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Sample records for human blood transcriptional

  1. The transcriptional landscape of age in human peripheral blood

    NARCIS (Netherlands)

    M.J. Peters (Marjolein); R. Joehanes (Roby); L.C. Pilling (Luke); C. Schurmann (Claudia); K.N. Conneely (Karen N.); J.E. Powell (Joseph); E. Reinmaa (Eva); G.L. Sutphin (George L.); A. Zhernakova (Alexandra); K. Schramm (Katharina); Y.A. Wilson (Yana A.); S. Kobes (Sayuko); T. Tukiainen (Taru); Y.F.M. Ramos (Yolande); H.H.H. Göring (Harald H.); M. Fornage (Myriam); Y. Liu (YongMei); S.A. Gharib (Sina); B.E. Stranger (Barbara); P.L. de Jager (Philip); A. Aviv (Abraham); D. Levy (Daniel); J. Murabito (Joanne); P.J. Munson (Peter J.); T. Huan (Tianxiao); A. Hofman (Albert); A.G. Uitterlinden (André); F. Rivadeneira Ramirez (Fernando); J. van Rooij (Jeroen); L. Stolk (Lisette); L. Broer (Linda); M.M.P.J. Verbiest (Michael); M. Jhamai (Mila); P.P. Arp (Pascal); A. Metspalu (Andres); L. Tserel (Liina); L. Milani (Lili); N.J. Samani (Nilesh); P. Peterson (Pärt); S. Kasela (Silva); V. Codd (Veryan); A. Peters (Annette); C.K. Ward-Caviness (Cavin K.); C. Herder (Christian); M. Waldenberger (Melanie); M. Roden (Michael); P. Singmann (Paula); S. Zeilinger (Sonja); T. Illig (Thomas); G. Homuth (Georg); H.J. Grabe (Hans Jörgen); H. Völzke (Henry); L. Steil (Leif); T. Kocher (Thomas); A. Murray (Anna); D. Melzer (David); H. Yaghootkar (Hanieh); S. Bandinelli; E.K. Moses (Eric); J.W. Kent (Jack); J.E. Curran (Joanne); M.P. Johnson (Matthew); S. Williams-Blangero (Sarah); H.J. Westra (Harm-Jan); A.F. McRae (Allan F.); J.A. Smith (Jennifer A); S.L.R. Kardia (Sharon); I. Hovatta (Iiris); M. Perola (Markus); S. Ripatti (Samuli); V. Salomaa (Veikko); A.K. Henders (Anjali); N.G. Martin (Nicholas); A.K. Smith (Alicia K.); D. Mehta (Divya); E.B. Binder (Elisabeth B.); K.M. Nylocks (K. Maria); E.M. Kennedy (Elizabeth M.); T. Klengel (Torsten); J. Ding (Jingzhong); A. Suchy-Dicey (Astrid); D. Enquobahrie; J. Brody (Jennifer); J.I. Rotter (Jerome I.); Y.-D.I. Chen (Yii-Der I.); J.J. Houwing-Duistermaat (Jeanine); M. Kloppenburg (Margreet); P.E. Slagboom (Eline); Q. Helmer (Quinta); W. den Hollander (Wouter); S. Bean (Shannon); T. Raj (Towfique); N. Bakhshi (Noman); Q.P. Wang (Qiao Ping); L.J. Oyston (Lisa J.); B.M. Psaty (Bruce); R.P. Tracy (Russell); G.W. Montgomery (Grant); S.T. Turner (Stephen); J. Blangero (John); I. Meulenbelt (Ingrid); K.J. Ressler (Kerry); J. Yang (Jian); L. Franke (Lude); J. Kettunen (Johannes); P.M. Visscher (Peter); G.G. Neely (G. Gregory); R. Korstanje (Ron); R.L. Hanson (Robert L.); H. Prokisch (Holger); L. Ferrucci (Luigi); T. Esko (Tõnu); A. Teumer (Alexander); J.B.J. van Meurs (Joyce); A.D. Johnson (Andrew D.); M.A. Nalls (Michael); D.G. Hernandez (Dena); M.R. Cookson (Mark); R.J. Gibbs (Raphael J.); J. Hardy (John); A. Ramasamy (Adaikalavan); A.B. Zonderman (Alan B.); A. Dillman (Allissa); B. Traynor (Bryan); C. Smith (Colin); D.L. Longo (Dan L.); D. Trabzuni (Danyah); J.C. Troncoso (Juan); M.P. van der Brug (Marcel); M.E. Weale (Michael); R. O'Brien (Richard); R. Johnson (Robert); R. Walker (Robert); R.H. Zielke (Ronald H.); S. Arepalli (Sampath); M. Ryten (Mina); A. Singleton

    2015-01-01

    textabstractDisease incidences increase with age, but the molecular characteristics of ageing that lead to increased disease susceptibility remain inadequately understood. Here we perform a whole-blood gene expression meta-analysis in 14,983 individuals of European ancestry (including replication)

  2. The transcriptional landscape of age in human peripheral blood

    NARCIS (Netherlands)

    Peters, Marjolein J.; Joehanes, Roby; Pilling, Luke C.; Schurmann, Claudia; Conneely, Karen N.; Powell, Joseph; Reinmaa, Eva; Sutphin, George L.; Zhernakova, Alexandra; Schramm, Katharina; Wilson, Yana A.; Kobes, Sayuko; Tukiainen, Taru; Ramos, Yolande F.; Goering, Harald H. H.; Fornage, Myriam; Liu, Yongmei; Gharib, Sina A.; Stranger, Barbara E.; De Jager, Philip L.; Aviv, Abraham; Levy, Daniel; Murabito, Joanne M.; Munson, Peter J.; Huan, Tianxiao; Hofman, Albert; Uitterlinden, Andre G.; Rivadeneira, Fernando; van Rooij, Jeroen; Stolk, Lisette; Broer, Linda; Verbiest, Michael M. P. J.; Jhamai, Mila; Arp, Pascal; Metspalu, Andres; Tserel, Liina; Milani, Lili; Samani, Nilesh J.; Peterson, Paert; Kasela, Silva; Codd, Veryan; Peters, Annette; Ward-Caviness, Cavin K.; Herder, Christian; Waldenberger, Melanie; Roden, Michael; Singmann, Paula; Zeilinger, Sonja; Westra, Harm-Jan; Franke, Lude

    2015-01-01

    Disease incidences increase with age, but the molecular characteristics of ageing that lead to increased disease susceptibility remain inadequately understood. Here we perform a whole-blood gene expression meta-analysis in 14,983 individuals of European ancestry (including replication) and identify

  3. Is human blood a good surrogate for brain tissue in transcriptional studies?

    Directory of Open Access Journals (Sweden)

    van den Berg Leonard H

    2010-10-01

    Full Text Available Abstract Background Since human brain tissue is often unavailable for transcriptional profiling studies, blood expression data is frequently used as a substitute. The underlying hypothesis in such studies is that genes expressed in brain tissue leave a transcriptional footprint in blood. We tested this hypothesis by relating three human brain expression data sets (from cortex, cerebellum and caudate nucleus to two large human blood expression data sets (comprised of 1463 individuals. Results We found mean expression levels were weakly correlated between the brain and blood data (r range: [0.24,0.32]. Further, we tested whether co-expression relationships were preserved between the three brain regions and blood. Only a handful of brain co-expression modules showed strong evidence of preservation and these modules could be combined into a single large blood module. We also identified highly connected intramodular "hub" genes inside preserved modules. These preserved intramodular hub genes had the following properties: first, their expression levels tended to be significantly more heritable than those from non-preserved intramodular hub genes (p -90; second, they had highly significant positive correlations with the following cluster of differentiation genes: CD58, CD47, CD48, CD53 and CD164; third, a significant number of them were known to be involved in infection mechanisms, post-transcriptional and post-translational modification and other basic processes. Conclusions Overall, we find transcriptome organization is poorly preserved between brain and blood. However, the subset of preserved co-expression relationships characterized here may aid future efforts to identify blood biomarkers for neurological and neuropsychiatric diseases when brain tissue samples are unavailable.

  4. Dissecting interferon-induced transcriptional programs in human peripheral blood cells.

    Directory of Open Access Journals (Sweden)

    Simon J Waddell

    2010-03-01

    Full Text Available Interferons are key modulators of the immune system, and are central to the control of many diseases. The response of immune cells to stimuli in complex populations is the product of direct and indirect effects, and of homotypic and heterotypic cell interactions. Dissecting the global transcriptional profiles of immune cell populations may provide insights into this regulatory interplay. The host transcriptional response may also be useful in discriminating between disease states, and in understanding pathophysiology. The transcriptional programs of cell populations in health therefore provide a paradigm for deconvoluting disease-associated gene expression profiles.We used human cDNA microarrays to (1 compare the gene expression programs in human peripheral blood mononuclear cells (PBMCs elicited by 6 major mediators of the immune response: interferons alpha, beta, omega and gamma, IL12 and TNFalpha; and (2 characterize the transcriptional responses of purified immune cell populations (CD4+ and CD8+ T cells, B cells, NK cells and monocytes to IFNgamma stimulation. We defined a highly stereotyped response to type I interferons, while responses to IFNgamma and IL12 were largely restricted to a subset of type I interferon-inducible genes. TNFalpha stimulation resulted in a distinct pattern of gene expression. Cell type-specific transcriptional programs were identified, highlighting the pronounced response of monocytes to IFNgamma, and emergent properties associated with IFN-mediated activation of mixed cell populations. This information provides a detailed view of cellular activation by immune mediators, and contributes an interpretive framework for the definition of host immune responses in a variety of disease settings.

  5. Transcriptional profiling of human brain endothelial cells reveals key properties crucial for predictive in vitro blood-brain barrier models.

    Directory of Open Access Journals (Sweden)

    Eduard Urich

    Full Text Available Brain microvascular endothelial cells (BEC constitute the blood-brain barrier (BBB which forms a dynamic interface between the blood and the central nervous system (CNS. This highly specialized interface restricts paracellular diffusion of fluids and solutes including chemicals, toxins and drugs from entering the brain. In this study we compared the transcriptome profiles of the human immortalized brain endothelial cell line hCMEC/D3 and human primary BEC. We identified transcriptional differences in immune response genes which are directly related to the immortalization procedure of the hCMEC/D3 cells. Interestingly, astrocytic co-culturing reduced cell adhesion and migration molecules in both BECs, which possibly could be related to regulation of immune surveillance of the CNS controlled by astrocytic cells within the neurovascular unit. By matching the transcriptome data from these two cell lines with published transcriptional data from freshly isolated mouse BECs, we discovered striking differences that could explain some of the limitations of using cultured BECs to study BBB properties. Key protein classes such as tight junction proteins, transporters and cell surface receptors show differing expression profiles. For example, the claudin-5, occludin and JAM2 expression is dramatically reduced in the two human BEC lines, which likely explains their low transcellular electric resistance and paracellular leakiness. In addition, the human BEC lines express low levels of unique brain endothelial transporters such as Glut1 and Pgp. Cell surface receptors such as LRP1, RAGE and the insulin receptor that are involved in receptor-mediated transport are also expressed at very low levels. Taken together, these data illustrate that BECs lose their unique protein expression pattern outside of their native environment and display a more generic endothelial cell phenotype. A collection of key genes that seems to be highly regulated by the local

  6. Identification of the key differential transcriptional responses of human whole blood following TLR2 or TLR4 ligation in-vitro.

