients, tissue mRNA levels, protein expression and urinary excretion were evaluated for all col(IV)alpha chains, from col(IV)alpha1 to col(IV)alpha5. The renal specimens were examined to get a semiquantitative score of the acute and chronic activity of the histological lesions. Urines obtained from 13 healthy subjects and 10 normal renal tissue samples were used as controls. Urinary levels of col(IV)alpha1, col(IV)alpha3, col(IV)alpha5 chains were significantly higher in patients than in controls [p < 0.01 for all], while only col(IV)alpha1 and col(IV)alpha3 urinary excretion correlated with the degree of chronic histological damage [col(IV)alpha1 R = 0.44, p < 0.001; col(IV)alpha3: R = 0.47, p < 0.001]. Compared with controls, patients showed a renal expression of mRNA for col(IV)alpha5 chain significantly higher [p = 0.001], while having a significantly lower protein expression of col(IV)alpha3, col(IV)alpha4 and col(IV)alpha5 chains [p < 0.01 for all]. Patients with chronic acquired glomerulopathies show important alterations in the col(IV)alpha chain network mimicking some molecular features of the X-linked Alport's syndrome. Further studies are needed to show whether urinary levels of the col(IV)alpha chains may be used as markers for monitoring renal injury. Copyright 2007 S. Karger AG, Basel.

  5. Role of miRNAs and alternative mRNA 3'-end cleavage and polyadenylation of their mRNA targets in cardiomyocyte hypertrophy.

    PubMed

    Soetanto, R; Hynes, C J; Patel, H R; Humphreys, D T; Evers, M; Duan, G; Parker, B J; Archer, S K; Clancy, J L; Graham, R M; Beilharz, T H; Smith, N J; Preiss, T

    2016-05-01

    miRNAs play critical roles in heart disease. In addition to differential miRNA expression, miRNA-mediated control is also affected by variable miRNA processing or alternative 3'-end cleavage and polyadenylation (APA) of their mRNA targets. To what extent these phenomena play a role in the heart remains unclear. We sought to explore miRNA processing and mRNA APA in cardiomyocytes, and whether these change during cardiac hypertrophy. Thoracic aortic constriction (TAC) was performed to induce hypertrophy in C57BL/6J mice. RNA extracted from cardiomyocytes of sham-treated, pre-hypertrophic (2 days post-TAC), and hypertrophic (7 days post-TAC) mice was subjected to small RNA- and poly(A)-test sequencing (PAT-Seq). Differential expression analysis matched expectations; nevertheless we identified ~400 mRNAs and hundreds of noncoding RNA loci as altered with hypertrophy for the first time. Although multiple processing variants were observed for many miRNAs, there was little change in their relative proportions during hypertrophy. PAT-Seq mapped ~48,000 mRNA 3'-ends, identifying novel 3' untranslated regions (3'UTRs) for over 7000 genes. Importantly, hypertrophy was associated with marked changes in APA with a net shift from distal to more proximal mRNA 3'-ends, which is predicted to decrease overall miRNA repression strength. We independently validated several examples of 3'UTR proportion change and showed that alternative 3'UTRs associate with differences in mRNA translation. Our work suggests that APA contributes to altered gene expression with the development of cardiomyocyte hypertrophy and provides a rich resource for a systems-level understanding of miRNA-mediated regulation in physiological and pathological states of the heart. Copyright © 2016 Elsevier B.V. All rights reserved.

  6. Self-amplifying mRNA vaccines.

    PubMed

    Brito, Luis A; Kommareddy, Sushma; Maione, Domenico; Uematsu, Yasushi; Giovani, Cinzia; Berlanda Scorza, Francesco; Otten, Gillis R; Yu, Dong; Mandl, Christian W; Mason, Peter W; Dormitzer, Philip R; Ulmer, Jeffrey B; Geall, Andrew J

    2015-01-01

    This chapter provides a brief introduction to nucleic acid-based vaccines and recent research in developing self-amplifying mRNA vaccines. These vaccines promise the flexibility of plasmid DNA vaccines with enhanced immunogenicity and safety. The key to realizing the full potential of these vaccines is efficient delivery of nucleic acid to the cytoplasm of a cell, where it can amplify and express the encoded antigenic protein. The hydrophilicity and strong net negative charge of RNA impedes cellular uptake. To overcome this limitation, electrostatic complexation with cationic lipids or polymers and physical delivery using electroporation or ballistic particles to improve cellular uptake has been evaluated. This chapter highlights the rapid progress made in using nonviral delivery systems for RNA-based vaccines. Initial preclinical testing of self-amplifying mRNA vaccines has shown nonviral delivery to be capable of producing potent and robust innate and adaptive immune responses in small animals and nonhuman primates. Historically, the prospect of developing mRNA vaccines was uncertain due to concerns of mRNA instability and the feasibility of large-scale manufacturing. Today, these issues are no longer perceived as barriers in the widespread implementation of the technology. Currently, nonamplifying mRNA vaccines are under investigation in human clinical trials and can be produced at a sufficient quantity and quality to meet regulatory requirements. If the encouraging preclinical data with self-amplifying mRNA vaccines are matched by equivalently positive immunogenicity, potency, and tolerability in human trials, this platform could establish nucleic acid vaccines as a versatile new tool for human immunization. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Conditioned Fear Inhibits c-fos mRNA Expression in the Central Extended Amygdala

    PubMed Central

    Day, Heidi E.W.; Kryskow, Elisa M.; Nyhuis, Tara J.; Herlihy, Lauren; Campeau, Serge

    2008-01-01

    We have shown previously that unconditioned stressors inhibit neurons of the lateral/capsular division of the central nucleus of the amygdala (CEAl/c) and oval division of the bed nucleus of the stria terminalis (BSTov), which form part of the central extended amygdala. The current study investigated whether conditioned fear inhibits c-fos mRNA expression in these regions. Male rats were trained either to associate a visual stimulus (light) with footshock or were exposed to the light alone. After training, animals were replaced in the apparatus, and 2 hours later injected remotely, via a catheter, with amphetamine (2 mg/kg i.p.), to induce c-fos mRNA and allow inhibition of expression to be measured. The rats were then presented with 15 visual stimuli over a 30 minute period. As expected, fear conditioned animals that were not injected with amphetamine, had extremely low levels of c-fos mRNA in the central extended amygdala. In contrast, animals that were trained with the light alone (no fear conditioning) and were injected with amphetamine had high levels of c-fos mRNA in the CEAl/c and BSTov. Animals that underwent fear-conditioning, and were re-exposed to the conditioned stimulus after amphetamine injection had significantly reduced levels of c-fos mRNA in both the BSTov and CEAl/c, compared to the non-conditioned animals. These data suggest that conditioned fear can inhibit neurons of the central extended amygdala. Because these neurons are GABAergic, and project to the medial CEA (an amygdaloid output region), this may be a novel mechanism whereby conditioned fear potentiates amygdaloid output. PMID:18634767

  8. Effect of raclopride on dopamine D2 receptor mRNA expression in rat brain.

    PubMed

    Kopp, J; Lindefors, N; Brené, S; Hall, H; Persson, H; Sedvall, G

    1992-01-01

    Prolonged treatment with dopamine D2 receptor antagonists is known to elevate the density of dopamine D2 receptor binding sites in caudate-putamen and nucleus accumbens in rat and human brain. In this study we used the dopamine D2 receptor antagonist raclopride (3 mumol/kg, s.c.) to determine if a single injection or daily administration of this drug for up to 18 days changed the expression of dopamine D2 receptor mRNA in rat caudate-putamen and accumbens as measured by in situ hybridization. A single injection of raclopride did not significantly change the numerical density of dopamine D2 receptor mRNA-expressing neurons in any of the regions examined. A daily administration of raclopride for 18 days resulted in a 31% increase in the number of cells expressing detectable amounts of dopamine D2 receptor mRNA in dorsolateral caudate-putamen and in a 20% increase in the area of silver grains over individual hybridization-positive neurons in this brain region measured on emulsion-dipped slides. The region-specific increase in the D2 receptor mRNA level in dorsolateral caudate-putamen was confirmed by measurement of the hybridization signal on X-ray film autoradiograms. The levels of D2 receptor mRNA remained unchanged in medial caudate-putamen and accumbens after 18 days' treatment. The region-selective increase in dopamine D2 receptor mRNA expression in dorsolateral caudate-putamen indicates a differential regulation of dopamine D2 receptor mRNA expression in a subpopulation of caudate-putamen neurons by this neuroleptic. We suggest that the increase in dopamine D2 receptor density in caudate-putamen known to follow prolonged dopamine D2 receptor blockade to some extent is regulated at the level of gene expression.

  9. Sex differences in spatiotemporal expression of AR, ERα, and ERβ mRNA in the perinatal mouse brain.

    PubMed

    Mogi, Kazutaka; Takanashi, Haruka; Nagasawa, Miho; Kikusui, Takefumi

    2015-01-01

    It has been shown that every masculinized function might be organized by a particular contribution of androgens vs. estrogens in a critical time window. Here, we aimed to investigate the sex differences in brain testosterone levels and in the spatiotemporal dynamics of steroid receptor mRNA expression in perinatal mice, by using enzyme immunoassay and real-time PCR, respectively. We found that testosterone levels in the forebrain transiently increased around birth in male mice. During the perinatal period, levels of androgen receptor mRNA in the hypothalamus (hypo) and prefrontal cortex (PFC) were higher in male mice than in female mice. Estrogen receptor α (ERα) mRNA levels in the hypo and hippocampus were higher in male mice than in female mice before birth. In contrast, ERβ mRNA expression in the PFC was higher in female mice immediately after birth. These spatiotemporal sex differences in steroid receptor expression might contribute to organizing sex differences of not only reproductive function, but also anxiety, stress responses, and cognition in mice. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  10. Neurotrophin-3 mRNA a putative target of miR21 following status epilepticus.

    PubMed

    Risbud, Rashmi M; Lee, Carolyn; Porter, Brenda E

    2011-11-18

    Status epilepticus induces a cascade of protein expression changes contributing to the subsequent development of epilepsy. By identifying the cascade of molecular changes that contribute to the development of epilepsy we hope to be able to design therapeutics for preventing epilepsy. MicroRNAs influence gene expression by altering mRNA stability and/or translation and have been implicated in the pathology of multiple diseases. MiR21 and its co-transcript miR21, microRNAs produced from either the 5' or 3' ends of the same precursor RNA strand, are increased in the hippocampus following status epilepticus. We have identified a miR21 binding site, in the 3' UTR of neurotrophin-3 that inhibits translation. Neurotrophin-3 mRNA levels decrease in the hippocampus following SE concurrent with the increase in miR21. MiR21 levels in cultured hippocampal neurons inversely correlate with neurotrophin-3 mRNA levels. Treatment of hippocampal neuronal cultures with excess K(+)Cl(-), a depolarizing agent mimicking the episode of status epilepticus, also results in an increase in miR21 and a decrease in neurotrophin-3 mRNA. MiR21 is a candidate for regulating neurotrophin-3 signaling in the hippocampus following status epilepticus. Copyright © 2011 Elsevier B.V. All rights reserved.

  11. The mTOR kinase inhibitor rapamycin decreases iNOS mRNA stability in astrocytes

    PubMed Central

    2011-01-01

    Background Reactive astrocytes are capable of producing a variety of pro-inflammatory mediators and potentially neurotoxic compounds, including nitric oxide (NO). High amounts of NO are synthesized following up-regulation of inducible NO synthase (iNOS). The expression of iNOS is tightly regulated by complex molecular mechanisms, involving both transcriptional and post-transcriptional processes. The mammalian target of rapamycin (mTOR) kinase modulates the activity of some proteins directly involved in post-transcriptional processes of mRNA degradation. mTOR is a serine-threonine kinase that plays an evolutionarily conserved role in the regulation of cell growth, proliferation, survival, and metabolism. It is also a key regulator of intracellular processes in glial cells. However, with respect to iNOS expression, both stimulatory and inhibitory actions involving the mTOR pathway have been described. In this study the effects of mTOR inhibition on iNOS regulation were evaluated in astrocytes. Methods Primary cultures of rat cortical astrocytes were activated with different proinflammatory stimuli, namely a mixture of cytokines (TNFα, IFNγ, and IL-1β) or by LPS plus IFNγ. Rapamycin was used at nM concentrations to block mTOR activity and under these conditions we measured its effects on the iNOS promoter, mRNA and protein levels. Functional experiments to evaluate iNOS activity were also included. Results In this experimental paradigm mTOR activation did not significantly affect astrocyte iNOS activity, but mTOR pathway was involved in the regulation of iNOS expression. Rapamycin did not display any significant effects under basal conditions, on either iNOS activity or its expression. However, the drug significantly increased iNOS mRNA levels after 4 h incubation in presence of pro-inflammatory stimuli. This stimulatory effect was transient, since no differences in either iNOS mRNA or protein levels were detected after 24 h. Interestingly, reduced levels of i

  12. Exaptive origins of regulated mRNA decay in eukaryotes.

    PubMed

    Hamid, Fursham M; Makeyev, Eugene V

    2016-09-01

    Eukaryotic gene expression is extensively controlled at the level of mRNA stability and the mechanisms underlying this regulation are markedly different from their archaeal and bacterial counterparts. We propose that two such mechanisms, nonsense-mediated decay (NMD) and motif-specific transcript destabilization by CCCH-type zinc finger RNA-binding proteins, originated as a part of cellular defense against RNA pathogens. These branches of the mRNA turnover pathway might have been used by primeval eukaryotes alongside RNA interference to distinguish their own messages from those of RNA viruses and retrotransposable elements. We further hypothesize that the subsequent advent of "professional" innate and adaptive immunity systems allowed NMD and the motif-triggered mechanisms to be efficiently repurposed for regulation of endogenous cellular transcripts. This scenario explains the rapid emergence of archetypical mRNA destabilization pathways in eukaryotes and argues that other aspects of post-transcriptional gene regulation in this lineage might have been derived through a similar exaptation route. © 2016 The Authors BioEssays Published by WILEY Periodicals, Inc.

  13. Effect of Alpha-Hederin, the active constituent of Nigella sativa, on miRNA-126, IL-13 mRNA levels and inflammation of lungs in ovalbumin-sensitized male rats

    PubMed Central

    Fallahi, Maryam; Keyhanmanesh, Rana; Khamaneh, Amir Mahdi; Ebrahimi Saadatlou, Mohammad Ali; Saadat, Saeideh; Ebrahimi, Hadi

    2016-01-01

    Objective: In previous studies the therapeutic effects of Nigella sativa have been demonstrated on asthmatic animals. In the present study, the preventive effect of single dose of alpha-hederin, its active constituent, has been evaluated on lung inflammation and some inflammatory mediators in lungs of ovalbumin sensitized rat in order to elicit its mechanism. Materials and Methods: Forty rats were randomly grouped in 4 groups; control (C), sensitized (S), sensitized pretreated groups with thymoquinone (3 mg/kg i.p., S+TQ) and alpha-hederin (0.02 mg/kg i.p., S+AH). Levels of IL-13 mRNA and miRNA-126 in lung tissue and its pathological changes in each group were assessed. Results: Elevated levels of miRNA-126, IL-13 mRNA and pathological changes were observed in the sensitized group compared to the control group (p<0.001 to p<0.05). All of these factors were significantly reduced in S+TQ and S+AH groups in comparison to S group (p<0.001 to p<0.05). Although alpha-hederin decreased the levels of miRNA-126, IL-13 mRNA and pathological changes in comparison with thymoquinone, the results were statistically not significant. Conclusion: The results suggested that alpha-hederin had preventive effect on sensitized rats like thymoquinone. It may intervene in miRNA-126 expression, which consequently could interfere with IL-13 secretion pathway leading to a reduction in inflammatory responses. PMID:27247924

  14. Cloning of a long HIV-1 readthrough transcript and detection of an increased level of early growth response protein-1 (Egr-1) mRNA in chronically infected U937 cells.

    PubMed

    Dron, M; Hameau, L; Benboudjema, L; Guymarho, J; Cajean-Feroldi, C; Rizza, P; Godard, C; Jasmin, C; Tovey, M G; Lang, M C

    1999-01-01

    To identify the pathways involved in HIV-1 modification of cellular gene expression, chronically infected U937 cells were screened by mRNA differential display. A chimeric transcript consisting of the 3' end of the LTR of a HIV-1 provirus, followed by 3.7 kb of cellular RNA was identified suggesting that long readthrough transcription might be one of the mechanisms by which gene expression could be modified in individual infected cells. Such a phenomenon may also be the first step towards the potential transduction of cellular sequences. Furthermore, the mRNA encoding for the transcription factor Egr-1 was detected as an over-represented transcript in infected cells. Northern blot analysis confirmed the increase of Egr-1 mRNA content in both HIV-1 infected promonocytic U937 cells and T cell lines such as Jurkat and CEM. Interestingly a similar increase of Egr-1 mRNA has previously been reported to occur in HTLV-1 and HTLV-2 infected T cell lines. Despite the consistent increase in the level of Egr-1 mRNA, the amount of the encoded protein did not appear to be modified in HIV-1 infected cells, suggesting an increased turn over of the protein in chronically infected cells.

  15. Patterns of mRNA and protein expression during minus-lens compensation and recovery in tree shrew sclera.

    PubMed

    Gao, Hong; Frost, Michael R; Siegwart, John T; Norton, Thomas T

    2011-04-12

    To increase our understanding of the mechanisms that remodel the sclera during the development of lens-induced myopia, when the sclera responds to putative "go" signals of retinal origin, and during recovery from lens-induced myopia, when the sclera responds to retinally-derived "stop" signals. Seven groups of tree shrews were used to examine mRNA levels during minus lens compensation and recovery. Starting 24 days after eye opening (days of visual experience [VE]) lens compensation animals wore a monocular -5D lens for 1, 4, or 11 days. Recovery animals wore the -5D lens for 11 days, which was then removed for 1 or 4 days. Normal animals were examined at 24 and 38 days of VE. All groups contained 8 animals. Scleral mRNA levels were examined in the treated and contralateral control eyes with quantitative real-time polymerase chain reaction (qPCR) for 27 genes divided into four categories: 1) signaling molecules, 2) matricellular proteins, 3) metalloproteinases (MPs) and tissue inhibitors of metalloproteinases (TIMPs), and 4) cell adhesion and other proteins. Four groups (n=5 per group) were used to examine protein levels. One group wore a -5D lens for 4 days. A second group recovered for 4 days after 11 days of -5D lens treatment. Two groups were used to examine age-matched normal protein levels at 28 and 39 days of VE. The levels of six scleral proteins that showed differential mRNA expression were examined with quantitative western blots. Nineteen of the genes showed differential (treated eye versus control eye) expression of mRNA levels in at least one group of animals. Which genes showed differential expression differed after 1 and 4 days of compensation and after 1 or 4 days of recovery. The mRNA level for one gene, a disintegrin and metalloproteinase with thrombospondin motifs 1 (ADAMTS1), was upregulated in the treated eyes after 1 day of compensation. After 4 days, transforming growth factor beta receptor 3 (TGFBR3), transforming growth factor

  16. mRNA export: threading the needle

    PubMed Central

    Gaouar, Ouassila; Germain, Hugo

    2013-01-01

    After mRNA biogenesis, several proteins interact with the messenger to ensure its proper export to the cytoplasm. Some of these proteins will bind RNA early on, at the onset of transcription by RNA polymerase II holoenzyme, while others will join later for downstream processing steps, such as poly-adenylation or splicing, or may direct mRNA ribonucleoprotein particle migration to the nucleopore. We recently discovered that Arabidopsis plant knockout for the protein MOS11 (MODIFIER OF SNC1, 11) partially suppresses autoimmune responses observed in the TNL-type [TIR/NBS/LRR (Toll-interleukin-like receptor/nucleotide-binding site/C-terminal leucine-rich repeat)] R gene gain-of-function variant snc1 (suppressor of npr1-1, constitutive 1). This suppression of resistance to pathogens appears to be caused by a decrease in nuclear mRNA export in mos11-1 snc1 plants. In humans, the putative ortholog of MOS11, CIP29 (29-kDa cytokine-induced protein), interacts with three proteins that are also involved in mRNA export: DDX39 (DEAD-box RNA helicase), TAF15 of the FUS family (FUSED IN SARCOMA), and ALY (ALWAYS EARLY), a protein implicated in mRNA export in mammalian systems. These proteins have received very little attention in plants. Here, we will discuss their particularities and role in mRNA export and biotic stress. PMID:23526740

  17. Clinical values of AFP, GPC3 mRNA in peripheral blood for prediction of hepatocellular carcinoma recurrence following OLT: AFP, GPC3 mRNA for prediction of HCC.

    PubMed

    Wang, Yuliang; Shen, Zhongyang; Zhu, Zhijun; Han, Ruifa; Huai, Mingsheng

    2011-03-01

    Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide. Annually, about 200,000 patients died of HCC in China. Liver transplantation (LT) holds great theoretical appeal in treating HCC. However, the high recurrence rate after transplantation is the most important limiting factor for long-term survival. To assess the value of alpha-fetoprotein (AFP) messenger RNA (mRNA), Glypican-3 (GPC3) mRNA-expressing cells in the peripheral blood (PB) for prediction of HCC recurrence following orthotopic liver transplantation (OLT). 29 patients with HCC who underwent OLT with a minimum clinical follow-up of 12 months were included in this retrospective study. We detected AFP mRNA, GPC3 mRNA-expressing cells in the PB by TaqMan real-time reverse transcriptase-polymerase chain reaction (RT-PCR), pre-, intra- and post-operatively. The early recurrence of patients was evaluated. 8 (28%), 15 (52%), and 9 (31%) patients had AFP mRNA detected pre-, intra-, and post-operatively, respectively. With 12 months of follow-up, HCC recurred in 7 (24%) patients. Univariate analysis revealed that positive pre- and post-operative AFP mRNA, TNM stage as well as vascular invasion were significant predictors for the HCC recurrence. Multivariate analysis revealed that being positive for AFP mRNA pre-operatively remained a significant risk factor for HCC recurrence after OLT. GPC3 mRNA was expressed in all PB samples. There was no significant difference in the expression levels of GPC3 mRNA between the HCC and control groups. There were no significant differences in GPC3 mRNA expression values between those patients with and without tumor recurrence. The pre-operative detection of circulating AFP mRNA-expressing cells could be a useful predictor for HCC recurrence following OLT. GPC3 mRNA-expressing cells in PB seem to have no diagnostic value.

  18. Subchronic cadmium exposure upregulates the mRNA level of genes associated to hepatic lipid metabolism in adult female CD1 mice.

    PubMed

    Zhang, Jun; Wang, Yan; Fu, Lin; Feng, Yu-Jie; Ji, Yan-Li; Wang, Hua; Xu, De-Xiang

    2018-07-01

    Cadmium (Cd) is a persistent environmental and occupational contaminant that accumulates in humans and shows adverse effects on health. Accumulating evidence reveals that environmental Cd exposure is associated with hepatic lipid accumulation and metabolic alterations in adult male mice. However, whether Cd exposure induces hepatic lipid accumulation and metabolic alterations in female mice remains poorly understood. In the present study, we aimed to investigate the effects of Cd exposure on insulin resistance, hepatic lipid accumulation and associated metabolic pathways. Female CD1 mice were administrated with CdCl 2 (10 and 100 mg l -1 ) by drinking water. We found that Cd exposure did not induce obesity, insulin resistance and hepatic lipid accumulation. By contrary, mice in the Cd-100 mg l -1 group presented a significant reduction of the glucose area under the curve during the glucose tolerance test. However, there was a significant elevation in the mRNA level of Fasn and Scd-1, which were critical genes during hepatic fatty acid synthesis. Moreover, hepatic Fabp1 and Fabp4, two genes for hepatic fatty acid uptake were upregulated in Cd-treated mice. Of interest, Lpl, a key gene for hepatic lipoprotein lysis, was also upregulated in Cd-treated mice. Collectively, our results suggest that Cd exposure upregulated mRNA level of genes related to hepatic lipid metabolism although there was no insulin resistance and hepatic lipid accumulation shown in the present study. Copyright © 2018 John Wiley & Sons, Ltd.

  19. Nerve Growth Factor Increases mRNA Levels for the Prion Protein and the β -amyloid Protein Precursor in Developing Hamster Brain

    NASA Astrophysics Data System (ADS)

    Mobley, William C.; Neve, Rachael L.; Prusiner, Stanley B.; McKinley, Michael P.

    1988-12-01

    Deposition of amyloid filaments serves as a pathologic hallmark for some neurodegenerative disorders. The prion protein (PrP) is found in amyloid of animals with scrapie and humans with Creutzfeldt-Jakob disease; the β protein is present in amyloid deposits in Alzheimer disease and Down syndrome patients. These two proteins are derived from precursors that in the brain are expressed primarily in neurons and are membrane bound. We found that gene expression for PrP and the β -protein precursor (β -PP) is regulated in developing hamster brain. Specific brain regions showed distinct patterns of ontogenesis for PrP and β -PP mRNAs. The increases in PrP and β -PP mRNAs in developing basal forebrain coincided with an increase in choline acetyltransferase activity, raising the possibility that these markers might be coordinately controlled in cholinergic neurons and regulated by nerve growth factor (NGF). Injections of NGF into the brains of neonatal hamsters increased both PrP and β -PP mRNA levels. Increased PrP and β -PP mRNA levels induced by NGF were confined to regions that contain NGF-responsive cholinergic neurons and were accompanied by elevations in choline acetyltransferase. It remains to be established whether or not exogenous NGF acts to increase PrP and β -PP gene expression selectively in forebrain cholinergic neurons in the developing hamster and endogenous NGF regulates expression of these genes.

  20. Testosterone Regulates NUCB2 mRNA Expression in Male Mouse Hypothalamus and Pituitary Gland

    PubMed Central

    Seon, Sojeong; Jeon, Daun; Kim, Heejeong; Chung, Yiwa; Choi, Narae; Yang, Hyunwon

    2017-01-01

    ABSTRACT Nesfatin-1/NUCB2 is known to take part in the control of the appetite and energy metabolism. Recently, many reports have shown nesfatin-1/NUCB2 expression and function in various organs. We previously demonstrated that nesfatin-1/NUCB2 expression level is higher in the pituitary gland compared to other organs and its expression is regulated by 17β-estradiol and progesterone secreted from the ovary. However, currently no data exist on the expression of nesfatin-1/NUCB2 and its regulation mechanism in the pituitary of male mouse. Therefore, we examined whether nesfatin-1/NUCB2 is expressed in the male mouse pituitary and if its expression is regulated by testosterone. As a result of PCR and western blotting, we found that a large amount of nesfatin-1/NUCB2 was expressed in the pituitary and hypothalamus. The NUCB2 mRNA expression level in the pituitary was decreased after castration, but not in the hypothalamus. In addition, its mRNA expression level in the pituitary was increased after testosterone treatment in the castrated mice, whereas, the expression level in the hypothalamus was significantly decreased after the treatment with testosterone. The in vitro experiment to elucidate the direct effect of testosterone on NUCB2 mRNA expression showed that NUCB2 mRNA expression was significantly decreased with testosterone in cultured hypothalamus tissue, but increased with testosterone in cultured pituitary gland. The present study demonstrated that nesfatin-1/NUCB2 was highly expressed in the male mouse pituitary and was regulated by testosterone. This data suggests that reproductive-endocrine regulation through hypothalamus-pituitary-testis axis may contribute to NUCB2 mRNA expression in the mouse hypothalamus and pituitary gland. PMID:28484746

  1. Arsenic Induces Polyadenylation of Canonical Histone mRNA by Down-regulating Stem-Loop-binding Protein Gene Expression*

    PubMed Central

    Brocato, Jason; Fang, Lei; Chervona, Yana; Chen, Danqi; Kiok, Kathrin; Sun, Hong; Tseng, Hsiang-Chi; Xu, Dazhong; Shamy, Magdy; Jin, Chunyuan; Costa, Max

    2014-01-01

    The replication-dependent histone genes are the only metazoan genes whose messenger RNA (mRNA) does not terminate with a poly(A) tail at the 3′-end. Instead, the histone mRNAs display a stem-loop structure at their 3′-end. Stem-loop-binding protein (SLBP) binds the stem-loop and regulates canonical histone mRNA metabolism. Here we report that exposure to arsenic, a carcinogenic metal, decreased cellular levels of SLBP by inducing its proteasomal degradation and inhibiting SLBP transcription via epigenetic mechanisms. Notably, arsenic exposure dramatically increased polyadenylation of canonical histone H3.1 mRNA possibly through down-regulation of SLBP expression. The polyadenylated H3.1 mRNA induced by arsenic was not susceptible to normal degradation that occurs at the end of S phase, resulting in continued presence into mitosis, increased total H3.1 mRNA, and increased H3 protein levels. Excess expression of canonical histones have been shown to increase sensitivity to DNA damage as well as increase the frequency of missing chromosomes and induce genomic instability. Thus, polyadenylation of canonical histone mRNA following arsenic exposure may contribute to arsenic-induced carcinogenesis. PMID:25266719

  2. Protein structure and the sequential structure of mRNA: alpha-helix and beta-sheet signals at the nucleotide level.

    PubMed

    Brunak, S; Engelbrecht, J

    1996-06-01

    A direct comparison of experimentally determined protein structures and their corresponding protein coding mRNA sequences has been performed. We examine whether real world data support the hypothesis that clusters of rare codons correlate with the location of structural units in the resulting protein. The degeneracy of the genetic code allows for a biased selection of codons which may control the translational rate of the ribosome, and may thus in vivo have a catalyzing effect on the folding of the polypeptide chain. A complete search for GenBank nucleotide sequences coding for structural entries in the Brookhaven Protein Data Bank produced 719 protein chains with matching mRNA sequence, amino acid sequence, and secondary structure assignment. By neural network analysis, we found strong signals in mRNA sequence regions surrounding helices and sheets. These signals do not originate from the clustering of rare codons, but from the similarity of codons coding for very abundant amino acid residues at the N- and C-termini of helices and sheets. No correlation between the positioning of rare codons and the location of structural units was found. The mRNA signals were also compared with conserved nucleotide features of 16S-like ribosomal RNA sequences and related to mechanisms for maintaining the correct reading frame by the ribosome.

  3. CELFish ways to modulate mRNA decay

    PubMed Central

    St. Louis, Irina Vlasova; Dickson, Alexa M.; Bohjanen, Paul R.; Wilusz, Carol J.

    2013-01-01

    The CELF family of RNA-binding proteins regulates many steps of mRNA metabolism. Although their best characterized function is in pre-mRNA splice site choice, CELF family members are also powerful modulators of mRNA decay. In this review we focus on the different modes of regulation that CELF proteins employ to mediate mRNA decay by binding to GU-rich elements. After starting with an overview of the importance of CELF proteins during development and disease pathogenesis, we then review the mRNA networks and cellular pathways these proteins regulate and the mechanisms by which they influence mRNA decay. Finally, we discuss how CELF protein activity is modulated during development and in response to cellular signals. We conclude by highlighting the priorities for new experiments in this field. PMID:23328451

  4. An mRNA decapping mutant deficient in P body assembly limits mRNA stabilization in response to osmotic stress.

    PubMed

    Huch, Susanne; Nissan, Tracy

    2017-03-14

    Yeast is exposed to changing environmental conditions and must adapt its genetic program to provide a homeostatic intracellular environment. An important stress for yeast in the wild is high osmolarity. A key response to this stress is increased mRNA stability primarily by the inhibition of deadenylation. We previously demonstrated that mutations in decapping activators (edc3∆ lsm4∆C), which result in defects in P body assembly, can destabilize mRNA under unstressed conditions. We wished to examine whether mRNA would be destabilized in the edc3∆ lsm4∆C mutant as compared to the wild-type in response to osmotic stress, when P bodies are intense and numerous. Our results show that the edc3∆ lsm4∆C mutant limits the mRNA stability in response to osmotic stress, while the magnitude of stabilization was similar as compared to the wild-type. The reduced mRNA stability in the edc3∆ lsm4∆C mutant was correlated with a shorter PGK1 poly(A) tail. Similarly, the MFA2 mRNA was more rapidly deadenylated as well as significantly stabilized in the ccr4∆ deadenylation mutant in the edc3∆ lsm4∆C background. These results suggest a role for these decapping factors in stabilizing mRNA and may implicate P bodies as sites of reduced mRNA degradation.

  5. Gnrh mRNA expression in the brain of cooperatively breeding female Damaraland mole-rats.

    PubMed

    Voigt, Cornelia; Bennett, Nigel C

    2017-04-01

    The Damaraland mole-rat ( Fukomys damarensis ) is a eusocial, subterranean rodent, in which breeding is limited to a single reproductive pair within each colony. Non-reproductive females, while in the confines of the colony, exhibit socially induced infertility. Anovulation is thought to be caused by a disruption in the normal gonadotropin-releasing hormone (GNRH) secretion from the hypothalamus. To assess whether social suppression is associated with altered Gnrh mRNA expression in the brain, we investigated the distribution and gene expression levels by means of in situ hybridization in female breeders and non-breeders from field captured colonies of the Damaraland mole-rat. We found expression of Gnrh mRNA as a loose network in several forebrain areas of female Damaraland mole-rats with the majority of labelling in the preoptic and anterior hypothalamus. The distribution matched previous findings using immunocytochemistry in this and other social mole-rat species. Quantification of the hybridisation signal revealed no difference between breeding and non-breeding females in the average optical density of the hybridization signal and the size of the total area covered by Gnrh mRNA. However, analysis along the rostro-caudal axis revealed significantly elevated Gnrh mRNA expression in the rostral preoptic region of breeders compared to non-breeders, whereas the latter had increased Gnrh mRNA expression at the caudal level of the anterior hypothalamus. This study indicates that social suppression affects the expression of Gnrh mRNA in female Damaraland mole-rats. Furthermore, differential regulation occurs within different neuron subpopulations. © 2017 Society for Reproduction and Fertility.

  6. Sodium 4-phenylbutyrate downregulates HSC70 expression by facilitating mRNA degradation.

    PubMed

    Rubenstein, R C; Lyons, B M

    2001-07-01

    Intracellular trafficking of the DeltaF508 cystic fibrosis transmembrane conductance regulator (CFTR) is repaired by sodium 4-phenylbutyrate (4PBA) by an undetermined mechanism. 4PBA downregulates protein and mRNA expression of the heat shock cognate protein HSC70 (the constitutively expressed member of the 70-kDa heat shock protein family) by approximately 40-50% and decreases formation of a HSC70-DeltaF508 CFTR complex that may be important in the intracellular degradation of DeltaF508 CFTR. We examined the potential mechanisms by which 4PBA decreases HSC70 mRNA and protein expression. In IB3-1 cells, 1 mM 4PBA did not alter the activity of the Chinese hamster ovary HSC70 promoter or of a human HSC70 promoter fragment in luciferase reporter assays nor did it alter HSC70 mRNA synthesis in nuclear runoff assays. In contrast, preincubation with 4PBA increased the rate of HSC70 mRNA degradation by approximately 40%. The initial rate of 35S-HSC70 protein synthesis in 4PBA-treated IB3-1 cells was reduced by approximately 40%, consistent with the steady-state mRNA level, whereas its rate of degradation was unaltered by 4PBA. 4PBA also reduced the steady-state accumulation of (35)S-HSC70 by approximately 40%. These data suggest that 4PBA decreases the expression of HSC70 mRNA and protein by inducing cellular adaptations that result in the decreased stability of HSC70 mRNA.