    Directory of Open Access Journals (Sweden)

    Simon Blankley

    Full Text Available The use of human whole blood for transcriptomic analysis has potential advantages over the use of isolated immune cells for studying the transcriptional response to pathogens and their products. Whole blood stimulation can be carried out in a laboratory without the expertise or equipment to isolate immune cells from blood, with the added advantage of being able to undertake experiments using very small volumes of blood. Toll like receptors (TLRs are a family of pattern recognition receptors which recognise highly conserved microbial products. Using the TLR2 ligand (Pam3CSK4 and the TLR4 ligand (LPS, human whole blood was stimulated for 0, 1, 3, 6, 12 or 24 hours at which times mRNA was isolated and a comparative microarray was undertaken. A common NFκB transcriptional programme was identified following both TLR2 and TLR4 ligation which peaked at between 3 to 6 hours including upregulation of many of the NFκB family members. In contrast an interferon transcriptional response was observed following TLR4 but not TLR2 ligation as early as 1 hour post stimulation and peaking at 6 hours. These results recapitulate the findings observed in previously published studies using isolated murine and human myeloid cells indicating that in vitro stimulated human whole blood can be used to interrogate the early transcriptional kinetic response of innate cells to TLR ligands. Our study demonstrates that a transcriptomic analysis of mRNA isolated from human whole blood can delineate both the temporal response and the key transcriptional differences following TLR2 and TLR4 ligation.

  7. The effects of high dose ionizing radiation on transcriptional regulation and paracrine signaling in human peripheral blood mononuclear cells

    International Nuclear Information System (INIS)

    Beer, L.

    2015-01-01

    While it has long been accepted that direct cell-cell interactions and the replacement of injured tissues with injected cells exerts therapeutic effects, it is currently believed that, in addition, paracrine factors released from different cell types activate cytoprotective and regenerative processes. Cells are now seen as bioreactors that produce and release soluble factors which might be used as therapeutics. We have previously shown that peripheral blood mononuclear cells (PBMCs) release a plethora of paracrine factors that enhance wound healing, attenuate myocardial damage following acute myocardial infarction, abolish microvascular obstruction, improve neurological outcome after acute ischemic stroke and spinal cord injury and protect mice from experimental autoimmune myocarditis. These PBMC derived paracrine factors may exert their effects via the induction of cytoprotective pathways, augmentation of angiogenesis, induction of NO-depended vasodilation and inhibition of VASP dependent platelet aggregation, as well as driving auto-reactive CD4+ cells into apoptosis. To enhance the cellular secretory capacity, treatments which induce stress responses, such as hypoxic preconditioning or ionizing irradiation (IR), have been developed. Although these effects have been evaluated in several disease states there is little data available on the cellular effects of ionizing irradiation on human PBMCs and their secretome. In this study, we have thus undertaken to investigate the effects of IR on human PBMCs in terms of the induction of transcriptional changes and release of pleiotropic paracrine factors. There are three primary aims of this doctoral thesis: 1. To investigate cellular processes activated or repressed in human PBMCs following high dose ionizing radiation (60Gy) and high density cell cultivation (25*10"6 cells/ml) for up to 24 hours. 2. To identify paracrine factors released from these cells using a multi-methodical biochemical/bioinformatics approach. 3

  8. 1,25-Dihydroxyvitamin D3 inhibits cytokine production by human blood monocytes at the post-transcriptional level

    DEFF Research Database (Denmark)

    Müller, K; Haahr, P M; Diamant, M

    1992-01-01

    was not caused by impaired production of mRNA. Taken together, the study demonstrates a vitamin D-induced inhibitory effect of LPS-driven monokine production, which is most likely a vitamin D-receptor mediated phenomenon exerted at a post-transcriptional, presecretory level. Impaired monokine production may...... be of importance in 1,25-(OH)2D3-mediated inhibition of lymphocyte functions in vitro.......1,25-Dihydroxyvitamin D3 [1,25-(OH)2D3] inhibits lymphocyte proliferation and production of antibodies and lymphokines such as interleukin (IL)-2 and interferon gamma. These lymphocyte functions are dependent upon cytokines, including IL-1 alpha, IL-1 beta, IL-6 and tumour necrosis factor alpha...

  9. A module of human peripheral blood mononuclear cell transcriptional network containing primitive and differentiation markers is related to specific cardiovascular health variables.

    Directory of Open Access Journals (Sweden)

    Leni Moldovan

    Full Text Available Peripheral blood mononuclear cells (PBMCs, including rare circulating stem and progenitor cells (CSPCs, have important yet poorly understood roles in the maintenance and repair of blood vessels and perfused organs. Our hypothesis was that the identities and functions of CSPCs in cardiovascular health could be ascertained by analyzing the patterns of their co-expressed markers in unselected PBMC samples. Because gene microarrays had failed to detect many stem cell-associated genes, we performed quantitative real-time PCR to measure the expression of 45 primitive and tissue differentiation markers in PBMCs from healthy and hypertensive human subjects. We compared these expression levels to the subjects' demographic and cardiovascular risk factors, including vascular stiffness. The tested marker genes were expressed in all of samples and organized in hierarchical transcriptional network modules, constructed by a bottom-up approach. An index of gene expression in one of these modules (metagene, defined as the average standardized relative copy numbers of 15 pluripotency and cardiovascular differentiation markers, was negatively correlated (all p<0.03 with age (R2 = -0.23, vascular stiffness (R2 = -0.24, and central aortic pressure (R2 = -0.19 and positively correlated with body mass index (R2 = 0.72, in women. The co-expression of three neovascular markers was validated at the single-cell level using mRNA in situ hybridization and immunocytochemistry. The overall gene expression in this cardiovascular module was reduced by 72±22% in the patients compared with controls. However, the compactness of both modules was increased in the patients' samples, which was reflected in reduced dispersion of their nodes' degrees of connectivity, suggesting a more primitive character of the patients' CSPCs. In conclusion, our results show that the relationship between CSPCs and vascular function is encoded in modules of the PBMCs transcriptional

  10. Transforming growth factor beta-1 and interleukin-17 gene transcription in peripheral blood mononuclear cells and the human response to infection.

    LENUS (Irish Health Repository)

    White, Mary

    2012-02-01

    INTRODUCTION: The occurrence of severe sepsis may be associated with deficient pro-inflammatory cytokine production. Transforming growth factor beta-1 (TGFbeta-1) predominantly inhibits inflammation and may simultaneously promote IL-17 production. Interleukin-17 (IL-17) is a recently described pro-inflammatory cytokine, which may be important in auto-immunity and infection. We investigated the hypothesis that the onset of sepsis is related to differential TGFbeta-1 and IL-17 gene expression. METHODS: A prospective observational study in a mixed intensive care unit (ICU) and hospital wards in a university hospital. Patients (59) with severe sepsis; 15 patients with gram-negative bacteraemia but without critical illness and 10 healthy controls were assayed for TGFbeta-1, IL-17a, IL-17f, IL-6 and IL-1beta mRNA in peripheral blood mononuclear cells (PBMC) by quantitative real-time PCR and serum protein levels by ELISA. RESULTS: TGFbeta-1 mRNA levels are reduced in patients with bacteraemia and sepsis compared with controls (p=0.02). IL-6 mRNA levels were reduced in bacteraemic patients compared with septic patients and controls (p=0.008). IL-1beta mRNA levels were similar in all groups, IL-17a and IL-17f mRNA levels are not detectable in peripheral blood mononuclear cells. IL-6 protein levels were greater in patients with sepsis than bacteraemic and control patients (p<0.0001). Activated TGFbeta-1 and IL-17 protein levels were similar in all groups. IL-1beta protein was not detectable in the majority of patients. CONCLUSIONS: Down regulation of TGFbeta-1 gene transcription was related to the occurrence of infection but not the onset of sepsis. Interleukin-17 production in PBMC may not be significant in the human host response to infection.

  11. Overlapping transcription structure of human cytomegalovirus

    Indian Academy of Sciences (India)

    Transcription of human cytomegalovirus UL/b′ region has been studied extensively for some genes. In this study, transcripts of the UL140 and UL141, two of the UL/b′ genes, were identified in late RNAs of three HCMV isolates using Northern blot hybridization, cDNA library screening and RACE-PCR. At least three ...

  12. Overlapping transcription structure of human cytomegalovirus ...

    Indian Academy of Sciences (India)

    2013-01-21

    Jan 21, 2013 ... Transcription of human cytomegalovirus UL/b′ region has been studied extensively for some genes. In this study, transcripts of the UL140 and UL141, two of the UL/b′ genes, were identified in late RNAs of three HCMV isolates using Northern blot hybridization, cDNA library screening and RACE-PCR.

  13. Over-expression of Oct4 and Sox2 transcription factors enhances differentiation of human umbilical cord blood cells in vivo

    International Nuclear Information System (INIS)

    Guseva, Daria; Rizvanov, Albert A.; Salafutdinov, Ilnur I.; Kudryashova, Nezhdana V.; Palotás, András; Islamov, Rustem R.

    2014-01-01

    Highlights: • Gene and cell-based therapies comprise innovative aspects of regenerative medicine. • Genetically modified hUCB-MCs enhanced differentiation of cells in a mouse model of ALS. • Stem cells successfully transformed into micro-glial and endothelial lines in spinal cords. • Over-expressing oct4 and sox2 also induced production of neural marker PGP9.5. • Formation of new nerve cells, secreting trophic factors and neo-vascularisation could improve symptoms in ALS. - Abstract: Gene and cell-based therapies comprise innovative aspects of regenerative medicine. Even though stem cells represent a highly potential therapeutic strategy, their wide-spread exploitation is marred by ethical concerns, potential for malignant transformation and a plethora of other technical issues, largely restricting their use to experimental studies. Utilizing genetically modified human umbilical cord blood mono-nuclear cells (hUCB-MCs), this communication reports enhanced differentiation of transplants in a mouse model of amyotrophic lateral sclerosis (ALS). Over-expressing Oct4 and Sox2 induced production of neural marker PGP9.5, as well as transformation of hUCB-MCs into micro-glial and endothelial lines in ALS spinal cords. In addition to producing new nerve cells, providing degenerated areas with trophic factors and neo-vascularisation might prevent and even reverse progressive loss of moto-neurons and skeletal muscle paralysis

  14. Over-expression of Oct4 and Sox2 transcription factors enhances differentiation of human umbilical cord blood cells in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Guseva, Daria [Kazan State Medical University, Kazan, Republic of Tatarstan (Russian Federation); Hannover Medical School, Hannover (Germany); Rizvanov, Albert A.; Salafutdinov, Ilnur I.; Kudryashova, Nezhdana V. [Kazan Federal University, Kazan, Republic of Tatarstan (Russian Federation); Palotás, András, E-mail: palotas@asklepios-med.eu [Kazan Federal University, Kazan, Republic of Tatarstan (Russian Federation); Asklepios-Med (Private Medical Practice and Research Center), Szeged (Hungary); Islamov, Rustem R., E-mail: islamru@yahoo.com [Kazan State Medical University, Kazan, Republic of Tatarstan (Russian Federation)

    2014-09-05

    Highlights: • Gene and cell-based therapies comprise innovative aspects of regenerative medicine. • Genetically modified hUCB-MCs enhanced differentiation of cells in a mouse model of ALS. • Stem cells successfully transformed into micro-glial and endothelial lines in spinal cords. • Over-expressing oct4 and sox2 also induced production of neural marker PGP9.5. • Formation of new nerve cells, secreting trophic factors and neo-vascularisation could improve symptoms in ALS. - Abstract: Gene and cell-based therapies comprise innovative aspects of regenerative medicine. Even though stem cells represent a highly potential therapeutic strategy, their wide-spread exploitation is marred by ethical concerns, potential for malignant transformation and a plethora of other technical issues, largely restricting their use to experimental studies. Utilizing genetically modified human umbilical cord blood mono-nuclear cells (hUCB-MCs), this communication reports enhanced differentiation of transplants in a mouse model of amyotrophic lateral sclerosis (ALS). Over-expressing Oct4 and Sox2 induced production of neural marker PGP9.5, as well as transformation of hUCB-MCs into micro-glial and endothelial lines in ALS spinal cords. In addition to producing new nerve cells, providing degenerated areas with trophic factors and neo-vascularisation might prevent and even reverse progressive loss of moto-neurons and skeletal muscle paralysis.

  15. Colon cancer associated transcripts in human cancers.

    Science.gov (United States)

    Chen, Yincong; Xie, Haibiao; Gao, Qunjun; Zhan, Hengji; Xiao, Huizhong; Zou, Yifan; Zhang, Fuyou; Liu, Yuchen; Li, Jianfa

    2017-10-01

    Long non-coding RNAs serve as important regulators in complicated cellular activities, including cell differentiation, proliferation and death. Dysregulation of long non-coding RNAs occurs in the formation and progression of cancers. The family of colon cancer associated transcripts, long non-coding RNAs colon cancer associated transcript-1 and colon cancer associated transcript-2 are known as oncogenes involved in various cancers. Colon cancer associated transcript-1 is a novel lncRNA located in 8q24.2, and colon cancer associated transcript-2 maps to the 8q24.21 region encompassing rs6983267. Colon cancer associated transcripts have close associations with clinical characteristics, such as lymph node metastasis, high TNM stage and short overall survival. Knockdown of them can reverse the malignant phenotypes of cancer cells, including proliferation, migration, invasion and apoptosis. Moreover, they can increase the expression level of c-MYC and oncogenic microRNAs via activating a series of complex mechanisms. In brief, the family of colon cancer associated transcripts may serve as potential biomarkers or therapeutic targets for human cancers. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  16. Mitochondrial transcription factor A protects human retinal ...

    African Journals Online (AJOL)

    Purpose: To investigate the impact of mitochondrial transcription factor A (TFAM), as a modulator of NF-κB, on proliferation of hypoxia-induced human retinal endothelial cell (HREC), and the probable mechanism. Methods: After exposure to hypoxia (1 % O2) for 5 days, cell proliferation and cell cycle of HREC were ...

  17. Blood Transcriptional Signatures for Disease Progression in a Rat Model of Osteoarthritis

    Directory of Open Access Journals (Sweden)

    Michał Korostyński

    2017-01-01

    Full Text Available Biomarkers of osteoarthritis (OA that can accurately diagnose the disease at the earliest stage would significantly support efforts to develop treatments for prevention and early intervention. We have sought to determine the time course of alterations in peripheral blood gene expression profile associated with the development of OA. Blood samples were collected from a tail vein of individual rats with monosodium iodoacetate- (MIA- induced OA (2, 14, 21, and 28 days after the treatment. We used whole-genome microarrays to reveal OA-related transcriptional alterations of 72 transcripts. Three main groups of coexpressed genes revealed diverse time-dependent profiles of up- and downregulation. Functional links that connect expression of the gradually downregulated genes to the G13 signaling pathway were indicated. The mRNA abundance levels of the identified transcripts were further analyzed in publicly available gene expression dataset obtained from a GARP study cohort of OA patients. We revealed three-gene signature differentially expressed in both rat and human blood (TNK2, KCTD2, and WDR37. The alterations in expression of the selected transcripts in peripheral blood samples of the patients indicate heterogeneity of the OA profiles potentially related to disease progress and severity of clinical symptoms. Our study identifies several potential stage-specific biomarkers of OA progression.

  18. The Human Blood Metabolome-Transcriptome Interface

    Science.gov (United States)

    Schramm, Katharina; Adamski, Jerzy; Gieger, Christian; Herder, Christian; Carstensen, Maren; Peters, Annette; Rathmann, Wolfgang; Roden, Michael; Strauch, Konstantin; Suhre, Karsten; Kastenmüller, Gabi; Prokisch, Holger; Theis, Fabian J.

    2015-01-01

    Biological systems consist of multiple organizational levels all densely interacting with each other to ensure function and flexibility of the system. Simultaneous analysis of cross-sectional multi-omics data from large population studies is a powerful tool to comprehensively characterize the underlying molecular mechanisms on a physiological scale. In this study, we systematically analyzed the relationship between fasting serum metabolomics and whole blood transcriptomics data from 712 individuals of the German KORA F4 cohort. Correlation-based analysis identified 1,109 significant associations between 522 transcripts and 114 metabolites summarized in an integrated network, the ‘human blood metabolome-transcriptome interface’ (BMTI). Bidirectional causality analysis using Mendelian randomization did not yield any statistically significant causal associations between transcripts and metabolites. A knowledge-based interpretation and integration with a genome-scale human metabolic reconstruction revealed systematic signatures of signaling, transport and metabolic processes, i.e. metabolic reactions mainly belonging to lipid, energy and amino acid metabolism. Moreover, the construction of a network based on functional categories illustrated the cross-talk between the biological layers at a pathway level. Using a transcription factor binding site enrichment analysis, this pathway cross-talk was further confirmed at a regulatory level. Finally, we demonstrated how the constructed networks can be used to gain novel insights into molecular mechanisms associated to intermediate clinical traits. Overall, our results demonstrate the utility of a multi-omics integrative approach to understand the molecular mechanisms underlying both normal physiology and disease. PMID:26086077

  19. The Human Blood Metabolome-Transcriptome Interface.

    Directory of Open Access Journals (Sweden)

    Jörg Bartel

    2015-06-01

    Full Text Available Biological systems consist of multiple organizational levels all densely interacting with each other to ensure function and flexibility of the system. Simultaneous analysis of cross-sectional multi-omics data from large population studies is a powerful tool to comprehensively characterize the underlying molecular mechanisms on a physiological scale. In this study, we systematically analyzed the relationship between fasting serum metabolomics and whole blood transcriptomics data from 712 individuals of the German KORA F4 cohort. Correlation-based analysis identified 1,109 significant associations between 522 transcripts and 114 metabolites summarized in an integrated network, the 'human blood metabolome-transcriptome interface' (BMTI. Bidirectional causality analysis using Mendelian randomization did not yield any statistically significant causal associations between transcripts and metabolites. A knowledge-based interpretation and integration with a genome-scale human metabolic reconstruction revealed systematic signatures of signaling, transport and metabolic processes, i.e. metabolic reactions mainly belonging to lipid, energy and amino acid metabolism. Moreover, the construction of a network based on functional categories illustrated the cross-talk between the biological layers at a pathway level. Using a transcription factor binding site enrichment analysis, this pathway cross-talk was further confirmed at a regulatory level. Finally, we demonstrated how the constructed networks can be used to gain novel insights into molecular mechanisms associated to intermediate clinical traits. Overall, our results demonstrate the utility of a multi-omics integrative approach to understand the molecular mechanisms underlying both normal physiology and disease.

  20. Curated compendium of human transcriptional biomarker data.

    Science.gov (United States)

    Golightly, Nathan P; Bell, Avery; Bischoff, Anna I; Hollingsworth, Parker D; Piccolo, Stephen R

    2018-04-17

    One important use of genome-wide transcriptional profiles is to identify relationships between transcription levels and patient outcomes. These translational insights can guide the development of biomarkers for clinical application. Data from thousands of translational-biomarker studies have been deposited in public repositories, enabling reuse. However, data-reuse efforts require considerable time and expertise because transcriptional data are generated using heterogeneous profiling technologies, preprocessed using diverse normalization procedures, and annotated in non-standard ways. To address this problem, we curated 45 publicly available, translational-biomarker datasets from a variety of human diseases. To increase the data's utility, we reprocessed the raw expression data using a uniform computational pipeline, addressed quality-control problems, mapped the clinical annotations to a controlled vocabulary, and prepared consistently structured, analysis-ready data files. These data, along with scripts we used to prepare the data, are available in a public repository. We believe these data will be particularly useful to researchers seeking to perform benchmarking studies-for example, to compare and optimize machine-learning algorithms' ability to predict biomedical outcomes.

  1. Transcriptional blood signatures distinguish pulmonary tuberculosis, pulmonary sarcoidosis, pneumonias and lung cancers.

    Science.gov (United States)

    Bloom, Chloe I; Graham, Christine M; Berry, Matthew P R; Rozakeas, Fotini; Redford, Paul S; Wang, Yuanyuan; Xu, Zhaohui; Wilkinson, Katalin A; Wilkinson, Robert J; Kendrick, Yvonne; Devouassoux, Gilles; Ferry, Tristan; Miyara, Makoto; Bouvry, Diane; Valeyre, Dominique; Dominique, Valeyre; Gorochov, Guy; Blankenship, Derek; Saadatian, Mitra; Vanhems, Phillip; Beynon, Huw; Vancheeswaran, Rama; Wickremasinghe, Melissa; Chaussabel, Damien; Banchereau, Jacques; Pascual, Virginia; Ho, Ling-Pei; Lipman, Marc; O'Garra, Anne

    2013-01-01

    New approaches to define factors underlying the immunopathogenesis of pulmonary diseases including sarcoidosis and tuberculosis are needed to develop new treatments and biomarkers. Comparing the blood transcriptional response of tuberculosis to other similar pulmonary diseases will advance knowledge of disease pathways and help distinguish diseases with similar clinical presentations. To determine the factors underlying the immunopathogenesis of the granulomatous diseases, sarcoidosis and tuberculosis, by comparing the blood transcriptional responses in these and other pulmonary diseases. We compared whole blood genome-wide transcriptional profiles in pulmonary sarcoidosis, pulmonary tuberculosis, to community acquired pneumonia and primary lung cancer and healthy controls, before and after treatment, and in purified leucocyte populations. An Interferon-inducible neutrophil-driven blood transcriptional signature was present in both sarcoidosis and tuberculosis, with a higher abundance and expression in tuberculosis. Heterogeneity of the sarcoidosis signature correlated significantly with disease activity. Transcriptional profiles in pneumonia and lung cancer revealed an over-abundance of inflammatory transcripts. After successful treatment the transcriptional activity in tuberculosis and pneumonia patients was significantly reduced. However the glucocorticoid-responsive sarcoidosis patients showed a significant increase in transcriptional activity. 144-blood transcripts were able to distinguish tuberculosis from other lung diseases and controls. Tuberculosis and sarcoidosis revealed similar blood transcriptional profiles, dominated by interferon-inducible transcripts, while pneumonia and lung cancer showed distinct signatures, dominated by inflammatory genes. There were also significant differences between tuberculosis and sarcoidosis in the degree of their transcriptional activity, the heterogeneity of their profiles and their transcriptional response to treatment.