  7. Changes in beta-actin mRNA expression in remodeling canine myocardium.

    PubMed

    Carlyle, W C; Toher, C A; Vandervelde, J R; McDonald, K M; Homans, D C; Cohn, J N

    1996-01-01

    Beta-actin, a cytoskeletal protein important in the maintenance of cytoarchitecture, has long been thought to be expressed constitutively in myocardial tissue. As such, beta-actin mRNA has been used as a control gene in a wide range of experiments. However, we have uncovered consistent changes in beta-actin mRNA expression in canine myocardium remodeling as a result of insult to the left ventricle. The experimental canine models used were either DC shock damage to the left ventricle or volume overload resulting from severe mitral regurgitation. The remodeling process in both canine models is characterized by an increase in left ventricular mass. PCR amplification using primers designed to selectively amplify the 3' end and a portion of the 3' untranslated region of beta-actin mRNA resulted in the generation of a 297 base pair product predominant only in normal canine myocardium and a 472 base pair product that became increasingly prominent from 1 to 30 days after DC shock damage to the left ventricle and from 10 to 90 days after creation of mitral regurgitation. Northern analysis showed a three-fold increase in beta-actin mRNA after either DC shock or creation of mitral regurgitation. Western analysis revealed an early increase in beta-actin protein followed by an apparent decrease to below baseline levels. These observations suggest that changes in beta-actin mRNA expression accompany the structural alterations that occur in response to myocardial damage. Whether or not the changes in beta-actin mRNA expression play a role in mediating these structural alterations remains to be determined.

  8. Determining if an mRNA is a Substrate of Nonsense-Mediated mRNA Decay in Saccharomyces cerevisiae.

    PubMed

    Johansson, Marcus J O

    2017-01-01

    Nonsense-mediated mRNA decay (NMD) is a conserved eukaryotic quality control mechanism which triggers decay of mRNAs harboring premature translation termination codons. In this chapter, I describe methods for monitoring the influence of NMD on mRNA abundance and decay rates in Saccharomyces cerevisiae. The descriptions include detailed methods for growing yeast cells, total RNA isolation, and Northern blotting. Although the chapter focuses on NMD, the methods can be easily adapted to assess the effect of other mRNA decay pathways.

  9. Hyperresponsive febrile reactions to interleukin (IL) 1α and IL-1β, and altered brain cytokine mRNA and serum cytokine levels, in IL-1β-deficient mice

    PubMed Central

    Alheim, Katarina; Chai, Zhen; Fantuzzi, Giamila; Hasanvan, Homa; Malinowsky, David; Di Santo, Elena; Ghezzi, Pietro; Dinarello, Charles A.; Bartfai, Tamas

    1997-01-01

    IL-1β is an endogenous pyrogen that is induced during systemic lipopolysaccharide (LPS)- or IL-1-induced fever. We have examined the fever and cytokine responses following i.p. injection of IL-1 agonists, IL-1α and IL-1β, and compared these with response to LPS (i.p.) in wild-type and IL-1β-deficient mice. The IL-1β deficient mice appear to have elevated body temperature but exhibit a normal circadian temperature cycle. Exogenously injected IL-1β, IL-1α, or LPS induced hyperresponsive fevers in the IL-1β-deficient mice. We also observed phenotypic differences between wild-type and IL-1β-deficient mice in hypothalamic basal mRNA levels for IL-1α and IL-6, but not for IL-1β-converting enzyme or IL-1 receptor type I or type II. The IL-1α mRNA levels were down-regulated, whereas the IL-6 mRNA levels were up-regulated in the hypothalamus of IL-1β-deficient mice as compared with wild-type mice. The IL-1β-deficient mice also responded to LPS challenge with significantly higher serum corticosterone and with lower serum tumor necrosis factor type α levels than the wild-type mice. The data suggest that, in the redundant cascade of proinflammatory cytokines, IL-1β plays an important but not obligatory role in fever induction by LPS or IL-1α, as well as in the induction of serum tumor necrosis factor type α and corticosterone responses either by LPS or by IL-1α or IL-1β. PMID:9122256

  10. p62/SQSTM1 enhances breast cancer stem-like properties by stabilizing MYC mRNA

    PubMed Central

    Xu, L-Z; Li, S-S; Zhou, W; Kang, Z-J; Zhang, Q-X; Kamran, M; Xu, J; Liang, D-P; Wang, C-L; Hou, Z-J; Wan, X-B; Wang, H-J; Lam, E W-F; Zhao, Z-W; Liu, Q

    2017-01-01

    Aberrant p62 overexpression has been implicated in breast cancer development. Here, we found that p62 expression was elevated in breast cancer stem cells (BCSCs), including CD44+CD24− fractions, mammospheres, ALDH1+ populations and side population cells. Indeed, short-hairpin RNA (shRNA)-mediated knockdown of p62 impaired breast cancer cells from self-renewing under anchorage-independent conditions, whereas ectopic overexpression of p62 enhanced the self-renewal ability of breast cancer cells in vitro. Genetic depletion of p62 robustly inhibited tumor-initiating frequencies, as well as growth rates of BCSC-derived tumor xenografts in immunodeficient mice. Consistently, immunohistochemical analysis of clinical breast tumor tissues showed that high p62 expression levels were linked to poorer clinical outcome. Further gene expression profiling analysis revealed that p62 was positively correlated with MYC expression level, which mediated the function of p62 in promoting breast cancer stem-like properties. MYC mRNA level was reduced upon p62 deletion by siRNA and increased with p62 overexpression in breast cancer cells, suggesting that p62 positively regulated MYC mRNA. Interestingly, p62 did not transactivate MYC promoter. Instead, p62 delayed the degradation of MYC mRNA by repressing the expression of let-7a and let-7b, thus promoting MYC mRNA stabilization at the post-transcriptional level. Consistently, let-7a and let-7b mimics attenuated p62-mediated MYC mRNA stabilization. Together, these findings unveiled a previously unappreciated role of p62 in the regulation of BCSCs, assigning p62 as a promising therapeutic target for breast cancer treatments. PMID:27345399

  11. Human Cytomegalovirus Strategies to Maintain and Promote mRNA Translation

    PubMed Central

    Vincent, Heather A.; Ziehr, Benjamin; Moorman, Nathaniel J.

    2016-01-01

    mRNA translation requires the ordered assembly of translation initiation factors and ribosomal subunits on a transcript. Host signaling pathways regulate each step in this process to match levels of protein synthesis to environmental cues. In response to infection, cells activate multiple defenses that limit viral protein synthesis, which viruses must counteract to successfully replicate. Human cytomegalovirus (HCMV) inhibits host defenses that limit viral protein expression and manipulates host signaling pathways to promote the expression of both host and viral proteins necessary for virus replication. Here we review key regulatory steps in mRNA translation, and the strategies used by HCMV to maintain protein synthesis in infected cells. PMID:27089357

  12. Nitric oxide signaling pathway regulates potassium chloride cotransporter-1 mRNA expression in vascular smooth muscle cells.

    PubMed

    Di Fulvio, M; Lauf, P K; Adragna, N C

    2001-11-30

    Rat vascular smooth muscle cells (VSMCs) express at least two mRNAs for K-Cl cotransporters (KCC): KCC1 and KCC3. cGMP-dependent protein kinase I regulates KCC3 mRNA expression in these cells. Here, we show evidence implicating the nitric oxide (NO)/cGMP signaling pathway in the expression of KCC1 mRNA, considered to be the major cell volume regulator. VSMCs, expressing soluble guanylyl cyclase (sGC) and PKG-I isoforms showed a time- and concentration-dependent increase in KCC1 mRNA levels after treatment with sodium nitroprusside as demonstrated by semiquantitative RT-PCR. sGC-dependent regulation of KCC1 mRNA expression was confirmed using YC-1, a NO-independent sGC stimulator. The sGC inhibitor LY83583 blocked the effects of sodium nitroprusside and YC-1. Moreover, 8-Br-cGMP increased KCC1 mRNA expression in a concentration- and time-dependent fashion. The 8-Br-cGMP effect was partially blocked by KT5823 but not by actinomycin D. However, actinomycin D and cycloheximide increased basal KCC1 mRNA in an additive manner, suggesting different mechanisms of action for both drugs. These findings suggest that in VSMCs, the NO/cGMP-signaling pathway participates in KCC1 mRNA regulation at the post-transcriptional level.

  13. Effects of Janus kinase inhibitor tofacitinib on circulating serum amyloid A and interleukin-6 during treatment for rheumatoid arthritis

    PubMed Central

    Migita, K; Izumi, Y; Jiuchi, Y; Kozuru, H; Kawahara, C; Izumi, M; Sakai, T; Nakamura, M; Motokawa, S; Nakamura, T; Kawakami, A

    2014-01-01

    The Janus kinase inhibitor tofacitinib is currently being investigated as a disease-modifying agent in rheumatoid arthritis (RA). We investigated the in-vivo effects of tofacitinib treatment for 4 weeks on elevated circulating acute-phase serum amyloid (SAA) levels in 14 Japanese patients with RA. SAA levels fell from 110·5 ± 118·5 μg/ml (mean ± standard deviation) at treatment initiation to 15·3 ± 13·3 μg/ml after 4 weeks treatment with tofacitinib. The reduction in SAA levels was greater in patients receiving tofacitinib plus methotrexate compared with those receiving tofacitinib monotherapy. Tofacitinib was also associated with reduced serum interleukin (IL)-6, but had no effect on serum levels of soluble IL-6 receptor. Patients were divided into groups with adequate (normalization) and inadequate SAA responses (without normalization). Serum IL-6 levels were reduced more in the group with adequate SAA response compared with those with inadequate SAA response. These results suggest that tofacitinib down-regulates the proinflammatory cytokine, IL-6, accompanied by reduced serum SAA levels in patients with active RA. The ability to regulate elevated serum IL-6 and SAA levels may explain the anti-inflammatory activity of tofacitinib. PMID:24665995

  14. [Houttuynia Cordata induces expression of human beta-defensin-2 mRNA in pulmonary epithelial cells in vitro].

    PubMed

    Luo, Li; Dong, Bi-rong; Teng, Li-hua

    2008-07-01

    To explore the effects of Houttuynia Cordata on expression of human beta-defensin-2 (HBD-2) in pulmonary epithelial cells (SPC-A-1) in vitro; and to observe the correlationship between the level of HBD-2 mRNA and the concentrations or treatment times of Houttuynia Cordata. The SPC-A-1 cells were cultured with different concentrations of Houttuynia Cordata in vitro, including 0, 12.5, 25, 50, 100 and 200 microg/ml. And then, the SPC-A-1 cells were cultured with the optimal concentration of Houttuynia Cordata in different lengths of time, including 1, 2, 4, 8, 16 and 24 hours. After the treatment, the mRNA level of HBD-2 in pulmonary epithelial cells was detected by means of semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). After being cultured with Houttuynia Cordata, the expression of HBD-2 mRNA had positive correlation with the stimulus concentrations (rs=0.829, P=0.042) and stimulus time (rs=0.914, P=0.003). The highest expression of HBD-2 mRNA was induced by 100 microg/ml Houttuynia Cordata after 8-hour treatment. In comparison with the normal control group and the interleukin-1beta group, 100 microg/ml Houttuynia Cordata could significantly up-regulate the expression of HBD-2 mRNA in SPC-A-1 cells after 8-hour treatment (P<0.01). Houttuynia Cordata can up-regulate expression of HBD-2 mRNA in SPC-A-1 cells, and the highest expression level of HBD-2 mRNA can be obtained by culture with 100 microg/ml Houttuynia Cordata for 8 hours.

  15. Regulation of adeno-associated virus gene expression in 293 cells: control of mRNA abundance and translation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Trempe, J.P.; Carter, B.J.

    1988-01-01

    The authors studied the effects of the adeno-associated virus (AAV) rep gene on the control of gene expression from the AAV p/sub 40/ promoter in 293 cells in the absence of an adenovirus coinfection. AAV vectors containing the chloramphenicol acetyltransferase (cat) gene were used to measure the levels of cat expression and steady-state mRNA from p/sub 40/. When the rep gene was present in cis or in trans, cat expression from p/sub 40/ was decreased 3- to 10-fold, but there was a 2- to 10-fold increase in the level of p/sub 40/ mRNA. Conversely, cat expression increased and the p/submore » 40/ mRNA level decreased in the absence of the rep gene. Both wild-type and carboxyl-terminal truncated Rep proteins were capable of eliciting both effects. These data suggest two roles for the pleiotropic AAV rep gene: as a translational inhibitor and as a positive regulator of p/sub 40/ mRNA levels. They also provide additional evidence for a cis-acting negative regulatory region which decreases RNA from the AAV p/sub 5/ promoter in a fashion independent of rep.« less

  16. Expression of klotho mRNA and protein in rat brain parenchyma from early postnatal development into adulthood

    PubMed Central

    Clinton, Sarah M.; Glover, Matthew E.; Maltare, Astha; Laszczyk, Ann M.; Mehi, Stephen J.; Simmons, Rebecca K.; King, Gwendalyn D.

    2013-01-01

    Without the age-regulating protein klotho, mouse lifespan is shortened and the rapid onset of age-related disorders occurs. Conversely, overexpression of klotho extends mouse lifespan. Klotho is most abundant in kidney and expressed in a limited number of other organs, including the brain, where klotho levels are highest in choroid plexus. Reports vary on where klotho is expressed within the brain parenchyma, and no data is available as to whether klotho levels change across postnatal development. We used in situ hybridization to map klotho mRNA expression in the developing and adult rat brain and report moderate, widespread expression across grey matter regions. mRNA expression levels in cortex, hippocampus, caudate putamen, and amygdala decreased during the second week of life and then gradually rose to adult levels by postnatal day 21. Immunohistochemistry revealed a protein expression pattern similar to the mRNA results, with klotho protein expressed widely throughout the brain. Klotho protein co-localized with both the neuronal marker NeuN, as well as, oligodendrocyte marker olig2. These results provide the first anatomical localization of klotho mRNA and protein in rat brain parenchyma and demonstrate that klotho levels vary during early postnatal development. PMID:23838326

  17. Relationship between Sustained Reductions in Plasma Lipid and Lipoprotein Concentrations with Apheresis and Plasma Levels and mRNA Expression of PTX3 and Plasma Levels of hsCRP in Patients with HyperLp(a)lipoproteinemia

    PubMed Central

    Stefanutti, Claudia; Mazza, Fabio; Steiner, Michael; Watts, Gerald F.; De Nève, Joel; Pasqualetti, Daniela; Paal, Juergen

    2016-01-01

    The effect of lipoprotein apheresis (Direct Adsorption of Lipids, DALI) (LA) on plasma levels of pentraxin 3 (PTX3), an inflammatory marker that reflects coronary plaque vulnerability, and expression of PTX3 mRNA was evaluated in patients with hyperLp(a)lipoproteinemia and angiographically defined atherosclerosis/coronary artery disease. Eleven patients, aged 55 ± 9.3 years (mean ± SD), were enrolled in the study. PTX3 soluble protein levels in plasma were unchanged by 2 sessions of LA; however, a downregulation of mRNA expression for PTX3 was observed, starting with the first session of LA (p < 0.001). The observed reduction was progressively increased in the interval between the first and second LA sessions to achieve a maximum decrease by the end of the second session. A statistically significantly greater treatment-effect correlation was observed in patients undergoing weekly treatments, compared with those undergoing treatment every 15 days. A progressive reduction in plasma levels of C-reactive protein was also seen from the first session of LA, with a statistically significant linear correlation for treatment-effect in the change in plasma levels of this established inflammatory marker (R 2 = 0.99; p < 0.001). Our findings suggest that LA has anti-inflammatory and endothelium protective effects beyond its well-established efficacy in lowering apoB100-containing lipoproteins. PMID:26903710

  18. Vaspin plasma concentrations and mRNA expressions in patients with stable and unstable angina pectoris.

    PubMed

    Li, Hai Ling; Peng, Wen Hui; Cui, Shi Tao; Lei, Hou; Wei, Yi Dong; Li, Wei Ming; Xu, Ya Wei

    2011-09-01

    Vaspin was a recently identified adipokine, playing a protective role in many metabolic diseases. The present study aimed to investigate the association between vaspin plasma level and stable angina pectoris (SAP) and unstable angina pectoris (UAP). A total of 88 patients with angiographically-proved coronary artery disease (CAD) (SAP 47, UAP 41) and 103 control subjects without cardiovascular diseases were enrolled in this study. Circulating vaspin, mRNA expression of vaspin in peripheral blood mononuclear cells (PBMC), clinical parameters, lipid profile and high-sensitivity C-reactive protein (hsCRP) were assayed. The severity of CAD was also assessed according to the number of vessels diseased. There are significant differences in circulating vaspin levels and mRNA levels of PBMC between SAP and UAP groups (SAP 0.91±0.95 ng/mL and UAP 0.43±0.38 ng/mL, p<0.01 in circulating vaspin level; SAP 1.19±0.85 and UAP 0.82±0.56, p<0.05 in mRNA level of PBMC). An inverse correlation between the number of diseased vessels and plasma vaspin concentration was observed (r=-0.350, p<0.01) in the CAD group. Construction of receiver operating characteristic curves confirmed that vaspin plasma concentrations significantly differentiated CAD patients (area under the curve=0.684, p<0.001), as well as UAP (area under the curve=0.640, p<0.05). Decreased vaspin plasma levels and mRNA levels in PBMC were observed in patients with UAP. Low vaspin concentrations correlate with CAD severity. The findings suggested that vaspin could serve as a novel biomarker of CAD as well as UAP.

  19. Inhibition of pyrimidine synthesis reverses viral virulence factor-mediated block of mRNA nuclear export

    PubMed Central

    Zhang, Liang; Das, Priyabrata; Schmolke, Mirco; Manicassamy, Balaji; Wang, Yaming; Deng, Xiaoyi; Cai, Ling; Tu, Benjamin P.; Forst, Christian V.; Roth, Michael G.; Levy, David E.; García-Sastre, Adolfo; de Brabander, Jef; Phillips, Margaret A.

    2012-01-01

    The NS1 protein of influenza virus is a major virulence factor essential for virus replication, as it redirects the host cell to promote viral protein expression. NS1 inhibits cellular messenger ribonucleic acid (mRNA) processing and export, down-regulating host gene expression and enhancing viral gene expression. We report in this paper the identification of a nontoxic quinoline carboxylic acid that reverts the inhibition of mRNA nuclear export by NS1, in the absence or presence of the virus. This quinoline carboxylic acid directly inhibited dihydroorotate dehydrogenase (DHODH), a host enzyme required for de novo pyrimidine biosynthesis, and partially reduced pyrimidine levels. This effect induced NXF1 expression, which promoted mRNA nuclear export in the presence of NS1. The release of NS1-mediated mRNA export block by DHODH inhibition also occurred in the presence of vesicular stomatitis virus M (matrix) protein, another viral inhibitor of mRNA export. This reversal of mRNA export block allowed expression of antiviral factors. Thus, pyrimidines play a necessary role in the inhibition of mRNA nuclear export by virulence factors. PMID:22312003

  20. Decreased TIM-3 mRNA expression in peripheral blood mononuclear cells from nephropathy patients.

    PubMed

    Cai, X Z; Liu, N; Qiao, Y; Du, S Y; Chen, Y; Chen, D; Yu, S; Jiang, Y

    2015-06-12

    Increasing evidence shows that TIM-1 and TIM-3 in-fluence chronic autoimmune diseases, and their expression levels in immune cells from nephritic patients are still unknown. Real-time transcription-polymerase chain reaction analysis was used to deter-mine expression levels of TIM-1 and TIM-3 mRNA in peripheral blood mononuclear cells (PBMCs) from 36 patients with minimal change glo-merulopathy (MCG), 65 patients with lupus nephritis (LN), 78 patients with IgA nephropathy (IgAN), 55 patients with membranous nephropa-thy (MN), 22 patients with crescentic glomerulonephritis (CGN), 26 patients with anaphylactoid purpura nephritis (APN), and 63 healthy controls. TIM-3 mRNA expression significantly decreased in PBMCs from nephritic patients (LN, P < 0.0001; MCG, P < 0.0001; MN, P = 0.0031; CGN, P = 0.0464; IgAN, P = 0.0002; APN, P = 0.0392) com-pared with healthy controls. In contrast, there was no significant differ-ence in TIM-1 mRNA expression between the patients and the healthy controls. Our results suggest that insufficient expression of TIM-3 mRNA may be involved in the pathogenesis of nephropathy.

  1. Disruption of the Axonal Trafficking of Tyrosine Hydroxylase mRNA Impairs Catecholamine Biosynthesis in the Axons of Sympathetic Neurons.

    PubMed

    Aschrafi, Armaz; Gioio, Anthony E; Dong, Lijin; Kaplan, Barry B

    2017-01-01

    Tyrosine hydroxylase (TH) is the enzyme that catalyzes the rate-limiting step in the biosynthesis of the catecholamine neurotransmitters. In a previous communication, evidence was provided that TH mRNA is trafficked to the axon, where it is locally translated. In addition, a 50-bp sequence element in the 3'untranslated region (3'UTR) of TH mRNA was identified that directs TH mRNA to distal axons (i.e., zip-code). In the present study, the hypothesis was tested that local translation of TH plays an important role in the biosynthesis of the catecholamine neurotransmitters in the axon and/or presynaptic nerve terminal. Toward this end, a targeted deletion of the axonal transport sequence element was developed, using the lentiviral delivery of the CRISPR/Cas9 system, and two guide RNA (gRNA) sequences flanking the 50-bp cis- acting regulatory element in rat superior cervical ganglion (SCG) neurons. Deletion of the axonal transport element reduced TH mRNA levels in the distal axons and reduced the axonal protein levels of TH and TH activity as measured by phosphorylation of SER40 in SCG neurons. Moreover, deletion of the zip-code diminished the axonal levels of dopamine (DA) and norepinephrine (NE). Conversely, the local translation of exogenous TH mRNA in the distal axon enhanced TH levels and activity, and elevated axonal NE levels. Taken together, these results provide direct evidence to support the hypothesis that TH mRNA trafficking and local synthesis of TH play an important role in the synthesis of catecholamines in the axon and presynaptic terminal.

  2. Neutrophil elastase increases MUC5AC mRNA and protein expression in respiratory epithelial cells.

    PubMed

    Voynow, J A; Young, L R; Wang, Y; Horger, T; Rose, M C; Fischer, B M

    1999-05-01

    Chronic neutrophil-predominant inflammation and hypersecretion of mucus are common pathophysiological features of cystic fibrosis, chronic bronchitis, and viral- or pollution-triggered asthma. Neutrophils release elastase, a serine protease, that causes increased mucin production and secretion. The molecular mechanisms of elastase-induced mucin production are unknown. We hypothesized that as part of this mechanism, elastase upregulates expression of a major respiratory mucin gene, MUC5AC. A549, a human lung carcinoma cell line that expresses MUC5AC mRNA and protein, and normal human bronchial epithelial cells in an air-liquid interface culture were stimulated with neutrophil elastase. Neutrophil elastase increased MUC5AC mRNA levels in a time-dependent manner in both cell culture systems. Neutrophil elastase treatment also increased MUC5AC protein levels in A549 cells. The mechanism of MUC5AC gene regulation by elastase was determined in A549 cells. The induction of MUC5AC gene expression required serine protease activity; other classes of proteases had no effect on MUC5AC gene expression. Neutrophil elastase increased MUC5AC mRNA levels by enhancing mRNA stability. This is the first report of mucin gene regulation by this mechanism.

  3. c-fms mRNA is regulated posttranscriptionally by 1,25(OH)2D3 in HL-60 cells.

    PubMed

    Biskobing, D M; Fan, D; Rubin, J

    1997-09-01

    Macrophage colony-stimulating factor (MCSF) is required for normal osteoclast and macrophage development. The receptor for MCSF (c-fms) is expressed on the pluripotent precursor and mature osteoclasts and macrophages. We have previously shown in myelomonocytic HL-60 cells that phorbol myristate acetate (PMA) upregulates c-fms mRNA expression. This induction of c-fms is inhibited by 1,25(OH)2D3. The major regulatory control of c-fms mRNA levels by PMA has been identified as posttranscriptional. However, a role of transcript elongation in controlling levels of c-fms mRNA has also been suggested. To better understand the 1,25(OH)2D3 regulation of c-fms mRNA expression we studied nuclear run on, mRNA stability, and transcript elongation in HL-60 cells treated with 10 ng/ml phorbol myristate acetate, 10 nM 1,25(OH)2D3 alone or combined. We demonstrated by nuclear run on that c-fms was constitutively transcribed in 1,25(OH)2D3 as well as control and PMA-treated cells. Transcript elongation was evaluated by RT-PCR for exon 2 or exon 3. Both exons were minimally expressed in control and 1,25(OH)2D3-treated cells, and increased in PMA-treated cells; this increased expression was inhibited by the addition of 1,25(OH)2D3. These results fail to show differential transcript elongation. Measurement of mRNA stability demonstrated decreased mRNA half-life to 5 hours in cells treated with PMA and 1,25(OH)2D3 compared with a half-life of 8 hours in cells treated with PMA alone. Our findings demonstrate that c-fms is regulated by 1,25(OH)2D3 at the posttranscriptional level by changes in mRNA stability. This gives the cell the ability to respond to local signals with rapid changes in c-fms levels altering the ability of the cell to respond to MCSF.

  4. Sequence-engineered mRNA Without Chemical Nucleoside Modifications Enables an Effective Protein Therapy in Large Animals

    PubMed Central

    Thess, Andreas; Grund, Stefanie; Mui, Barbara L; Hope, Michael J; Baumhof, Patrick; Fotin-Mleczek, Mariola; Schlake, Thomas

    2015-01-01

    Being a transient carrier of genetic information, mRNA could be a versatile, flexible, and safe means for protein therapies. While recent findings highlight the enormous therapeutic potential of mRNA, evidence that mRNA-based protein therapies are feasible beyond small animals such as mice is still lacking. Previous studies imply that mRNA therapeutics require chemical nucleoside modifications to obtain sufficient protein expression and avoid activation of the innate immune system. Here we show that chemically unmodified mRNA can achieve those goals as well by applying sequence-engineered molecules. Using erythropoietin (EPO) driven production of red blood cells as the biological model, engineered Epo mRNA elicited meaningful physiological responses from mice to nonhuman primates. Even in pigs of about 20 kg in weight, a single adequate dose of engineered mRNA encapsulated in lipid nanoparticles (LNPs) induced high systemic Epo levels and strong physiological effects. Our results demonstrate that sequence-engineered mRNA has the potential to revolutionize human protein therapies. PMID:26050989

  5. Intraovarian expression of GnRH-1 and gonadotropin mRNA and protein levels in Siberian hamsters during the estrus cycle and photoperiod induced regression/recrudescence

    PubMed Central

    Shahed, Asha; Young, Kelly A.

    2010-01-01

    The hypothalamic-pituitary-gonadal (HPG) axis is the key reproductive regulator in vertebrates. While gonadotropin releasing hormone (GnRH), follicle stimulating (FSH), and luteinizing (LH) hormones are primarily produced in the hypothalamus and pituitary, they can be synthesized in the gonads, suggesting an intraovarian GnRH-gonadotropin axis. Because these hormones are critical for follicle maturation and steroidogenesis, we hypothesized that this intraovarian axis may be important in photoperiod-induced ovarian regression/recrudescence in seasonal breeders. Thus, we investigated GnRH-1 and gonadotropin mRNA and protein expression in Siberian hamster ovaries during (1) the estrous cycle; where ovaries from cycling long day hamsters (LD;16L:8D) were collected at proestrus, estrus, diestrus I, and diestrus II and (2) during photoperiod induced regression/ recrudescence; where ovaries were collected from hamsters exposed to 14wks of LD, short days (SD;8L:16D), or 8wks post-transfer to LD after 14wks SD (PT). GnRH-1, LHβ, FSHβ, and common α subunit mRNA expression was observed in cycling ovaries. GnRH-1 expression peaked at diestrus I compared to other stages (p<0.05). FSHβ and LHβ mRNA levels peaked at proestrus and diestrus I (p<0.05), with no change in the α subunit across the cycle (p>0.05). SD exposure decreased ovarian mass and plasma estradiol concentrations (p<0.05) and increased GnRH-1, LHβ, FSHβ, and α subunit mRNA expression as compared to LD and, except for LH, compared to PT (p<0.05). GnRH and gonadotropin protein was also dynamically expressed across the estrous cycle and photoperiod exposure. The presence of cycling intraovarian GnRH-1 and gonadotropin mRNA suggests that these hormones may be locally involved in ovarian maintenance during SD regression and/or could potentially serve to prime ovaries for rapid recrudescence. PMID:20955709

  6. Arc mRNA induction in striatal efferent neurons associated with response learning.

    PubMed

    Daberkow, D P; Riedy, M D; Kesner, R P; Keefe, K A

    2007-07-01

    The dorsal striatum is involved in motor-response learning, but the extent to which distinct populations of striatal efferent neurons are differentially involved in such learning is unknown. Activity-regulated, cytoskeleton-associated (Arc) protein is an effector immediate-early gene implicated in synaptic plasticity. We examined arc mRNA expression in striatopallidal vs. striatonigral efferent neurons in dorsomedial and dorsolateral striatum of rats engaged in reversal learning on a T-maze motor-response task. Male Sprague-Dawley rats learned to turn right or left for 3 days. Half of the rats then underwent reversal training. The remaining rats were yoked to rats undergoing reversal training, such that they ran the same number of trials but ran them as continued-acquisition trials. Brains were removed and processed using double-label fluorescent in situ hybridization for arc and preproenkephalin (PPE) mRNA. In the reversal, but not the continued-acquisition, group there was a significant relation between the overall arc mRNA signal in dorsomedial striatum and the number of trials run, with rats reaching criterion in fewer trials having higher levels of arc mRNA expression. A similar relation was seen between the numbers of PPE(+) and PPE(-) neurons in dorsomedial striatum with cytoplasmic arc mRNA expression. Interestingly, in behaviourally activated animals significantly more PPE(-) neurons had cytoplasmic arc mRNA expression. These data suggest that Arc in both striatonigral and striatopallidal efferent neurons is involved in striatal synaptic plasticity mediating motor-response learning in the T-maze and that there is differential processing of arc mRNA in distinct subpopulations of striatal efferent neurons.

  7. Cytokine mRNA expression in normal skin of various age populations before and after engraftment onto nude mice.

    PubMed

    Gilhar, A; Ullmann, Y; Shalagino, R; Weisinger, G

    1998-01-01

    Whether the impact of skin biological age on cytokine expression is a result of this tissue's proliferation potential or not is an important issue in dermatology. We investigated these questions by monitoring cytokine marker mRNA expression from human skin samples from healthy groups of individuals. The skin samples studied represented three age groups: fetal (17-21 weeks), young (18-35 years) and aged (76-88 years). Furthermore, upon skin transplantation of tissue from different age groups onto nude mice, we investigated whether cytokine marker RNA levels would change or normalize. Interestingly, both TNF-alpha and P53 mRNA showed a similar pattern of expression. Both were significantly higher in fetal skin (p < 0.0001 and p < 0.05, respectively), and no difference was noted between aged versus young skin. In contrast to this, IL1-alpha mRNA was expressed at its lowest and highest levels in fetal and young skin, respectively. Following skin transplantation, cytokines and P53 mRNA expression were normalized to similar levels in all age groups. This study implies that when cytokine expression was determined directly at the mRNA level, post-natal expression was not significantly different at either age group. Furthermore, it seems that the environmental conditions surrounding the grafted human skin found on nude mice encouraged normalization of donor cytokine expression.

  8. Time-dependent effects of intermittent hydrostatic pressure on articular chondrocyte type II collagen and aggrecan mRNA expression.

    PubMed

    Smith, R L; Lin, J; Trindade, M C; Shida, J; Kajiyama, G; Vu, T; Hoffman, A R; van der Meulen, M C; Goodman, S B; Schurman, D J; Carter, D R

    2000-01-01

    The normal loading of joints during daily activities causes the articular cartilage to be exposed to high levels of intermittent hydrostatic pressure. This study quantified effects of intermittent hydrostatic pressure on expression of mRNA for important extracellular matrix constituents. Normal adult bovine articular chondrocytes were isolated and tested in primary culture, either as high-density monolayers or formed aggregates. Loaded cells were exposed to 10 MPa of intermittent hydrostatic pressure at a frequency of 1 Hz for periods of 2, 4, 8, 12, and 24 hrs. Other cells were intermittently loaded for a period of 4 hrs per day for 4 days. Semiquantitative reverse transcription polymerase chain reaction assays were used to assess mRNA signal levels for collagen types II and I and aggrecan. The results showed that type II collagen mRNA signal levels exhibited a biphasic pattern, with an initial increase of approximately five-fold at 4 and 8 hrs that subsequently decreased by 24 hrs. In contrast, aggrecan mRNA signal increased progressively up to three-fold throughout the loading period. Changing the loading profile to 4 hrs per day for 4 days increased the mRNA signal levels for type II collagen nine-fold and for aggrecan twenty-fold when compared to unloaded cultures. These data suggest that specific mechanical loading protocols may be required to optimally promote repair and regeneration of diseased joints.

  9. Simulated microgravity reduces mRNA levels of multidrug resistance genes 4 and 5 in non-metastatic human melanoma cells

    NASA Astrophysics Data System (ADS)

    Eiermann, Peter; Tsiockas, Wasiliki; Hauslage, Jens; Hemmersbach, Ruth; Gerzer, Rupert; Ivanova, Krassimira

    mRNA levels of sGC α and β were down-regulated by about 31% and 22%, respectively. Thus, the reduced expression of MRP4/5 could be related to the decrease in mRNA levels for the sGC subunits. In addition, the long-term exposure to simulated microgravity did not alter cellular morphology. Taken together, the results of our studies indicate that the expression of MRP4/5 in non-metastatic melanoma cells is inversely regulated by hypergravity and simulated microgravity. Finally, a reduced expression of MRP4 and MRP5 may increase the availability of drugs in cells and influence astronaut medication.