  2. A human transcription factor in search mode.

    Science.gov (United States)

    Hauser, Kevin; Essuman, Bernard; He, Yiqing; Coutsias, Evangelos; Garcia-Diaz, Miguel; Simmerling, Carlos

    2016-01-08

    Transcription factors (TF) can change shape to bind and recognize DNA, shifting the energy landscape from a weak binding, rapid search mode to a higher affinity recognition mode. However, the mechanism(s) driving this conformational change remains unresolved and in most cases high-resolution structures of the non-specific complexes are unavailable. Here, we investigate the conformational switch of the human mitochondrial transcription termination factor MTERF1, which has a modular, superhelical topology complementary to DNA. Our goal was to characterize the details of the non-specific search mode to complement the crystal structure of the specific binding complex, providing a basis for understanding the recognition mechanism. In the specific complex, MTERF1 binds a significantly distorted and unwound DNA structure, exhibiting a protein conformation incompatible with binding to B-form DNA. In contrast, our simulations of apo MTERF1 revealed significant flexibility, sampling structures with superhelical pitch and radius complementary to the major groove of B-DNA. Docking these structures to B-DNA followed by unrestrained MD simulations led to a stable complex in which MTERF1 was observed to undergo spontaneous diffusion on the DNA. Overall, the data support an MTERF1-DNA binding and recognition mechanism driven by intrinsic dynamics of the MTERF1 superhelical topology. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  3. An anatomic transcriptional atlas of human glioblastoma.

    Science.gov (United States)

    Puchalski, Ralph B; Shah, Nameeta; Miller, Jeremy; Dalley, Rachel; Nomura, Steve R; Yoon, Jae-Guen; Smith, Kimberly A; Lankerovich, Michael; Bertagnolli, Darren; Bickley, Kris; Boe, Andrew F; Brouner, Krissy; Butler, Stephanie; Caldejon, Shiella; Chapin, Mike; Datta, Suvro; Dee, Nick; Desta, Tsega; Dolbeare, Tim; Dotson, Nadezhda; Ebbert, Amanda; Feng, David; Feng, Xu; Fisher, Michael; Gee, Garrett; Goldy, Jeff; Gourley, Lindsey; Gregor, Benjamin W; Gu, Guangyu; Hejazinia, Nika; Hohmann, John; Hothi, Parvinder; Howard, Robert; Joines, Kevin; Kriedberg, Ali; Kuan, Leonard; Lau, Chris; Lee, Felix; Lee, Hwahyung; Lemon, Tracy; Long, Fuhui; Mastan, Naveed; Mott, Erika; Murthy, Chantal; Ngo, Kiet; Olson, Eric; Reding, Melissa; Riley, Zack; Rosen, David; Sandman, David; Shapovalova, Nadiya; Slaughterbeck, Clifford R; Sodt, Andrew; Stockdale, Graham; Szafer, Aaron; Wakeman, Wayne; Wohnoutka, Paul E; White, Steven J; Marsh, Don; Rostomily, Robert C; Ng, Lydia; Dang, Chinh; Jones, Allan; Keogh, Bart; Gittleman, Haley R; Barnholtz-Sloan, Jill S; Cimino, Patrick J; Uppin, Megha S; Keene, C Dirk; Farrokhi, Farrokh R; Lathia, Justin D; Berens, Michael E; Iavarone, Antonio; Bernard, Amy; Lein, Ed; Phillips, John W; Rostad, Steven W; Cobbs, Charles; Hawrylycz, Michael J; Foltz, Greg D

    2018-05-11

    Glioblastoma is an aggressive brain tumor that carries a poor prognosis. The tumor's molecular and cellular landscapes are complex, and their relationships to histologic features routinely used for diagnosis are unclear. We present the Ivy Glioblastoma Atlas, an anatomically based transcriptional atlas of human glioblastoma that aligns individual histologic features with genomic alterations and gene expression patterns, thus assigning molecular information to the most important morphologic hallmarks of the tumor. The atlas and its clinical and genomic database are freely accessible online data resources that will serve as a valuable platform for future investigations of glioblastoma pathogenesis, diagnosis, and treatment. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  4. From reverse transcription to human brain tumors

    Directory of Open Access Journals (Sweden)

    Dmitrenko V. V.

    2013-05-01

    Full Text Available Reverse transcriptase from avian myeloblastosis virus (AMV was the subject of the study, from which the investi- gations of the Department of biosynthesis of nucleic acids were started. Production of AMV in grams quantities and isolation of AMV reverse transcriptase were established in the laboratory during the seventies of the past cen- tury and this initiated research on the cDNA synthesis, cloning and investigation of the structure and functions of the eukaryotic genes. Structures of salmon insulin and insulin-like growth factor (IGF family genes and their transcripts were determined during long-term investigations. Results of two modern techniques, microarray-ba- sed hybridization and SAGE, were used for the identification of the genes differentially expressed in astrocytic gliomas and human normal brain. Comparison of SAGE results on the genes overexpressed in glioblastoma with the results of microarray analysis revealed a limited number of common genes. 105 differentially expressed genes, common to both methods, can be included in the list of candidates for the molecular typing of glioblastoma. The first experiments on the classification of glioblastomas based on the data of the 20 genes expression were conducted by using of artificial neural network analysis. The results of these experiments showed that the expression profiles of these genes in 224 glioblastoma samples and 74 normal brain samples could be according to the Koho- nen’s maps. The CHI3L1 and CHI3L2 genes of chitinase-like cartilage protein were revealed among the most overexpressed genes in glioblastoma, which could have prognostic and diagnostic potential. Results of in vitro experiments demonstrated that both proteins, CHI3L1 and CHI3L2, may initiate the phosphorylation of ERK1/ ERK2 and AKT kinases leading to the activation of MAPK/ERK1/2 and PI3K/AKT signaling cascades in human embryonic kidney 293 cells, human glioblastoma U87MG, and U373 cells. The new human cell line

  5. Blood-informative transcripts define nine common axes of peripheral blood gene expression.

    Directory of Open Access Journals (Sweden)

    Marcela Preininger

    Full Text Available We describe a novel approach to capturing the covariance structure of peripheral blood gene expression that relies on the identification of highly conserved Axes of variation. Starting with a comparison of microarray transcriptome profiles for a new dataset of 189 healthy adult participants in the Emory-Georgia Tech Center for Health Discovery and Well-Being (CHDWB cohort, with a previously published study of 208 adult Moroccans, we identify nine Axes each with between 99 and 1,028 strongly co-regulated transcripts in common. Each axis is enriched for gene ontology categories related to sub-classes of blood and immune function, including T-cell and B-cell physiology and innate, adaptive, and anti-viral responses. Conservation of the Axes is demonstrated in each of five additional population-based gene expression profiling studies, one of which is robustly associated with Body Mass Index in the CHDWB as well as Finnish and Australian cohorts. Furthermore, ten tightly co-regulated genes can be used to define each Axis as "Blood Informative Transcripts" (BITs, generating scores that define an individual with respect to the represented immune activity and blood physiology. We show that environmental factors, including lifestyle differences in Morocco and infection leading to active or latent tuberculosis, significantly impact specific axes, but that there is also significant heritability for the Axis scores. In the context of personalized medicine, reanalysis of the longitudinal profile of one individual during and after infection with two respiratory viruses demonstrates that specific axes also characterize clinical incidents. This mode of analysis suggests the view that, rather than unique subsets of genes marking each class of disease, differential expression reflects movement along the major normal Axes in response to environmental and genetic stimuli.

  6. Blood-informative transcripts define nine common axes of peripheral blood gene expression.

    Science.gov (United States)

    Preininger, Marcela; Arafat, Dalia; Kim, Jinhee; Nath, Artika P; Idaghdour, Youssef; Brigham, Kenneth L; Gibson, Greg

    2013-01-01

    We describe a novel approach to capturing the covariance structure of peripheral blood gene expression that relies on the identification of highly conserved Axes of variation. Starting with a comparison of microarray transcriptome profiles for a new dataset of 189 healthy adult participants in the Emory-Georgia Tech Center for Health Discovery and Well-Being (CHDWB) cohort, with a previously published study of 208 adult Moroccans, we identify nine Axes each with between 99 and 1,028 strongly co-regulated transcripts in common. Each axis is enriched for gene ontology categories related to sub-classes of blood and immune function, including T-cell and B-cell physiology and innate, adaptive, and anti-viral responses. Conservation of the Axes is demonstrated in each of five additional population-based gene expression profiling studies, one of which is robustly associated with Body Mass Index in the CHDWB as well as Finnish and Australian cohorts. Furthermore, ten tightly co-regulated genes can be used to define each Axis as "Blood Informative Transcripts" (BITs), generating scores that define an individual with respect to the represented immune activity and blood physiology. We show that environmental factors, including lifestyle differences in Morocco and infection leading to active or latent tuberculosis, significantly impact specific axes, but that there is also significant heritability for the Axis scores. In the context of personalized medicine, reanalysis of the longitudinal profile of one individual during and after infection with two respiratory viruses demonstrates that specific axes also characterize clinical incidents. This mode of analysis suggests the view that, rather than unique subsets of genes marking each class of disease, differential expression reflects movement along the major normal Axes in response to environmental and genetic stimuli.

  7. A brief history of human blood groups.

    Science.gov (United States)

    Farhud, Dariush D; Zarif Yeganeh, Marjan

    2013-01-01

    The evolution of human blood groups, without doubt, has a history as old as man himself. There are at least three hypotheses about the emergence and mutation of human blood groups. Global distribution pattern of blood groups depends on various environmental factors, such as disease, climate, altitude, humidity etc. In this survey, the collection of main blood groups ABO and Rh, along with some minor groups, are presented. Several investigations of blood groups from Iran, particularly a large sampling on 291857 individuals from Iran, including the main blood groups ABO and Rh, as well as minor blood groups such as Duffy, Lutheran, Kell, KP, Kidd, and Xg, have been reviewed.

  8. Quantification of BCR-ABL transcripts in peripheral blood cells and ...

    African Journals Online (AJOL)

    Purpose: To investigate the feasibility of using peripheral blood plasma samples as surrogates for blood cell sampling for quantification of breakpoint cluster region-Abelson oncogene (BCR-ABL) transcript levels to monitor treatment responses in chronic myeloid leukemia (CML) patients. Methods: Peripheral blood samples ...

  9. Novel transcriptional networks regulated by CLOCK in human neurons.

    Science.gov (United States)

    Fontenot, Miles R; Berto, Stefano; Liu, Yuxiang; Werthmann, Gordon; Douglas, Connor; Usui, Noriyoshi; Gleason, Kelly; Tamminga, Carol A; Takahashi, Joseph S; Konopka, Genevieve

    2017-11-01

    The molecular mechanisms underlying human brain evolution are not fully understood; however, previous work suggested that expression of the transcription factor CLOCK in the human cortex might be relevant to human cognition and disease. In this study, we investigated this novel transcriptional role for CLOCK in human neurons by performing chromatin immunoprecipitation sequencing for endogenous CLOCK in adult neocortices and RNA sequencing following CLOCK knockdown in differentiated human neurons in vitro. These data suggested that CLOCK regulates the expression of genes involved in neuronal migration, and a functional assay showed that CLOCK knockdown increased neuronal migratory distance. Furthermore, dysregulation of CLOCK disrupts coexpressed networks of genes implicated in neuropsychiatric disorders, and the expression of these networks is driven by hub genes with human-specific patterns of expression. These data support a role for CLOCK-regulated transcriptional cascades involved in human brain evolution and function. © 2017 Fontenot et al.; Published by Cold Spring Harbor Laboratory Press.

  10. Human cytomegalovirus (HCMV) induces human endogenous retrovirus (HERV) transcription.

    Science.gov (United States)

    Assinger, Alice; Yaiw, Koon-Chu; Göttesdorfer, Ingmar; Leib-Mösch, Christine; Söderberg-Nauclér, Cecilia

    2013-11-12

    Emerging evidence suggests that human cytomegalovirus (HCMV) is highly prevalent in tumours of different origin. This virus is implied to have oncogenic and oncomodulatory functions, through its ability to control host gene expression. Human endogenous retroviruses (HERV) are also frequently active in tumours of different origin, and are supposed to contribute as cofactors to cancer development. Due to the high prevalence of HCMV in several different tumours, and its ability to control host cell gene expression, we sought to define whether HCMV may affect HERV transcription. Infection of 3 established cancer cell lines, 2 primary glioblastoma cells, endothelial cells from 3 donors and monocytes from 4 donors with HCMV (strains VR 1814 or TB40/F) induced reverse transcriptase (RT) activity in all cells tested, but the response varied between donors. Both, gammaretrovirus-related class I elements HERV-T, HERV-W, HERV-F and ERV-9, and betaretrovirus-related class II elements HML-2 - 4 and HML-7 - 8, as well as spuma-virus related class III elements of the HERV-L group were up-regulated in response to HCMV infection in GliNS1 cells. Up-regulation of HERV activity was more pronounced in cells harbouring active HCMV infection, but was also induced by UV-inactivated virus. The effect was only slightly affected by ganciclovir treatment and was not controlled by the IE72 or IE86 HCMV genes. Within this brief report we show that HCMV infection induces HERV transcriptional activity in different cell types.

  11. Characterization of human mesothelin transcripts in ovarian and pancreatic cancer

    International Nuclear Information System (INIS)

    Muminova, Zhanat E; Strong, Theresa V; Shaw, Denise R

    2004-01-01

    Mesothelin is an attractive target for cancer immunotherapy due to its restricted expression in normal tissues and high level expression in several tumor types including ovarian and pancreatic adenocarcinomas. Three mesothelin transcript variants have been reported, but their relative expression in normal tissues and tumors has been poorly characterized. The goal of the present study was to clarify which mesothelin transcript variants are commonly expressed in human tumors. Human genomic and EST nucleotide sequences in the public databases were used to evaluate sequences reported for the three mesothelin transcript variants in silico. Subsequently, RNA samples from normal ovary, ovarian and pancreatic carcinoma cell lines, and primary ovarian tumors were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and nucleotide sequencing to directly identify expressed transcripts. In silico comparisons of genomic DNA sequences with available EST sequences supported expression of mesothelin transcript variants 1 and 3, but there were no sequence matches for transcript variant 2. Newly-derived nucleotide sequences of RT-PCR products from tissues and cell lines corresponded to mesothelin transcript variant 1. Mesothelin transcript variant 2 was not detected. Transcript variant 3 was observed as a small percentage of total mesothelin amplification products from all studied cell lines and tissues. Fractionation of nuclear and cytoplasmic RNA indicated that variant 3 was present primarily in the nuclear fraction. Thus, mesothelin transcript variant 3 may represent incompletely processed hnRNA. Mesothelin transcript variant 1 represents the predominant mature mRNA species expressed by both normal and tumor cells. This conclusion should be important for future development of cancer immunotherapies, diagnostic tests, and gene microarray studies targeting mesothelin

  12. Transcriptional and post-transcriptional regulation of nucleotide excision repair genes in human cells

    Energy Technology Data Exchange (ETDEWEB)

    Lefkofsky, Hailey B. [Translational Oncology Program, University of Michigan Medical School, Ann Arbor, MI (United States); Veloso, Artur [Translational Oncology Program, University of Michigan Medical School, Ann Arbor, MI (United States); Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI (United States); Bioinformatics Program, Department of Computational Medicine and Bioinformatics, University of Michigan, Ann Arbor, MI (United States); Ljungman, Mats, E-mail: ljungman@umich.edu [Translational Oncology Program, University of Michigan Medical School, Ann Arbor, MI (United States); Department of Radiation Oncology, University of Michigan Medical School, Ann Arbor, MI (United States); Department of Environmental Health Sciences, School of Public Health, University of Michigan, Ann Arbor, MI (United States)

    2015-06-15

    Nucleotide excision repair (NER) removes DNA helix-distorting lesions induced by UV light and various chemotherapeutic agents such as cisplatin. These lesions efficiently block the elongation of transcription and need to be rapidly removed by transcription-coupled NER (TC-NER) to avoid the induction of apoptosis. Twenty-nine genes have been classified to code for proteins participating in nucleotide excision repair (NER) in human cells. Here we explored the transcriptional and post-transcriptional regulation of these NER genes across 13 human cell lines using Bru-seq and BruChase-seq, respectively. Many NER genes are relatively large in size and therefore will be easily inactivated by UV-induced transcription-blocking lesions. Furthermore, many of these genes produce transcripts that are rather unstable. Thus, these genes are expected to rapidly lose expression leading to a diminished function of NER. One such gene is ERCC6 that codes for the CSB protein critical for TC-NER. Due to its large gene size and high RNA turnover rate, the ERCC6 gene may act as dosimeter of DNA damage so that at high levels of damage, ERCC6 RNA levels would be diminished leading to the loss of CSB expression, inhibition of TC-NER and the promotion of cell death.

  13. Transcriptional response to hypoxia in human tumors.

    NARCIS (Netherlands)

    Lal, A.; Peters, H.; Croix, B. St.; Haroon, Z.A.; Dewhirst, M.W.; Strausberg, R.L.; Kaanders, J.H.A.M.; Kogel, A.J. van der; Riggins, G.J.

    2001-01-01

    BACKGROUND: The presence of hypoxic regions within solid tumors is associated with a more malignant tumor phenotype and worse prognosis. To obtain a blood supply and protect against cellular damage and death, oxygen-deprived cells in tumors alter gene expression, resulting in resistance to therapy.

  14. Transcription factors: normal and malignant development of blood cells

    National Research Council Canada - National Science Library

    Ravid, Katya; Licht, Jonathan

    2001-01-01

    ... and the Development of the Erythroid Lineage James J. Bieker 71 II TRANSCRIPTION FACTORS AND THE MYELOID LINEAGE 85 6 RUNX1(AML1) and CBFB: Genes Required for the Development of All Definitive Hematopoietic Lineages 87 Nancy A. Speck and Elaine Dzierzak 7 PU.1 and the Development of the Myeloid Lineage Daniel G. Tenen 103 vvi CONTENTS 8 CCAAT/Enhancer-...

  15. Identification of identical transcript changes in liver and whole blood during acetaminophen toxicity

    Directory of Open Access Journals (Sweden)

    Liwen eZhang

    2012-09-01

    Full Text Available Abstract The ability to identify mechanisms underlying drug-induced liver injury (DILI in man has been hampered by the difficulty in obtaining liver tissue from patients. It has recently been proposed that whole blood toxicogenomics may provide a noninvasive means for mechanistic studies of human DILI. However, it remains unclear to what extent changes in whole blood transcriptome mirror those in liver mechanistically linked to hepatotoxicity. To address this question, we applied the program Extracting Patterns and Identifying co-expressed Genes (EPIG to publically available toxicogenomic data obtained from rats treated with both toxic and subtoxic doses of acetaminophen (APAP. In a training set of animals, we identified genes (760 at 6 h and 185 at 24 h post dose with similar patterns of expression in blood and liver during APAP induced hepatotoxicity. The pathways represented in the coordinately regulated genes largely involved mitochondrial and immune functions. The identified expression signatures were then evaluated in a separate set of animals for discernment of APAP exposure level or APAP induced hepatotoxicity. At 6 h, the gene sets from liver and blood had equally sufficient classification of APAP exposure levels. At 24 h when toxicity was evident, the gene sets did not perform well in evaluating APAP exposure doses, but provided accurate classification of dose-independent liver injury that was evaluated by serum ALT elevation in the blood. Only thirty eight genes were common to both the 6 and 24h gene sets, but these genes had the same capability as the parent gene sets to discern the exposure level and degree of liver injury. Some of the parallel transcript changes reflect pathways that are relevant to APAP hepatotoxicity, including mitochondria and immune functions. However, the extent to which these changes reflect similar mechanisms of action in both tissues remains to be determined.