  10. Activation of Tyrosine Hydroxylase mRNA Translation by cAMP in Midbrain Dopaminergic Neurons

    PubMed Central

    Chen, Xiqun; Xu, Lu; Radcliffe, Pheona; Sun, Baoyong; Tank, A. William

    2009-01-01

    During prolonged stress or chronic treatment with neurotoxins, robust compensatory mechanisms occur which maintain sufficient levels of catecholamine neurotransmitters in terminal regions. One of these mechanisms is the up-regulation of tyrosine hydroxylase (TH), the enzyme that controls catecholamine biosynthesis. In neurons of the periphery and locus coeruleus, this up-regulation is associated with an initial induction of TH mRNA. In contrast, this induction either does not occur or is nominal in mesencephalic dopamine neurons. The reasons for this lack of compensatory TH mRNA induction remain obscure, because so little is known about the regulation of TH expression in these neurons. In this report we test whether activation of the cAMP signaling pathway regulates TH gene expression in two rodent models of midbrain dopamine neurons, ventral midbrain organotypic slice cultures and MN9D cells. Our results demonstrate that elevation of cAMP leads to induction of TH protein and TH activity in both model systems; however, TH mRNA levels are not up-regulated by cAMP. The induction of TH protein is the result of a novel post-transcriptional mechanism that activates TH mRNA translation. This translational activation is mediated by sequences within the 3′UTR of TH mRNA. Our results support a model in which cAMP induces or activates trans-factors that interact with the TH mRNA 3′UTR to increase TH protein synthesis. An understanding of this novel regulatory mechanism may help to explain the control of TH gene expression and consequently dopamine biosynthesis in midbrain neurons under different physiological and pathological conditions. PMID:18349104

  11. Global miRNA expression and correlation with mRNA levels in primary human bone cells

    PubMed Central

    Laxman, Navya; Rubin, Carl-Johan; Mallmin, Hans; Nilsson, Olle; Pastinen, Tomi; Grundberg, Elin; Kindmark, Andreas

    2015-01-01

    MicroRNAs (miRNAs) are important post-transcriptional regulators that have recently introduced an additional level of intricacy to our understanding of gene regulation. The aim of this study was to investigate miRNA–mRNA interactions that may be relevant for bone metabolism by assessing correlations and interindividual variability in miRNA levels as well as global correlations between miRNA and mRNA levels in a large cohort of primary human osteoblasts (HOBs) obtained during orthopedic surgery in otherwise healthy individuals. We identified differential expression (DE) of 24 miRNAs, and found 9 miRNAs exhibiting DE between males and females. We identified hsa-miR-29b, hsa-miR-30c2, and hsa-miR-125b and their target genes as important modulators of bone metabolism. Further, we used an integrated analysis of global miRNA–mRNA correlations, mRNA-expression profiling, DE, bioinformatics analysis, and functional studies to identify novel target genes for miRNAs with the potential to regulate osteoblast differentiation and extracellular matrix production. Functional studies by overexpression and knockdown of miRNAs showed that, the differentially expressed miRNAs hsa-miR-29b, hsa-miR-30c2, and hsa-miR-125b target genes highly relevant to bone metabolism, e.g., collagen, type I, α1 (COL1A1), osteonectin (SPARC), Runt-related transcription factor 2 (RUNX2), osteocalcin (BGLAP), and frizzled-related protein (FRZB). These miRNAs orchestrate the activities of key regulators of osteoblast differentiation and extracellular matrix proteins by their convergent action on target genes and pathways to control the skeletal gene expression. PMID:26078267

  12. Towards a molecular level understanding of the sulfanilamide-soil organic matter-interaction.

    PubMed

    Ahmed, Ashour A; Thiele-Bruhn, Sören; Leinweber, Peter; Kühn, Oliver

    2016-07-15

    Sorption experiments of sulfanilamide (SAA) on well-characterized samples of soil size-fractions were combined with the modeling of SAA-soil-interaction via quantum chemical calculations. Freundlich unit capacities were determined in batch experiments and it was found that they increase with the soil organic matter (SOM) content according to the order fine silt > medium silt > clay > whole soil > coarse silt > sand. The calculated binding energies for mass-spectrometrically quantified sorption sites followed the order ionic species > peptides > carbohydrates > phenols and lignin monomers > lignin dimers > heterocyclic compounds > fatty acids > sterols > aromatic compounds > lipids, alkanes, and alkenes. SAA forms H-bonds through its polar centers with the polar SOM sorption sites. In contrast dispersion and π-π-interactions predominate the interaction of the SAA aromatic ring with the non-polar moieties of SOM. Moreover, the dipole moment, partial atomic charges, and molecular volume of the SOM sorption sites are the main physical properties controlling the SAA-SOM-interaction. Further, reasonable estimates of the Freundlich unit capacities from the calculated binding energies have been established. Consequently, we suggest using this approach in forthcoming studies to disclose the interactions of a wide range of organic pollutants with SOM. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. Double-Stranded RNA-Binding Protein Regulates Vascular Endothelial Growth Factor mRNA Stability, Translation, and Breast Cancer Angiogenesis▿

    PubMed Central

    Vumbaca, Frank; Phoenix, Kathryn N.; Rodriguez-Pinto, Daniel; Han, David K.; Claffey, Kevin P.

    2008-01-01

    Vascular endothelial growth factor (VEGF) is a key angiogenic factor expressed under restricted nutrient and oxygen conditions in most solid tumors. The expression of VEGF under hypoxic conditions requires transcription through activated hypoxia-inducible factor 1 (HIF-1), increased mRNA stability, and facilitated translation. This study identified double-stranded RNA-binding protein 76/NF90 (DRBP76/NF90), a specific isoform of the DRBP family, as a VEGF mRNA-binding protein which plays a key role in VEGF mRNA stability and protein synthesis under hypoxia. The DRBP76/NF90 protein binds to a human VEGF 3′ untranslated mRNA stability element. RNA interference targeting the DRBP76/NF90 isoform limited hypoxia-inducible VEGF mRNA and protein expression with no change in HIF-1-dependent transcriptional activity. Stable repression of DRBP76/NF90 in MDA-MB-435 breast cancer cells demonstrated reduced polysome-associated VEGF mRNA levels under hypoxic conditions and reduced mRNA stability. Transient overexpression of the DRBP76/NF90 protein increased both VEGF mRNA and protein levels synthesized under normoxic and hypoxic conditions. Cells with stable repression of the DRBP76/NF90 isoform showed reduced tumorigenic and angiogenic potential in an orthotopic breast tumor model. These data demonstrate that the DRBP76/NF90 isoform facilitates VEGF expression by promoting VEGF mRNA loading onto polysomes and translation under hypoxic conditions, thus promoting breast cancer growth and angiogenesis in vivo. PMID:18039850

  14. Measurement of funny current (I(f)) channel mRNA in human atrial tissue: correlation with left atrial filling pressure and atrial fibrillation.

    PubMed

    Lai, L P; Su, M J; Lin, J L; Tsai, C H; Lin, F Y; Chen, Y S; Hwang, J J; Huang, S K; Tseng, Y Z; Lien, W P

    1999-07-01

    The funny current (I(f)) contributes to phase IV spontaneous depolarization in cardiac pacemaker tissue. Enhanced I(f) activity in myocardial tissue may lead to increased automaticity and therefore tachyarrhythmia. We measured the amount of I(f) activity in the messenger ribonucleic acid (mRNA) in human atrial tissue and correlated the mRNA amount to left atrial filling pressure and atrial fibrillation (AF). A total of 34 patients undergoing open heart surgery were included (15 men and 19 women, aged 55+/-10 years). Atrial tissue was obtained from the right atrial free wall, the right atrial appendage, the left atrial free wall, and the left atrial appendage, respectively. The mRNA amount of the I(f) channel was measured by reverse transcription polymerase chain reaction and was normalized to the mRNA levels of glyceraldehyde 3-phosphate dehydrogenase. We found that the I(f) channel mRNA was present at all the atrial sampling sites. A higher left atrial filling pressure, an indicator of congestive heart failure, was associated with a higher I(f) mRNA level (r2 = 0.446, P < 0.01 by linear regression). We also found that the mRNA amount was significantly higher in patients with AF than in patients without AF (1.68+/-0.49 vs 1.27+/-0.43; P < 0.05). Age, sex, right atrial filling pressure, left atrial dimension, and left ventricular ejection fraction had no significant effect on the mRNA level. The mRNA of the I(f) channel is present in the free-wall area and appendage area from both atria. Increased left atrial filling pressure and clinical AF are associated with increased I(f) mRNA level.

  15. Group II intron inhibits conjugative relaxase expression in bacteria by mRNA targeting

    PubMed Central

    Piazza, Carol Lyn; Smith, Dorie

    2018-01-01

    Group II introns are mobile ribozymes that are rare in bacterial genomes, often cohabiting with various mobile elements, and seldom interrupting housekeeping genes. What accounts for this distribution has not been well understood. Here, we demonstrate that Ll.LtrB, the group II intron residing in a relaxase gene on a conjugative plasmid from Lactococcus lactis, inhibits its host gene expression and restrains the naturally cohabiting mobile element from conjugative horizontal transfer. We show that reduction in gene expression is mainly at the mRNA level, and results from the interaction between exon-binding sequences (EBSs) in the intron and intron-binding sequences (IBSs) in the mRNA. The spliced intron targets the relaxase mRNA and reopens ligated exons, causing major mRNA loss. Taken together, this study provides an explanation for the distribution and paucity of group II introns in bacteria, and suggests a potential force for those introns to evolve into spliceosomal introns. PMID:29905149

  16. Group II intron inhibits conjugative relaxase expression in bacteria by mRNA targeting.

    PubMed

    Qu, Guosheng; Piazza, Carol Lyn; Smith, Dorie; Belfort, Marlene

    2018-06-15

    Group II introns are mobile ribozymes that are rare in bacterial genomes, often cohabiting with various mobile elements, and seldom interrupting housekeeping genes. What accounts for this distribution has not been well understood. Here, we demonstrate that Ll.LtrB, the group II intron residing in a relaxase gene on a conjugative plasmid from Lactococcus lactis , inhibits its host gene expression and restrains the naturally cohabiting mobile element from conjugative horizontal transfer. We show that reduction in gene expression is mainly at the mRNA level, and results from the interaction between exon-binding sequences (EBSs) in the intron and intron-binding sequences (IBSs) in the mRNA. The spliced intron targets the relaxase mRNA and reopens ligated exons, causing major mRNA loss. Taken together, this study provides an explanation for the distribution and paucity of group II introns in bacteria, and suggests a potential force for those introns to evolve into spliceosomal introns. © 2018, Qu et al.

  17. Seasonal Relationship between Gonadotropin, Growth Hormone, and Estrogen Receptor mRNA Expression in the Pituitary Gland of Largemouth Bass

    PubMed Central

    Martyniuk, Christopher J; Kroll, Kevin J.; Porak, Wesley F.; Steward, Cheree; Grier, Harry J.; Denslow, Nancy D.

    2011-01-01

    The objectives of this study were to investigate the seasonal changes in pituitary gonadotropins, growth hormone (GH), and estrogen receptor (ER) isoform mRNA in wild female and male largemouth bass (LMB) (Micropterus salmoides) from an unpolluted habitat to better understand reproductive physiology in this ecologically important species. Female pituitary luteinizing hormone (LH) β subunit and follicle-stimulating hormone (FSH) β subunit mRNA showed significant seasonal variation with levels peaking from January to April and were lowest from May through August. Male LMB showed more variation in gonadotropin subunit expression from month to month. Females had approximately 2–3 times higher gonadotropin mRNA levels in the pituitary when compared to males. All three gonadotropin mRNAs in females were positively correlated to gonadosomatic index (GSI), but only LHβ mRNA was correlated to GSI in males. Gonadotropin mRNA expression also increased with increasing oocyte and sperm maturation. Gonadotropin β subunit mRNA expression was positively correlated to GH mRNA in both sexes. The expression of all three ER isoforms was significantly correlated to each other in both sexes. The concurrent increase in all three ER mRNA isoforms with increasing gonadotropin mRNA in females and males suggests a prominent role for E2 feedback on pituitary gonadotropin synthesis in both sexes and that each of the three ER isoforms are likely to play a role in the pituitary during teleost reproduction. PMID:19416730

  18. Seasonal relationship between gonadotropin, growth hormone, and estrogen receptor mRNA expression in the pituitary gland of largemouth bass.

    PubMed

    Martyniuk, Christopher J; Kroll, Kevin J; Porak, Wesley F; Steward, Cheree; Grier, Harry J; Denslow, Nancy D

    2009-09-15

    The objectives of this study were to investigate the seasonal changes in pituitary gonadotropins, growth hormone (GH), and estrogen receptor (ER) isoform mRNA in wild female and male largemouth bass (LMB) (Micropterus salmoides) from an unpolluted habitat to better understand reproductive physiology in this ecologically important species. Female pituitary luteinizing hormone (LH) beta subunit and follicle stimulating hormone (FSH) beta subunit mRNA showed significant seasonal variation with levels peaking from January to April and were lowest from May to August. Male LMB showed more variation in gonadotropin subunit expression from month to month. Females had approximately 2-3 times higher gonadotropin mRNA levels in the pituitary when compared to males. All three gonadotropin mRNAs in females were positively correlated to gonadosomatic index (GSI), but only LHbeta mRNA was correlated to GSI in males. Gonadotropin mRNA expression also increased with increasing oocyte and sperm maturation. Gonadotropin beta subunit mRNA expression was positively correlated to GH mRNA in both sexes. The expression of all three ER isoforms was significantly correlated to each other in both sexes. The concurrent increase in all three ER mRNA isoforms with increasing gonadotropin mRNA in females and males suggests a prominent role for E2 feedback on pituitary gonadotropin synthesis in both sexes and that each of the three ER isoforms are likely to play a role in the pituitary during teleost reproduction.

  19. The cytoplasmic mRNA degradation factor Pat1 is required for rRNA processing

    PubMed Central

    Muppavarapu, Mridula; Huch, Susanne; Nissan, Tracy

    2016-01-01

    ABSTRACT Pat1 is a key cytoplasmic mRNA degradation factor, the loss of which severely increases mRNA half-lives. Several recent studies have shown that Pat1 can enter the nucleus and can shuttle between the nucleus and the cytoplasm. As a result, many nuclear roles have been proposed for Pat1. In this study, we analyzed four previously suggested nuclear roles of Pat1 and show that Pat1 is not required for efficient pre-mRNA splicing or pre-mRNA decay in yeast. However, lack of Pat1 results in accumulation of pre-rRNA processing intermediates. Intriguingly, we identified a novel genetic relationship between Pat1 and the rRNA decay machinery, specifically the exosome and the TRAMP complex. While the pre-rRNA processing intermediates that accumulate in the pat1 deletion mutant are, at least to some extent, recognized as aberrant by the rRNA degradation machinery, it is unlikely that these accumulations are the cause of their synthetic sick relationship. Here, we show that the dysregulation of the levels of mRNAs related to ribosome biogenesis could be the cause of the accumulation of the pre-rRNA processing intermediates. Although our results support a role for Pat1 in transcription, they nevertheless suggest that the primary cause of the dysregulated mRNA levels is most likely due to Pat1's role in mRNA decapping and mRNA degradation. PMID:26918764

  20. The cytoplasmic mRNA degradation factor Pat1 is required for rRNA processing.

    PubMed

    Muppavarapu, Mridula; Huch, Susanne; Nissan, Tracy

    2016-01-01

    Pat1 is a key cytoplasmic mRNA degradation factor, the loss of which severely increases mRNA half-lives. Several recent studies have shown that Pat1 can enter the nucleus and can shuttle between the nucleus and the cytoplasm. As a result, many nuclear roles have been proposed for Pat1. In this study, we analyzed four previously suggested nuclear roles of Pat1 and show that Pat1 is not required for efficient pre-mRNA splicing or pre-mRNA decay in yeast. However, lack of Pat1 results in accumulation of pre-rRNA processing intermediates. Intriguingly, we identified a novel genetic relationship between Pat1 and the rRNA decay machinery, specifically the exosome and the TRAMP complex. While the pre-rRNA processing intermediates that accumulate in the pat1 deletion mutant are, at least to some extent, recognized as aberrant by the rRNA degradation machinery, it is unlikely that these accumulations are the cause of their synthetic sick relationship. Here, we show that the dysregulation of the levels of mRNAs related to ribosome biogenesis could be the cause of the accumulation of the pre-rRNA processing intermediates. Although our results support a role for Pat1 in transcription, they nevertheless suggest that the primary cause of the dysregulated mRNA levels is most likely due to Pat1's role in mRNA decapping and mRNA degradation.

  1. Are we missing a mineralocorticoid in teleost fish? Effects of cortisol, deoxycorticosterone and aldosterone on osmoregulation, gill Na+,K+-ATPase activity and isoform mRNA levels in Atlantic salmon

    USGS Publications Warehouse

    McCormick, S.D.; Regish, A.; O'Dea, M. F.; Shrimpton, J.M.

    2008-01-01

    It has long been held that cortisol, acting through a single receptor, carries out both glucocorticoid and mineralocorticoid actions in teleost fish. The recent finding that fish express a gene with high sequence similarity to the mammalian mineralocorticoid receptor (MR) suggests the possibility that a hormone other than cortisol carries out some mineralocorticoid functions in fish. To test for this possibility, we examined the effect of in vivo cortisol, 11-deoxycorticosterone (DOC) and aldosterone on salinity tolerance, gill Na+,K+-ATPase (NKA) activity and mRNA levels of NKA α1a and α1b in Atlantic salmon. Cortisol treatment for 6–14 days resulted in increased, physiological levels of cortisol, increased gill NKA activity and improved salinity tolerance (lower plasma chloride after a 24 h seawater challenge), whereas DOC and aldosterone had no effect on either NKA activity or salinity tolerance. NKA α1a and α1b mRNA levels, which increase in response to fresh water and seawater acclimation, respectively, were both upregulated by cortisol, whereas DOC and aldosterone were without effect. Cortisol, DOC and aldosterone had no effect on gill glucocorticoid receptor GR1, GR2 and MR mRNA levels, although there was some indication of possible upregulation of GR1 by cortisol (p = 0.07). The putative GR blocker RU486 inhibited cortisol-induced increases in salinity tolerance, NKA activity and NKA α1a and α1b transcription, whereas the putative MR blocker spironolactone had no effect. The results provide support that cortisol, and not DOC or aldosterone, is involved in regulating the mineralocorticoid functions of ion uptake and salt secretion in teleost fish.

  2. A selection of reference genes and early-warning mRNA biomarkers for environmental monitoring using Mytilus spp. as sentinel species.

    PubMed

    Lacroix, C; Coquillé, V; Guyomarch, J; Auffret, M; Moraga, D

    2014-09-15

    mRNA biomarkers are promising tools for environmental health assessment and reference genes are needed to perform relevant qPCR analyses in tissue samples of sentinel species. In the present study, potential reference genes and mRNA biomarkers were tested in the gills and digestive glands of native and caged mussels (Mytilus spp.) exposed to harbor pollution. Results highlighted the difficulty to find stable reference genes in wild, non-model species and suggested the use of normalization indices instead of single genes as they exhibit a higher stability. Several target genes were found differentially expressed between mussel groups, especially in gills where cyp32, π-gst and CuZn-sod mRNA levels could be biomarker candidates. Multivariate analyses confirmed the ability of mRNA levels to highlight site-effects and suggested the use of several combined markers instead of individual ones. These findings support the use of qPCR technology and mRNA levels as early-warning biomarkers in marine monitoring programs. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. Sensitivity of mRNA Translation

    PubMed Central

    Poker, Gilad; Margaliot, Michael; Tuller, Tamir

    2015-01-01

    Using the dynamic mean-field approximation of the totally asymmetric simple exclusion process (TASEP), we investigate the effect of small changes in the initiation, elongation, and termination rates along the mRNA strand on the steady-state protein translation rate. We show that the sensitivity of mRNA translation is equal to the sensitivity of the maximal eigenvalue of a symmetric, nonnegative, tridiagonal, and irreducible matrix. This leads to new analytical results as well as efficient numerical schemes that are applicable for large-scale models. Our results show that in the usual endogenous case, when initiation is more rate-limiting than elongation, the sensitivity of the translation rate to small mutations rapidly increases towards the 5′ end of the ORF. When the initiation rate is high, as may be the case for highly expressed and/or heterologous optimized genes, the maximal sensitivity is with respect to the elongation rates at the middle of the mRNA strand. We also show that the maximal possible effect of a small increase/decrease in any of the rates along the mRNA is an increase/decrease of the same magnitude in the translation rate. These results are in agreement with previous molecular evolutionary and synthetic biology experimental studies. PMID:26238363

  4. Disruption of the Axonal Trafficking of Tyrosine Hydroxylase mRNA Impairs Catecholamine Biosynthesis in the Axons of Sympathetic Neurons

    PubMed Central

    Gioio, Anthony E.

    2017-01-01

    Abstract Tyrosine hydroxylase (TH) is the enzyme that catalyzes the rate-limiting step in the biosynthesis of the catecholamine neurotransmitters. In a previous communication, evidence was provided that TH mRNA is trafficked to the axon, where it is locally translated. In addition, a 50-bp sequence element in the 3′untranslated region (3’UTR) of TH mRNA was identified that directs TH mRNA to distal axons (i.e., zip-code). In the present study, the hypothesis was tested that local translation of TH plays an important role in the biosynthesis of the catecholamine neurotransmitters in the axon and/or presynaptic nerve terminal. Toward this end, a targeted deletion of the axonal transport sequence element was developed, using the lentiviral delivery of the CRISPR/Cas9 system, and two guide RNA (gRNA) sequences flanking the 50-bp cis-acting regulatory element in rat superior cervical ganglion (SCG) neurons. Deletion of the axonal transport element reduced TH mRNA levels in the distal axons and reduced the axonal protein levels of TH and TH activity as measured by phosphorylation of SER40 in SCG neurons. Moreover, deletion of the zip-code diminished the axonal levels of dopamine (DA) and norepinephrine (NE). Conversely, the local translation of exogenous TH mRNA in the distal axon enhanced TH levels and activity, and elevated axonal NE levels. Taken together, these results provide direct evidence to support the hypothesis that TH mRNA trafficking and local synthesis of TH play an important role in the synthesis of catecholamines in the axon and presynaptic terminal. PMID:28630892

  5. [Effect of citric acid stimulation on salivary alpha-amylase, total protein, salivary flow rate and pH value in Pi deficiency children].

    PubMed

    Yang, Ze-min; Chen, Long-hui; Lin, Jing; Zhang, Min; Yang, Xiao-rong; Chen, Wei-wen

    2015-02-01

    To compare the effect of citric acid stimulation on salivary alpha-amylase (sAA), total protein (TP), salivary flow rate, and pH value between Pi deficiency (PD) children and healthy children, thereby providing evidence for Pi controlling saliva theory. Twenty PD children were recruited, and 29 healthy children were also recruited at the same time. Saliva samples from all subjects were collected before and after citric acid stimulation. The sAA activity and amount, TP contents, salivary flow rate, and pH value were determined and compared. (1) Citric acid stimulation was able to significantly increase salivary flow rate, pH value, sAA activities, sAA specific activity and sAA amount (including glycosylated and non-glycosylated sAA amount) in healthy children (P<0.05), while it could markedly increase salivary flow rate, pH value, and glycosylated sAA levels in PD children (P<0.05); (2) Although there was no statistical difference in determined salivary indices between the two groups (P>0.05), salivary indices except salivary flow rate and glycosylated sAA levels decreased more in PD children. There was statistical difference in sAA activity ratio, sAA specific activity ratio, and the ratio of glycosylated sAA levels between PD children and healthy children (P<0.05). PD children had decreased response to citric acid stimulation.

  6. Orotate phosphoribosyl transferase mRNA expression and the response of cholangiocarcinoma to 5-fluorouracil

    PubMed Central

    Hahnvajanawong, Chariya; Chaiyagool, Jariya; Seubwai, Wunchana; Bhudhisawasdi, Vajarabhongsa; Namwat, Nisana; Khuntikeo, Narong; Sripa, Banchob; Pugkhem, Ake; Tassaneeyakul, Wichittra

    2012-01-01

    AIM: To determine whether expression of certain enzymes related to 5-fluorouracil (5-FU) metabolism predicts 5-FU chemosensitivity in cholangiocarcinoma (CCA). METHODS: The histoculture drug response assay (HDRA) was performed using surgically resected CCA tissues. Tumor cell viability was determined morphologically with hematoxylin and eosin- and terminal deoxynucleotide transferase-mediated dUTP nick-end labeling-stained tissues. The mRNA expression of thymidine phosphorylase (TP), orotate phosphoribosyl transferase (OPRT), thymidylate synthase (TS), and dihydropyrimidine dehydrogenase (DPD) was determined with real-time reverse transcriptase-polymerase chain reaction. The levels of gene expression and the sensitivity to 5-FU were evaluated. RESULTS: Twenty-three CCA tissues were obtained from patients who had been diagnosed with intrahepatic CCA and who underwent surgical resection at Srinagarind Hospital, Khon Kaen University from 2007 to 2009. HDRA was used to determine the response of these CCA tissues to 5-FU. Based on the dose-response curve, 200 μg/mL 5-FU was selected as the test concentration. The percentage of inhibition index at the median point was selected as the cut-off point to differentiate the responding and non-responding tumors to 5-FU. When the relationship between TP, OPRT, TS and DPD mRNA expression levels and the sensitivity of CCA tissues to 5-FU was examined, only OPRT mRNA expression was significantly correlated with the response to 5-FU. The mean expression level of OPRT was significantly higher in the responder group compared to the non-responder group (0.41 ± 0.25 vs 0.22 ± 0.12, P < 0.05). CONCLUSION: OPRT mRNA expression may be a useful predictor of 5-FU chemosensitivity of CCA. Whether OPRT mRNA could be used to predict the success of 5-FU chemotherapy in CCA patients requires confirmation in patients. PMID:22912546

  7. Heat Shock Response in Yeast Involves Changes in Both Transcription Rates and mRNA Stabilities

    PubMed Central

    Castells-Roca, Laia; García-Martínez, José; Moreno, Joaquín; Herrero, Enrique; Bellí, Gemma; Pérez-Ortín, José E.

    2011-01-01

    We have analyzed the heat stress response in the yeast Saccharomyces cerevisiae by determining mRNA levels and transcription rates for the whole transcriptome after a shift from 25°C to 37°C. Using an established mathematical algorithm, theoretical mRNA decay rates have also been calculated from the experimental data. We have verified the mathematical predictions for selected genes by determining their mRNA decay rates at different times during heat stress response using the regulatable tetO promoter. This study indicates that the yeast response to heat shock is not only due to changes in transcription rates, but also to changes in the mRNA stabilities. mRNA stability is affected in 62% of the yeast genes and it is particularly important in shaping the mRNA profile of the genes belonging to the environmental stress response. In most cases, changes in transcription rates and mRNA stabilities are homodirectional for both parameters, although some interesting cases of antagonist behavior are found. The statistical analysis of gene targets and sequence motifs within the clusters of genes with similar behaviors shows that both transcriptional and post-transcriptional regulons apparently contribute to the general heat stress response by means of transcriptional factors and RNA binding proteins. PMID:21364882

  8. IGF-1R mRNA expression is increased in obese children.

    PubMed

    Ricco, Rafaela Cristina; Ricco, Rubens Garcia; Queluz, Mariangela Carletti; de Paula, Mariana Teresa Sarti; Atique, Patricia Volpon; Custódio, Rodrigo José; Tourinho Filho, Hugo; Del Roio Liberatori, Raphael; Martinelli, Carlos Eduardo

    2018-04-01

    Obese children are often taller than age-matched subjects. Reports on GH and IGF-I levels in obese individuals are controversial, with normal and reduced GH-IGF-I levels having been reported in this group of patients. Thus, the aim of this study was to analyse insulin-like growth factor type 1 receptor (IGF-IR) mRNA expression in obese children. Forty-seven pre-pubertal children were included in this study: 29 were obese and taller than their target height, and 18 were normal eutrophic controls. Fasting blood samples were collected for IGF-IR mRNA expression in isolated lymphocytes and serum IGF-I, ALS, IGFBP-3, and IGFBP-1 concentration analysis. Relative IGF-IR gene expression (2 -ΔΔCT ) was significantly (P=0.025) higher in obese children (median 1.87) than in controls (1.15). Fourteen of the 29 obese subjects showed 2 -ΔΔCT values greater than or equal to 2, while only 2 individuals in the control group showed values above 2 (P=0.01). Obese children showed significantly (P=0.01) higher IGF-I concentrations than the control group (237ng/ml and 144ng/ml, respectively). Among obese patients, 65.5% had IGF-I values above the 75 percentile of the control group (P=0.02). ALS concentration was significantly (P=0.04) higher in the obese group, while IGFBP-3 levels were similar in obese and control children. IGFBP-1 concentration was lower in obese children, while insulin levels and HOMA-IR index were higher than in controls. The higher IGF-IR mRNA expression observed in obese children, associated with the higher IGF-I and ALS and the lower IGFBP-1 levels, suggest that the higher stature observed in these children may be due to increased IGF-I bioactivity. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Increase of prolactin mRNA in the rat hypothalamus after intracerebroventricular injection of VIP or PACAP.

    PubMed

    Bredow, S; Kacsóh, B; Obál, F; Fang, J; Krueger, J M

    1994-10-17

    Vasoactive intestinal peptide (VIP), the structurally homologous pituitary adenylate cyclase-activating peptide (PACAP) and the pituitary hormone, prolactin (PRL) enhance rapid eye movement sleep (REMS). VIP and PACAP are both inducers of PRL gene expression and release in the pituitary gland. Little is known about PRL regulation in the brain although it is hypothesized that the REMS-promoting activity of i.c.v. administered VIP may be mediated via the activation of cerebral PRL. To test whether VIP or PACAP in fact increase intracerebral mRNA, the peptides (VIP: 30 or 300 pmol; PACAP: 220 pmol) were injected i.c.v. into rats at dark onset. 1 h later, cDNA was synthesized from purified hypothalamic mRNA. Standardized amounts were analysed for PRL using the polymerase chain reaction followed by Southern blotting and hybridization. Compared with beta-actin mRNA levels, both VIP and PACAP increased PRL mRNA levels in a dose-dependent fashion though VIP was more effective on a molar basis. The previously reported alternatively spliced PRL mRNA (lacking exon 4) was not detected. The data support the hypothesis that the REMS-promoting activity of central VIP and PACAP might be mediated by cerebral PRL.

  10. Induction of cysteine-rich motor neuron 1 mRNA expression in vascular endothelial cells.

    PubMed

    Nakashima, Yukiko; Takahashi, Satoru

    2014-08-22

    Cysteine-rich motor neuron 1 (CRIM1) is expressed in vascular endothelial cells and plays a crucial role in angiogenesis. In this study, we investigated the expression of CRIM1 mRNA in human umbilical vein endothelial cells (HUVECs). CRIM1 mRNA levels were not altered in vascular endothelial growth factor (VEGF)-stimulated monolayer HUVECs or in cells in collagen gels without VEGF. In contrast, the expression of CRIM1 mRNA was elevated in VEGF-stimulated cells in collagen gels. The increase in CRIM1 mRNA expression was observed even at 2h when HUVECs did not form tubular structures in collagen gels. Extracellular signal-regulated kinase (Erk) 1/2, Akt and focal adhesion kinase (FAK) were activated by VEGF in HUVECs. The VEGF-induced expression of CRIM1 mRNA was significantly abrogated by PD98059 or PF562271, but was not affected by LY294002. These results demonstrate that CRIM1 is an early response gene in the presence of both angiogenic stimulation (VEGF) and environmental (extracellular matrix) factors, and Erk and FAK might be involved in the upregulation of CRIM1 mRNA expression in vascular endothelial cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  11. Impairment of FOS mRNA stabilization following translation arrest in granulocytes from myelodysplastic syndrome patients.

    PubMed

    Feng, Xiaomin; Shikama, Yayoi; Shichishima, Tsutomu; Noji, Hideyoshi; Ikeda, Kazuhiko; Ogawa, Kazuei; Kimura, Hideo; Takeishi, Yasuchika; Kimura, Junko

    2013-01-01

    Although quantitative and qualitative granulocyte defects have been described in myelodysplastic syndromes (MDS), the underlying molecular basis of granulocyte dysfunction in MDS is largely unknown. We recently found that FOS mRNA elevation under translation-inhibiting stimuli was significantly smaller in granulocytes from MDS patients than in healthy individuals. The aim of this study is to clarify the cause of the impaired FOS induction in MDS. We first examined the mechanisms of FOS mRNA elevation using granulocytes from healthy donors cultured with the translation inhibitor emetine. Emetine increased both transcription and mRNA stability of FOS. p38 MAPK inhibition abolished the emetine-induced increase of FOS transcription but did not affect FOS mRNA stabilization. The binding of an AU-rich element (ARE)-binding protein HuR to FOS mRNA containing an ARE in 3'UTR was increased by emetine, and the knockdown of HuR reduced the FOS mRNA stabilizing effect of emetine. We next compared the emetine-induced transcription and mRNA stabilization of FOS between MDS patients and healthy controls. Increased rates of FOS transcription by emetine were similar in MDS and controls. In the absence of emetine, FOS mRNA decayed to nearly 17% of initial levels in 45 min in both groups. In the presence of emetine, however, 76.7±19.8% of FOS mRNA remained after 45 min in healthy controls, versus 37.9±25.5% in MDS (P<0.01). To our knowledge, this is the first report demonstrating attenuation of stress-induced FOS mRNA stabilization in MDS granulocytes.

  12. Quantifying Temporal Autocorrelations for the Expression of Geobacter species mRNA Gene Transcripts at Variable Ammonium Levels during in situ U(VI) Bioremediation

    NASA Astrophysics Data System (ADS)

    Mouser, P. J.

    2010-12-01

    In order to develop decision-making tools for the prediction and optimization of subsurface bioremediation strategies, we must be able to link the molecular-scale activity of microorganisms involved in remediation processes with biogeochemical processes observed at the field-scale. This requires the ability to quantify changes in the in situ metabolic condition of dominant microbes and associate these changes to fluctuations in nutrient levels throughout the bioremediation process. It also necessitates a need to understand the spatiotemporal variability of the molecular-scale information to develop meaningful parameters and constraint ranges in complex bio-physio-chemical models. The expression of three Geobacter species genes (ammonium transporter (amtB), nitrogen fixation (nifD), and a housekeeping gene (recA)) were tracked at two monitoring locations that differed significantly in ammonium (NH4+) concentrations during a field-scale experiment where acetate was injected into the subsurface to simulate Geobacteraceae in a uranium-contaminated aquifer. Analysis of amtB and nifD mRNA transcript levels indicated that NH4+ was the primary form of fixed nitrogen during bioremediation. Overall expression levels of amtB were on average 8-fold higher at NH4+ concentrations of 300 μM or more than at lower NH4+ levels (average 60 μM). The degree of temporal correlation in Geobacter species mRNA expression levels was calculated at both locations using autocorrelation methods that describe the relationship between sample semi-variance and time lag. At the monitoring location with lower NH4+, a temporal correlation lag of 8 days was observed for both amtB and nifD transcript patterns. At the location where higher NH4+ levels were observed, no discernable temporal correlation lag above the sampling frequency (approximately every 2 days) was observed for amtB or nifD transcript fluctuations. Autocorrelation trends in recA expression levels at both locations indicated that

  13. Serum amyloid protein A concentration in cryopyrin-associated periodic syndromes patients treated with interleukin-1 beta antagonist.

    PubMed

    Pastore, Serena; Paloni, Giulia; Caorsi, Roberta; Ronfani, Luca; Taddio, Andrea; Lepore, Loredana

    2014-01-01

    Cryopyrin-associated periodic syndromes (CAPS) are a group of chronic, relapsing autoinflammatory disorders which may be complicated by systemic AA amyloidosis. The aim of our study was to evaluate serum amyloid protein A (SAA) level in CAPS patients treated with Interleukin-1beta (IL-1β) antagonist and to correlate its level with treatment response. All patients of CAPS Italian Register treated with IL-1β inhibitor were enrolled. SAA levels before starting therapy, and at last visit were evaluated. Patients were then divided in complete responders and partial responders. Twenty-five patients were enrolled. SAA level before starting therapy was increased (median 118.5 mg/L, IQR 96.4-252.8; normal value <6.4 mg/L), while at last visit SAA was significantly reduced (median 4.3 mg/L, IQR 2.3-12.7) (p<0.001). However 12 patients still presented SAA levels beyond normal range, 10/25 patients (40%) showed a complete response to treatment. Conversely, 15 patients presented only a partial response, of which 12 for increased SAA value and 3 for increased CRP value. Patients with partial response had SAA values significantly higher than patients with complete response (median 12.6 mg/L; IQR 8.3-20.0 vs. 2.7 mg/L; IQR 1.6-4.1, p<0.001). Our results confirm the long term efficacy of anti IL-1β treatment in CAPS and the decrease of SAA levels; however 48% of patients still presented SAA elevation despite treatment. The real risk of these patients in developing amyloidosis is not clear but the persistent increase of SAA needs a close follow-up.

  14. Promoter methylation, mRNA expression of goat tumor‑associated genes and mRNA expression of DNA methyltransferase in enzootic nasal tumors.

    PubMed

    Quan, Zifang; Ye, Ni; Hao, Zhongxiang; Wen, Caifang; Liao, Hong; Zhang, Manli; Luo, Lu; Cao, Sanjie; Wen, Xintian; Wu, Rui; Yan, Qigui

    2015-10-01

    The aim of the present study was to investigate the promoter methylation status and mRNA expression of goat tumor‑associated genes, in addition to the mRNA expression of DNA methyltransferase genes in enzootic nasal tumors (ENT). Methylation‑specific polymerase chain reaction and SYBR Green reverse transcription‑quantitative polymerase chain reaction were used to detect the methylation status and the mRNA expression levels of DNA methyltransferases (DNMTs), O6‑methylguanine‑DNA methyltransferase (MGMT), the tumor suppressor genes P73, P53, GADD45G, CHFR and THBS1, the transcription factor CEBPA, the proto‑oncogenes KRAS, NRAS and C‑myc and EGFR in 24 nasal tumor tissue samples and 20 normal nasal epithelia tissue samples. The associations between promoter methylation and DNMT, and promoter methylation and mRNA expression of the genes were analyzed. The results indicated that the expression levels of DNMT1 increased by 56% compared with those in normal nasal epithelial tissues, while MGMT, DNMT3a and DNMT3b had similar expression levels in the two tissue types. The expression levels of P53 decreased by 36.8% and those of THBS1 by 43%, while C‑myc increased by 2.9‑fold and CEBPA by 2‑fold compared with that in normal nasal epithelial tissues. GADD45G, P73, CHFR and NRAS were observed to have similar expression levels in the two tissue types. However, no expression was observed for EGFR and KRAS. CHFR, GADD45G and THBS1 were identified to be methylated in tumor suppressor genes. The methylation expression rate of the CHFR gene was ~60% in the two tissue types and for THBS1 it was 100% in the nasal tumor tissues as opposed to 20% in the normal nasal epithelial tissues. The exhaustive methylation expression rate of GADD45G was 62.5% and the partial methylation expression rate was 37.5% in nasal tumor tissue, while no methylation was observed in normal nasal epithelial tissues. C‑myc was the only gene identified to be methylated amongst proto

  15. Generation of human induced pluripotent stem cells using non-synthetic mRNA.

    PubMed

    Rohani, L; Fabian, C; Holland, H; Naaldijk, Y; Dressel, R; Löffler-Wirth, H; Binder, H; Arnold, A; Stolzing, A

    2016-05-01

    Here we describe some of the crucial steps to generate induced pluripotent stem cells (iPSCs) using mRNA transfection. Our approach uses a V. virus-derived capping enzyme instead of a cap-analog, ensuring 100% proper cap orientation for in vitro transcribed mRNA. V. virus' 2'-O-Methyltransferase enzyme creates a cap1 structure found in higher eukaryotes and has higher translation efficiency compared to other methods. Use of the polymeric transfection reagent polyethylenimine proved superior to other transfection methods. The mRNA created via this method did not trigger an intracellular immune response via human IFN-gamma (hIFN-γ) or alpha (hIFN-α) release, thus circumventing the use of suppressors. Resulting mRNA and protein were expressed at high levels for over 48h, thus obviating daily transfections. Using this method, we demonstrated swift activation of pluripotency associated genes in human fibroblasts. Low oxygen conditions further facilitated colony formation. Differentiation into different germ layers was confirmed via teratoma assay. Reprogramming with non-synthetic mRNA holds great promise for safe generation of iPSCs of human origin. Using the protocols described herein we hope to make this method more accessible to other groups as a fast, inexpensive, and non-viral reprogramming approach. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  16. Ammonia exposure affects the mRNA and protein expression levels of certain Rhesus glycoproteins in the gills of climbing perch.

    PubMed

    Chen, Xiu L; Zhang, Biyan; Chng, You R; Ong, Jasmine L Y; Chew, Shit F; Wong, Wai P; Lam, Siew H; Nakada, Tsutomu; Ip, Yuen K

    2017-08-15

    The freshwater climbing perch, Anabas testudineus , is an obligate air-breathing and euryhaline teleost capable of active ammonia excretion and tolerant of high concentrations of environmental ammonia. As Rhesus glycoproteins (RhGP/Rhgp) are known to transport ammonia, this study aimed to obtain the complete cDNA coding sequences of various rhgp isoforms from the gills of A. testudineus , and to determine their mRNA and protein expression levels during 6 days of exposure to 100 mmol l -1 NH 4 Cl. The subcellular localization of Rhgp isoforms in the branchial epithelium was also examined in order to elucidate the type of ionocyte involved in active ammonia excretion. Four rhgp ( rhag , rhbg , rhcg1 and rhcg2 ) had been identified from the gills of A. testudineus They had conserved amino acid residues for NH 4 + binding, NH 4 + deprotonation, channel gating and lining of the vestibules. Despite inwardly directed NH 3 and NH 4 + gradients, there were significant increases in the mRNA expression levels of the four branchial rhgp in A. testudineus at certain time points during 6 days of ammonia exposure, with significant increases in the protein abundances of Rhag and Rhcg2 on day 6. Immunofluorescence microscopy revealed a type of ammonia-inducible Na + /K + -ATPase α1c-immunoreactive ionocyte with apical Rhag and basolateral Rhcg2 in the gills of fish exposed to ammonia for 6 days. Hence, active ammonia excretion may involve NH 4 + entering the ionocyte through the basolateral Rhcg2 and being excreted through the apical Rhag, driven by a transapical membrane electrical potential generated by the apical cystic fibrosis transmembrane conductance regulator Cl - channel, as suggested previously. © 2017. Published by The Company of Biologists Ltd.

  17. Increased Expression of Interleukin-18 mRNA is Associated with Carotid Artery Stenosis

    PubMed

    Arapi, Berk; Bayoğlu, Burcu; Cengiz, Müjgan; Dirican, Ahmet; Deser, Serkan Burç; Junusbekov, Yerik; Arslan, Caner

    2018-05-29

    Carotid artery stenosis is the atherosclerotic narrowing of the proximal internal carotid artery and one of the primary causes of stroke. Elevated expression of the pleiotropic proinflammatory cytokine interleukin-18 has been demonstrated in human atherosclerotic plaques. To investigate whether the mRNA expression levels of interleukin-18 and interleukin-18-binding protein and interleukin-18 −137 G/C (rs187238) variants are associated with carotid artery stenosis development. Case-control study. The mRNA expression levels of interleukin-18 and interleukin-18-binding protein and interleukin-18 rs187238 variants were evaluated by quantitative real-time polymerase chain reaction and real-time polymerase chain reaction, respectively, in the peripheral blood mononuclear cells of 70 patients with carotid artery stenosis (36 symptomatic, 34 asymptomatic) and 75 healthy controls. Interleukin-18 mRNA expression was significantly increased in carotid artery stenosis patients compared to that in healthy controls (p=0.01). However, no significant difference was observed between interleukin-18-binding protein mRNA expression levels in patients with carotid artery stenosis and those in controls (p=0.101). Internal carotid artery stenosis severity was significantly higher in symptomatic patients than that in asymptomatic patients (p<0.001). A significant relationship was identified between interleukin-18 expression and internal carotid artery stenosis severity in patients with carotid artery stenosis (p=0.051). Interleukin-18 rs187238 polymorphism genotype frequencies did not significantly differ between patients with carotid artery stenosis and controls (p=0.246). A significant difference was identified between interleukin-18-binding protein gene expression and symptomatic and asymptomatic patients (p=0.026), but there was no difference in interleukin-18 expression between the symptomatic and asymptomatic subgroups (p=0.397). Interleukin-18 mRNA expression may affect carotid

  18. Changes in mRNA expression for gluconeogenic enzymes in liver of dairy cattle during the transition to lactation.

    PubMed

    Greenfield, R B; Cecava, M J; Donkin, S S

    2000-06-01

    The objective of this study was to profile phosphoenolpyruvate carboxykinase (PEPCK) and pyruvate carboxylase (PC) mRNA expression in the liver of dairy cattle during the peripartum transition and determine changes in abundance of these mRNA in response to protein fed during the prepartum period. Thirty-eight multiparous Holstein cows were fed diets containing either 12% crude protein (CP) and 26% rumen undegradable protein (RUP), 16% CP and 26% RUP, 16% CP and 33% RUP, or 16% CP and 40% RUP on a dry-matter basis beginning 28 d before expected calving. After calving, all cows were fed a common diet through 56 d in milk (DIM). Northern analysis of RNA from liver biopsy samples obtained on days -28, -14, +1, +28, and +56 relative to calving indicated that PC and PEPCK mRNA expression were responsive to onset of lactation but not to prepartum protein or RUP concentration. Abundance of PEPCK mRNA was similar at -28, -14, and +1 DIM but was elevated by +28 and +56 DIM relative to precalving levels. Liver PC mRNA abundance was elevated on +1 DIM, remained elevated through 28 DIM, and declined to precalving levels by 56 DIM. The activity of PC enzyme was correlated (r2 = 0.89) with PC mRNA abundance. The data demonstrate increased abundance of PC mRNA during the early transition period followed by increased abundance of PEPCK mRNA during the postpartum period and suggest increased potential metabolism of lactate, pyruvate, and amino acids that contribute to the liver pyruvate pool.

  19. Time- and dose-dependent differential regulation of copper-zinc superoxide dismutase and manganese superoxide dismutase enzymatic activity and mRNA level by vitamin E in rat blood cells.

    PubMed

    Hajiani, Maliheh; Razi, Farideh; Golestani, Aboualfazl; Frouzandeh, Mehdi; Owji, Ali Akbar; Khaghani, Shahnaz; Ghannadian, Naghmeh; Shariftabrizi, Ahmad; Pasalar, Parvin

    2012-01-01

    Vitamin E is the most important lipid-soluble antioxidant. Recently, it has been proposed as a gene regulator, and its gene modulation effects have been observed at different levels of gene expression and cell signaling. This study was performed to investigate the effects of vitamin E on the activity and expression of the most important endogenous antioxidant enzyme, superoxide dismutase (SOD), in rat plasma. Twenty-eight male Sprauge-Dawley rats were divided into four groups: control group and three dosing groups. The control group received the vehicle (liquid paraffin), and the dosing groups received twice-weekly intraperitoneal injections of 10, 30, and 100 mg/kg of vitamin E ((±)-α-Tocopherol) for 6 weeks. Quantitative real-time reverse transcription-polymerase chain reaction and enzyme assays were used to assess the levels of Cu/Zn-SOD and Mn-SOD mRNA and enzyme activity levels in blood cells at 0, 2, 4, and 6 weeks following vitamin E administration. Catalase enzyme activity and total antioxidant capacity were also assessed in plasma at the same time intervals. Mn-SOD activity was significantly increased in the 100 and 30 mg/kg dosing groups after 4 and 6 weeks, with corresponding significant increase in their mRNA levels. Cu/Zn-SOD activity was not significantly changed in response to vitamin E administration at any time points, whereas Cu/Zn-SOD mRNA levels were significantly increased after longer time points with high doses (30 and 100 mg/kg) of vitamin E. Catalase enzyme activity was transiently but significantly increased after 4 weeks of vitamin E treatment in 30 and 100 mg/kg dosing groups. Total antioxidant status was significantly increased after 4 and 6 weeks in the 100 mg/kg dosing group. Only the chronic administration of higher doses of alpha-tocopherol is associated with the increased activity and expression of Mn-SOD in rats. Cu/Zn-SOD activity and expression does not dramatically change in response to vitamin E.

  20. Expression of CD30 mRNA, CD30L mRNA and a variant form of CD30 mRNA in restimulated peripheral blood mononuclear cells (PBMC) of patients with helminthic infections resembling a Th2 disease

    PubMed Central

    Kilwinski, J; Berger, T; Mpalaskas, J; Reuter, S; Flick, W; Kern, P

    1999-01-01

    It has been proposed that CD30, a member of the tumour necrosis factor (TNF) receptor superfamily, is preferentially up-regulated on Th2-type human T cells. In order to investigate a correlation between infection with Echinococcus multilocularis and CD30 expression, we analysed regulation of CD30 mRNA, a variant form of CD30 mRNA (CD30v) and CD30 ligand (CD30L) mRNA expression on PBMC from patients with alveolar echinococcosis (AE) using reverse transcriptase-polymerase chain reaction (RT-PCR). In PBMC of patients with AE as well as healthy donors, spontaneous expression of CD30L mRNA and the CD30v mRNA could be detected. However, the intact form of CD30 mRNA could be detected neither in freshly isolated PBMC of patients nor in PBMC of healthy individuals. Expression of CD30L mRNA and the variant form of CD30 mRNA was frequently detected at individual time points during 72 h of culture of PBMC stimulated with crude Echinococcus antigen. In contrast to CD30v or CD30L mRNA expression, induction of CD30 mRNA expression was detected only in three out of six (50%) healthy donors and in 10 out of 21 (48%) patients with alveolar echinococcosis after 72 h of incubation. As a control, mitogenic stimulation of PBMC of both healthy individuals and infected patients led to expression of intact CD30 mRNA within 24 h of culture. These data demonstrate the different expression of two different forms of CD30 mRNA in PBMC of human individuals. The specific induction of CD30 expression is correlated only in rare cases with the clinical status of patients with AE, indicating the lack of a general induction of CD30 mRNA in this Th2-type-dominated helminthic disease. The data provide further evidence that the CD30 receptor is not an exclusive marker for a Th2-type response. PMID:9933429

  1. Urinary podocyte-associated mRNA levels correlate with proximal tubule dysfunction in early diabetic nephropathy of type 2 diabetes mellitus.

    PubMed

    Petrica, Ligia; Ursoniu, Sorin; Gadalean, Florica; Vlad, Adrian; Gluhovschi, Gheorghe; Dumitrascu, Victor; Vlad, Daliborca; Gluhovschi, Cristina; Velciov, Silvia; Bob, Flaviu; Matusz, Petru; Milas, Oana; Secara, Alina; Simulescu, Anca; Popescu, Roxana

    2017-01-01

    The study assessed mRNA expression of podocyte-associated molecules in urinary sediments of patients with type 2 diabetes mellitus (DM) in relation to urinary podocytes, biomarkers of podocyte injury and of proximal tubule (PT) dysfunction. A total of 76 patients with type 2 DM and 20 healthy subjects were enrolled in a cross-sectional study, and assessed concerning urinary podocytes, urinary mRNA of podocyte-associated genes, urinary biomarkers of podocyte damage and of PT dysfunction. We found significant differences between urinary mRNA of podocyte-associated molecules in relation with albuminuria stage. In multivariable regression analysis, urinary mRNA of nephrin, podocin, alpha-actinin-4, CD2-associated protein, glomerular epithelial protein 1 (GLEPP1), ADAM 10, and NFκB correlated directly with urinary podocytes, albuminuria, urinary alpha 1 -microglobulin, urinary kidney-injury molecule-1, nephrinuria, urinary vascular endothelial growth factor, urinary advanced glycation end-products (AGE), and indirectly with eGFR (p < 0.0001, R 2  = 0.808; p < 0.0001, R 2  = 0.825; p < 0.0001, R 2  = 0.805; p < 0.0001, R 2  = 0.663; p < 0.0001, R 2  = 0.726; p < 0.0001, R 2  = 0.720; p < 0.0001, R 2  = 0.724). In patients with type 2 DM there is an association between urinary mRNA of podocyte-associated molecules, biomarkers of podocyte damage, and of PT dysfunction. GLEPP1, ADAM10, and NFκB may be considered additional candidate molecules indicative of early diabetic nephropathy. AGE could be involved in this association.

  2. Characterization of rat calcitonin mRNA.

    PubMed Central

    Amara, S G; David, D N; Rosenfeld, M G; Roos, B A; Evans, R M

    1980-01-01

    A chimeric plasmic containing cDNA complementary to rat calcitonin mRNA has been constructed. Partial sequence analysis shows that the insert contains a nucleotide sequence encoding the complete amino acid sequence of calcitonin. Two basic amino acids precede and three basic amino acids follow the hormone sequence, suggesting that calcitonin is generated by the proteolytic cleavage of a larger precursor in a manner analogous to that of other small polypeptide hormones. The COOH-terminal proline, known to be amidated in the secreted hormone, is followed by a glycine in the precursor. The cloned calcitonin DNA was used to characterize the expression of calcitonin mRNA. Cytoplasmic mRNAs from calcitonin-producing rat medullary thyroid carcinoma lines and from normal rat thyroid glands contain a single species, 1050 nucleotides long, whch hybridizes to the cloned calcitonin cDNA. The concentration of calcitonin mRNA sequences is greater in those tumors that produce larger amounts of immunoreactive calcitonin. RNAs from other endocrine tissues, including anterior and neurointermediate lobes of rat pituitary, contain no detectable calcitonin mRNA. Images PMID:6933496

  3. mRNA Cap Methyltransferase, RNMT-RAM, Promotes RNA Pol II-Dependent Transcription.

    PubMed

    Varshney, Dhaval; Lombardi, Olivia; Schweikert, Gabriele; Dunn, Sianadh; Suska, Olga; Cowling, Victoria H

    2018-05-01

    mRNA cap addition occurs early during RNA Pol II-dependent transcription, facilitating pre-mRNA processing and translation. We report that the mammalian mRNA cap methyltransferase, RNMT-RAM, promotes RNA Pol II transcription independent of mRNA capping and translation. In cells, sublethal suppression of RNMT-RAM reduces RNA Pol II occupancy, net mRNA synthesis, and pre-mRNA levels. Conversely, expression of RNMT-RAM increases transcription independent of cap methyltransferase activity. In isolated nuclei, recombinant RNMT-RAM stimulates transcriptional output; this requires the RAM RNA binding domain. RNMT-RAM interacts with nascent transcripts along their entire length and with transcription-associated factors including the RNA Pol II subunits SPT4, SPT6, and PAFc. Suppression of RNMT-RAM inhibits transcriptional markers including histone H2BK120 ubiquitination, H3K4 and H3K36 methylation, RNA Pol II CTD S5 and S2 phosphorylation, and PAFc recruitment. These findings suggest that multiple interactions among RNMT-RAM, RNA Pol II factors, and RNA along the transcription unit stimulate transcription. Copyright © 2018 The Author(s). Published by Elsevier Inc. All rights reserved.

  4. A small RNA activates CFA synthase by isoform-specific mRNA stabilization

    PubMed Central

    Fröhlich, Kathrin Sophie; Papenfort, Kai; Fekete, Agnes; Vogel, Jörg

    2013-01-01

    Small RNAs use a diversity of well-characterized mechanisms to repress mRNAs, but how they activate gene expression at the mRNA level remains not well understood. The predominant activation mechanism of Hfq-associated small RNAs has been translational control whereby base pairing with the target prevents the formation of an intrinsic inhibitory structure in the mRNA and promotes translation initiation. Here, we report a translation-independent mechanism whereby the small RNA RydC selectively activates the longer of two isoforms of cfa mRNA (encoding cyclopropane fatty acid synthase) in Salmonella enterica. Target activation is achieved through seed pairing of the pseudoknot-exposed, conserved 5′ end of RydC to an upstream region of the cfa mRNA. The seed pairing stabilizes the messenger, likely by interfering directly with RNase E-mediated decay in the 5′ untranslated region. Intriguingly, this mechanism is generic such that the activation is equally achieved by seed pairing of unrelated small RNAs, suggesting that this mechanism may be utilized in the design of RNA-controlled synthetic circuits. Physiologically, RydC is the first small RNA known to regulate membrane stability. PMID:24141880

  5. A small RNA activates CFA synthase by isoform-specific mRNA stabilization.

    PubMed

    Fröhlich, Kathrin Sophie; Papenfort, Kai; Fekete, Agnes; Vogel, Jörg

    2013-11-13

    Small RNAs use a diversity of well-characterized mechanisms to repress mRNAs, but how they activate gene expression at the mRNA level remains not well understood. The predominant activation mechanism of Hfq-associated small RNAs has been translational control whereby base pairing with the target prevents the formation of an intrinsic inhibitory structure in the mRNA and promotes translation initiation. Here, we report a translation-independent mechanism whereby the small RNA RydC selectively activates the longer of two isoforms of cfa mRNA (encoding cyclopropane fatty acid synthase) in Salmonella enterica. Target activation is achieved through seed pairing of the pseudoknot-exposed, conserved 5' end of RydC to an upstream region of the cfa mRNA. The seed pairing stabilizes the messenger, likely by interfering directly with RNase E-mediated decay in the 5' untranslated region. Intriguingly, this mechanism is generic such that the activation is equally achieved by seed pairing of unrelated small RNAs, suggesting that this mechanism may be utilized in the design of RNA-controlled synthetic circuits. Physiologically, RydC is the first small RNA known to regulate membrane stability.

  6. Stability of the Osmoregulated Promoter-Derived proP mRNA Is Posttranscriptionally Regulated by RNase III in Escherichia coli

    PubMed Central

    Lim, Boram

    2015-01-01

    ABSTRACT The enzymatic activity of Escherichia coli endo-RNase III determines the stability of a subgroup of mRNA species, including bdm, betT, and proU, whose protein products are associated with the cellular response to osmotic stress. Here, we report that the stability of proP mRNA, which encodes a transporter of osmoprotectants, is controlled by RNase III in response to osmotic stress. We observed that steady-state levels of proP mRNA and ProP protein are inversely correlated with cellular RNase III activity and, in turn, affect the proline uptake capacity of the cell. In vitro and in vivo analyses of proP mRNA revealed RNase III cleavage sites in a stem-loop within the 5′ untranslated region present only in proP mRNA species synthesized from the osmoregulated P1 promoter. Introduction of nucleotide substitutions in the cleavage site identified inhibited the ribonucleolytic activity of RNase III on proP mRNA, increasing the steady-state levels and half-life of the mRNA. In addition, decreased RNase III activity coincided with a significant increase in both the half-life and abundance of proP mRNA under hyperosmotic stress conditions. Analysis of the RNA bound to RNase III via in vivo cross-linking and immunoprecipitation indicated that this phenomenon is related to the decreased RNA binding capacity of RNase III. Our findings suggest the existence of an RNase III-mediated osmoregulatory network that rapidly balances the expression levels of factors associated with the cellular response to osmotic stress in E. coli. IMPORTANCE Our results demonstrate that RNase III activity on proP mRNA degradation is downregulated in Escherichia coli cells under osmotic stress. In addition, we show that the downregulation of RNase III activity is associated with decreased RNA binding capacity of RNase III under hyperosmotic conditions. In particular, our findings demonstrate a link between osmotic stress and RNase III activity, underscoring the growing importance of

  7. Rapid regulation of brain-derived neurotrophic factor mRNA within eye-specific circuits during ocular dominance column formation.

    PubMed

    Lein, E S; Shatz, C J

    2000-02-15

    The neurotrophin brain-derived neurotrophic factor (BDNF) has emerged as a candidate retrograde signaling molecule for geniculocortical axons during the formation of ocular dominance columns. Here we examined whether neuronal activity can regulate BDNF mRNA in eye-specific circuits in the developing cat visual system. Dark-rearing throughout the critical period for ocular dominance column formation decreases levels of BDNF mRNA within primary visual cortex, whereas short-term (2 d) binocular blockade of retinal activity with tetrodotoxin (TTX) downregulates BDNF mRNA within the lateral geniculate nucleus (LGN) and visual cortical areas. Brief (6 hr to 2 d) monocular TTX blockade during the critical period and also in adulthood causes downregulation in appropriate eye-specific laminae in the LGN and ocular dominance columns within primary visual cortex. Monocular TTX blockade at postnatal day 23 also downregulates BDNF mRNA in a periodic fashion, consistent with recent observations that ocular dominance columns can be detected at these early ages by physiological methods. In contrast, 10 d monocular TTX during the critical period does not cause a lasting decrease in BDNF mRNA expression in columns pertaining to the treated eye, consistent with the nearly complete shift in physiological response properties of cortical neurons in favor of the unmanipulated eye known to result from long-term monocular deprivation. These observations demonstrate that BDNF mRNA levels can provide an accurate "molecular readout" of the activity levels of cortical neurons and are consistent with a highly local action of BDNF in strengthening and maintaining active synapses during ocular dominance column formation.

  8. Effects of hypothermia and cerebral ischemia on cold-inducible RNA-binding protein mRNA expression in rat brain.

    PubMed

    Liu, Aijun; Zhang, Zhiwen; Li, Anmin; Xue, Jinghui

    2010-08-06

    CIRP (cold-inducible RNA-binding protein) mRNA is highly expressed in hypothermic conditions in mammalian cells, and the relationship between CIRP and neuroprotection for cerebral ischemia under hypothermia has been focused upon. At present, however, the expression characteristics of CIRP under hypothermia and cerebral ischemia in vivo are not clearly elucidated. In this study, CIRP mRNA expression in various regions of rat brain was examined by reverse transcriptase polymerase chain reaction (RT-PCR). CIRP expression levels were found to be similar in the hippocampus and cortex. Real-time quantitative PCR analysis revealed increasing CIRP mRNA expression in the cortex during the 24-h observation period following treatment with hypothermia or cerebral ischemia, with a greater increase in the hypothermia group. When cerebral ischemia was induced following hypothermia, CIRP mRNA expression in the cortex again showed a significant increasing tendency, but ischemia delayed the appearance of this increase. To reveal the relationship between CIRP and energy metabolism in the rat brain, lactate and pyruvate concentrations in the cortex of the rats treated with hypothermia, ischemia and ischemia after hypothermia were determined by spectrophotometric assay, and levels of phosphofructokinas-1 (PFK-1), the major regulatory enzyme of the glycolytic pathway, in the rat cortex in the three groups was also analyzed by Western blot. Using linear correlation, lactate and pyruvate concentrations, and PFK-1 levels, were each analyzed in the three groups in association with CIRP mRNA expression levels. The analysis did not reveal any correlation between the three metabolic parameters and CIRP mRNA expression induced by hypothermia, suggesting that while playing a role in neuroprotection under hypothermia, CIRP does not affect cerebral energy metabolism. Copyright 2010. Published by Elsevier B.V.

  9. Validation of serum amyloid α as an independent biomarker for progression-free and overall survival in metastatic renal cell cancer patients.

    PubMed

    Vermaat, Joost S; Gerritse, Frank L; van der Veldt, Astrid A; Roessingh, Wijnand M; Niers, Tatjana M; Oosting, Sjoukje F; Sleijfer, Stefan; Roodhart, Jeanine M; Beijnen, Jos H; Schellens, Jan H; Gietema, Jourik A; Boven, Epie; Richel, Dick J; Haanen, John B; Voest, Emile E

    2012-10-01

    We recently identified apolipoprotein A2 (ApoA2) and serum amyloid α (SAA) as independent prognosticators in metastatic renal cell carcinoma (mRCC) patients, thereby improving the accuracy of the Memorial-Sloan Kettering Cancer Center (MSKCC) model. Validate these results prospectively in a separate cohort of mRCC patients treated with tyrosine kinase inhibitors (TKIs). For training we used 114 interferon-treated mRCC patients (inclusion 2001-2006). For validation we studied 151 TKI-treated mRCC patients (inclusion 2003-2009). Using Cox proportional hazards regression analysis, SAA and ApoA2 were associated with progression-free survival (PFS) and overall survival (OS). In 72 TKI-treated patients, SAA levels were analyzed longitudinally as a potential early marker for treatment effect. Baseline ApoA2 and SAA levels significantly predicted PFS and OS in the training and validation cohorts. Multivariate analysis identified SAA in both separate patient sets as a robust and independent prognosticator for PFS and OS. In contrast to our previous findings, ApoA2 interacted with SAA in the validation cohort and did not contribute to a better predictive accuracy than SAA alone and was therefore excluded from further analysis. According to the tertiles of SAA levels, patients were categorized in three risk groups, demonstrating accurate risk prognostication. SAA as a single biomarker showed equal prognostic accuracy when compared with the multifactorial MSKCC risk mode. Using receiver operating characteristic analysis, SAA levels >71 ng/ml were designated as the optimal cut-off value in the training cohort, which was confirmed for its significant sensitivity and specificity in the validation cohort. Applying SAA >71 ng/ml as an additional risk factor significantly improved the predictive accuracy of the MSKCC model in both independent cohorts. Changes in SAA levels after 6-8 wk of TKI treatment had no value in predicting treatment outcome. SAA but not ApoA2 was shown to be

  10. Identification of a Serum amyloid A gene and the association of SNPs with Vibrio-resistance and growth traits in the clam Meretrix meretrix.

    PubMed

    Zou, Linhu; Liu, Baozhong

    2015-04-01

    Serum amyloid A (SAA), an acute response protein as well as an apolipoprotein, is considered to play crucial roles in both innate immunity and lipid metabolism. In this study, a SAA gene (MmSAA) was identified in the clam Meretrix meretrix. The full length DNA of MmSAA was 1407bp, consisting of three exons and two introns. The distribution of MmSAA in clam tissues was examined with the highest expression in hepatopancreas. In response to the Vibrio parahaemolyticus challenge, MmSAA mRNA showed significantly higher expression at 24 h post-challenge in experimental clams (P < 0.05). Forty-eight single nucleotide polymorphisms (SNPs) in the DNA partial sequence of MmSAA were discovered and examined for their association with Vibrio-resistance and growth traits, respectively. The single SNP association analysis indicated that five single SNPs (g.42, g.72, g.82, g.147 and g.165) were significantly associated with Vibrio-resistance (P < 0.05). Haplotype analysis produced additional support for association with the Chi-square values 6.393 (P = 0.012). Among the five selected SNPs, the effect of a missense mutation (g.82, A → G) was detected by site-directed mutagenesis with fusion expression of protein assay, and the result showed that the recombinant plasmids containing wild-type pET30a-MmSAA had more inhibition effect than the mutant ones on the growth rate of the host bacteria. In addition, four growth traits of the clams in 09G3SPSB population were recorded and the SNP g.176 was found to be significantly associated with the growth traits with the Global score value 0.790 (P = 0.015). Our findings suggested that common genetic variation in MmSAA might contribute to the risk of susceptibility to Vibrio infection and might be associated with the growth traits in the clams M. meretrix, and more works are still needed to validate these SNPs as potential markers for actual selective breeding. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. mRNA trans-splicing in gene therapy for genetic diseases.

    PubMed

    Berger, Adeline; Maire, Séverine; Gaillard, Marie-Claude; Sahel, José-Alain; Hantraye, Philippe; Bemelmans, Alexis-Pierre

    2016-07-01

    Spliceosome-mediated RNA trans-splicing, or SMaRT, is a promising strategy to design innovative gene therapy solutions for currently intractable genetic diseases. SMaRT relies on the correction of mutations at the post-transcriptional level by modifying the mRNA sequence. To achieve this, an exogenous RNA is introduced into the target cell, usually by means of gene transfer, to induce a splice event in trans between the exogenous RNA and the target endogenous pre-mRNA. This produces a chimeric mRNA composed partly of exons of the latter, and partly of exons of the former, encoding a sequence free of mutations. The principal challenge of SMaRT technology is to achieve a reaction as complete as possible, i.e., resulting in 100% repairing of the endogenous mRNA target. The proof of concept of SMaRT feasibility has already been established in several models of genetic diseases caused by recessive mutations. In such cases, in fact, the repair of only a portion of the mutant mRNA pool may be sufficient to obtain a significant therapeutic effect. However in the case of dominant mutations, the target cell must be freed from the majority of mutant mRNA copies, requiring a highly efficient trans-splicing reaction. This likely explains why only a few examples of SMaRT approaches targeting dominant mutations are reported in the literature. In this review, we explain in details the mechanism of trans-splicing, review the different strategies that are under evaluation to lead to efficient trans-splicing, and discuss the advantages and limitations of SMaRT. WIREs RNA 2016, 7:487-498. doi: 10.1002/wrna.1347 For further resources related to this article, please visit the WIREs website. © 2016 The Authors. WIREs RNA published by Wiley Periodicals, Inc.