  16. Cyclin D3 interacts with human activating transcription factor 5 and potentiates its transcription activity

    International Nuclear Information System (INIS)

    Liu Wenjin; Sun Maoyun; Jiang Jianhai; Shen Xiaoyun; Sun Qing; Liu Weicheng; Shen Hailian; Gu Jianxin

    2004-01-01

    The Cyclin D3 protein is a member of the D-type cyclins. Besides serving as cell cycle regulators, D-type cyclins have been reported to be able to interact with several transcription factors and modulate their transcriptional activations. Here we report that human activating transcription factor 5 (hATF5) is a new interacting partner of Cyclin D3. The interaction was confirmed by in vivo coimmunoprecipitation and in vitro binding analysis. Neither interaction between Cyclin D1 and hATF5 nor interaction between Cyclin D2 and hATF5 was observed. Confocal microscopy analysis showed that Cyclin D3 could colocalize with hATF5 in the nuclear region. Cyclin D3 could potentiate hATF5 transcriptional activity independently of its Cdk4 partner. But Cyclin D1 and Cyclin D2 had no effect on hATF5 transcriptional activity. These data provide a new clue to understand the new role of Cyclin D3 as a transcriptional regulator

  17. Effect of laser on human blood

    International Nuclear Information System (INIS)

    Abdalsamad, Amuna Nagash Mohammed

    2016-03-01

    In this work, the effect of He-Ne (632.8 nm), N2 (337.1 nm), LED (450 nm) on the human blood, and blood component was studied by using CBC machine ( complete blood count) and UV -visible spectroscopy. Blood samples platoon A+ were collected and irradiated for different periods of time (10 minute, 20 minute, and 30 minute), to varied types of light source ( He- Ne laser and LED). Blood parameters of samples were measured by using complete Blood Count Machine (CBC). The absorption spectrum of blood samples were examined by using UV-visible spectrometer. The obtained results have shown different values of Complete Blood Counts and absorption spectrum due to different laser types and periods of time. We conclude that the laser light has clear effect on blood samples. (Author)

  18. Sensitive detection of viral transcripts in human tumor transcriptomes.

    Directory of Open Access Journals (Sweden)

    Sven-Eric Schelhorn

    Full Text Available In excess of 12% of human cancer incidents have a viral cofactor. Epidemiological studies of idiopathic human cancers indicate that additional tumor viruses remain to be discovered. Recent advances in sequencing technology have enabled systematic screenings of human tumor transcriptomes for viral transcripts. However, technical problems such as low abundances of viral transcripts in large volumes of sequencing data, viral sequence divergence, and homology between viral and human factors significantly confound identification of tumor viruses. We have developed a novel computational approach for detecting viral transcripts in human cancers that takes the aforementioned confounding factors into account and is applicable to a wide variety of viruses and tumors. We apply the approach to conducting the first systematic search for viruses in neuroblastoma, the most common cancer in infancy. The diverse clinical progression of this disease as well as related epidemiological and virological findings are highly suggestive of a pathogenic cofactor. However, a viral etiology of neuroblastoma is currently contested. We mapped 14 transcriptomes of neuroblastoma as well as positive and negative controls to the human and all known viral genomes in order to detect both known and unknown viruses. Analysis of controls, comparisons with related methods, and statistical estimates demonstrate the high sensitivity of our approach. Detailed investigation of putative viral transcripts within neuroblastoma samples did not provide evidence for the existence of any known human viruses. Likewise, de-novo assembly and analysis of chimeric transcripts did not result in expression signatures associated with novel human pathogens. While confounding factors such as sample dilution or viral clearance in progressed tumors may mask viral cofactors in the data, in principle, this is rendered less likely by the high sensitivity of our approach and the number of biological replicates

  19. Quantification of photocatalytic oxygenation of human blood.

    Science.gov (United States)

    Subrahmanyam, Aryasomayajula; Thangaraj, Paul R; Kanuru, Chandrasekhar; Jayakumar, Albert; Gopal, Jayashree

    2014-04-01

    Photocatalytic oxygenation of human blood is an emerging concept based on the principle of photocatalytic splitting of water into oxygen and hydrogen. This communication reports: (i) a design of a photocatalytic cell (PC) that separates the blood from UV (incident) radiation source, (ii) a pH, temperature and flow controlled circuit designed for quantifying the oxygenation of human blood by photocatalysis and (iii) measuring the current efficacy of ITO/TiO2 nano thin films in oxygenating human blood in a dynamic circuit in real time. The average increase in oxygen saturation was around 5% above baseline compared to control (p<0.0005). We believe this is one of the first attempts to quantify photocatalytic oxygenation of human blood under controlled conditions. Copyright © 2013 IPEM. Published by Elsevier Ltd. All rights reserved.

  20. Characterisation of CDKL5 Transcript Isoforms in Human and Mouse.

    Science.gov (United States)

    Hector, Ralph D; Dando, Owen; Landsberger, Nicoletta; Kilstrup-Nielsen, Charlotte; Kind, Peter C; Bailey, Mark E S; Cobb, Stuart R

    2016-01-01

    Mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5) cause early onset infantile spasms and subsequent severe developmental delay in affected children. Deleterious mutations have been reported to occur throughout the CDKL5 coding region. Several studies point to a complex CDKL5 gene structure in terms of exon usage and transcript expression. Improvements in molecular diagnosis and more extensive research into the neurobiology of CDKL5 and pathophysiology of CDKL5 disorders necessitate an updated analysis of the gene. In this study, we have analysed human and mouse CDKL5 transcript patterns both bioinformatically and experimentally. We have characterised the predominant brain isoform of CDKL5, a 9.7 kb transcript comprised of 18 exons with a large 6.6 kb 3'-untranslated region (UTR), which we name hCDKL5_1. In addition we describe new exonic regions and a range of novel splice and UTR isoforms. This has enabled the description of an updated gene model in both species and a standardised nomenclature system for CDKL5 transcripts. Profiling revealed tissue- and brain development stage-specific differences in expression between transcript isoforms. These findings provide an essential backdrop for the diagnosis of CDKL5-related disorders, for investigations into the basic biology of this gene and its protein products, and for the rational design of gene-based and molecular therapies for these disorders.

  1. Characterisation of CDKL5 Transcript Isoforms in Human and Mouse.

    Directory of Open Access Journals (Sweden)

    Ralph D Hector

    Full Text Available Mutations in the X-linked Cyclin-Dependent Kinase-Like 5 gene (CDKL5 cause early onset infantile spasms and subsequent severe developmental delay in affected children. Deleterious mutations have been reported to occur throughout the CDKL5 coding region. Several studies point to a complex CDKL5 gene structure in terms of exon usage and transcript expression. Improvements in molecular diagnosis and more extensive research into the neurobiology of CDKL5 and pathophysiology of CDKL5 disorders necessitate an updated analysis of the gene. In this study, we have analysed human and mouse CDKL5 transcript patterns both bioinformatically and experimentally. We have characterised the predominant brain isoform of CDKL5, a 9.7 kb transcript comprised of 18 exons with a large 6.6 kb 3'-untranslated region (UTR, which we name hCDKL5_1. In addition we describe new exonic regions and a range of novel splice and UTR isoforms. This has enabled the description of an updated gene model in both species and a standardised nomenclature system for CDKL5 transcripts. Profiling revealed tissue- and brain development stage-specific differences in expression between transcript isoforms. These findings provide an essential backdrop for the diagnosis of CDKL5-related disorders, for investigations into the basic biology of this gene and its protein products, and for the rational design of gene-based and molecular therapies for these disorders.

  2. 5' diversity of human hepatic PXR (NR1I2) transcripts and identification of the major transcription initiation site.

    Science.gov (United States)

    Kurose, Kouichi; Koyano, Satoru; Ikeda, Shinobu; Tohkin, Masahiro; Hasegawa, Ryuichi; Sawada, Jun-Ichi

    2005-05-01

    The human pregnane X receptor (PXR) is a crucial regulator of the genes encoding several major cytochrome P450 enzymes and transporters, such as CYP3A4 and MDR1, but its own transcriptional regulation remains unclear. To elucidate the transcriptional mechanisms of human PXR gene, we first endeavored to identify the transcription initiation site of human PXR using 5'-RACE. Five types of 5'-variable transcripts (a, b, c, d, and e) with common exon 2 sequence were found, and comparison of these sequences with the genomic sequence suggested that their 5' diversity is derived from initiation by alternative promoters and alternative splicing. None of the exons found in our study contain any new in-frame coding regions. Newly identified introns IVS-a and IVS-b were found to have CT-AC splice sites that do not follow the GT-AG rule of conventional donor and acceptor splice sites. Of the five types of 5' variable transcripts identified, RT-PCR showed that type-a was the major transcript type. Four transcription initiation sites (A-D) for type-a transcript were identified by 5'-RACE using GeneRacer RACE Ready cDNA (human liver) constructed by the oligo-capping method. Putative TATA boxes were located approximately 30 bp upstream from the transcriptional start sites of the major transcript (C) and the longest minor transcript (A) expressed in the human liver. These results indicate that the initiation of transcription of human PXR is more complex than previously reported.

  3. Transcriptional changes in blood after aerobic interval training in patients with the metabolic syndrome.

    Science.gov (United States)

    Bye, Anja; Tjønna, Arnt E; Stølen, Tomas O; Røsbjørgen, Ragnhild E N; Wisløff, Ulrik

    2009-02-01

    Regular physical activity has beneficial effects on the metabolic syndrome. Eleven metabolic syndrome patients performing 16 weeks of aerobic interval training, significantly reduced their risk of cardiovascular disease, in terms of improved VO2max, endothelial function, blood pressure, insulin signaling, and plasma lipid composition. The knowledge on underlying mechanism of exercise-induced improvements is sparse, and a broad spectrum of methods is needed to gain more insight. The aim was, for the first time, to determine whether transcriptional changes occur in blood cells of metabolic syndrome patients after participating in an exercise program. Blood was collected in PAXgene and EDTA tubes before and after 16 weeks of exercise. RNA was extracted and run on microarrays. Eleven biological processes and molecular functions were upregulated after exercise, whereas seven were downregulated. Blood clotting, cell adhesion, and steroid metabolism were among the downregulated processes, whereas steroid hormone-mediated signaling was upregulated. Downregulated protein levels of arginase 1 and von Willebrand factor confirmed microarray results. Increased transcription of genes involved in steroid hormone-mediated signaling, decreased levels of arginase 1, and reduced transcription of genes involved in cell adhesion, and blood clotting are likely to be involved in exercise-induced improvements of endothelial function, and improved cardiovascular risk profile of metabolic syndrome patients. These findings have provided new insights on exercise-induced improvement of cardiovascular health.

  4. Transcriptional changes common to human cocaine, cannabis and phencyclidine abuse.

    Directory of Open Access Journals (Sweden)

    Elin Lehrmann

    2006-12-01

    Full Text Available A major goal of drug abuse research is to identify and understand drug-induced changes in brain function that are common to many or all drugs of abuse. As these may underlie drug dependence and addiction, the purpose of the present study was to examine if different drugs of abuse effect changes in gene expression that converge in common molecular pathways. Microarray analysis was employed to assay brain gene expression in postmortem anterior prefrontal cortex (aPFC from 42 human cocaine, cannabis and/or phencyclidine abuse cases and 30 control cases, which were characterized by toxicology and drug abuse history. Common transcriptional changes were demonstrated for a majority of drug abuse cases (N = 34, representing a number of consistently changed functional classes: Calmodulin-related transcripts (CALM1, CALM2, CAMK2B were decreased, while transcripts related to cholesterol biosynthesis and trafficking (FDFT1, APOL2, SCARB1, and Golgi/endoplasmic reticulum (ER functions (SEMA3B, GCC1 were all increased. Quantitative PCR validated decreases in calmodulin 2 (CALM2 mRNA and increases in apolipoprotein L, 2 (APOL2 and semaphorin 3B (SEMA3B mRNA for individual cases. A comparison between control cases with and without cardiovascular disease and elevated body mass index indicated that these changes were not due to general cellular and metabolic stress, but appeared specific to the use of drugs. Therefore, humans who abused cocaine, cannabis and/or phencyclidine share a decrease in transcription of calmodulin-related genes and increased transcription related to lipid/cholesterol and Golgi/ER function. These changes represent common molecular features of drug abuse, which may underlie changes in synaptic function and plasticity that could have important ramifications for decision-making capabilities in drug abusers.