  12. Nuclear receptor and target gene mRNA abundance in duodenum and colon of dogs with chronic enteropathies.

    PubMed

    Greger, D L; Gropp, F; Morel, C; Sauter, S; Blum, J W

    2006-11-01

    Nuclear receptors (NR), such as constitutive androstane receptor (CAR), pregnane X receptor (PXR) and peroxisome proliferator-associated receptors alpha and gamma (PPARalpha, PPARgamma) are mediators of inflammation and may be involved in inflammatory bowel disease (IBD) and food responsive diarrhea (FRD) of dogs. The present study compared mRNA abundance of NR and NR target genes [multi drug-resistance gene-1 (MDR1), multiple drug-resistance-associated proteins (MRD2, MRD3), cytochrome P450 (CYP3A12), phenol-sulfating phenol sulfotransferase (SULT1A1) and glutathione-S-transferase (GST A3-3)] in biopsies obtained from duodenum and colon of dogs with IBD and FRD and healthy control dogs (CON; n=7 per group). Upon first presentation of dogs, mRNA levels of PPARalpha, PPARgamma, CAR, PXR and RXRalpha in duodenum as well as PPARgamma, CAR, PXR and RXRalpha in colon were not different among groups (P>0.10). Although mRNA abundance of PPARalpha in colon of dogs with FRD was similar in both IBD and CON (P>0.10), PPARalpha mRNA abundance was higher in IBD than CON (P<0.05). Levels of mRNA of MDR1 in duodenum were higher in FRD than IBD (P<0.05) or CON (P<0.001). Compared with CON, abundances of mRNA for MRP2, CYP3A12 and SULT1A1 were higher in both FRD and IBD than CON (P<0.05). Differences in mRNA levels of PPARalpha and MRP2 in colon and MDR1, MRP2, CYP3A12 and SULT1A1 in duodenum may be indicative for enteropathy in FRD and (or) IBD dogs relative to healthy dogs. More importantly, increased expression of MDR1 in FRD relative to IBD in duodenum may be a useful diagnostic marker to distinguish dogs with FRD from dogs with IBD.

  13. Correlation between expressions of ERCC1/TS mRNA and effects of gastric cancer to chemotherapy in the short term.

    PubMed

    Chen, Liqi; Li, Guoli; Li, Jieshou; Fan, Chaogang; Xu, Jian; Wu, Bo; Liu, Kun; Zhang, Caihua

    2013-04-01

    To study the correlation between expression levels of ERCC1/TS mRNA and the susceptibility of preoperative chemotherapy for patients with gastric cancer. A total of forty cases with advanced gastric cancer of T3-4N1-2M0 were treated with preoperative chemotherapy according to FLEEOX regimen based on endarterial-intravenous coadministration. Sufficient, fresh gastric tissue specimens were obtained with the help of gastroscope, and the expression levels of ERCC1/TS mRNA were detected by qRT-PCR before chemotherapy. The chemotherapeutic response was evaluated with Choi Criteria after chemotherapy, and pathologic remission extent was observed after surgery. The correlation between the expression levels of ERCC1/TS mRNA before chemotherapy and the chemotherapeutic effect based on imageology and pathology was analyzed. The response rate of Chemotherapy in this cohort was 80.0 % based on imageology and 51.43 % based on pathology. The expression levels of ERCC1/TS mRNA were significantly associated with imageology remission extent (P = 0.033, P = 0.025) and pathologic remission extent (P = 0.044, P = 0.016), respectively. The chemotherapeutic effect on patients with low-expression levels of ERCC1/TS mRNA was better. From the perspective of pathology and imageology evaluating the preoperative chemotherapeutic response for patients with gastric cancer, ERCC1 and TS were used as the molecular predictors and provided prognostic information in this study.

  14. PKA- and PKC-dependent regulation of angiopoietin 2 mRNA in human granulosa lutein cells.

    PubMed

    Witt, P S; Pietrowski, D; Keck, C

    2004-02-01

    New blood vessels develop from preexisting vessels in response to growth factors or hypoxic conditions. Recent studies have shown that angiopoietin 2 (ANGPT-2) plays an important role in the modulation of angiogenesis and vasculogenesis in humans and mice. The signaling pathways that lead to the regulation of ANGPT-2 are largely unclear. Here, we report that protein kinase C and protein kinase A activators (ADMB, 8-Cl-cAMP) increased the mRNA levels of ANGPT-2 in human Granulosa cells, whereas PKC and PKA Inhibitors (Rp-cAMP, GO 6983) decreased markedly the level of ANGPT-2 mRNA. Due to varying specificity of the modulators for certain protein kinases subunits, we conclude that the conventional PKCs, but not PKC alpha and beta1, the atypical PKCs and the PKA I, are involved in the regulation of ANGPT-2. These findings may help to explain the role of both PKA and PKC dependent signaling cascades in the regulation of ANGPT-2 mRNA.

  15. Interrelations between translation and general mRNA degradation in yeast.

    PubMed

    Huch, Susanne; Nissan, Tracy

    2014-01-01

    Messenger RNA (mRNA) degradation is an important element of gene expression that can be modulated by alterations in translation, such as reductions in initiation or elongation rates. Reducing translation initiation strongly affects mRNA degradation by driving mRNA toward the assembly of a decapping complex, leading to decapping. While mRNA stability decreases as a consequence of translational inhibition, in apparent contradiction several external stresses both inhibit translation initiation and stabilize mRNA. A key difference in these processes is that stresses induce multiple responses, one of which stabilizes mRNAs at the initial and rate-limiting step of general mRNA decay. Because this increase in mRNA stability is directly induced by stress, it is independent of the translational effects of stress, which provide the cell with an opportunity to assess its response to changing environmental conditions. After assessment, the cell can store mRNAs, reinitiate their translation or, alternatively, embark on a program of enhanced mRNA decay en masse. Finally, recent results suggest that mRNA decay is not limited to non-translating messages and can occur when ribosomes are not initiating but are still elongating on mRNA. This review will discuss the models for the mechanisms of these processes and recent developments in understanding the relationship between translation and general mRNA degradation, with a focus on yeast as a model system. © 2014 The Authors. WIREs RNA published by John Wiley & Sons, Ltd.

  16. Cup regulates oskar mRNA stability during oogenesis.

    PubMed

    Broyer, Risa M; Monfort, Elena; Wilhelm, James E

    2017-01-01

    The proper regulation of the localization, translation, and stability of maternally deposited transcripts is essential for embryonic development in many organisms. These different forms of regulation are mediated by the various protein subunits of the ribonucleoprotein (RNP) complexes that assemble on maternal mRNAs. However, while many of the subunits that regulate the localization and translation of maternal transcripts have been identified, relatively little is known about how maternal mRNAs are stockpiled and stored in a stable form to support early development. One of the best characterized regulators of maternal transcripts is Cup - a broadly conserved component of the maternal RNP complex that in Drosophila acts as a translational repressor of the localized message oskar. In this study, we have found that loss of cup disrupts the localization of both the oskar mRNA and its associated proteins to the posterior pole of the developing oocyte. This defect is not due to a failure to specify the oocyte or to disruption of RNP transport. Rather, the localization defects are due to a drop in oskar mRNA levels in cup mutant egg chambers. Thus, in addition to its role in regulating oskar mRNA translation, Cup also plays a critical role in controlling the stability of the oskar transcript. This suggests that Cup is ideally positioned to coordinate the translational control function of the maternal RNP complex with its role in storing maternal transcripts in a stable form. Published by Elsevier Inc.

  17. Up-regulated EMMPRIN/CD147 protein expression might play a role in colorectal carcinogenesis and its subsequent progression without an alteration of its glycosylation and mRNA level.

    PubMed

    Zheng, Hua-chuan; Wang, Wei; Xu, Xiao-yan; Xia, Pu; Yu, Miao; Sugiyama, Toshiro; Takano, Yasuo

    2011-04-01

    Extracellular matrix metalloproteinase inducer (EMMPRIN) was reported to involve in the invasion and metastasis of malignancies by regulating the expression of vascular endothelial growth factor (VEGF) in stromal and cancer cells. The study aimed to clarify the role of EMMPRIN expression in tumorigenesis and progression of colorectal carcinomas (CRC). EMMPRIN expression was examined on tissue microarray containing colorectal carcinomas, adenoma and non-neoplastic mucosa (NNM) by immunohistochemistry and in situ hybridization (ISH). Colorectal carcinoma cell lines (DLD-1, HCT-15, SW480 and WiDr) and tissues were studied for EMMPRIN expression by Western blot or RT-PCR, followed by sequencing. All carcinoma cell lines showed EMMPRIN expression at both mRNA and protein levels. Two synonymous mutations were found in carcinoma cell lines at codon109 (GCT → GCC: Ala) or 179 (GAT → GAC: Asp). Frozen CRC tissues displayed higher EMMPRIN expression than paired NNM (P < 0.05). EMMPRIN expression was immunohistochemically stronger in colorectal high-grade adenoma, adenocarcinoma and metastatic carcinoma than non-neoplastic superficial epithelium and low-grade adenoma (P < 0.05). In contrast, its mRNA level was similar from colorectal NNM, adenoma to adenocarcinoma by ISH, in line with the findings of RT-PCR (P > 0.05). Immunohistochemically, EMMPRIN expression was positively correlated with tumor size, depth of invasion, vascular or lymphatic invasion, grade of infiltration (INF), ki-67 and VEGF expression of CRCs (P < 0.05). Among them, depth of invasion was an independent associated factor for EMMPRIN expression in CRCs (P < 0.05). Up-regulated EMMPRIN protein expression might contribute to colorectal carcinogenesis without the alteration of its glycosylation and mRNA level. Aberrant EMMPRIN protein expression might promote growth or invasion of CRCs possibly through increased ki-67 expression and inducible angiogenesis via up-regulating VEGF expression.

  18. Selenium Deficiency Affects the mRNA Expression of Inflammatory Factors and Selenoprotein Genes in the Kidneys of Broiler Chicks.

    PubMed

    Zhang, Jiu-Li; Xu, Bo; Huang, Xiao-Dan; Gao, Yu-Hong; Chen, Yu; Shan, An-Shan

    2016-05-01

    The aim of this study was to investigate the influence of Se deficiency on the transcription of inflammatory factors and selenoprotein genes in the kidneys of broiler chicks. One hundred fifty 1-day-old broiler chicks were randomly assigned to two groups fed with either a low-Se diet (L group, 0.033 mg/kg Se) or an adequate Se diet (C group, 0.2 mg/kg Se). The levels of uric acid (UA) and creatinine (Cr) in the serum and the mRNA levels of 6 inflammatory factors and 25 selenoprotein genes in the kidneys were measured as the clinical signs of Se deficiency occurred at 20 days old. The results indicated that the contents of UA and Cr in the serum increased in L group (p < 0.05), and the mRNA levels of the inflammatory factors (NF-κB, iNOS, COX-2, and TNF-α) increased in L group (p < 0.05). Meanwhile, the mRNA levels of PTGEs and HO-1 were not changed. In addition, 25 selenoprotein transcripts displayed ubiquitous expression in the kidneys of the chicks. The mRNA levels of 14 selenoprotein genes (Dio1, Dio2, GPx3, Sepp1, SelH, SelI, SelK, Sepn1, SelO, SelW, Sep15, SelT, SelU, and SelS) decreased, and 9 selenoprotein genes (GPx1, GPx2, GPx4, SelPb, Txnrd1, Txnrd2, Txnrd3, SPS2, and SelM) increased in L group (p < 0.05), but the Dio3 and Sepx1 mRNA levels did not change. The results indicated that Se deficiency resulted in kidney dysfunction, activation of the NF-κB pathway, and a change in selenoprotein gene expression. The changes of inflammatory factor and selenoprotein gene expression levels were directly related to the abnormal renal functions induced by Se deficiency.

  19. Cloning of guinea pig IL-4: reduced IL-4 mRNA after vaccination or Mycobacterium tuberculosis infection.

    PubMed

    Jeevan, Amminikutty; Yoshimura, Teizo; Ly, Lan H; Dirisala, Vijaya R; McMurray, David N

    2011-01-01

    Interleukin-4 (IL-4), a pleiotropic cytokine produced by T-helper type 2 (Th2) cells, is involved in promoting humoral immune responses, allergic reactions and asthma. Previous studies suggested an important role for IL-4 in susceptibility to pulmonary tuberculosis; however, the role of IL-4 has not been studied in the guinea pig, a highly relevant model for this disease. In the present study, we cloned a cDNA for guinea pig IL-4 and examined, for the first time, mRNA expression by real-time RT-PCR in cultured guinea pig cells. High levels of IL-4 mRNA expression were detected in spleen T cells of naïve animals after in vitro stimulation with PMA plus ionomycin for 4-24 h. The expression of IL-4 mRNA was low in spleen and lymph node cells immunized with ovalbumin (OVA) plus Complete Freund's Adjuvant (CFA) in response to OVA (Th1), but significantly higher in the guinea pigs immunized with OVA plus alum (Th2). BCG vaccination reduced the expression of IL-4 mRNA in both spleen and lung digest cells compared to naïve guinea pigs, while levels of IFN-γ were similar in both groups. Furthermore, lung cells from Mycobacterium tuberculosis-infected guinea pigs stimulated in vitro with PPD or MPT64 showed low levels of IL-4 mRNA expression. Thus, BCG vaccination or M. tuberculosis infection modulates IL-4 mRNA expression in the guinea pig. Cloning of guinea pig IL-4 will allow us to address the role of IL-4 in vaccine-induced resistance to pulmonary TB in a highly relevant animal model. Copyright © 2010 Elsevier Ltd. All rights reserved.

  20. Restriction of the Xenopus DEADSouth mRNA to the primordial germ cells is ensured by multiple mechanisms.

    PubMed

    Yamaguchi, Takeshi; Kataoka, Kensuke; Watanabe, Kenji; Orii, Hidefumi

    2014-02-01

    DEADSouth mRNA encoding the RNA helicase DDX25 is a component of the germ plasm in Xenopus laevis. We investigated the mechanisms underlying its specific mRNA expression in primordial germ cells (PGCs). Based on our previous findings of several microRNA miR-427 recognition elements (MREs) in the 3' untranslated region of the mRNA, we first examined whether DEADSouth mRNA was degraded by miR-427 targeting in somatic cells. Injection of antisense miR-427 oligomer and reporter mRNA for mutated MREs revealed that DEADSouth mRNA was potentially degraded in somatic cells via miR-427 targeting, but not in PGCs after the mid-blastula transition (MBT). The expression level of miR-427 was very low in PGCs, which probably resulted in the lack of miR-427-mediated degradation. In addition, the DEADSouth gene was expressed zygotically after MBT. Thus, the predominant expression of DEADSouth mRNA in the PGCs is ensured by multiple mechanisms including zygotic expression and prohibition from miR-427-mediated degradation. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  1. mRNA Translation Gone Awry: Translation Fidelity and Neurological Disease.

    PubMed

    Kapur, Mridu; Ackerman, Susan L

    2018-03-01

    Errors during mRNA translation can lead to a reduction in the levels of functional proteins and an increase in deleterious molecules. Advances in next-generation sequencing have led to the discovery of rare genetic disorders, many caused by mutations in genes encoding the mRNA translation machinery, as well as to a better understanding of translational dynamics through ribosome profiling. We discuss here multiple neurological disorders that are linked to errors in tRNA aminoacylation and ribosome decoding. We draw on studies from genetic models, including yeast and mice, to enhance our understanding of the translational defects observed in these diseases. Finally, we emphasize the importance of tRNA, their associated enzymes, and the inextricable link between accuracy and efficiency in the maintenance of translational fidelity. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Software Assurance Competency Model

    DTIC Science & Technology

    2013-03-01

    COTS) software , and software as a service ( SaaS ). L2: Define and analyze risks in the acquisition of contracted software , COTS software , and SaaS ...2010a]: Application of technologies and processes to achieve a required level of confidence that software systems and services function in the...

  3. Experiment K-6-11. Actin mRNA and cytochrome c mRNA concentrations in the tricepts brachia muscle of rats

    NASA Technical Reports Server (NTRS)

    Booth, F. W.; Morrison, P. R.; Thomason, D. B.; Oganov, V. S.

    1990-01-01

    It is well known that some skeletal muscles atrophy as a result of weightlessness (Steffen and Musacchia 1986) and as a result of hindlimb suspension (Tischler et al., 1985, Thomason et al., 1987). Because the content of protein is determined by the rates of protein synthesis and degradation, a decrease in protein synthesis rate, or an increase in the protein degradation, or changes in both could produce the atrophy. Indeed, an increased protein degradation (Tischler et al., 1985) and a decreased protein synthesis (Thomason et al., 1988) have been observed in skeletal muscles of suspended hindlimbs of rats. Any decrease in protein synthesis rate could be caused by decreases in mRNA concentrations. Such decreases in the concentration and content of alpha-actin mRNA and cytochrome c mRNA have been noted in skeletal muscles of hindlimb suspended rats (Babij and Booth, 1988). From these findings researchers hypothesized that alpha-actin mRNA and cytochrome c mRNA would decrease in the triceps brachia muscle of Cosmos 1887 rats.

  4. Matrin 3 binds and stabilizes mRNA.

    PubMed

    Salton, Maayan; Elkon, Ran; Borodina, Tatiana; Davydov, Aleksey; Yaspo, Marie-Laure; Halperin, Eran; Shiloh, Yosef

    2011-01-01

    Matrin 3 (MATR3) is a highly conserved, inner nuclear matrix protein with two zinc finger domains and two RNA recognition motifs (RRM), whose function is largely unknown. Recently we found MATR3 to be phosphorylated by the protein kinase ATM, which activates the cellular response to double strand breaks in the DNA. Here, we show that MATR3 interacts in an RNA-dependent manner with several proteins with established roles in RNA processing, and maintains its interaction with RNA via its RRM2 domain. Deep sequencing of the bound RNA (RIP-seq) identified several small noncoding RNA species. Using microarray analysis to explore MATR3's role in transcription, we identified 77 transcripts whose amounts depended on the presence of MATR3. We validated this finding with nine transcripts which were also bound to the MATR3 complex. Finally, we demonstrated the importance of MATR3 for maintaining the stability of several of these mRNA species and conclude that it has a role in mRNA stabilization. The data suggest that the cellular level of MATR3, known to be highly regulated, modulates the stability of a group of gene transcripts.

  5. Matrin 3 Binds and Stabilizes mRNA

    PubMed Central

    Salton, Maayan; Elkon, Ran; Borodina, Tatiana; Davydov, Aleksey; Yaspo, Marie-Laure; Halperin, Eran; Shiloh, Yosef

    2011-01-01

    Matrin 3 (MATR3) is a highly conserved, inner nuclear matrix protein with two zinc finger domains and two RNA recognition motifs (RRM), whose function is largely unknown. Recently we found MATR3 to be phosphorylated by the protein kinase ATM, which activates the cellular response to double strand breaks in the DNA. Here, we show that MATR3 interacts in an RNA-dependent manner with several proteins with established roles in RNA processing, and maintains its interaction with RNA via its RRM2 domain. Deep sequencing of the bound RNA (RIP-seq) identified several small noncoding RNA species. Using microarray analysis to explore MATR3′s role in transcription, we identified 77 transcripts whose amounts depended on the presence of MATR3. We validated this finding with nine transcripts which were also bound to the MATR3 complex. Finally, we demonstrated the importance of MATR3 for maintaining the stability of several of these mRNA species and conclude that it has a role in mRNA stabilization. The data suggest that the cellular level of MATR3, known to be highly regulated, modulates the stability of a group of gene transcripts. PMID:21858232

  6. The developmental course of salivary alpha-amylase and cortisol from 12 to 36 months: Relations with early poverty and later behavior problems.

    PubMed

    Hill-Soderlund, Ashley L; Holochwost, Steven J; Willoughby, Michael T; Granger, Douglas A; Gariépy, Jean-Louis; Mills-Koonce, W Roger; Cox, Martha J

    2015-02-01

    This study examined the development of baseline autonomic nervous system (ANS) and hypothalamic-pituitary-adrenal (HPA) physiological activity from 12 to 36 months as well as antecedents (poverty) and consequents (behavior problems) of individual differences in physiological development. Children (N=179; 50% poor; 56% African American; 52% male) provided saliva samples at 12, 18, 24, 30, and 36 months of age. Latent growth curve models indicated that nonlinear change was evident for both sAA and cortisol, with sAA increasing and cortisol decreasing with age. Children residing in poor households exhibited lower initial levels of sAA, but not cortisol. African-American children showed slightly smaller decreases in cortisol over time. Initial levels of sAA predicted higher levels of internalizing behaviors at 36 months and both initial levels of and total change in sAA predicted higher levels of externalizing behaviors at 36 months. There was no evidence that sAA or cortisol mediated the relationship between poverty and later behavior problems. Copyright © 2014 Elsevier Ltd. All rights reserved.

  7. Diet-induced weight loss is associated with decreases in plasma serum amyloid a and C-reactive protein independent of dietary macronutrient composition in obese subjects.

    PubMed

    O'Brien, Kevin D; Brehm, Bonnie J; Seeley, Randy J; Bean, Judy; Wener, Mark H; Daniels, Stephen; D'Alessio, David A

    2005-04-01

    Elevated levels of serum amyloid A (SAA) and C-reactive protein (CRP) have been associated with increased cardiovascular risk. Although levels of CRP decrease with weight loss, it is not known whether SAA decreases with weight loss or whether dietary macronutrient composition affects levels of either SAA or CRP. SAA and CRP levels were measured retrospectively on baseline and 3-month plasma samples from 41 obese (mean body mass index 33.63 +/- 1.86 kg/m2) women completing a randomized trial comparing a low-fat diet (n = 19) and a very low-carbohydrate diet (n = 22). For the 41 participants, there were significant decreases from baseline to 3 months in both LogSAA (P = 0.049) and LogCRP (P = 0.035). The very low-carbohydrate dieters had a significantly greater decrease in LogSAA (P = 0.04), but their weight loss also was significantly greater (-7.6 +/- 3.2 vs. -4.3 +/- 3.5 kg, P < 0.01). In this study, the decreases in inflammatory markers correlated significantly with weight loss (r = 0.44, P = 0.004 vs. LogSAA and r = 0.35, P = 0.03 vs. LogCRP). Also, change in LogSAA correlated with change in insulin resistance (r = 0.35, P = 0.03). Thus, in otherwise healthy, obese women, weight loss was associated with significant decreases in both SAA and CRP. These effects were proportional to the amount of weight lost but independent of dietary macronutrient composition.

  8. Interrelations between translation and general mRNA degradation in yeast

    PubMed Central

    Huch, Susanne; Nissan, Tracy

    2014-01-01

    Messenger RNA (mRNA) degradation is an important element of gene expression that can be modulated by alterations in translation, such as reductions in initiation or elongation rates. Reducing translation initiation strongly affects mRNA degradation by driving mRNA toward the assembly of a decapping complex, leading to decapping. While mRNA stability decreases as a consequence of translational inhibition, in apparent contradiction several external stresses both inhibit translation initiation and stabilize mRNA. A key difference in these processes is that stresses induce multiple responses, one of which stabilizes mRNAs at the initial and rate-limiting step of general mRNA decay. Because this increase in mRNA stability is directly induced by stress, it is independent of the translational effects of stress, which provide the cell with an opportunity to assess its response to changing environmental conditions. After assessment, the cell can store mRNAs, reinitiate their translation or, alternatively, embark on a program of enhanced mRNA decay en masse. Finally, recent results suggest that mRNA decay is not limited to non-translating messages and can occur when ribosomes are not initiating but are still elongating on mRNA. This review will discuss the models for the mechanisms of these processes and recent developments in understanding the relationship between translation and general mRNA degradation, with a focus on yeast as a model system. How to cite this article: WIREs RNA 2014, 5:747–763. doi: 10.1002/wrna.1244 PMID:24944158

  9. Optimal Down Regulation of mRNA Translation

    NASA Astrophysics Data System (ADS)

    Zarai, Yoram; Margaliot, Michael; Tuller, Tamir

    2017-01-01

    Down regulation of mRNA translation is an important problem in various bio-medical domains ranging from developing effective medicines for tumors and for viral diseases to developing attenuated virus strains that can be used for vaccination. Here, we study the problem of down regulation of mRNA translation using a mathematical model called the ribosome flow model (RFM). In the RFM, the mRNA molecule is modeled as a chain of n sites. The flow of ribosomes between consecutive sites is regulated by n + 1 transition rates. Given a set of feasible transition rates, that models the outcome of all possible mutations, we consider the problem of maximally down regulating protein production by altering the rates within this set of feasible rates. Under certain conditions on the feasible set, we show that an optimal solution can be determined efficiently. We also rigorously analyze two special cases of the down regulation optimization problem. Our results suggest that one must focus on the position along the mRNA molecule where the transition rate has the strongest effect on the protein production rate. However, this rate is not necessarily the slowest transition rate along the mRNA molecule. We discuss some of the biological implications of these results.

  10. Expression and significance of cyclooxygenase-2 mRNA in benign and malignant ascites

    PubMed Central

    Lu, Jing; Li, Xiao-Feng; Kong, Li-Xia; Ma, Lin; Liao, Su-Huan; Jiang, Chang-You

    2013-01-01

    AIM: To investigate the mRNA expression of cyclooxygensae-2 (COX-2) in benign and malignant ascites, and to explore the difference in COX-2 mRNA expression among different diseases. METHODS: A total of 36 samples were collected from the Fifth Affiliated Hospital of Sun Yat-Sen University and divided into two experimental groups: benign ascites (n = 21) and malignant ascites (n = 15). Benign ascites included cirrhotic ascites (n = 10) and tuberculous ascites (n = 5). Malignant ascites included oophoroma (n = 7), cancer of colon (n = 5), cancer of the liver (n = 6), gastric cancer (n = 2), and bladder carcinoma (n = 1). The mRNA expression of COX-2 in ascites was examined with reverse transcriptase polymerase chain reaction (RT-PCR) technology, and the positive rate of COX-2 mRNA was compared between different diseases. RESULTS: The positive rate of COX-2 mRNA in malignant ascites was 42.9% (9/21), which was significantly higher than in benign ascites, 6.7% (1/15), difference being significant between these two groups (χ2 = 4.051, P = 0.044). The proportion of the positive rate in the malignant ascites was as follows: ovarian cancers 57.1% (4/7), colon cancer 40.0% (2/5), liver cancer 33.3% (2/6), gastric cancer 50.0% (1/2), and bladder cancer 0.00% (0/1). However, there was no significant difference in COX-2 mRNA expression among various tumors with malignant ascites (χ2 = 1.614, P = 0.806). Among the benign ascites, COX-2 mRNA levels were different between the tuberculous ascites (0/5) and cirrhotic ascites (1/10), but there was no significant difference (P = 1.000). CONCLUSION: COX-2 mRNA, detected by RT-PCR, is useful in the differential diagnosis of benign and malignant ascites, which also has potential value in the clinical diagnosis of tumors. PMID:24187465

  11. Influenza A viruses suppress cyclooxygenase-2 expression by affecting its mRNA stability.

    PubMed

    Dudek, Sabine Eva; Nitzsche, Katja; Ludwig, Stephan; Ehrhardt, Christina

    2016-06-06

    Infection with influenza A viruses (IAV) provokes activation of cellular defence mechanisms contributing to the innate immune and inflammatory response. In this process the cyclooxygenase-2 (COX-2) plays an important role in the induction of prostaglandin-dependent inflammation. While it has been reported that COX-2 is induced upon IAV infection, in the present study we observed a down-regulation at later stages of infection suggesting a tight regulation of COX-2 by IAV. Our data indicate the pattern-recognition receptor RIG-I as mediator of the initial IAV-induced COX-2 synthesis. Nonetheless, during on-going IAV replication substantial suppression of COX-2 mRNA and protein synthesis could be detected, accompanied by a decrease in mRNA half-life. Interestingly, COX-2 mRNA stability was not only imbalanced by IAV replication but also by stimulation of cells with viral RNA. Our results reveal tristetraprolin (TTP), which is known to bind COX-2 mRNA and promote its rapid degradation, as regulator of COX-2 expression in IAV infection. During IAV replication and viral RNA accumulation TTP mRNA synthesis was induced, resulting in reduced COX-2 levels. Accordingly, the down-regulation of TTP resulted in increased COX-2 protein expression after IAV infection. These findings indicate a novel IAV-regulated cellular mechanism, contributing to the repression of host defence and therefore facilitating viral replication.

  12. Increased expression of ADAM 9 and ADAM 15 mRNA in pancreatic cancer.

    PubMed

    Yamada, Daisuke; Ohuchida, Kenoki; Mizumoto, Kazuhiro; Ohhashi, Seiji; Yu, Jun; Egami, Takuya; Fujita, Hayato; Nagai, Eishi; Tanaka, Masao

    2007-01-01

    A disintegrin and metalloproteases (ADAMs) comprise a multifunctional family of membrane-anchored proteins. ADAM 9 and ADAM 15 are involved in cell migration and invasion. Expression of ADAM 9 and ADAM 15 was reported to be altered in several types of cancer. Quantitative real-time reverse transcription-polymerase chain reaction was performed to measure the expression of ADAM 9 mRNA in bulk pancreatic tissues. Results showed no significant difference in the expression of ADAM 9 mRNA between pancreatic cancer and non-neoplastic pancreas. Primary cultured pancreatic fibroblasts also expressed ADAM 9 mRNA. Therefore, a laser microdissection and pressure catapulting technique was employed to isolate cancer cells from tumor tissues. The expression of ADAM 9 and ADAM 15 mRNA was measured in microdissected samples (cancer cells, n = 11; normal epithelial cells, n = 13 for ADAM 9; cancer cells, n = 9; normal epithelial cells, n = 9 for ADAM 15). Pancreatic cancer cells expressed significantly higher levels of ADAM 9 and ADAM 15 mRNA than did normal pancreatic epithelial cells (p = 0.016 for ADAM 9; p = 0.004 for ADAM 15). ADAM 9 and ADAM 15 are involved in pancreatic cancer. Microdissection-based analysis appears to be indispensable for the accurate analysis of the expression of certain ADAM family members in pancreatic cancer.

  13. miR-128 inhibits telomerase activity by targeting TERT mRNA

    PubMed Central

    Guzman, Herlinda; Sanders, Katie; Idica, Adam; Bochnakian, Aurore; Jury, Douglas; Daugaard, Iben; Zisoulis, Dimitrios G; Pedersen, Irene Munk

    2018-01-01

    Telomerase is a unique cellular reverse transcriptase (RT) essential for maintaining telomere stability and required for the unlimited proliferation of cancer cells. The limiting determinant of telomerase activity is the catalytic component TERT, and TERT expression is closely correlated with telomerase activity and cancer initiation and disease progression. For this reason the regulation of TERT levels in the cell is of great importance. microRNAs (miRs) function as an additional regulatory level in cells, crucial for defining expression boundaries, proper cell fate decisions, cell cycle control, genome integrity, cell death and metastasis. We performed an anti-miR library screen to identity novel miRs, which participate in the control of telomerase. We identified the tumor suppressor miR (miR-128) as a novel endogenous telomerase inhibitor and determined that miR-128 significantly reduces the mRNA and protein levels of Tert in a panel of cancer cell lines. We further evaluated the mechanism by which miR-128 regulates TERT and demonstrated that miR-128 interacts directly with the coding sequence of TERT mRNA in both HeLa cells and teratoma cells. Interestingly, the functional miR-128 binding site in TERT mRNA, is conserved between TERT and the other cellular reverse transcriptase encoded by Long Interspersed Elements-1 (LINE-1 or L1), which can also contribute to the oncogenic phenotype of cancer. This finding supports the novel idea that miRs may function in parallel pathways to inhibit tumorigenesis, by regulating a group of enzymes (such as RT) by targeting conserved binding sites in the coding region of both enzymes. PMID:29568354

  14. The relative expression levels of insulin-like growth factor 1 and myostatin mRNA in the asynchronous development of skeletal muscle in ducks during early development.

    PubMed

    Hu, Yan; Liu, Hongxiang; Shan, Yanju; Ji, Gaige; Xu, Wenjuan; Shu, Jingting; Li, Huifang

    2015-08-10

    Genetic selection is a powerful tool for modifying poultry muscle yield. Insulin-like growth factor I (IGF-I) and myostatin (MSTN) are important regulators of muscle growth, especially in the myogenesis stage. This study compared the developmental pattern of the pectoralis major (PM) and lateral gastrocnemius (LM) muscles, mRNA expression characterization of IGF-I and MSTN-A and their correlation between 14 days in ovo and 1 week post-hatch in two Chinese local duck breeds. During early development, the growth of duck PM and LM followed an asynchronous pattern. Variations in PM growth rate observed with development followed the relative variations of MSTN and IGF-I expression; however, the same behavior was not observed in LM. Moreover, the profile of IGF-I expression in duck skeletal muscles indicated that genetic selection for high meat-yield poultry has altered the temporal expression of IGF-I and affected cellular characteristics and mass by hatch in a PM-specific manner. The MSTN-A expression profile showed synchronization with the growth of skeletal muscle and peaks of myofiber proliferation. The expression patterns of IGF-I and MSTN suggest that duck pectoralis fibers are prioritized for proliferation in embryogenesis. The IGF-1/MSTN-A mRNA ratios in PM and LM presented very similar trends in the changes of myofiber characteristics, and differences in the IGF-1/MSTN-A mRNA ratio in PM between the two breeds corresponded to the timing of differences in PM mass between the varieties. Our results support the hypothesis that relative levels of IGF-I and MSTN mRNA may participate in ordering muscle growth rates with selected development. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. hnRNP A2/B1 interacts with influenza A viral protein NS1 and inhibits virus replication potentially through suppressing NS1 RNA/protein levels and NS1 mRNA nuclear export

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wang, Yimeng; Zhou, Jianhong; Du, Yuchun, E-mail: ydu@uark.edu

    The NS1 protein of influenza viruses is a major virulence factor and exerts its function through interacting with viral/cellular RNAs and proteins. In this study, we identified heterogeneous nuclear ribonucleoprotein A2/B1 (hnRNP A2/B1) as an interacting partner of NS1 proteins by a proteomic method. Knockdown of hnRNP A2/B1 by small interfering RNA (siRNA) resulted in higher levels of NS vRNA, NS1 mRNA, and NS1 protein in the virus-infected cells. In addition, we demonstrated that hnRNP A2/B1 proteins are associated with NS1 and NS2 mRNAs and that knockdown of hnRNP A2/B1 promotes transport of NS1 mRNA from the nucleus to themore » cytoplasm in the infected cells. Lastly, we showed that knockdown of hnRNP A2/B1 leads to enhanced virus replication. Our results suggest that hnRNP A2/B1 plays an inhibitory role in the replication of influenza A virus in host cells potentially through suppressing NS1 RNA/protein levels and NS1 mRNA nucleocytoplasmic translocation. - Highlights: • Cellular protein hnRNP A2/B1 interacts with influenza viral protein NS1. • hnRNP A2/B1 suppresses the levels of NS1 protein, vRNA and mRNA in infected cells. • hnRNP A2/B1 protein is associated with NS1 and NS2 mRNAs. • hnRNP A2/B1 inhibits the nuclear export of NS1 mRNAs. • hnRNP A2/B1 inhibits influenza virus replication.« less

  16. Olive Leaf Extract Elevates Hepatic PPAR α mRNA Expression and Improves Serum Lipid Profiles in Ovariectomized Rats.

    PubMed

    Yoon, Leena; Liu, Ya-Nan; Park, Hyunjin; Kim, Hyun-Sook

    2015-07-01

    We hypothesized that olive leaf extract might alleviate dyslipidemia resulting from estrogen deficiency. Serum lipid profile and mRNA expression of the related genes in the liver and adipose tissue were analyzed after providing olive leaf extract (200 or 400 mg/kg body weight; n=7 for each group) to ovariectomized rats for 10 weeks. After 10 weeks' administration, the rats in the olive leaf extract-administered groups showed significantly lower levels of serum triglyceride and very-low-density lipoprotein (VLDL)-cholesterol compared with the rats in the control group, whereas the administration of olive leaf extract did not significantly change the elevated low-density lipoprotein cholesterol levels. In addition, administration of high dose of olive leaf extract significantly decreased the liver triglyceride and increased serum estradiol levels. mRNA expressions of peroxisome proliferator-activated receptor alpha (PPAR α) and acyl-CoA oxidase (ACO) were not affected by ovariectomy, however, administration of olive leaf extract significantly increased both PPAR α and ACO mRNA expression. Expression of adiponectin mRNA in adipose tissue was significantly decreased in the ovariectomized control group. Rats administered low-dose olive leaf extract showed significantly elevated adiponectin mRNA expression compared with rats in the ovariectomized control group. Even though dose-dependent effects were not observed in most of the measurements, these results suggest that genes involved in lipid metabolism may be regulated by olive leaf extract administration in ovariectomized rats.

  17. [Effect of Panax notoginseng saponins on liver drug metablic enzyme activity, mRNA and protein expressions in rats].

    PubMed

    Chen, Yan-Jin; Wang, Yu-Guang; Ma, Zeng-Chun; Xiao, Cheng-Rong; Tan, Hong-Ling; Liang, Qian-De; Tang, Xiang-Lin; Zhao, Yong-Hong; Wang, Dong-Gen; Gao, Yue

    2014-10-01

    To study the effect of Panax notoginseng saponins (PNS) on liver drug metabolic enzyme activity, mRNA and protein expressions in rats. Male Wistar rats were randomly divided into nine groups. After administration of the test drugs, their liver microsomes, liver total RNA and total protein were extracted to detect the regulating effect of PNS on liver drug metabolic enzyme activity-related subtype enzymatic activity, mRNA and protein expression by substrate probe, quantitative PCR and Western Blot technology. The result of this experiment was that PNS could significantly induce CYP1A2 and CYP2E1 enzyme activity, mRNA expression, CYP2E1 protein expression level. PNS significantly induced CYP3A mRNA expression, but with no significant effect in CYP3A enzyme activity level. PNS had no significant effect CYP1A1 and CYP2B mRNA expressions and enzyme activity levels. PNS had selective regulations on different P450 subtypes, and the major subtypes were CYP1A2 and CYP2E1. In clinical practice, particularly in the combination with CYP1A2 and CYP2E1 metabolism-related drugs, full consideration shall be given to the possible drug interactions in order to avoid potential toxic and side effects. Meanwhile, whether the induction effect of CYP2E1 gets involved in ginsenoside's effect incavenging free radicals deserves further studies.

  18. Incorporation of mRNA in Lamellar Lipid Matrices for Parenteral Administration.

    PubMed

    Ziller, Antje; Nogueira, Sara S; Hühn, Eva; Funari, Sergio S; Brezesinski, Gerald; Hartmann, Hermann; Sahin, Ugur; Haas, Heinrich; Langguth, Peter

    2018-02-05

    Insertion of high molecular weight messenger RNA (mRNA) into lyotropic lipid phases as model systems for controlled release formulations for the mRNA was investigated. Low fractions of 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) were used as an anchor to load the mRNA into a lamellar lipid matrix. Dispersions of zwitterionic lipid in the aqueous phase in the presence of increasing fractions of mRNA and cationic lipid were prepared, and the molecular organization was investigated as a function of mRNA and cationic lipid fraction. Insertion of both cationic lipid and mRNA was clearly proven from the physicochemical characteristics. The d-spacing of the lipid bilayers, as determined by small-angle X-ray scattering (SAXS) measurements, responded sensitively to the amount of inserted DOTAP and mRNA. A concise model of the insertion of the mRNA in the lipid matrices was derived, indicating that the mRNA was accommodated in the aqueous slab between lipid bilayers. Depending on the DOTAP and mRNA fraction, a different excess of water was present in this slab. Results from further physicochemical characterization, including determination of free and bound mRNA, zeta potential, and calorimetry data, were in line with this assumption. The structure of these concentrated lipid/mRNA preparations was maintained upon dilution. The functionality of the inserted mRNA was proven by cell culture experiments using C2C12 murine myoblast cells with the luciferase-encoding mRNA. The described lipid phases as carriers for the mRNA may be applicable for different routes of local administration, where control of the release kinetics and the form of the released mRNA (bound or free) is required.

  19. Clinical significance of serum circulating insulin-like growth factor-1 (IGF-1) mRNA in hepatocellular carcinoma.

    PubMed

    Karabulut, S; Duranyıldız, D; Tas, F; Gezer, U; Akyüz, F; Serilmez, M; Ozgür, E; Yasasever, C T; Vatansever, S; Aykan, N F

    2014-03-01

    The principal aim of our study was to investigate the usefulness of serum protein and circulating mRNA of insulin-like growth factor-1 (IGF-1) as a diagnostic and prognostic tool in hepatocellular carcinoma (HCC). Fifty-four HCC patients and age- and sex-matched 20 healthy controls were enrolled into this study. Pretreatment serum IGF-1 and IGF-1 mRNA were determined by the solid-phase sandwich ELISA and quantitative RT-PCR method, respectively. The median age at diagnosis was 60 years, range 36-77 years; where majority of group were male (n = 48, 88.8%). All patients had cirrhotic history. Forty-six percent (n = 25) of patients had Child-Pugh score A, 30% (n = 16) had score B or C. All of the patients were treated with local therapies and none of them received sorafenib. The baseline serum IGF-1 mRNA levels were significantly higher in HCC patients than in the control group (p = 0.04), whereas no significant difference was observed for IGF-1 protein levels between the two group (p = 0.18). Patients with history of HBV infection, who were not treated, and who received multiple palliative treatment for HCC had higher serum IGF-1 mRNA levels (p = 0.03, 0.03, and 0.05, respectively). Poor performance status (p < 0.001), viral etiology of cirrhosis (p = 0.03), larger tumor size (p = 0.01), lower serum hemoglobin levels (p = 0.03), and not be treated for HCC (p = 0.001) related to worse survival. However, neither serum IGF-1 nor serum IGF-1 mRNA had significantly adverse effect on survival (p = 0.53 and 0.42, respectively).

  20. Expression of dopamine D2 receptor and choline acetyltransferase mRNA in the dopamine deafferented rat caudate-putamen.

    PubMed

    Brené, S; Lindefors, N; Herrera-Marschitz, M; Persson, H

    1990-01-01

    In situ hybridization was used to study dopamine D2 receptor (D2R) and choline acetyltransferase (ChAT) mRNA expression in neurons of the rat forebrain, both on control animals and after a unilateral 6-hydroxydopamine (6-OHDA) lesion of midbrain dopamine neurons. D2R mRNA expressing neurons were seen in regions which are known to be heavily innervated by midbrain dopamine fibers such as caudate-putamen, nucleus accumbens and olfactory tubercle. ChAT mRNA expressing neurons were seen in caudate-putamen, nucleus accumbens and septal regions including vertical limb of the diagonal band. In caudate-putamen, approximately 55% of the medium sized neurons, which is the predominating neuronal cell-size in this region, were specifically labeled with the D2R probe. In addition, approximately 95% of the large size neurons in caudate-putamen were specifically labeled with both the D2R and ChAT probes, suggesting that most cholinergic neurons in the caudate-putamen express D2R mRNA. After a unilateral lesion of midbrain dopamine neurons, no change in the level of either D2R or ChAT mRNA were seen in the large size intrinsic cholinergic neurons in caudate-putamen. Similarly, no evidence was obtained for altered levels of D2R mRNA in medium size neurons in medial caudate-putamen, or nucleus accumbens. However, an increase in the number of medium size neurons expressing D2R mRNA was observed in the lateral part of the dopamine deafferented caudate-putamen. Thus, it appears that midbrain dopamine deafferentation causes an increase in D2R mRNA expression in a subpopulation of medium size neurons in the lateral caudate-putamen.

  1. T-lymphocyte cytokine mRNA expression in cystic echinococcosis.

    PubMed

    Fauser, S; Kern, P

    1997-04-01

    In the present study we investigated cytokine mRNA expression by peripheral blood mononuclear cells (PBMC) from patients with cystic echinococcosis (CE) after stimulation with different antigens. By using reverse transcriptase polymerase chain reaction (RT-PCR) we could demonstrate that restimulation with crude Echinococcus granulosus antigen (Eg-Ag) induced or enhanced Th2 cytokine mRNA expression, especially IL-5 (by using antigen from sheep cyst fluid) in 23 out of 26 investigated CE patients and IL-10 (by using antigen from camel cyst fluid) in 10 out of 10 investigated CE patients. In contrast, IL-5 mRNA expression was absent in PBMC of healthy controls after Eg-Ag stimulation. To determine the specificity of this reaction we stimulated PBMC from 11 CE patients with crude Echinococcus multilocularis antigen (Em-Ag) and PBMC from 8 CE patients with Toxocara canis antigen (Tc-Ag). We found that the PBMC of patients showed a similar mRNA cytokine pattern on stimulation with Em-Ag when compared with Eg-Ag stimulation. The cytokine mRNA pattern on stimulation with Tc-Ag, however, resembled the cytokine mRNA pattern of unstimulated PBMC. Furthermore, the stimulation of PBMC with crude Mycobacterium tuberculosis antigen (H37Ra) and purified protein derivative (PPD) of M. tuberculosis revealed distinct IL-5 mRNA expression in all investigated CE patients, whereas in healthy controls IL-5 mRNA expression was very weak or totally absent. Thus, our results indicate an induction of Th2 cytokine mRNA expression in CE patients, which is frequently observed in parasite infections. Interestingly, this response persists after stimulation with tuberculosis antigens, which normally induce Th1 response.

  2. Widespread promoter-mediated coordination of transcription and mRNA degradation

    PubMed Central

    2012-01-01

    Background Previous work showed that mRNA degradation is coordinated with transcription in yeast, and in several genes the control of mRNA degradation was linked to promoter elements through two different mechanisms. Here we show at the genomic scale that the coordination of transcription and mRNA degradation is promoter-dependent in yeast and is also observed in humans. Results We first demonstrate that swapping upstream cis-regulatory sequences between two yeast species affects both transcription and mRNA degradation and suggest that while some cis-regulatory elements control either transcription or degradation, multiple other elements enhance both processes. Second, we show that adjacent yeast genes that share a promoter (through divergent orientation) have increased similarity in their patterns of mRNA degradation, providing independent evidence for the promoter-mediated coupling of transcription to mRNA degradation. Finally, analysis of the differences in mRNA degradation rates between mammalian cell types or mammalian species suggests a similar coordination between transcription and mRNA degradation in humans. Conclusions Our results extend previous studies and suggest a pervasive promoter-mediated coordination between transcription and mRNA degradation in yeast. The diverse genes and regulatory elements associated with this coordination suggest that it is generated by a global mechanism of gene regulation and modulated by gene-specific mechanisms. The observation of a similar coupling in mammals raises the possibility that coupling of transcription and mRNA degradation may reflect an evolutionarily conserved phenomenon in gene regulation. PMID:23237624

  3. Effect of low levels of dietary available phosphorus on phosphorus utilization, bone mineralization, phosphorus transporter mRNA expression and performance in growing pigs.

    PubMed

    Pokharel, Bishwo B; Regassa, Alemu; Nyachoti, Charles M; Kim, Woo K

    2017-06-03

    A study was conducted to examine the effects of different dietary levels of available phosphorus (aP) on P excretion, bone mineralization, performance and the mRNA expression of sodium-dependent P transporters in growing pigs. Sixty-day old growing pigs (n = 54) with an average initial BW of 19.50 ± 1.11 kg were randomly allocated to a control diet (C) containing 0.23% available phosphorus (aP), T1 containing 0.17% aP and T2 containing 0.11% aP. There were 6 pens per treatment with 3 pigs per pen. Body weight and feed intake were measured weekly. At the end of each week, one pig from each pen was housed in a metabolic crate for 24 h to collect fecal and urine samples and then sacrificed to obtain third metacarpal (MC3) bones and jejunal and kidney samples. Bones were scanned by Dual Energy X-ray Absorptiometry (DEXA). Fecal and urine samples were sub-sampled and analyzed for P content. The expression of P transporter mRNA in jejunum and kidney samples was measured using quantitative real-time polymerase chain reaction (qRT-PCR). Data were analyzed using GLM procedure of the Statistical Analysis System (SAS Institute version 9.2). Pigs fed the T2 diet had reduced (P < 0.05) average daily gain (ADG) and gain to feed (G:F) compared to those fed the C diet during week 2. Overall, ADG and G:F were also reduced (P < 0.05) in pigs fed the T2 diet compared to those fed the C and T1 diets. Bone mineral density (BMD) and bone mineral content (BMC) were reduced (P < 0.05) in pigs fed the T2 diet compared to those fed the C diet throughout the experiment. At week 1, jejunal mRNA expression of Na (+)-dependent phosphate transporter 2 (SLC34A2) was increased (P < 0.01) in pigs fed the T2 diet compared to C diet. Renal mRNA expression of Na(+)-dependent phosphate transporter 1 (SLC34A1) and SLC34A3 were increased (P < 0.05) in pigs fed the T2 diet compared to those fed the C diet at week 2 and was accompanied by lower (P < 0.05) urinary P in pigs fed the T2 diet during week 2

  4. beta(2)microglobulin mRNA expression levels are prognostic for lymph node metastasis in colorectal cancer patients.

    PubMed

    Shrout, J; Yousefzadeh, M; Dodd, A; Kirven, K; Blum, C; Graham, A; Benjamin, K; Hoda, R; Krishna, M; Romano, M; Wallace, M; Garrett-Mayer, E; Mitas, M

    2008-06-17

    Colorectal cancer (CRC) is the fourth most common non-cutaneous malignancy in the United States and the second most frequent cause of cancer-related death. One of the most important determinants of CRC survival is lymph node metastasis. To determine whether molecular markers might be prognostic for lymph node metastases, we measured by quantitative real-time RT-PCR the expression levels of 15 cancer-associated genes in formalin-fixed paraffin-embedded primary tissues derived from stage I-IV CRC patients with (n=20) and without (n=18) nodal metastases. Using the mean of the 15 genes as an internal reference control, we observed that low expression of beta(2)microglobulin (B2M) was a strong prognostic indicator of lymph node metastases (area under the curve (AUC)=0.85; 95% confidence interval (CI)=0.69-0.94). We also observed that the expression ratio of B2M/Spint2 had the highest prognostic accuracy (AUC=0.87; 95% CI=0.71-0.96) of all potential two-gene combinations. Expression values of Spint2 correlated with the mean of the entire gene set at an R(2) value of 0.97, providing evidence that Spint2 serves not as an independent prognostic gene, but rather as a reliable reference control gene. These studies are the first to demonstrate a prognostic role of B2M at the mRNA level and suggest that low B2M expression levels might be useful for identifying patients with lymph node metastasis and/or poor survival.

  5. Influence of Starvation on Potential Ammonia-Oxidizing Activity and amoA mRNA Levels of Nitrosospira briensis

    PubMed Central

    Bollmann, Annette; Schmidt, Ingo; Saunders, Aaron M.; Nicolaisen, Mette H.

    2005-01-01

    The effect of short-term ammonia starvation on Nitrosospira briensis was investigated. The ammonia-oxidizing activity was determined in a concentrated cell suspension with a NOx biosensor. The apparent half-saturation constant [Km(app)] value of the NH3 oxidation of N. briensis was 3 μM NH3 for cultures grown both in continuous and batch cultures as determined by a NOx biosensor. Cells grown on the wall of the vessel had a lower Km(app) value of 1.8 μM NH3. Nonstarving cultures of N. briensis showed potential ammonia-oxidizing activities of between 200 to 250 μM N h−1, and this activity decreased only slowly during starvation up to 10 days. Within 10 min after the addition of fresh NH4+, 100% activity was regained. Parallel with activity measurements, amoA mRNA and 16S rRNA were investigated. No changes were observed in the 16S rRNA, but a relative decrease of amoA mRNA was observed during the starvation period. During resuscitation, an increase in amoA mRNA expression was detected simultaneously. The patterns of the soluble protein fraction of a 2-week-starved culture of N. briensis showed only small differences in comparison to a nonstarved control. From these results we conclude that N. briensis cells remain in a state allowing fast recovery of ammonia-oxidizing activity after addition of NH4+ to a starved culture. Maintaining cells in this kind of active state could be the survival strategy of ammonia-oxidizing bacteria in nature under fluctuating NH4+ availability. PMID:15746329

  6. Expression of calmodulin mRNA in rat olfactory neuroepithelium.

    PubMed

    Biffo, S; Goren, T; Khew-Goodall, Y S; Miara, J; Margolis, F L

    1991-04-01

    A calmodulin (CaM) cDNA was isolated by differential hybridization screening of a lambda gt10 library prepared from rat olfactory mucosa. This cDNA fragment, containing most of the open reading frame of the rat CaMI gene, was subcloned and used to characterize steady-state expression of CaM mRNA in rat olfactory neuroepithelium and bulb. Within the bulb mitral cells are the primary neuronal population expressing CaM mRNA. The major CaM mRNA expressed in the olfactory mucosa is 1.7 kb with smaller contributions from mRNAs of 4.0 and 1.4 kb. CaM mRNA was primarily associated with the olfactory neurons and, despite the cellular complexity of the tissue and the known involvement of CaM in diverse cellular processes, was only minimally evident in sustentacular cells, gland cells or respiratory epithelium. Following bulbectomy CaM mRNA declines in the olfactory neuroepithelium as does olfactory marker protein (OMP) mRNA. In contrast to the latter, CaM mRNA makes a partial recovery by one month after surgery. These results, coupled with those from in situ hybridization, indicate that CaM mRNA is expressed in both mature and immature olfactory neurons. The program regulating CaM gene expression in olfactory neurons is distinct from those controlling expression of B50/GAP43 in immature, or OMP in mature, neurons respectively.

  7. Cinnamaldehyde up-regulates the mRNA expression level of TRPV1 receptor potential ion channel protein and its function in primary rat DRG neurons in vitro.

    PubMed

    Sui, Feng; Lin, Na; Guo, Jian-You; Zhang, Chang-Bin; Du, Xin-Liang; Zhao, Bao-Sheng; Liu, Hong-Bin; Yang, Na; Li, Lan-Fang; Guo, Shu-Ying; Huo, Hai-Ru; Jiang, Ting-Liang

    2010-01-01

    Cinnamaldehyde (1) is a pharmacologically active ingredient isolated from cassia twig (Ramulus Cinnamomi), which is commonly used in herbal remedies to treat fever-related diseases. Both TRPV1 and TRPM8 ion channel proteins are abundantly expressed in sensory neurons, and are assumed to act as a thermosensor, with the former mediating the feeling of warmth and the latter the feeling of cold in the body. Both of them have recently been reported to be involved in thermoregulation. The purpose of this paper is to further uncover the antipyretic mechanisms of 1 by investigating its effects on the mRNA expression levels and functions of both TRPV1 and TRPM8. The results showed that 1 could up-regulate the mRNA expression levels of TRPV1 at both 37 and 39 degrees C, and its calcium-mediating function was significantly increased at 39 degrees C, all of which could not be blocked by pretreatment of the neuronal cells with ruthenium red, a general transient receptor potential (TRP) blocker, indicating that the action of 1 was achieved through a non-TRPA1 channel pathway. In conclusion, the findings in our in vitro studies might account for part of the peripheral molecular mechanisms for the antipyretic action of 1.

  8. The Behaviour of Fe Stable Isotopes Accompanying Fluid Migration in Subducted Serpentinite from the Zermatt-Saas Ophiolite of the Swiss Alps

    NASA Astrophysics Data System (ADS)

    Inglis, E.; Bouilhol, P.; Burton, K. W.; Debret, B.; Millet, M. A.; Williams, H. M.

    2016-12-01

    During subduction the destabilisation of hydrous serpentine group phases can generate significant fluid fluxes between the subducting slab and the overlying mantle wedge. Despite our knowledge of this, the exact process and nature of the fluids released during serpentinite devolatilisation remain poorly understood. This study presents new field observations alongside petrographic and geochemical data for metamorphic veins and host serpentinite from the Zermatt-Saas ophiolite from the Swiss Alps, which underwent high-pressure metamorphism during the Alpine orogeny. Samples were collected from the serpentinised ultramafic section of the Zermatt-Saas ophiolite, which is mainly comprised of variably foliated and sheared antigorite serpentine. High-pressure metamorphic veins hosted within the antigorite serpentinite, are observed within the least deformed part of the massif, occurring as cm scale laterally continuous channels or mm scale interconnected anastomosing networks. Preliminary high-precision Fe isotope data for the host antigorite serpentine yield a mean δ56Fe value of -0.09‰ ± 0.04‰ (n=3), notably lighter than previously measured Alpine and abyssal serpentinites (Debret et al., 2016). In contrast, samples of cm scale olivine-bearing veins display a mean δ56Fe value of 0.07 ± 0.05‰ (n=3), resolvably heavier than that of the host serpentinite. These preliminary results suggest preferential mobility of isotopically heavy Fe within the vein forming fluids, but at this stage it is unclear if this fluid is related to local devolatilisation of the host serpentinite or input from an external source. Debret et al., 2016. Isotopic evidence for iron mobility during subduction. Geology, v. 44, no. 3, pp. 215 -218.

  9. Cell cycle-dependent regulation of Aurora kinase B mRNA by the Microprocessor complex.

    PubMed

    Jung, Eunsun; Seong, Youngmo; Seo, Jae Hong; Kwon, Young-Soo; Song, Hoseok

    2014-03-28

    Aurora kinase B regulates the segregation of chromosomes and the spindle checkpoint during mitosis. In this study, we showed that the Microprocessor complex, which is responsible for the processing of the primary transcripts during the generation of microRNAs, destabilizes the mRNA of Aurora kinase B in human cells. The Microprocessor-mediated cleavage kept Aurora kinase B at a low level and prevented premature entrance into mitosis. The cleavage was reduced during mitosis leading to the accumulation of Aurora kinase B mRNA and protein. In addition to Aurora kinase B mRNA, the processing of other primary transcripts of miRNAs were also decreased during mitosis. We found that the cleavage was dependent on an RNA helicase, DDX5, and the association of DDX5 and DDX17 with the Microprocessor was reduced during mitosis. Thus, we propose a novel mechanism by which the Microprocessor complex regulates stability of Aurora kinase B mRNA and cell cycle progression. Copyright © 2014 Elsevier Inc. All rights reserved.

  10. Experimental induction of chicken amyloid A amyloidosis in white layer chickens by inoculation with inactivated vaccines.

    PubMed

    Habibi, Wazir Ahmad; Hirai, Takuya; Niazmand, Mohammad Hakim; Okumura, Naoko; Yamaguchi, Ryoji

    2017-10-01

    We investigated the amyloidogenic potential of inactivated vaccines and the localized production of serum amyloid A (SAA) at the injection site in white layer chickens. Hens in the treated group were injected intramuscularly three times with high doses of inactivated oil-emulsion Salmonella Enteritidis vaccine and multivalent viral and bacterial inactivated oil-emulsion vaccines at two-week intervals. Chickens in the control group did not receive any inoculum. In the treated group, emaciation and granulomas were present, while several chickens died between 4 and 6 weeks after the first injection. Hepatomegaly was seen at necropsy, and the liver parenchyma showed inconsistent discolouration with patchy green to yellowish-brown areas, or sometimes red-brown areas with haemorrhage. Amyloid deposition in the liver, spleen, duodenum, and at injection sites was demonstrated using haematoxylin and eosin staining, Congo red, and immunohistochemistry. The incidence of chicken amyloid A (AA) amyloidosis was 47% (28 of 60) in the treated group. In addition, RT-PCR was used to identify chicken SAA mRNA expression in the liver and at the injection sites. Furthermore, SAA mRNA was detected by in situ hybridization in fibroblasts at the injection sites, and also in hepatocytes. We believe that this is the first report of the experimental induction of systemic AA amyloidosis in white layer chickens following repeated inoculation with inactivated vaccines without the administration of amyloid fibrils or other amyloid-enhancing factors.

  11. Social defeat disrupts reward learning and potentiates striatal nociceptin/orphanin FQ mRNA in rats.

    PubMed

    Der-Avakian, Andre; D'Souza, Manoranjan S; Potter, David N; Chartoff, Elena H; Carlezon, William A; Pizzagalli, Diego A; Markou, Athina

    2017-05-01

    Mood disorders can be triggered by stress and are characterized by deficits in reward processing, including disrupted reward learning (the ability to modulate behavior according to past rewards). Reward learning is regulated by the anterior cingulate cortex (ACC) and striatal circuits, both of which are implicated in the pathophysiology of mood disorders. Here, we assessed in rats the effects of a potent stressor (social defeat) on reward learning and gene expression in the ACC, ventral tegmental area (VTA), and striatum. Adult male Wistar rats were trained on an operant probabilistic reward task (PRT) and then exposed to 3 days of social defeat before assessment of reward learning. After testing, the ACC, VTA, and striatum were dissected, and expression of genes previously implicated in stress was assessed. Social defeat blunted reward learning (manifested as reduced response bias toward a more frequently rewarded stimulus) and was associated with increased nociceptin/orphanin FQ (N/OFQ) peptide mRNA levels in the striatum and decreased Fos mRNA levels in the VTA. Moreover, N/OFQ peptide and nociceptin receptor mRNA levels in the ACC, VTA and striatum were inversely related to reward learning. The behavioral findings parallel previous data in humans, suggesting that stress similarly disrupts reward learning in both species. Increased striatal N/OFQ mRNA in stressed rats characterized by impaired reward learning is consistent with accumulating evidence that antagonism of nociceptin receptors, which bind N/OFQ, has antidepressant-like effects. These results raise the possibility that nociceptin systems represent a molecular substrate through which stress produces reward learning deficits in mood disorders.

  12. Identification of MSX1 and DCLK1 as mRNA Biomarkers for Colorectal Cancer Detection Through DNA Methylation Information.

    PubMed

    Sun, Ai-Jun; Gao, Hai-Bo; Liu, Gao; Ge, Heng-Fa; Ke, Zun-Ping; Li, Sen

    2017-07-01

    Colorectal cancer is the second most deadly malignancy in the United States. However, the currently screening options had their limitation. Novel biomarkers for colorectal cancer detections are necessary to reduce the mortality. The clinical information, mRNA expression levels and DNA methylation information of colorectal cancer were downloaded from TCGA. The patients were separated into training group and testing group based on their platforms for DNA methylation. Beta values of DNA methylation from tumor tissues and normal tissues were utilized to figure out the position that were differentially methylated. The expression levels of mRNA of thirteen genes, whose CpG islands were differentially methylated, were extracted from the RNA-Seq results from TCGA. The probabilities whether the mRNA was differentially expressed between tumor and normal samples were calculated using Student's t-test. Logistic regression and decision tree were built for cancer detection and their performances were evaluated by the area under the curve (AUC). Twenty-four genomic locations were differentially methylated, which could be mapped to eleven genes. Nine out of eleven genes had differentially expressed mRNA levels, which were used to build the model for cancer detection. The final detection models consisting of mRNA expression levels of these nine genes had great performances on both training group and testing group. The model that constructed in this study suggested MSX1 and DCLK1 might be used in colorectal cancer detection or as target of cancer therapies. J. Cell. Physiol. 232: 1879-1884, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  13. Assessment of potential biomarkers, metallothionein and vitellogenin mRNA expressions in various chemically exposed benthic Chironomus riparius larvae

    NASA Astrophysics Data System (ADS)

    Park, Kiyun; Kwak, Inn-Sil

    2012-12-01

    The objective of this study was conducted to identify the possibility of using Chironomus metallothionein (MT) and vitellogenin (VTG) as biomarkers of stress caused by endocrinedisrupting chemicals (EDCs), heavy metals, herbicides and veterinary antibiotics. We characterized the MT and VTG cDNA in Chironomus riparius and evaluated their mRNA expression profiles following exposure to different environmental pollutants. The gene expression analysis showed that the MT mRNA levels increased significantly after long-term exposure to cadmium (Cd), copper (Cu), Lead (Pb), di(2-ethylhexyl) phthalate (DEHP), and 2,4-dichlorophenoxyacetic acid (2,4-D). Moreover, the VTG mRNA expression increased significantly in C. riparius larvae exposed to BPA, NP, DEHP, Cd, 2,4-D and fenbendazole. Evaluation of the long-term effects of environmental pollutants revealed up regulation of Chironomus MT mRNA in response to DEHP exposure among EDCs, and the level of the VTG mRNA was increased significantly following treatment with Cd and herbicide 2,4-D at all concentrations in a dose-dependent manner. These results indicate that VTG could be used as a potential biomarker of herbicide and Cd as well as EDCs, while MT was a potential biomarker of heavy metals such as Cd, Cu, and Pb in aquatic environments.

  14. Ways and means of eukaryotic mRNA decay.

    PubMed

    Balagopal, Vidya; Fluch, Lydia; Nissan, Tracy

    2012-06-01

    Messenger RNA degradation is an important point of control for gene expression. Genome-wide studies on mRNA stability have demonstrated its importance in adaptation and stress response. Most of the key players in mRNA decay appear to have been identified. The study of these proteins brings insight into the mechanism of general and specific targeting of transcripts for degradation. Recruitment and assembly of mRNP complexes enhance and bring specificity to mRNA decay. mRNP complexes can form larger structures that have been found to be ubiquitous in nature. Discovery of P-Bodies, an archetype of this sort of aggregates, has generated interest in the question of where mRNA degrades. This is currently an open question under extensive investigation. This review will discuss in detail the recent developments in the regulation of mRNA decay focusing on yeast as a model system. This article is part of a Special Issue entitled: Nuclear Transport and RNA Processing. Copyright © 2012 Elsevier B.V. All rights reserved.

  15. UAP56 is a conserved crucial component of a divergent mRNA export pathway in Toxoplasma gondii.

    PubMed

    Serpeloni, Mariana; Jiménez-Ruiz, Elena; Vidal, Newton Medeiros; Kroeber, Constanze; Andenmatten, Nicole; Lemgruber, Leandro; Mörking, Patricia; Pall, Gurman S; Meissner, Markus; Ávila, Andréa R

    2016-11-01

    Nucleo-cytoplasmic RNA export is an essential post-transcriptional step to control gene expression in eukaryotic cells and is poorly understood in apicomplexan parasites. With the exception of UAP56, a component of TREX (Transcription Export) complex, other components of mRNA export machinery are not well conserved in divergent supergroups. Here, we use Toxoplasma gondii as a model system to functionally characterize TgUAP56 and its potential interaction factors. We demonstrate that TgUAP56 is crucial for mRNA export and that functional interference leads to significant accumulation of mRNA in the nucleus. It was necessary to employ bioinformatics and phylogenetic analysis to identify orthologs related to mRNA export, which show a remarkable low level of conservation in T. gondii. We adapted a conditional Cas9/CRISPR system to carry out a genetic screen to verify if these factors were involved in mRNA export in T. gondii. Only the disruption of TgRRM_1330 caused accumulation of mRNA in the nucleus as found with TgUAP56. This protein is potentially a divergent partner of TgUAP56, and provides insight into a divergent mRNA export pathway in apicomplexans. © 2016 The Authors. Molecular Microbiology Published by John Wiley & Sons Ltd.

  16. UAP56 is a conserved crucial component of a divergent mRNA export pathway in Toxoplasma gondii

    PubMed Central

    Serpeloni, Mariana; Jiménez‐Ruiz, Elena; Vidal, Newton Medeiros; Kroeber, Constanze; Andenmatten, Nicole; Lemgruber, Leandro; Mörking, Patricia; Pall, Gurman S.

    2016-01-01

    Summary Nucleo‐cytoplasmic RNA export is an essential post‐transcriptional step to control gene expression in eukaryotic cells and is poorly understood in apicomplexan parasites. With the exception of UAP56, a component of TREX (Transcription Export) complex, other components of mRNA export machinery are not well conserved in divergent supergroups. Here, we use Toxoplasma gondii as a model system to functionally characterize TgUAP56 and its potential interaction factors. We demonstrate that TgUAP56 is crucial for mRNA export and that functional interference leads to significant accumulation of mRNA in the nucleus. It was necessary to employ bioinformatics and phylogenetic analysis to identify orthologs related to mRNA export, which show a remarkable low level of conservation in T. gondii. We adapted a conditional Cas9/CRISPR system to carry out a genetic screen to verify if these factors were involved in mRNA export in T. gondii. Only the disruption of TgRRM_1330 caused accumulation of mRNA in the nucleus as found with TgUAP56. This protein is potentially a divergent partner of TgUAP56, and provides insight into a divergent mRNA export pathway in apicomplexans. PMID:27542978

  17. Differential expression of cytokeratin mRNA and protein in normal prostate, prostatic intraepithelial neoplasia, and invasive carcinoma.

    PubMed Central

    Yang, Y.; Hao, J.; Liu, X.; Dalkin, B.; Nagle, R. B.

    1997-01-01

    The expression of cytokeratin (CK) mRNA for CK5, -8, -14, -16, and -19 was investigated in normal prostate, prostatic intraepithelial neoplasia (PIN) lesions, and invasive carcinoma using in situ hybridization. Protein localization was carried out in adjacent sections using immunohistochemistry and correlated with mRNA expression. Snap-frozen human prostate samples including 22 examples of normal glands, 20 cases of PIN lesions, and 12 cases of invasive carcinoma were examined. CK5 and -14 mRNA and protein were prominently expressed only in the basal cells of normal glands and PIN lesions. CK14 mRNA was absent in the luminal cells of the most of the PIN lesions but was seen at a low level in some PIN lesions. CK14 protein was not detected in any PIN lesion, suggesting that, if the cell that makes up the PIN lesions is derived from a basal cell, CK14 translation is depressed although a low level of CK14 mRNA may persist. CK8 mRNA and protein were constitutively expressed in all epithelia of normal and abnormal prostate tissues. CK19 mRNA and protein were persistently expressed in both basal and luminal cells of the tubular portion of normal glands as well as PIN lesions, but were expressed heterogeneously in both basal and luminal cells of normal alveoli. CK16 mRNA was expressed in a similar pattern as CK19, but CK16 protein was not detected either in normal or in abnormal prostate tissues. In conclusion, the expression of CK19 in PIN lesions is similar to its tubular expression and would support an origin of PIN lesions from this structure rather than the alveolar portion of the glands. The similar cytokeratin expression between PIN lesions and invasive carcinoma further supports the concept that PIN is a precursor lesion of invasive carcinoma. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 PMID:9033282

  18. Evaluation of immune and stress status in harbour porpoises (Phocoena phocoena): can hormones and mRNA expression levels serve as indicators to assess stress?