  5. Bone blood flow and metabolism in humans

    DEFF Research Database (Denmark)

    Heinonen, Ilkka; Kemppainen, Jukka; Kaskinoro, Kimmo

    2012-01-01

    Human bone blood flow and metabolism during physical exercise remains poorly characterised. In the present study we measured femoral bone blood flow and glucose uptake in young healthy subjects by positron emission tomography in three separate protocols. In six women, blood flow was measured...... in femoral bone at rest and during one leg intermittent isometric exercise with increasing exercise intensities. In nine men, blood flow in femur was determined at rest and during dynamic one leg exercise, and two other physiological perturbations: moderate systemic hypoxia (14 O(2) ) at rest and during...... exercise, and during intra-femoral infusion of high-dose adenosine. Bone glucose uptake was measured at rest and during dynamic one leg exercise in five men. The results indicate that isometric exercise increased femoral bone blood flow from rest (1.8 ± 0.6 ml/100g/min) to low intensity exercise (4.1 ± 1...

  6. Transcriptional correlates of disease outcome in anticoagulant-treated non-human primates infected with ebolavirus.

    Directory of Open Access Journals (Sweden)

    Sara Garamszegi

    Full Text Available Ebola virus (EBOV infection in humans and non-human primates (NHPs is highly lethal, and there is limited understanding of the mechanisms associated with pathogenesis and survival. Here, we describe a transcriptomic analysis of NHPs that survived lethal EBOV infection, compared to NHPs that did not survive. It has been previously demonstrated that anticoagulant therapeutics increase the survival rate in EBOV-infected NHPs, and that the characteristic transcriptional profile of immune response changes in anticoagulant-treated NHPs. In order to identify transcriptional signatures that correlate with survival following EBOV infection, we compared the mRNA expression profile in peripheral blood mononuclear cells from EBOV-infected NHPs that received anticoagulant treatment, to those that did not receive treatment. We identified a small set of 20 genes that are highly confident predictors and can accurately distinguish between surviving and non-surviving animals. In addition, we identified a larger predictive signature of 238 genes that correlated with disease outcome and treatment; this latter signature was associated with a variety of host responses, such as the inflammatory response, T cell death, and inhibition of viral replication. Notably, among survival-associated genes were subsets of genes that are transcriptionally regulated by (1 CCAAT/enhancer-binding protein alpha, (2 tumor protein 53, and (3 megakaryoblastic leukemia 1 and myocardin-like protein 2. These pathways merit further investigation as potential transcriptional signatures of host immune response to EBOV infection.

  7. Haemopoietic progenitor cells in human peripheral blood

    International Nuclear Information System (INIS)

    Zwaan, F.E.

    1980-01-01

    The purpose of the investigation reported is to purify haemopoietic progenitor cells from human peripheral blood using density gradient centrifugation in order to isolate a progenitor cell fraction without immunocompetent cells. The purification technique of peripheral blood flow colony forming unit culture (CFU-c) by means of density gradient centrifugation and a combined depletion of various rosettes is described. The results of several 'in vitro' characteristics of purified CFU-c suspensions and of the plasma clot diffusion chamber culture technique are presented. Irradiation studies revealed that for both human bone marrow and peripheral blood the CFU-c were less radioresistant than clusters. Elimination of monocytes (and granulocytes) from the test suspensions induced an alteration in radiosensitivity pararmeters. The results obtained with the different techniques are described by analysing peripheral progenitor cell activity in myeloproliferative disorders. (Auth.)

  8. Transcriptional profiling of putative human epithelial stem cells

    Directory of Open Access Journals (Sweden)

    Koçer Salih S

    2008-07-01

    Full Text Available Abstract Background Human interfollicular epidermis is sustained by the proliferation of stem cells and their progeny, transient amplifying cells. Molecular characterization of these two cell populations is essential for better understanding of self renewal, differentiation and mechanisms of skin pathogenesis. The purpose of this study was to obtain gene expression profiles of alpha 6+/MHCI+, transient amplifying cells and alpha 6+/MHCI-, putative stem cells, and to compare them with existing data bases of gene expression profiles of hair follicle stem cells. The expression of Major Histocompatibility Complex (MHC class I, previously shown to be absent in stem cells in several tissues, and alpha 6 integrin were used to isolate MHCI positive basal cells, and MHCI low/negative basal cells. Results Transcriptional profiles of the two cell populations were determined and comparisons made with published data for hair follicle stem cell gene expression profiles. We demonstrate that presumptive interfollicular stem cells, alpha 6+/MHCI- cells, are enriched in messenger RNAs encoding surface receptors, cell adhesion molecules, extracellular matrix proteins, transcripts encoding members of IFN-alpha family proteins and components of IFN signaling, but contain lower levels of transcripts encoding proteins which take part in energy metabolism, cell cycle, ribosome biosynthesis, splicing, protein translation, degradation, DNA replication, repair, and chromosome remodeling. Furthermore, our data indicate that the cell signaling pathways Notch1 and NF-κB are downregulated/inhibited in MHC negative basal cells. Conclusion This study demonstrates that alpha 6+/MHCI- cells have additional characteristics attributed to stem cells. Moreover, the transcription profile of alpha 6+/MHCI- cells shows similarities to transcription profiles of mouse hair follicle bulge cells known to be enriched for stem cells. Collectively, our data suggests that alpha 6+/MHCI- cells

  9. Human-Phosphate-Binding-Protein inhibits HIV-1 gene transcription and replication

    Directory of Open Access Journals (Sweden)

    Candolfi Ermanno

    2011-07-01

    Full Text Available Abstract The Human Phosphate-Binding protein (HPBP is a serendipitously discovered lipoprotein that binds phosphate with high affinity. HPBP belongs to the DING protein family, involved in various biological processes like cell cycle regulation. We report that HPBP inhibits HIV-1 gene transcription and replication in T cell line, primary peripherical blood lymphocytes and primary macrophages. We show that HPBP is efficient in naïve and HIV-1 AZT-resistant strains. Our results revealed HPBP as a new and potent anti HIV molecule that inhibits transcription of the virus, which has not yet been targeted by HAART and therefore opens new strategies in the treatment of HIV infection.

  10. Cocoa procyanidins and human cytokine transcription and secretion.

    Science.gov (United States)

    Mao, T; Van De Water, J; Keen, C L; Schmitz, H H; Gershwin, M E

    2000-08-01

    We examined whether cocoa, in its isolated procyanidin fractions (monomer through decamer), would modulate cytokine production at the levels of transcription and protein secretion in both resting and phytohemagglutinin (PHA)-stimulated peripheral blood mononuclear cells (PBMC). In resting cells, interleukin (IL)-1beta and IL-4 gene expression from cocoa-treated cells varied markedly among the subjects tested. However, at the protein level, the larger fractions (pentamer through decamer) stimulated a dramatic increase in IL-1beta concentration (up to ninefold) with increasing degree of polymerization. Similarly, these larger fractions augmented IL-4 concentration by as much as 2 pg/ml, whereas the control displayed levels nearly undetectable. In the presence of PHA, gene expression also seemed to be most affected by the larger procyanidin fractions. The pentameric through decameric fractions increased IL-1 beta expression by 7-19% compared with PHA control, whereas the hexameric through decameric fractions significantly inhibited PHA-induced IL-4 transcription in the range of 71-86%. This observation at the transcription level for IL-1 beta was reflected at the protein level in PHA-stimulated PBMC. Significant reductions in mitogen-induced IL-4 production were also seen at the protein level with the hexamer, heptamer and octamer. Individual oligomeric cocoa fractions were unstimulatory for IL-2 in resting PBMC. However, when induced with PHA, the pentamer, hexamer and heptamer fractions caused a 61-73% inhibition in IL-2 gene expression. This study offers additional data for the consideration of the health benefits of dietary polyphenols from a wide variety of foods, including those benefits associated specifically with cocoa and chocolate consumption.

  11. Lea blood group antigen on human platelets

    International Nuclear Information System (INIS)

    Dunstan, R.A.; Simpson, M.B.; Rosse, W.F.

    1985-01-01

    One- and two-stage radioligand assays were used to determine if human platelets possess the Lea antigen. Goat IgG anti-Lea antibody was purified by multiple adsorptions with Le(a-b-) human red blood cells, followed by affinity chromatography with synthetic Lea substance and labeling with 125 I. Human IgG anti-Lea antibody was used either in a two stage radioassay with 125 I-labeled mouse monoclonal IgG anti-human IgG as the second antibody or, alternatively, purified by Staph protein A chromatography, labeled with 125 I, and used in a one-stage radioassay. Platelets from donors of appropriate red blood cell phenotypes were incubated with the antisera, centrifuged through phthalate esters, and assayed in a gamma scintillation counter. Dose response and saturation curve analysis demonstrate the presence of Lewis a antigen on platelets from Lea+ donors. Furthermore, platelets from an Le(a-b-) donor incubated in Le (a+b-) plasma adsorb Lea antigen in a similar manner to red blood cells. The clinical significance of these antigens in platelet transfusion remains undefined

  12. Mutations and binding sites of human transcription factors

    KAUST Repository

    Kamanu, Frederick Kinyua

    2012-06-01

    Mutations in any genome may lead to phenotype characteristics that determine ability of an individual to cope with adaptation to environmental challenges. In studies of human biology, among the most interesting ones are phenotype characteristics that determine responses to drug treatments, response to infections, or predisposition to specific inherited diseases. Most of the research in this field has been focused on the studies of mutation effects on the final gene products, peptides, and their alterations. Considerably less attention was given to the mutations that may affect regulatory mechanism(s) of gene expression, although these may also affect the phenotype characteristics. In this study we make a pilot analysis of mutations observed in the regulatory regions of 24,667 human RefSeq genes. Our study reveals that out of eight studied mutation types, insertions are the only one that in a statistically significant manner alters predicted transcription factor binding sites (TFBSs). We also find that 25 families of TFBSs have been altered by mutations in a statistically significant manner in the promoter regions we considered. Moreover, we find that the related transcription factors are, for example, prominent in processes related to intracellular signaling; cell fate; morphogenesis of organs and epithelium; development of urogenital system, epithelium, and tube; neuron fate commitment. Our study highlights the significance of studying mutations within the genes regulatory regions and opens way for further detailed investigations on this topic, particularly on the downstream affected pathways. 2012 Kamanu, Medvedeva, Schaefer, Jankovic, Archer and Bajic.

  13. A high parasite density environment induces transcriptional changes and cell death in Plasmodium falciparum blood stages.