    PubMed Central

    2013-01-01

    Background The harbour porpoise is exposed to increasing pressure caused by anthropogenic activities in its marine environment. Numerous offshore wind farms are planned or under construction in the North and Baltic Seas, which will increase underwater noise during both construction and operation. A better understanding of how anthropogenic impacts affect the behaviour, health, endocrinology, immunology and physiology of the animals is thus needed. The present study compares levels of stress hormones and mRNA expression of cytokines and acute-phase proteins in blood samples of harbour porpoises exposed to different levels of stress during handling, in rehabilitation or permanent human care. Free-ranging harbour porpoises, incidentally caught in pound nets in Denmark, were compared to harbour porpoises in rehabilitation at SOS Dolfijn in Harderwijk, the Netherlands, and individuals permanently kept in human care in the Dolfinarium Harderwijk and Fjord & Belt Kerteminde, Denmark. Blood samples were investigated for catecholamines, adrenaline, noradrenaline and dopamine, as well as for adrenocorticotropic hormone (ACTH), cortisol, metanephrine and normetanephrine. mRNA expression levels of relevant cell mediators (cytokines IL-10 and TNFα, acute-phase proteins haptoglobin and C-reactive protein and the heat shock protein HSP70) were measured using real-time PCR. Results Biomarker expression levels varied between free-ranging animals and porpoises in human care. Hormone and cytokine ranges showed correlations to each other and to the health status of investigated harbour porpoises. Hormone concentrations were higher in free-ranging harbour porpoises than in animals in human care. Adrenaline can be used as a parameter for the initial reaction to acute stress situations; noradrenaline, dopamine, ACTH and cortisol are more likely indicators for the following minutes of acute stress. There is evidence for different correlations between production of normetanephrine

  19. Occupational Toluene Exposure Induces Cytochrome P450 2E1 mRNA Expression in Peripheral Lymphocytes

    PubMed Central

    Mendoza-Cantú, Ania; Castorena-Torres, Fabiola; de León, Mario Bermúdez; Cisneros, Bulmaro; López-Carrillo, Lizbeth; Rojas-García, Aurora E.; Aguilar-Salinas, Alberto; Manno, Maurizio; Albores, Arnulfo

    2006-01-01

    Print workers are exposed to organic solvents, of which the systemic toxicant toluene is a main component. Toluene induces expression of cytochrome P450 2E1 (CYP2E1), an enzyme involved in its own metabolism and that of other protoxicants, including some procarcinogens. Therefore, we investigated the association between toluene exposure and the CYP2E1 response, as assessed by mRNA content in peripheral lymphocytes or the 6-hydroxychlorzoxazone (6OH-CHZ)/chlorzoxazone (CHZ) quotient (known as CHZ metabolic ratio) in plasma, and the role of genotype (5′-flanking region RsaI/PstI polymorphic sites) in 97 male print workers. The geometric mean (GM) of toluene concentration in the air was 52.80 ppm (10–760 ppm); 54% of the study participants were exposed to toluene concentrations that exceeded the maximum permissible exposure level (MPEL). The GM of urinary hippuric acid at the end of a work shift (0.041 g/g creatinine) was elevated relative to that before the shift (0.027 g/g creatinine; p < 0.05). The GM of the CHZ metabolic ratio was 0.33 (0–9.3), with 40% of the subjects having ratios below the GM. However, the average CYP2E1 mRNA level in peripheral lymphocytes was 1.07 (0.30–3.08), and CYP2E1 mRNA levels within subjects correlated with the toluene exposure ratio (environmental toluene concentration:urinary hippuric acid concentration) (p = 0.014). Genotype did not alter the association between the toluene exposure ratio and mRNA content. In summary, with further validation, CYP2E1 mRNA content in peripheral lymphocytes could be a sensitive and noninvasive biomarker for the continuous monitoring of toluene effects in exposed persons. PMID:16581535

  20. Colonization by non-pathogenic bacteria alters mRNA expression of cytochromes P450 in originally germ-free mice.

    PubMed

    Jourová, L; Anzenbacher, P; Lišková, B; Matušková, Z; Hermanová, P; Hudcovic, T; Kozáková, H; Hrnčířová, L; Anzenbacherová, E

    2017-11-01

    Gut microbiota provides a wide range of beneficial function for the host and has an immense effect on the host's health state. It has also been shown that gut microbiome is often involved in the biotransformation of xenobiotics; however, the molecular mechanisms of the interaction between the gut bacteria and the metabolism of drugs by the host are still unclear. To investigate the effect of microbial colonization on messenger RNA (mRNA) expression of liver cytochromes P450 (CYPs), the main drug-metabolizing enzymes, we used germ-free (GF) mice, lacking the intestinal flora and mice monocolonized by non-pathogenic bacteria Lactobacillus plantarum NIZO2877 or probiotic bacteria Escherichia coli Nissle 1917 compared to specific pathogen-free (SPF) mice. Our results show that the mRNA expression of Cyp1a2 and Cyp2e1 was significantly increased, while the expression of Cyp3a11 mRNA was decreased under GF conditions compared to the SPF mice. The both bacteria L. plantarum NIZO2877 and E. coli Nissle 1917 given to the GF mice decreased the level of Cyp1a2 mRNA and normalized it to the control level. On the other hand, the colonization by these bacteria had no effect on the expression of Cyp3a11 mRNA in the liver of the GF mice (which remained decreased). Surprisingly, monocolonization with chosen bacterial strains has shown a different effect on the expression of Cyp2e1 mRNA in GF mice. Increased level of Cyp2e1 expression observed in the GF mice was found also in mice colonized by L. plantarum NIZO2877 ; however, the colonization with probiotic E. coli Nissle 1917 caused a decrease in Cyp2e1 expression and partially restored the SPF mice conditions.

  1. Seasonal changes in hormone-sensitive and lipoprotein lipase mRNA concentrations in marmot white adipose tissue.

    PubMed

    Wilson, B E; Deeb, S; Florant, G L

    1992-02-01

    White adipose tissue (WAT) and plasma samples were obtained from yellow-bellied marmots (Marmota flaviventris) throughout the year. Mean plasma triacylglycerol (TG), free fatty acids (FFAs), and glycerol were determined. There was a clear increase in FFAs and decrease in mean TG and glycerol during the hibernation period when animals were fasting, suggesting increased lipolysis. RNA was isolated from WAT biopsies at four times in the year: spring, summer, fall, and winter. There were significant changes in the relative levels of mRNA for lipoprotein lipase (LPL) and hormone-sensitive lipase (HSL) during the body mass cycle of the marmot. The relative levels of LPL mRNA are high during the mass gain phase of the year and that of HSL mRNA are high during the fasting period when endogenous lipid is utilized. These results suggest that the genes for LPL and HSL are regulated seasonally to control the adipose mass depot in marmots.

  2. Relationship between mRNA secondary structure and sequence variability in Chloroplast genes: possible life history implications.

    PubMed

    Krishnan, Neeraja M; Seligmann, Hervé; Rao, Basuthkar J

    2008-01-28

    Synonymous sites are freer to vary because of redundancy in genetic code. Messenger RNA secondary structure restricts this freedom, as revealed by previous findings in mitochondrial genes that mutations at third codon position nucleotides in helices are more selected against than those in loops. This motivated us to explore the constraints imposed by mRNA secondary structure on evolutionary variability at all codon positions in general, in chloroplast systems. We found that the evolutionary variability and intrinsic secondary structure stability of these sequences share an inverse relationship. Simulations of most likely single nucleotide evolution in Psilotum nudum and Nephroselmis olivacea mRNAs, indicate that helix-forming propensities of mutated mRNAs are greater than those of the natural mRNAs for short sequences and vice-versa for long sequences. Moreover, helix-forming propensity estimated by the percentage of total mRNA in helices increases gradually with mRNA length, saturating beyond 1000 nucleotides. Protection levels of functionally important sites vary across plants and proteins: r-strategists minimize mutation costs in large genes; K-strategists do the opposite. Mrna length presumably predisposes shorter mRNAs to evolve under different constraints than longer mRNAs. The positive correlation between secondary structure protection and functional importance of sites suggests that some sites might be conserved due to packing-protection constraints at the nucleic acid level in addition to protein level constraints. Consequently, nucleic acid secondary structure a priori biases mutations. The converse (exposure of conserved sites) apparently occurs in a smaller number of cases, indicating a different evolutionary adaptive strategy in these plants. The differences between the protection levels of functionally important sites for r- and K-strategists reflect their respective molecular adaptive strategies. These converge with increasing domestication levels of

  3. Coordinated modulation of albumin synthesis and mRNA levels in cultured hepatoma cells by hydrocortisone and cyclic AMP analogs

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Brown, P.C.; Papaconstantinou, J.

    The treatment of Hepa-2 cells, a permanent mouse hepatoma cell line, for 72 h with hydrocortisone (10/sup -6/ M), N/sup 6/,O/sup 2/-dibutyryl cyclic AMP (10/sup -3/ M), or 8-bromo cyclic AMP(10/sup -3/ M) results in a 2-, 3-, or 4-fold increase, respectively, in rates of synthesis and secretion of mouse serum albumin. Simultaneous treatment with hydrocortisone and N/sup 6/,O/sup 2/-dibutyryl cyclic AMP results in a 10-fold stimulation in these parameters, an effect that is significantly more than additive for the two compounds tested. The number of albumin mRNA sequences, determined by hybridization of total cell RNA to albumin complementary DNA,more » was increased in direct proportion to the increases in albumin synthesis in all experiments. The relative rate of albumin synthesis approaches in vivo levels in cells treated simultaneously with hydrocortisone and N/sup 6/,O/sup 2/-dibutyryl cyclic AMP. We propose that these factors may be necessary to maintain the maximal level of differentiated function in the continuous culture of Hepa-2 cells.« less

  4. Wig1 prevents cellular senescence by regulating p21 mRNA decay through control of RISC recruitment

    PubMed Central

    Kim, Bong Cho; Lee, Hyung Chul; Lee, Je-Jung; Choi, Chang-Min; Kim, Dong-Kwan; Lee, Jae Cheol; Ko, Young-Gyu; Lee, Jae-Seon

    2012-01-01

    Premature senescence, a key strategy used to suppress carcinogenesis, can be driven by p53/p21 proteins in response to various stresses. Here, we demonstrate that Wig1 plays a critical role in this process through regulation of p21 mRNA stability. Wig1 controls the association of Argonaute2 (Ago2), a central component of the RNA-induced silencing complex (RISC), with target p21 mRNA via binding of the stem-loop structure near the microRNA (miRNA) target site. Depletion of Wig1 prohibited miRNA-mediated p21 mRNA decay and resulted in premature senescence. Wig1 plays an essential role in cell proliferation, as demonstrated in tumour xenografts in mice, and Wig1 and p21 mRNA levels are inversely correlated in human normal and cancer tissues. Together, our data indicate a novel role of Wig1 in RISC target accessibility, which is a key step in RNA-mediated gene silencing. In addition, these findings indicate that fine-tuning of p21 levels by Wig1 is essential for the prevention of cellular senescence. PMID:23085987

  5. Wig1 prevents cellular senescence by regulating p21 mRNA decay through control of RISC recruitment.

    PubMed

    Kim, Bong Cho; Lee, Hyung Chul; Lee, Je-Jung; Choi, Chang-Min; Kim, Dong-Kwan; Lee, Jae Cheol; Ko, Young-Gyu; Lee, Jae-Seon

    2012-11-14

    Premature senescence, a key strategy used to suppress carcinogenesis, can be driven by p53/p21 proteins in response to various stresses. Here, we demonstrate that Wig1 plays a critical role in this process through regulation of p21 mRNA stability. Wig1 controls the association of Argonaute2 (Ago2), a central component of the RNA-induced silencing complex (RISC), with target p21 mRNA via binding of the stem-loop structure near the microRNA (miRNA) target site. Depletion of Wig1 prohibited miRNA-mediated p21 mRNA decay and resulted in premature senescence. Wig1 plays an essential role in cell proliferation, as demonstrated in tumour xenografts in mice, and Wig1 and p21 mRNA levels are inversely correlated in human normal and cancer tissues. Together, our data indicate a novel role of Wig1 in RISC target accessibility, which is a key step in RNA-mediated gene silencing. In addition, these findings indicate that fine-tuning of p21 levels by Wig1 is essential for the prevention of cellular senescence.

  6. Putative Role of Serum Amyloid-A and Proinflammatory Cytokines as Biomarkers for Behcet's Disease

    PubMed Central

    Lopalco, Giuseppe; Lucherini, Orso Maria; Vitale, Antonio; Talarico, Rosaria; Lopalco, Antonio; Galeazzi, Mauro; Lapadula, Giovanni; Cantarini, Luca; Iannone, Florenzo

    2015-01-01

    Abstract Behcet's disease (BD) is a multisystemic disorder of unknown etiology characterized by relapsing oral–genital ulcers, uveitis, and involvement of vascular, gastrointestinal, neurological, and musculoskeletal system. Although disease pathogenesis is still unclear, both innate and adaptive immunity have shown to play a pivotal role, and multiple proinflammatory cytokines seem to be involved in different pathogenic pathways that eventually lead to tissue damage. The aims of our study were to evaluate serum cytokines levels of IL-8, IL-18, IFN-α2a, IL-6, IFN-γ, CXCL10, CXCL11, CXCL9, and SAA levels in patients with BD, in comparison to healthy controls (HC), and to correlate their levels to disease activity. We included 78 serum samples obtained from 58 BD patients and analyzed a set of proinflammatory cytokines including IL-8, IL-18, IFN-α2a, IL-6, IFN-γ, CXCL10, CXCL11, and CXCL9 by multiplex bead analysis as well as SAA by enzyme-linked immunosorbent assay. Compared to HC, BD patients showed elevated cytokine levels of IL-8, IL-18, IFN-α2a, and IL-6, and low levels of CXCL11. BD patients with SAA serum levels >20 mg/L showed higher levels of proinflammatory markers than HC or group with SAA ≤20 mg/L. IL-18, IFN-α2a, and IL-6 were higher in BD group with SAA >20 mg/L than HC, while IL-8 and CXCL9 levels were higher than in patients with SAA ≤20 mg/L and HC. Active BD patients with SAA >20 mg/L exhibited elevated levels of inflammatory mediators, suggesting that may exist a relationship between SAA and proinflammatory cytokines in the intricate scenario of BD pathogenesis. PMID:26496336

  7. Posttranscriptional mRNA processing as a mechanism for regulation of human A1 adenosine receptor expression.

    PubMed Central

    Ren, H; Stiles, G L

    1994-01-01

    The human A1 adenosine receptor gene contains six exons with exons 1, 2, 3, 4, and part of 5 representing 5' untranslated regions. Reverse transcription-PCR with exon-specific primers showed two distinct transcripts containing either exons 3, 5, and 6 or exons 4, 5, and 6, with exons 3 and 4 being mutually exclusive. No mature mRNAs containing exons 1 and 2 have been detected. All human tissues that express any A1 receptors contain mRNA with exons 4, 5, and 6. Tissues which express high levels of A1 receptors contain mRNA with exons 3, 5, and 6. Exon 4 contains two upstream ATG codons whereas exon 3 contains none. COS cells transfected with expression vectors containing exon 4 (exons 1-6, 3-6, or Ex4-6) express much lower levels of A1 receptors than vectors without exon 4 (exons 3, 5, and 6). Mutation of upstream ATG codons in exon 4 leads to 3- to 7-fold increased A1 receptor expression, up to the level seen with the construct containing exons 3, 5, and 6. Thus, in human tissues "basal" levels of A1 receptors can be expressed by use of mRNA containing exons 4, 5, and 6, but when high levels are needed, alternative transcripts with exons 3, 5, and 6 are produced. Images PMID:8197148

  8. Expression of insulin-like growth factor I receptors at mRNA and protein levels during metamorphosis of Japanese flounder (Paralichthys olivaceus).

    PubMed

    Zhang, Junling; Shi, Zhiyi; Cheng, Qi; Chen, Xiaowu

    2011-08-01

    Insulin-like growth factor I (IGF-I) is an important regulator of fish growth and development, and its biological actions are initiated by binding to IGF-I receptor (IGF-IR). Our previous study has revealed that IGF-I could play an important role during metamorphosis of Japanese flounder, Paralichthys olivaceus. The analysis of IGF-IR expression thus helps further elucidate the IGF-I regulation of metamorphic processes. In this study, the spatial-temporal expression of two distinct IGF-IR mRNAs was investigated by real-time RT-PCR. The spatial distribution of two IGF-IR mRNAs in adult tissues is largely overlapped, but they exhibit distinct temporal expression patterns during larval development. A remarkable decrease in IGF-IR-2 mRNA was detected during metamorphosis. In contrast, a significant increase in IGF-IR-1 mRNA was determined from pre-metamorphosis to metamorphic completion. These indicate that they may play different function roles during the flounder metamorphosis. The levels and localization of IGF-IR proteins during larval development were further studied by Western blotting and immunohistochemistry. Immunoreactive IGF-IRs were detected throughout larval development, and the IGF-IR proteins displayed a relatively abundant expression during metamorphosis. Moreover, the IGF-IR proteins appeared in key tissues, such as thickened skin beneath the migrating eye, developing intestine, gills and kidney during metamorphosis. These results further suggest that the IGF-I system may be involved in metamorphic development of Japanese flounder. Copyright © 2011 Elsevier Inc. All rights reserved.

  9. Hepatic chemerin mRNA in morbidly obese patients with nonalcoholic fatty liver disease.

    PubMed

    Kajor, Maciej; Kukla, Michał; Waluga, Marek; Liszka, Łukasz; Dyaczyński, Michał; Kowalski, Grzegorz; Żądło, Dominika; Berdowska, Agnieszka; Chapuła, Mateusz; Kostrząb-Zdebel, Anna; Bułdak, Rafał J; Sawczyn, Tomasz; Hartleb, Marek

    The aim of this study was to investigate hepatic chemerin mRNA, serum chemerin concentration, and immunohistochemical staining for chemerin and and chemokine receptor-like 1 (CMKLR1) in hepatic tissue in 56 morbidly obese women with nonalcoholic fatty liver disease (NAFLD) and to search for a relationship with metabolic and histopathological features. Chemerin mRNA was assessed by quantitative real-time PCR, chemerin, and CMKLR1 immunohistochemical expression with specific antibodies, while serum chemerin concentration was assessed with commercially available enzyme-linked immunosorbent assays. Serum chemerin concentration reached 874.1 ±234.6 ng/ml. There was no difference in serum chemerin levels between patients with BMI < 40 kg/m2 and ≥ 40 kg/m2. Serum chemerin concentration tended to be higher in patients with hepatocyte ballooning, greater extent of steatosis, and definite nonalcoholic steatohepatitis (NASH). Liver chemerin mRNA was observed in all included patients and was markedly, but insignificantly, higher in those with BMI ≥ 40 kg/m2, hepatocyte ballooning, greater extent of steatosis, and definite NASH. Hepatic chemerin mRNA might be a predictor of hepatic steatosis, hepatocyte ballooning, and NAFLD activity score (NAS) but seemed not to be a primary driver regulating liver necroinflammatory activity and fibrosis. The lack of association between serum chemerin and hepatic chemerin mRNA may suggest that adipose tissue but not the liver is the main source of chemerin in morbidly obese women.

  10. [Experimental study on the Der f 1 mRNA molecules derived from dermatophagoides farinae for specific immunotherapy on murine model of asthma].

    PubMed

    Jiang, Yu-xin; Yin, Kang; Jin, Wen-jie; Wu, Lu-yi; Li, Chao-pin

    2014-08-01

    To investigate the effect of Der f 1 mRNA molecules for specific immunotherapy on murine model of asthma. Fifty BALB/c mice were randomly divided into 5 groups: PBS group, Der f 1 sensitization group, Der f 1 specific immunotherapy (SIT) group, beta-actin mRNA SIT group, and Derf 1 mRNA SIT group. On days 0, 7 and 14, mice in PBS group received PBS injection; mice in the other groups were intraperitoneally injected with 10 microg Derf 1. At day 21, the mice in the 4 experimental groups were challenged with a 30-min inhaled dose of Der f 1 (100 microg/ml) for 7 successive days. Two weeks after the final sensitization, the mice in the above five groups were im- munized by intradermal injection with PBS, 1 microg Der f 1, 10 microg Der f 1, 2 microg beta-actin mRNA, and 2 microg Der f 1 mRNA, respectively for 3 times at one-week intervals. Two weeks after the last intradermal injection, all mice were sacrificed and bronchoalveolar lavage fluid (BALF) was collected. ELISA was performed to detect the levels of IFN-gamma and IL-13 in BALF, the number of eosinophils in the BALF was recorded. Splenocytes were prepared, and cultured with Der f 1 al- lergen (10 Jg/ml) for 72 h. Splenocytes of PBS group was cultured without Derf 1 allergen. The levels of IFN-gamma and IL-13 in splenocyte culture supernatant were measured by ELISA, as well as serum antibody levels of total IgE, allergen- specific IgE (sIgE), sIgG1, and sIgG2a. Lung sections were stained in hematoxylin and eosin, and observed under the microsope. Except for PBS group, mice in the other 4 group showed symptoms of acute asthma attack. Com- pared with Derf 1 sensitization group [(897.56 +/- 105.73) pg/ml] and beta-actin mRNA SIT group [(219.47 +/- 64.72) pg/ml], the level of IFN-gamma in BALF from Der f 1 mRNA SIT group [(897.56 +/- 105.73) pg/ml] and Derfl SIT group [(864.48 +/- 70.62)pg/ml] significantly increased (P<0.01). However, the level of IL-13 in BALF from Derf 1 mRNA SIT group [(241.64 +/- 31.41) pg

  11. MiR-506 suppresses liver cancer angiogenesis through targeting sphingosine kinase 1 (SPHK1) mRNA.

    PubMed

    Lu, Zhanping; Zhang, Weiying; Gao, Shan; Jiang, Qiulei; Xiao, Zelin; Ye, Lihong; Zhang, Xiaodong

    MicroRNAs acting as oncogenes or tumor suppressor genes play crucial roles in human cancers. Sphingosine kinase 1 (SPHK1) and its metabolite sphingosine 1-phosphate (S1P) contribute to tumor angiogenesis. We have reported that the down-regulation of miR-506 targeting YAP mRNA results in the hepatocarcinogenesis. In the present study, we report a novel function of miR-506, which suppresses tumor angiogenesis through targeting SPHK1 mRNA in liver cancer. Bioinformatics analysis showed that miR-506 might target 3'-untranslated region (3'UTR) of SPHK1 mRNA. Then, we validated that by luciferase reporter gene assays. MiR-506 was able to reduce the expression of SPHK1 at the levels of mRNA and protein using reverse transcription-polymerase chain reaction and Western blot analysis in hepatoma HepG2 cells. Functionally, human umbilical vein endothelial cell (HUVEC) tube formation assays demonstrated that the forced miR-506 expression remarkably inhibited the production of S1P in the supernatant of hepatoma cells. The supernatant resulted in the inhibition of tumor angiogenesis. Interestingly, the supernatant with overexpression of SPHK1 could rescue the inhibition of angiogenesis of liver cancer mediated by miR-506. Anti-miR-506 increased the production of S1P in the supernatant of hepatoma cells, but the supernatant with silencing of SPHK1 abolished anti-miR-506-induced acceleration of tumor angiogenesis. Clinically, we observed that the levels of miR-506 were negatively related to those of SPHK1 mRNA in liver cancer tissues. Thus, we conclude that miR-506 depresses the angiogenesis of liver cancer through targeting 3'UTR of SPHK1 mRNA. Our finding provides new insights into the mechanism of tumor angiogenesis. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Staufen-mediated mRNA decay.

    PubMed

    Park, Eonyoung; Maquat, Lynne E

    2013-01-01

    Staufen1 (STAU1)-mediated mRNA decay (SMD) is an mRNA degradation process in mammalian cells that is mediated by the binding of STAU1 to a STAU1-binding site (SBS) within the 3'-untranslated region (3'-UTR) of target mRNAs. During SMD, STAU1, a double-stranded (ds) RNA-binding protein, recognizes dsRNA structures formed either by intramolecular base pairing of 3'-UTR sequences or by intermolecular base pairing of 3'-UTR sequences with a long-noncoding RNA (lncRNA) via partially complementary Alu elements. Recently, STAU2, a paralog of STAU1, has also been reported to mediate SMD. Both STAU1 and STAU2 interact directly with the ATP-dependent RNA helicase UPF1, a key SMD factor, enhancing its helicase activity to promote effective SMD. Moreover, STAU1 and STAU2 form homodimeric and heterodimeric interactions via domain-swapping. Because both SMD and the mechanistically related nonsense-mediated mRNA decay (NMD) employ UPF1; SMD and NMD are competitive pathways. Competition contributes to cellular differentiation processes, such as myogenesis and adipogenesis, placing SMD at the heart of various physiologically important mechanisms. Copyright © 2013 John Wiley & Sons, Ltd.

  13. Staufen-mediated mRNA decay

    PubMed Central

    Park, Eonyoung; Maquat, Lynne E.

    2013-01-01

    Staufen1 (STAU1)-mediated mRNA decay (SMD) is an mRNA degradation process in mammalian cells that is mediated by the binding of STAU1 to a STAU1-binding site (SBS) within the 3'-untranslated region (3'UTR) of target mRNAs. During SMD, STAU1, a double-stranded (ds) RNA-binding protein, recognizes dsRNA structures formed either by intramolecular base-pairing of 3'UTR sequences or by intermolecular base-pairing of 3'UTR sequences with a long noncoding RNA (lncRNA) via partially complementary Alu elements. Recently, STAU2, a paralog of STAU1, has also been reported to mediate SMD. Both STAU1 and STAU2 interact directly with the ATP-dependent RNA helicase UPF1, a key SMD factor, enhancing its helicase activity to promote effective SMD. Moreover, STAU1 and STAU2 form homodimeric and heterodimeric interactions via domain-swapping. Since both SMD and the mechanistically related nonsense-mediated mRNA decay (NMD) employ UPF1, SMD and NMD are competitive pathways. Competition contributes to cellular differentiation processes, such as myogenesis and adipogenesis, placing SMD at the heart of various physiologically important mechanisms. PMID:23681777

  14. Genome level analysis of rice mRNA 3′-end processing signals and alternative polyadenylation

    PubMed Central

    Shen, Yingjia; Ji, Guoli; Haas, Brian J.; Wu, Xiaohui; Zheng, Jianti; Reese, Greg J.; Li, Qingshun Quinn

    2008-01-01

    The position of a poly(A) site of eukaryotic mRNA is determined by sequence signals in pre-mRNA and a group of polyadenylation factors. To reveal rice poly(A) signals at a genome level, we constructed a dataset of 55 742 authenticated poly(A) sites and characterized the poly(A) signals. This resulted in identifying the typical tripartite cis-elements, including FUE, NUE and CE, as previously observed in Arabidopsis. The average size of the 3′-UTR was 289 nucleotides. When mapped to the genome, however, 15% of these poly(A) sites were found to be located in the currently annotated intergenic regions. Moreover, an extensive alternative polyadenylation profile was evident where 50% of the genes analyzed had more than one unique poly(A) site (excluding microheterogeneity sites), and 13% had four or more poly(A) sites. About 4% of the analyzed genes possessed alternative poly(A) sites at their introns, 5′-UTRs, or protein coding regions. The authenticity of these alternative poly(A) sites was partially confirmed using MPSS data. Analysis of nucleotide profile and signal patterns indicated that there may be a different set of poly(A) signals for those poly(A) sites found in the coding regions. Based on the features of rice poly(A) signals, an updated algorithm termed PASS-Rice was designed to predict poly(A) sites. PMID:18411206

  15. Temporal and Spatial Post-Transcriptional Regulation of Zebrafish Tie1 mRNA by Long Noncoding RNA During Brain Vascular Assembly.

    PubMed

    Chowdhury, Tamjid A; Koceja, Chris; Eisa-Beygi, Shahram; Kleinstiver, Benjamin P; Kumar, Suresh N; Lin, Chien-Wei; Li, Keguo; Prabhudesai, Shubhangi; Joung, J Keith; Ramchandran, Ramani

    2018-05-03

    Tie1 (tyrosine kinase containing immunoglobulin and epidermal growth factor homology 1), an endothelial and hematopoietic cell-specific receptor tyrosine kinase, is an important regulator of angiogenesis and critical for maintaining vascular integrity. The post-transcriptional regulation of tie1 mRNA expression is not understood, but it might partly explain Tie1's differential expression pattern in endothelium. Following up on our previous work that identified natural antisense transcripts from the tie1 locus- tie1 antisense ( tie1AS ), which regulates tie1 mRNA levels in zebrafish-we attempted to identify the mechanism of this regulation. Through in vitro and in vivo ribonucleoprotein binding studies, we demonstrated that tie1AS long noncoding RNA interacts with an RNA binding protein-embryonic lethal and abnormal vision Drosophila-like 1 (Elavl1)-that regulates tie1 mRNA levels. When we disrupted the interaction between tie1AS and Elavl1 by using constitutively active antisense morpholino oligonucleotides or photoactivatable morpholino oligonucleotides, tie1 mRNA levels increased between 26 and 31 hours post-fertilization, particularly in the head. This increase correlated with dilation of primordial midbrain channels, smaller eyes, and reduced ventricular space. We also observed these phenotypes when we used CRISPR (clustered regularly interspaced short palindromic repeats)-mediated CRISPRi (CRISPR-mediated interference) to knock down tie1AS . Treatment of the morpholino oligonucleotide-injected embryos with a small molecule that decreased tie1 mRNA levels rescued all 3 abnormal phenotypes. We identified a novel mode of temporal and spatial post-transcriptional regulation of tie1 mRNA. It involves long noncoding RNA, tie1AS, and Elavl1 (an interactor of tie1AS ). © 2018 American Heart Association, Inc.

  16. Increased expression of glutamic acid decarboxylase mRNA in rat substantia nigra after an ibotenic acid lesion in the caudate-putamen.

    PubMed

    Lindefors, N; Brené, S; Persson, H

    1990-04-01

    In situ hybridization histochemistry and RNA blots were used to study expression of glutamic acid decarboxylase (GAD) mRNA in rat caudate-nucleus and substantia nigra. In situ hybridization combined with computerized image analysis revealed that in the intact substantia nigra reticulata the cross-section area of GAD mRNA positive neurons were 25% larger in the dorsolateral part as compared with the ventromedial part. A unilateral ibotenic acid injection in caudate-putamen lesioned neurons, some of which project to the ipsilateral substantia nigra. An increased level of GAD mRNA was observed in substantia nigra ipsilateral to the lesion. Computerized image analysis of sections from in situ hybridization revealed an increase in the number of silver grains over GAD mRNA positive neurons in the dorsolateral substantia nigra reticulata ipsilateral to the lesion. However, no change was observed in the ventromedial part suggesting that GAD mRNA expression in this part of the nigra is less sensitive to inhibition by caudate-putamen afferents. In agreement with in situ experiments, RNA blots showed a 2-fold increased level of GAD mRNA in substantia nigra ipsilateral to the lesion. The increased GAD mRNA expression in the deafferented substantia nigra suggests a disinhibition of nigral GABA neurons, resulting in an increased utilization of GABA in these substantia nigra neurons.

  17. KSRP Modulation of GAP-43 mRNA Stability Restricts Axonal Outgrowth in Embryonic Hippocampal Neurons

    PubMed Central

    Bird, Clark W.; Gardiner, Amy S.; Bolognani, Federico; Tanner, Daniel C.; Chen, Ching-Yi; Lin, Wei-Jye; Yoo, Soonmoon; Twiss, Jeffery L.; Perrone- Bizzozero, Nora

    2013-01-01

    The KH-type splicing regulatory protein (KSRP) promotes the decay of AU-rich element (ARE)-containing mRNAs. Although KSRP is expressed in the nervous system, very little is known about its role in neurons. In this study, we examined whether KSRP regulates the stability of the ARE-containing GAP-43 mRNA. We found that KSRP destabilizes this mRNA by binding to its ARE, a process that requires the presence of its fourth KH domain (KH4). Furthermore, KSRP competed with the stabilizing factor HuD for binding to these sequences. We also examined the functional consequences of KSRP overexpression and knockdown on the differentiation of primary hippocampal neurons in culture. Overexpression of full length KSRP or KSRP without its nuclear localization signal hindered axonal outgrowth in these cultures, while overexpression of a mutant protein without the KH4 domain that has less affinity for binding to GAP-43′s ARE had no effect. In contrast, depletion of KSRP led to a rise in GAP-43 mRNA levels and a dramatic increase in axonal length, both in KSRP shRNA transfected cells and neurons cultured from Ksrp+/− and Ksrp −/−embryos. Finally we found that overexpression of GAP-43 rescued the axonal outgrowth deficits seen with KSRP overexpression, but only when cells were transfected with GAP-43 constructs containing 3′ UTR sequences targeting the transport of this mRNA to axons. Together, our results suggest that KSRP is an important regulator of mRNA stability and axonal length that works in direct opposition to HuD to regulate the levels of GAP-43 and other ARE-containing neuronal mRNAs. PMID:24244461

  18. Stimulation by thyroid-stimulating hormone and Grave's immunoglobulin G of vascular endothelial growth factor mRNA expression in human thyroid follicles in vitro and flt mRNA expression in the rat thyroid in vivo.

    PubMed

    Sato, K; Yamazaki, K; Shizume, K; Kanaji, Y; Obara, T; Ohsumi, K; Demura, H; Yamaguchi, S; Shibuya, M

    1995-09-01

    To elucidate the pathogenesis of thyroid gland hypervascularity in patients with Graves' disease, we studied the expression of mRNAs for vascular endothelial growth factor (VEGF) and its receptor, Flt family, using human thyroid follicles in vitro and thiouracil-fed rats in vivo. Human thyroid follicles, cultured in the absence of endothelial cells, secreted de novo-synthesized thyroid hormone in response to thyroid-stimulating hormone (TSH) and Graves' IgG. The thyroid follicles produced VEGF mRNA but not flt-1 mRNA. The expression of VEGF mRNA was enhanced by insulin, tumor-promoting phorbol ester, calcium ionophore, dibutyryl cAMP, TSH, and Graves' IgG. When rats were fed thiouracil for 4 wk, their serum levels of TSH were increased at day 3. VEGF mRNA was also increased on day 3, accompanied by an increase in flt family (flt-1 and KDR/ flk-1) mRNA expression. These in vitro and in vivo findings suggest that VEGF is produced by thyroid follicles in response to stimulators of TSH receptors, via the protein kinase A and C pathways. VEGF, a secretable angiogenesis factor, subsequently stimulates Flt receptors on endothelial cells in a paracrine manner, leading to their proliferation and producing hypervascularity of the thyroid gland, as seen in patients with Graves' disease.

  19. Post-burn hypertrophic scars are characterized by high levels of IL-1β mRNA and protein and TNF-α type I receptors.

    PubMed

    Salgado, Rosa M; Alcántara, Luz; Mendoza-Rodríguez, C Adriana; Cerbón, Marco; Hidalgo-González, Christian; Mercadillo, Patricia; Moreno, Luis M; Alvarez-Jiménez, Ricardo; Krötzsch, Edgar

    2012-08-01

    Post-burn hypertrophic scars are characterized by increased collagen synthesis and hyperplasia, and may be associated with erythema, pain, dysesthesia, pruritus, and skin border elevation. Although the etiopathogenesis of hypertrophic scarring remains unclear, proinflammatory and profibrogenic cytokines are known to play an important role in general skin dysfunction. This study assessed mRNA expression, proteins, and type I receptors of tumor necrosis factor-alpha (TNF-α) and interleukin 1-beta (IL-1β) in normal skin, normotrophic and post-burn hypertrophic scars. Skin biopsies were obtained from 10 hypertrophic and 9 normotrophic scars, and 4 normal skin sites. Only post-burn scars covering more than 10% of the body were included. Ex vivo histopathological analysis evaluated scar maturity, in situ hybridization assessed mRNA expression, and cytokine protein and cytokine/cell colocalization were performed using single- and double-label immunohistochemistry, respectively. IL-1β is overexpressed in hypertrophic scars at the post-transcriptional level, associated primarily with keratinocytes and CD1a(+) cells. Type I receptors for TNF-α are overexpressed in blood vessels of hypertrophic scars. The coordinated overexpression of IL-1β and TNF-α type I receptor may maintain the fibrogenic phenotypes of hypertrophic scars, even those in "remission". Copyright © 2011 Elsevier Ltd and ISBI. All rights reserved.

  20. Molecular effects of leptin on peroxisome proliferator activated receptor gamma (PPAR-γ) mRNA expression in rat's adipose and liver tissue.

    PubMed

    Abbasi, A; Moghadam, A A; Kahrarian, Z; Abbsavaran, R; Yari, K; Alizadeh, E

    2017-08-15

    Leptin is a 16-kDa peptide hormone secreted by adipose tissue that participates in the regulation of energy homeostasis. The aim of this study was to determine the effect of leptin injection on mRNA expression of peroxisome proliferator-activated receptor gamma (PPAR-γ) and comparison of PPAR-γ mRNA expression in rat's adipose and liver tissue. Twenty adult male rats were divided into the following groups: Group 1asa control (n=10) that did not receive any treatment. Group 2as a treatment (n=10) that received leptin (30 µg ⁄ kg BW) intraperitoneally (ip) for two successive days. Blood samples were taken before and one day after second leptin injection for triglyceride (TG), Free Fatty Acid (FFA), HLD-cholesterol, and LDL-cholesterol measurement. Total RNA was extractedfrom the adipose tissue and liver tissues of rats.  Adipose and liver tissue cells' cDNA was synthesized to characterize the expression of PPAR-γ. Gene expression of PPAR-γ mRNA was tested by RT- PCR technique. Results show leptin decreases expression of PPAR-γ on rat. Low levels of PPAR-γ mRNA were detected in adipose and liver tissues of treatment rats in comparison to control group. In treatment group, the level of PPAR-γ mRNA in liver tissue was very lower than the adipose tissue. The levels of HDL and FFA in treatment rats were increased whereas serum levels TG, VLDL and LDL were not changed. It is concluded that leptin signal with suppressing of PPAR-γ mRNA expression in rat's adipose and liver tissues can result in lipolysis instead of lipogenesis.

  1. Comparison of Glomerular and Podocyte mRNA Profiles in Streptozotocin-Induced Diabetes

    PubMed Central

    Fu, Jia; Wei, Chengguo; Lee, Kyung; Zhang, Weijia; He, Wu; Chuang, Peter

    2016-01-01

    Evaluating the mRNA profile of podocytes in the diabetic kidney may indicate genes involved in the pathogenesis of diabetic nephropathy. To determine if the podocyte-specific gene information contained in mRNA profiles of the whole glomerulus of the diabetic kidney accurately reflects gene expression in the isolated podocytes, we crossed Nos3−/− IRG mice with podocin-rtTA and TetON-Cre mice for enhanced green fluorescent protein labeling of podocytes before diabetic injury. Diabetes was induced by streptozotocin, and mRNA profiles of isolated glomeruli and sorted podocytes from diabetic and control mice were examined 10 weeks later. Expression of podocyte-specific markers in glomeruli was downregulated in diabetic mice compared with controls. However, expression of these markers was not altered in sorted podocytes from diabetic mice. When mRNA levels of glomeruli were corrected for podocyte number per glomerulus, the differences in podocyte marker expression disappeared. Analysis of the differentially expressed genes in diabetic mice also revealed distinct upregulated pathways in the glomeruli (mitochondrial function, oxidative stress) and in podocytes (actin organization). In conclusion, our data suggest reduced expression of podocyte markers in glomeruli is a secondary effect of reduced podocyte number, thus podocyte-specific gene expression detected in the whole glomerulus may not represent that in podocytes in the diabetic kidney. PMID:26264855

  2. Developmental expression of VGF mRNA in the prenatal and postnatal rat.

    PubMed

    Snyder, S E; Pintar, J E; Salton, S R

    1998-04-27

    VGF is a developmentally regulated, secretory peptide precursor that is expressed by neurons and neuroendocrine cells and that has its transcription and secretion induced rapidly by neurotrophins and by depolarization. To gain insight into the possible functions and regulation of VGF in vivo, we have characterized the distribution of VGF mRNA in the developing rat nervous system. VGF expression was first detectable at embryonic day 11.5 in the primordia of cranial, sympathetic, and dorsal root ganglia, and its distribution expanded throughout development to include significant expression throughout the brain, spinal cord, and retina of the adult rat. The earliest expression of VGF, therefore, appeared in the peripheral nervous system as developing neurons settled in their designated ganglia. In many regions of the brain, VGF mRNA levels were found to be highest during periods when axonal outgrowth and synaptogenesis predominate. Areas of the central nervous system that contain predominantly dividing cells never displayed any VGF mRNA expression, nor did the vast majority of nonneural tissues.

  3. mRNA vaccines — a new era in vaccinology

    PubMed Central

    Pardi, Norbert; Hogan, Michael J.; Porter, Frederick W.; Weissman, Drew

    2018-01-01

    mRNA vaccines represent a promising alternative to conventional vaccine approaches because of their high potency, capacity for rapid development and potential for low-cost manufacture and safe administration. However, their application has until recently been restricted by the instability and inefficient in vivo delivery of mRNA. Recent technological advances have now largely overcome these issues, and multiple mRNA vaccine platforms against infectious diseases and several types of cancer have demonstrated encouraging results in both animal models and humans. This Review provides a detailed overview of mRNA vaccines and considers future directions and challenges in advancing this promising vaccine platform to widespread therapeutic use. PMID:29326426

  4. Dietary cholesterol worsens adipose tissue macrophage accumulation and atherosclerosis in obese LDL receptor-deficient mice

    PubMed Central

    Subramanian, Savitha; Han, Chang Yeop; Chiba, Tsuyoshi; McMillen, Timothy S.; Wang, Shari A.; Haw, Antonio; Kirk, Elizabeth A.; O’Brien, Kevin D.; Chait, Alan

    2009-01-01

    Objective Chronic systemic inflammation accompanies obesity and predicts development of cardiovascular disease. Dietary cholesterol has been shown to increase inflammation and atherosclerosis in LDL receptor-deficient (LDLR-/-) mice. This study was undertaken to determine whether dietary cholesterol and obesity have additive effects on inflammation and atherosclerosis. Methods and Results LDLR-/- mice were fed chow, high fat, high carbohydrate (diabetogenic) diet without (DD) or with added cholesterol (DDC) for 24 weeks. Effects on adipose tissue, inflammatory markers and atherosclerosis were studied. Despite similar weight gain between DD and DDC groups, addition of dietary cholesterol increased insulin resistance relative to DD. Adipocyte hypertrophy, macrophage accumulation and local inflammation were observed in intra-abdominal adipose tissue in DD and DDC, but were significantly higher in the DDC group. Circulating levels of the inflammatory protein serum amyloid A (SAA) were 4.4-fold higher in DD animals and 15-fold higher in DDC animals than controls, suggesting chronic systemic inflammation. Hepatic SAA mRNA levels were similarly elevated. Atherosclerosis was increased in the DD-fed animals and further increased in the DDC group. Conclusions Obesity-induced macrophage accumulation in adipose tissue is exacerbated by dietary cholesterol. These local inflammatory changes in adipose tissue are associated with insulin resistance, systemic inflammation and increased atherosclerosis in this mouse model. PMID:18239153

  5. [Expression of heat shock protein 70 and its mRNA in career exposure to manganese].

    PubMed

    Chen, Wenwen; Shao, Hua; Chi, Mingfeng; Zhang, Zhihu; Shan, Yongle; Zou, Wei

    2015-10-01

    To analyze the expression levels of heat shock protein70 (HSPs70) and HSPs70 mRNA in different exposure to manganese, and research the neuroprotective effect on the career exposure to manganese. From 2008 to 2009, with cross-sectional study design, and in a locomotive and rolling stock works, by stratified random sampling method, the exposed sample consisted of 180 welders from different welding shops and 100 unexposed in the last three years, non-welder controls with age-matched workers of similar socioeconomic status from the same industry. The control workers had not been exposed to neurotoxic chemicals. The mRNA expressions of four different metabolic enzyme were detected by SYBR Green I quantitative real-time polymerase chain reaction. The expression levels of the two enzymes mRNA in different exposure to manganese were analyzed. The expressions of HSPs70 were detected by Western blot. The concentration of air manganese was determined by GFAAS. The average concentration of 8 h time (8h-TWA) was used to express the level of individual exposure to manganese, according to the air manganese workplace occupational exposure limit (8h-TWA=0.15 mg/m3), the exposed group is divided into high exposed group (>0.15 mg/m3) and low exposure group (<0.15 mg/m3). The individuals exposed to manganese dose of exposed group ((0.25±0.31) mg/m3) was higher than the control group ((0.06±0.02) mg/m3) (t=6.15, P=0.001); individuals exposed to manganese dose of high exposure group for (0.42±0.34) mg/m3, which was higher than low exposure group (0.09±0.07) mg/m3 (t=9.80, P=0.001). HSPs70 mRNA and protein of exposure group (5.65±0.21, 3.26±0.15) were higher than the reference group (0.41±0.03, 1.32±0.12) (t=18.91, t=8.68, P=0.001). HSP70 mRNA and protein of high exposure group (6.48±0.37, 3.67±0.26) were higher than the low exposure group (5.15±0.23, 3.02±0.19) (t=3.24, t=2.01, P=0.003, P=0.043). The expression of peripheral blood lymphocytes HSPs70 level and HSPs70 mRNA

  6. Reversible inhibition of PSD-95 mRNA translation by miR-125a, FMRP phosphorylation and mGluR signaling

    PubMed Central

    Muddashetty, Ravi S.; Nalavadi, Vijayalaxmi C.; Gross, Christina; Yao, Xiaodi; Xing, Lei; Laur, Oskar; Warren, Stephen T.; Bassell, Gary J.

    2011-01-01

    Summary The molecular mechanism how RISC and microRNAs selectively and reversibly regulate mRNA translation in response to receptor signaling is unknown but could provide a means for temporal and spatial control of translation. Here we show that miR-125a targeting PSD-95 mRNA allows reversible inhibition of translation and regulation by mGluR signaling. Inhibition of miR-125a increased PSD-95 levels in dendrites and altered dendritic spine morphology. Bidirectional control of PSD-95 expression depends on miR-125a and FMRP phosphorylation status. miR-125a levels at synapses and its association with AGO2 is reduced in Fmr1 KO. FMRP phosphorylation promotes the formation of an AGO2-miR-125a inhibitory complex on PSD-95 mRNA, whereas mGluR signaling of translation requires FMRP dephosphorylation and release of AGO2 from the mRNA. These findings reveal a novel mechanism whereby FMRP phosphorylation provides a reversible switch for AGO2 and microRNA to selectively regulate mRNA translation at synapses in response to receptor activation. PMID:21658607

  7. Expression of very low density lipoprotein receptor mRNA in circulating human monocytes: its up-regulation by hypoxia.

    PubMed

    Nakazato, K; Ishibashi, T; Nagata, K; Seino, Y; Wada, Y; Sakamoto, T; Matsuoka, R; Teramoto, T; Sekimata, M; Homma, Y; Maruyama, Y

    2001-04-01

    Although very low density lipoprotein (VLDL) receptor expression by macrophages has been shown in the vascular wall, it is not clear whether or not circulating monocytes express the VLDL receptor. We investigated the expression of VLDL receptor mRNA in human peripheral blood monocytes and monocyte-derived macrophages by reverse transcriptase polymerase chain reaction (RT-PCR) and nucleotide sequencing after subcloning of PCR product. VLDL receptor mRNA was detected both in peripheral blood monocytes and monocyte-derived macrophages. Expression of VLDL receptor mRNA was upregulated by hypoxia in monocytes, whereas treatment with oxidized LDL, interleukin-1beta or monocyte chemoattractant protein-1 did not affect the levels of VLDL receptor mRNA in monocytes and macrophages. The present study shows a novel response of VLDL receptor mRNA to hypoxia, suggesting a role for VLDL receptor in the metabolism of lipoproteins in the vascular wall and the development of atherosclerosis.

  8. Serum amyloid A secretion from monocytic leukaemia cell line THP-1 and cultured human peripheral monocytes.

    PubMed

    Yamada, T; Wada, A; Itoh, K; Igari, J

    2000-07-01

    Serum amyloid A (SAA), an acute-phase protein and a precursor of fibrous components in reactive amyloid deposits, is synthesized mainly in the liver under the stimulation of inflammation-related cytokines. In addition, the SAA gene is expressed in monocytes/macrophages, which are believed to play a central role in amyloid fibrillogenesis. Consequently, the pathogenic implication of SAA produced from these cells has been of major concern. Because SAA synthesis at the protein level in such cells has never been analyzed quantitatively, in this study an enzyme-linked immunosorbent assay was generated with a detection level sufficiently high to measure SAA concentrations in the culture supernatants of the human monocytic leukaemia cell line THP-1. SAA secretion by THP-1 with interleukin (IL)-1beta required the presence of dexamethasone as proposed previously. We also found that unidentified components in fetal calf serum (FCS) could induce SAA production by THP-1 in the presence of dexamethasone. These findings are in contrast to the results obtained from hepatoma cell line HepG2, in which IL-1beta alone could induce SAA secretion, while dexamethasone-supplemented FCS could not. The method was able to quantify SAA secreted from cultured human peripheral monocytes. The findings suggest that monocytes produce SAA in almost the same manner as THP-1. Thus, THP-1 cells can be utilized to investigate a distinctive manner of SAA production from monocytes.

  9. Chaperone Hsp27 Modulates AUF1 Proteolysis and AU-Rich Element-Mediated mRNA Degradation▿

    PubMed Central

    Knapinska, Anna M.; Gratacós, Frances M.; Krause, Christopher D.; Hernandez, Kristina; Jensen, Amber G.; Bradley, Jacquelyn J.; Wu, Xiangyue; Pestka, Sidney; Brewer, Gary

    2011-01-01

    AUF1 is an AU-rich element (ARE)-binding protein that recruits translation initiation factors, molecular chaperones, and mRNA degradation enzymes to the ARE for mRNA destruction. We recently found chaperone Hsp27 to be an AUF1-associated ARE-binding protein required for tumor necrosis factor alpha (TNF-α) mRNA degradation in monocytes. Hsp27 is a multifunctional protein that participates in ubiquitination of proteins for their degradation by proteasomes. A variety of extracellular stimuli promote Hsp27 phosphorylation on three serine residues—Ser15, Ser78, and Ser82—by a number of kinases, including the mitogen-activated protein (MAP) pathway kinases p38 and MK2. Activating either kinase stabilizes ARE mRNAs. Likewise, ectopic expression of phosphomimetic mutant forms of Hsp27 stabilizes reporter ARE mRNAs. Here, we continued to examine the contributions of Hsp27 to mRNA degradation. As AUF1 is ubiquitinated and degraded by proteasomes, we addressed the hypothesis that Hsp27 phosphorylation controls AUF1 levels to modulate ARE mRNA degradation. Indeed, selected phosphomimetic mutants of Hsp27 promote proteolysis of AUF1 in a proteasome-dependent fashion and render ARE mRNAs more stable. Our results suggest that the p38 MAP kinase (MAPK)-MK2–Hsp27 signaling axis may target AUF1 destruction by proteasomes, thereby promoting ARE mRNA stabilization. PMID:21245386

  10. [Impacts of the formula of Suoquanwan(SQW) on expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency].

    PubMed

    Cao, Hong-Ying; Wu, Qing-He; Huang, Ping; He, Jin-Yang

    2009-06-01

    To observe the impacts of the formula of Suoquanwan (SQW) on the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency. The model rats were induced by adenine (250 mg/kg) for 4 weeks, then treated respectively with SQW or dDAVP. The expression of AQP-2 mRNA and AVPR-V2 mRNA in kidney of Yang-deficiency model by realtime fluorescence quantitative PCR method were investigated. In model rats, the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney decreased, dDAVP and SQW high dose could increased the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney. The others had no influence on the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney. SQW can increase the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency.

  11. Effect of atrophy and contractions on myogenin mRNA concentration in chick and rat myoblast omega muscle cells

    NASA Technical Reports Server (NTRS)

    Krebs, J. M.; Denney, R. M.

    1997-01-01

    The skeletal rat myoblast omega (RMo) cell line forms myotubes that exhibit spontaneous contractions under appropriate conditions in culture. We examined if the RMo cells would provide a model for studying atrophy and muscle contraction. To better understand how to obtain contractile cultures, we examined levels of contraction under different growing conditions. The proliferation medium and density of plating affected the subsequent proportion of spontaneously contracting myotubes. Using a ribonuclease protection assay, we found that exponentially growing RMo myoblasts contained no detectable myogenin or herculin mRNA, while differentiating myoblasts contained high levels of myogenin mRNA but no herculin mRNA. There was no increase in myogenin mRNA concentration in either primary chick or RMo myotubes whose contractions were inhibited by depolarizing concentrations of potassium (K+). Thus, altered myogenin mRNA concentrations are not involved in atrophy of chick myotubes. Depolarizing concentrations of potassium inhibited spontaneous contractions in both RMo cultures and primary chick myotube cultures. However, we found that the myosin concentration of 6-d-old contracting RMo cells fed medium plus AraC was 11 +/- 3 micrograms myosin/microgram DNA, not significantly different from 12 +/- 4 micrograms myosin/microgram DNA (n = 3), the myosin concentration of noncontracting RMo cells (treated with 12 mM K+ for 6 d). Resolving how RMo cells maintained their myosin content when contraction is inhibited may be important for understanding atrophy.

  12. Light differentially regulates cell division and the mRNA abundance of pea nucleolin during de-etiolation

    NASA Technical Reports Server (NTRS)

    Reichler, S. A.; Balk, J.; Brown, M. E.; Woodruff, K.; Clark, G. B.; Roux, S. J.

    2001-01-01

    The abundance of plant nucleolin mRNA is regulated during de-etiolation by phytochrome. A close correlation between the mRNA abundance of nucleolin and mitosis has also been previously reported. These results raised the question of whether the effects of light on nucleolin mRNA expression were a consequence of light effects on mitosis. To test this we compared the kinetics of light-mediated increases in cell proliferation with that of light-mediated changes in the abundance of nucleolin mRNA using plumules of dark-grown pea (Pisum sativum) seedlings. These experiments show that S-phase increases 9 h after a red light pulse, followed by M-phase increases in the plumule leaves at 12 h post-irradiation, a time course consistent with separately measured kinetics of red light-induced increases in the expression of cell cycle-regulated genes. These increases in cell cycle-regulated genes are photoreversible, implying that the light-induced increases in cell proliferation are, like nucleolin mRNA expression, regulated via phytochrome. Red light stimulates increases in the mRNA for nucleolin at 6 h post-irradiation, prior to any cell proliferation changes and concurrent with the reported timing of phytochrome-mediated increases of rRNA abundance. After a green light pulse, nucleolin mRNA levels increase without increasing S-phase or M-phase. Studies in animals and yeast indicate that nucleolin plays a significant role in ribosome biosynthesis. Consistent with this function, pea nucleolin can rescue nucleolin deletion mutants of yeast that are defective in rRNA synthesis. Our data show that during de-etiolation, the increased expression of nucleolin mRNA is more directly regulated by light than by mitosis.

  13. The Anti-Apoptotic and Cardioprotective Effects of Salvianolic Acid A on Rat Cardiomyocytes following Ischemia/Reperfusion by DUSP-Mediated Regulation of the ERK1/2/JNK Pathway

    PubMed Central

    Chen, Qiuping; Zhu, Shasha; Liu, Yang; Pan, Defeng; Chen, Xiaohu; Li, Dongye

    2014-01-01

    The purpose of this study was to observe the effects of salvianolic acid A (SAA) pretreatment on the myocardium during ischemia/reperfusion (I/R) and to illuminate the interrelationships among dual specificity protein phosphatase (DUSP) 2/4/16, ERK1/2 and JNK pathways during myocardial I/R, with the ultimate goal of elucidating how SAA exerts cardioprotection against I/R injury (IRI). Wistar rats were divided into the following six groups: control group (CON), I/R group, SAA+I/R group, ERK1/2 inhibitor PD098059+I/R group (PD+I/R), PD+SAA+I/R group, and JNK inhibitor SP600125+I/R group (SP+I/R). The cardioprotective effects of SAA on the myocardium during I/R were investigated with a Langendorff device. Heart rate (HR), left ventricular systolic pressure (LVSP), left ventricular end-diastolic pressure (LVEDP), maximum rate of ventricular pressure rise and fall (±dp/dtmax), myocardial infarction areas (MIA), lactate dehydrogenase (LDH), and cardiomyocytes apoptosis were monitored. To determine the crosstalk betwee JNK and ERK1/2 via DUSP2/4/16 with SAA pretreatment, siRNA-DUSP2/4/16 were performed. The expression levels of Bcl-2, Bax, caspase 3, p-JNK, p-ERK1/2 and DUSP2/4/16 in cardiomyocytes were assayed by Western blot. Our results showed that LDH, MIA and cell apoptosis were decreased, and various parameters of heart function were improved by SAA pretreatment and SP application. In the I/R group, the expression levels of p-ERK1/2 and DUSP4/16 were not significantly different compared with the CON group, however, the protein expression levels of p-ERK1/2, Bcl-2 and DUSP4/16 were higher, while p-JNK, Bax, caspase 3 and DUSP2 levels were reduced among the SAA+I/R, PD+SAA+I/R and SP+I/R groups. The above indices were not significantly different between the SAA+I/R and SP+I/R groups. Compared with the SAA+I/R group, p-ERK1/2 was increased and p-JNK was decreased in the SAA+si-DUSP2+I/R, however, p-ERK was downregulated and p-JNK was upregulated in SAA+si-DUSP4+I

  14. BAG3 directly stabilizes Hexokinase 2 mRNA and promotes aerobic glycolysis in pancreatic cancer cells.

    PubMed

    An, Ming-Xin; Li, Si; Yao, Han-Bing; Li, Chao; Wang, Jia-Mei; Sun, Jia; Li, Xin-Yu; Meng, Xiao-Na; Wang, Hua-Qin

    2017-12-04

    Aerobic glycolysis, a phenomenon known historically as the Warburg effect, is one of the hallmarks of cancer cells. In this study, we characterized the role of BAG3 in aerobic glycolysis of pancreatic ductal adenocarcinoma (PDAC) and its molecular mechanisms. Our data show that aberrant expression of BAG3 significantly contributes to the reprogramming of glucose metabolism in PDAC cells. Mechanistically, BAG3 increased Hexokinase 2 (HK2) expression, the first key enzyme involved in glycolysis, at the posttranscriptional level. BAG3 interacted with HK2 mRNA, and the degree of BAG3 expression altered recruitment of the RNA-binding proteins Roquin and IMP3 to the HK2 mRNA. BAG3 knockdown destabilized HK2 mRNA via promotion of Roquin recruitment, whereas BAG3 overexpression stabilized HK2 mRNA via promotion of IMP3 recruitment. Collectively, our results show that BAG3 promotes reprogramming of glucose metabolism via interaction with HK2 mRNA in PDAC cells, suggesting that BAG3 may be a potential target in the aerobic glycolysis pathway for developing novel anticancer agents. © 2017 An et al.

  15. Antihyperalgesic activity of a novel nonpeptide bradykinin B1 receptor antagonist in transgenic mice expressing the human B1 receptor

    PubMed Central

    Fox, Alyson; Kaur, Satbir; Li, Bifang; Panesar, Moh; Saha, Uma; Davis, Clare; Dragoni, Ilaria; Colley, Sian; Ritchie, Tim; Bevan, Stuart; Burgess, Gillian; McIntyre, Peter

    2005-01-01

    We describe the properties of a novel nonpeptide kinin B1 receptor antagonist, NVP-SAA164, and demonstrate its in vivo activity in models of inflammatory pain in transgenic mice expressing the human B1 receptor. NVP-SAA164 showed high affinity for the human B1 receptor expressed in HEK293 cells (Ki 8 nM), and inhibited increases in intracellular calcium induced by desArg10kallidin (desArg10KD) (IC50 33 nM). While a similar high affinity was observed in monkey fibroblasts (Ki 7.7 nM), NVP-SAA164 showed no affinity for the rat B1 receptor expressed in Cos-7 cells. In transgenic mice in which the native B1 receptor was deleted and the gene encoding the human B1 receptor was inserted (hB1 knockin, hB1-KI), hB1 receptor mRNA was induced in tissues following LPS treatment. No mRNA encoding the mouse or human B1 receptor was detected in mouse B1 receptor knockout (mB1-KO) mice following LPS treatment. Freund's complete adjuvant-induced mechanical hyperalgesia was similar in wild-type and hB1-KI mice, but was significantly reduced in mB1-KO animals. Mechanical hyperalgesia induced by injection of the B1 agonist desArg10KD into the contralateral paw 24 h following FCA injection was similar in wild-type and hB1-KI mice, but was absent in mB1-KO animals. Oral administration of NVP-SAA164 produced a dose-related reversal of FCA-induced mechanical hyperalgesia and desArg10KD-induced hyperalgesia in hB1-KI mice, but was inactive against inflammatory pain in wild-type mice. These data demonstrate the use of transgenic technology to investigate the in vivo efficacy of species selective agents and show that NVP-SAA164 is a novel orally active B1 receptor antagonist, providing further support for the utility of B1 receptor antagonists in inflammatory pain conditions in man. PMID:15685199

  16. Conservation of mRNA secondary structures may filter out mutations in Escherichia coli evolution

    PubMed Central

    Chursov, Andrey; Frishman, Dmitrij; Shneider, Alexander

    2013-01-01

    Recent reports indicate that mutations in viral genomes tend to preserve RNA secondary structure, and those mutations that disrupt secondary structural elements may reduce gene expression levels, thereby serving as a functional knockout. In this article, we explore the conservation of secondary structures of mRNA coding regions, a previously unknown factor in bacterial evolution, by comparing the structural consequences of mutations in essential and nonessential Escherichia coli genes accumulated over 40 000 generations in the course of the ‘long-term evolution experiment’. We monitored the extent to which mutations influence minimum free energy (MFE) values, assuming that a substantial change in MFE is indicative of structural perturbation. Our principal finding is that purifying selection tends to eliminate those mutations in essential genes that lead to greater changes of MFE values and, therefore, may be more disruptive for the corresponding mRNA secondary structures. This effect implies that synonymous mutations disrupting mRNA secondary structures may directly affect the fitness of the organism. These results demonstrate that the need to maintain intact mRNA structures imposes additional evolutionary constraints on bacterial genomes, which go beyond preservation of structure and function of the encoded proteins. PMID:23783573

  17. Protamine mRNA ratio in stallion spermatozoa correlates with mare fecundity.

    PubMed

    Paradowska-Dogan, A; Fernandez, A; Bergmann, M; Kretzer, K; Mallidis, C; Vieweg, M; Waliszewski, P; Zitzmann, M; Weidner, W; Steger, K; Kliesch, S

    2014-07-01

    of spermatogenesis and as a corollary the level of protamine mRNA may reflect the quality of spermatogenesis and spermatozoa's fertilizing capacity. © 2014 American Society of Andrology and European Academy of Andrology.

  18. Glutathione S-transferase PI (GST-PI) mRNA expression and DNA methylation is involved in the pathogenesis and prognosis of NSCLC.

    PubMed

    Grimminger, Peter P; Maus, Martin K H; Schneider, Paul M; Metzger, Ralf; Hölscher, Arnulf H; Sugita, Hirofumi; Danenberg, Peter V; Alakus, Hakan; Brabender, Jan

    2012-10-01

    The aim of this study was to investigate the relevance of mRNA expression and DNA methylation of GST-PI in tumor and non-tumor lung tissue from NSCLC patients in terms of prognostic and pathogenetic value of this biomarker. Quantitative real-time PCR was used to measure mRNA expression and DNA methylation of GST-PI in paired tumor (T) and non-tumor (N) lung tissue of 91 NSCLC patients. Of all 91 patients 49% were stage I, 21% stage II and 30% stage IIIA. Forty-seven percent of the patients had squamous cell carcinoma, 36% adenocarcinoma and 17% large cell carcinoma. All patients were R0 resected. GST-PI mRNA expression could be measured in 100% in both (T and N) tissues; GST-PI DNA methylation was detected in 14% (N) and 14% (T). The median GST-PI mRNA expression in N was 7.83 (range: 0.01-19.43) and in T 13.15 (range: 0.01-116.8; p≤0.001). The median GST-PI methylation was not significantly different between T and N. No associations were seen between the mRNA expression or DNA methylation levels and clinical or histopathologic parameters such as gender, age, TNM stage, tumor histology and grading. The median survival of the investigated patients was 59.7 years (the median follow-up was 85.9 months). High GST-PI DNA methylation was significantly associated with a worse prognosis (p=0.041, log rank test). No correlation was found between the GST-PI DNA methylation levels and the correlating mRNA expression levels. GST-PI mRNA expression seems to be involved in the pathogenesis of NSCLC. High levels of GST-PI DNA methylation in tumor tissue of NSCLC patients have a potential as a biomarker identifying subpopulations with a more aggressive tumor biology. Quantitation of GST-PI DNA methylation may be a useful method to identify patients with a poor prognosis after curative resection and who will benefit from intensive adjuvant therapy. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  19. Effects of varying dietary iodine supplementation levels as iodide or iodate on thyroid status as well as mRNA expression and enzyme activity of antioxidative enzymes in tissues of grower/finisher pigs.

    PubMed

    Li, Qimeng; Mair, Christiane; Schedle, Karl; Hellmayr, Isabella; Windisch, Wilhelm

    2013-02-01

    The objective of this study was to investigate the influence of high dietary iodine supply and different iodine sources on thyroid status and oxidative stress in target tissues of the thyroid hormones in fattening pigs. Eighty castrates (body weight: 33.3 ± 0.4 kg) were randomly allotted into five different treatments: The control diet contained 150 μg I/kg as KI, the other feeding groups were supplemented with 4,000 μg I/kg (as KI and KIO(3)) and 10,000 μg I/kg (as KI and KIO(3)), respectively. The mRNA expression levels of sodium/iodide symporter (NIS) and key antioxidant enzymes (Cu/Zn SOD, CAT, GPx) were analyzed in thyroid gland, liver, kidney, muscle, and adipose tissue sampled during slaughter. Furthermore, antioxidant enzyme activities and the effect on lipid peroxidation (MDA) were determined in liver and muscle. In thyroid gland, a significant downregulation of NIS and Cu/Zn SOD mRNA expression was observed in high-iodine groups. In liver, a source effect on the mRNA expression of Cu/Zn SOD between KI and KIO(3) at 4,000 μg I/kg was shown. In contrast, not SOD but GPx activity was affected by iodine source with strongest downregulation in high KIO(3) group. In muscle, GPx activity was affected by both iodine source and dose, showing stronger downregulation in KI groups. In kidney and adipose tissue, oxidative stress parameters showed no or only unsystematic changes. However, variation in iodine supply had no effect on MDA concentrations. NIS expression was significantly decreased with increased iodine supplementation, which is to ensure the thyroid gland function. However, the alleviating effect of iodine supplementation observed in antioxidant enzyme mRNA expression and activity did not reflect on the lipid peroxide level.

  20. Molecular determinants of Cytochrome C oxidase IV mRNA axonal trafficking

    PubMed Central

    Kar, Amar N.; Vargas, Jose Norberto S.; Chen, Cai-Yun; Kowalak, Jeffrey A; Gioio, Anthony E.; Kaplan, Barry B.

    2017-01-01

    In previous studies, we identified a putative 38-nucleotide stem-loop structure (zipcode) in the 3′ untranslated region of the cytochrome c oxidase subunit IV (COXIV) mRNA that was necessary and sufficient for the axonal localization of the message in primary superior cervical ganglion (SCG) neurons. However, little is known about the proteins that interact with the COXIV-zipcode and regulate the axonal trafficking and local translation of the COXIV message. To identify proteins involved in the axonal transport of the COXIV mRNA, we used the biotinylated 38-nucleotide COXIV RNA zipcode as bait in the affinity purification of COXIV zipcode binding proteins. Gel-shift assays of the biotinylated COXIV zipcode indicated that the putative stem-loop structure functions as a nucleation site for the formation of ribonucleoprotein complexes. Mass spectrometric analysis of the COXIV zipcode ribonucleoprotein complex led to the identification of a large number RNA binding proteins, including fused in sarcoma/translated in liposarcoma (FUS/TLS), and Y-box protein 1 (YB-1). Validation experiments, using western analyses, confirmed the presence of the candidate proteins in the COXIV zipcode affinity purified complexes obtained from SCG axons. Immunohistochemical studies show that FUS, and YB-1 are present in SCG axons. Importantly, RNA immunoprecipitation studies show that FUS, and YB-1 interact with endogenous axonal COXIV transcripts. siRNA-mediated downregulation of the candidate proteins FUS and YB-1 expression in the cell-bodies diminishes the levels of COXIV mRNA in the axon, suggesting functional roles for these proteins in the axonal trafficking of COXIV mRNA. PMID:28161363