    Science.gov (United States)

    Chou, Evelyn S; Abidi, Sabia Z; Teye, Marian; Leliwa-Sytek, Aleksandra; Rask, Thomas S; Cobbold, Simon A; Tonkin-Hill, Gerry Q; Subramaniam, Krishanthi S; Sexton, Anna E; Creek, Darren J; Daily, Johanna P; Duffy, Michael F; Day, Karen P

    2018-03-01

    Transient regulation of Plasmodium numbers below the density that induces fever has been observed in chronic malaria infections in humans. This species transcending control cannot be explained by immunity alone. Using an in vitro system we have observed density dependent regulation of malaria population size as a mechanism to possibly explain these in vivo observations. Specifically, Plasmodium falciparum blood stages from a high but not low-density environment exhibited unique phenotypic changes during the late trophozoite (LT) and schizont stages of the intraerythrocytic cycle. These included in order of appearance: failure of schizonts to mature and merozoites to replicate, apoptotic-like morphological changes including shrinking, loss of mitochondrial membrane potential, and blebbing with eventual release of aberrant parasites from infected erythrocytes. This unique death phenotype was triggered in a stage-specific manner by sensing of a high-density culture environment. Conditions of glucose starvation, nutrient depletion, and high lactate could not induce the phenotype. A high-density culture environment induced rapid global changes in the parasite transcriptome including differential expression of genes involved in cell remodeling, clonal antigenic variation, metabolism, and cell death pathways including an apoptosis-associated metacaspase gene. This transcriptional profile was also characterized by concomitant expression of asexual and sexual stage-specific genes. The data show strong evidence to support our hypothesis that density sensing exists in P. falciparum. They indicate that an apoptotic-like mechanism may play a role in P. falciparum density regulation, which, as in yeast, has features quite distinguishable from mammalian apoptosis. Gene expression data are available in the GEO databases under the accession number GSE91188. © 2017 Federation of European Biochemical Societies.

  14. DNA methylation and transcriptional trajectories during human development and reprogramming of isogenic pluripotent stem cells

    NARCIS (Netherlands)

    Roost, Matthias S; Slieker, Roderick C; Bialecka, Monika; van Iperen, Liesbeth; Gomes Fernandes, Maria M; He, Nannan; Suchiman, H Eka D; Szuhai, Karoly; Carlotti, Françoise; de Koning, Eelco J P; Mummery, Christine L; Heijmans, Bastiaan T; Chuva de Sousa Lopes, Susana M

    2017-01-01

    Determining cell identity and maturation status of differentiated pluripotent stem cells (PSCs) requires knowledge of the transcriptional and epigenetic trajectory of organs during development. Here, we generate a transcriptional and DNA methylation atlas covering 21 organs during human fetal

  15. Whole Blood Transcriptional Profiling of Interferon-Inducible Genes Identifies Highly Upregulated IFI27 in Primary Myelofibrosis

    DEFF Research Database (Denmark)

    Skov, Vibe; Larsen, Thomas Stauffer; Thomassen, Mads

    2011-01-01

    focused upon the transcriptional profiling of interferon-associated genes in patients with essential thrombocythemia (ET) (n = 19), polycythemia vera (PV) (n = 41), and primary myelofibrosis (PMF) (n = 9). Using whole-blood transcriptional profiling and accordingly obtaining an integrated signature...

  16. Whole-blood transcriptional profiling of interferon-inducible genes identifies highly upregulated IFI27 in primary myelofibrosis

    DEFF Research Database (Denmark)

    Skov, Vibe; Larsen, Thomas Stauffer; Thomassen, Mads

    2011-01-01

    focused upon the transcriptional profiling of interferon-associated genes in patients with essential thrombocythemia (ET) (n = 19), polycythemia vera (PV) (n = 41), and primary myelofibrosis (PMF) (n = 9). Using whole-blood transcriptional profiling and accordingly obtaining an integrated signature...

  17. TcoF-DB: dragon database for human transcription co-factors and transcription factor interacting proteins

    KAUST Repository

    Schaefer, Ulf

    2010-10-21

    The initiation and regulation of transcription in eukaryotes is complex and involves a large number of transcription factors (TFs), which are known to bind to the regulatory regions of eukaryotic DNA. Apart from TF-DNA binding, protein-protein interaction involving TFs is an essential component of the machinery facilitating transcriptional regulation. Proteins that interact with TFs in the context of transcription regulation but do not bind to the DNA themselves, we consider transcription co-factors (TcoFs). The influence of TcoFs on transcriptional regulation and initiation, although indirect, has been shown to be significant with the functionality of TFs strongly influenced by the presence of TcoFs. While the role of TFs and their interaction with regulatory DNA regions has been well-studied, the association between TFs and TcoFs has so far been given less attention. Here, we present a resource that is comprised of a collection of human TFs and the TcoFs with which they interact. Other proteins that have a proven interaction with a TF, but are not considered TcoFs are also included. Our database contains 157 high-confidence TcoFs and additionally 379 hypothetical TcoFs. These have been identified and classified according to the type of available evidence for their involvement in transcriptional regulation and their presence in the cell nucleus. We have divided TcoFs into four groups, one of which contains high-confidence TcoFs and three others contain TcoFs which are hypothetical to different extents. We have developed the Dragon Database for Human Transcription Co-Factors and Transcription Factor Interacting Proteins (TcoF-DB). A web-based interface for this resource can be freely accessed at http://cbrc.kaust.edu.sa/tcof/ and http://apps.sanbi.ac.za/tcof/. © The Author(s) 2010.

  18. TcoF-DB: dragon database for human transcription co-factors and transcription factor interacting proteins

    KAUST Repository

    Schaefer, Ulf; Schmeier, Sebastian; Bajic, Vladimir B.

    2010-01-01

    The initiation and regulation of transcription in eukaryotes is complex and involves a large number of transcription factors (TFs), which are known to bind to the regulatory regions of eukaryotic DNA. Apart from TF-DNA binding, protein-protein interaction involving TFs is an essential component of the machinery facilitating transcriptional regulation. Proteins that interact with TFs in the context of transcription regulation but do not bind to the DNA themselves, we consider transcription co-factors (TcoFs). The influence of TcoFs on transcriptional regulation and initiation, although indirect, has been shown to be significant with the functionality of TFs strongly influenced by the presence of TcoFs. While the role of TFs and their interaction with regulatory DNA regions has been well-studied, the association between TFs and TcoFs has so far been given less attention. Here, we present a resource that is comprised of a collection of human TFs and the TcoFs with which they interact. Other proteins that have a proven interaction with a TF, but are not considered TcoFs are also included. Our database contains 157 high-confidence TcoFs and additionally 379 hypothetical TcoFs. These have been identified and classified according to the type of available evidence for their involvement in transcriptional regulation and their presence in the cell nucleus. We have divided TcoFs into four groups, one of which contains high-confidence TcoFs and three others contain TcoFs which are hypothetical to different extents. We have developed the Dragon Database for Human Transcription Co-Factors and Transcription Factor Interacting Proteins (TcoF-DB). A web-based interface for this resource can be freely accessed at http://cbrc.kaust.edu.sa/tcof/ and http://apps.sanbi.ac.za/tcof/. © The Author(s) 2010.

  19. The transcriptional profiling of human in vivo-generated plasma cells identifies selective imbalances in monoclonal gammopathies.

    Directory of Open Access Journals (Sweden)

    Luis M Valor

    Full Text Available Plasma cells (PC represent the heterogeneous final stage of the B cells (BC differentiation process. To characterize the transition of BC into PC, transcriptomes from human naïve BC were compared to those of three functionally-different subsets of human in vivo-generated PC: i tonsil PC, mainly consisting of early PC; ii PC released to the blood after a potent booster-immunization (mostly cycling plasmablasts; and, iii bone marrow CD138+ PC that represent highly mature PC and include the long-lived PC compartment. This transcriptional transition involves subsets of genes related to key processes for PC maturation: the already known protein processing, apoptosis and homeostasis, and of new discovery including histones, macromolecule assembly, zinc-finger transcription factors and neuromodulation. This human PC signature is partially reproduced in vitro and is conserved in mouse. Moreover, the present study identifies genes that define PC subtypes (e.g., proliferation-associated genes for circulating PC and transcriptional-related genes for tonsil and bone marrow PC and proposes some putative transcriptional regulators of the human PC signatures (e.g., OCT/POU, XBP1/CREB, E2F, among others. Finally, we also identified a restricted imbalance of the present PC transcriptional program in monoclonal gammopathies that correlated with PC malignancy.

  20. Gene expression analysis in human breast cancer associated blood vessels.

    Directory of Open Access Journals (Sweden)

    Dylan T Jones

    Full Text Available Angiogenesis is essential for solid tumour growth, whilst the molecular profiles of tumour blood vessels have been reported to be different between cancer types. Although presently available anti-angiogenic strategies are providing some promise for the treatment of some cancers it is perhaps not surprisingly that, none of the anti-angiogenic agents available work on all tumours. Thus, the discovery of novel anti-angiogenic targets, relevant to individual cancer types, is required. Using Affymetrix microarray analysis of laser-captured, CD31-positive blood vessels we have identified 63 genes that are upregulated significantly (5-72 fold in angiogenic blood vessels associated with human invasive ductal carcinoma (IDC of the breast as compared with blood vessels in normal human breast. We tested the angiogenic capacity of a subset of these genes. Genes were selected based on either their known cellular functions, their enriched expression in endothelial cells and/or their sensitivity to anti-VEGF treatment; all features implicating their involvement in angiogenesis. For example, RRM2, a ribonucleotide reductase involved in DNA synthesis, was upregulated 32-fold in IDC-associated blood vessels; ATF1, a nuclear activating transcription factor involved in cellular growth and survival was upregulated 23-fold in IDC-associated blood vessels and HEX-B, a hexosaminidase involved in the breakdown of GM2 gangliosides, was upregulated 8-fold in IDC-associated blood vessels. Furthermore, in silico analysis confirmed that AFT1 and HEX-B also were enriched in endothelial cells when compared with non-endothelial cells. None of these genes have been reported previously to be involved in neovascularisation. However, our data establish that siRNA depletion of Rrm2, Atf1 or Hex-B had significant anti-angiogenic effects in VEGF-stimulated ex vivo mouse aortic ring assays. Overall, our results provide proof-of-principle that our approach can identify a cohort of

  1. Regulation of bone blood flow in humans

    DEFF Research Database (Denmark)

    Heinonen, Ilkka; Boushel, Robert; Hellsten, Ylva

    2018-01-01

    of cyclooxygenase (COX) enzyme, thus prostaglandin (PG) synthesis on femoral bone marrow blood flow by positron emission tomography in healthy young men at rest and during one leg dynamic exercise. In an additional group of healthy men, the role of adenosine (ADO) in the regulation of BBF during exercise......The mechanisms that regulate bone blood flow (BBF) in humans are largely unknown. Animal studies suggest that nitric oxide (NO) could be involved and in the present study we investigated the effects of inhibition of nitric oxide synthase (NOS) alone and in combination with inhibition.......036), but did not affect BBF significantly during exercise (5.5±1.4 ml/100g/min, p=0.25). On the other hand, while combined NOS and COX inhibition did not cause any further reduction of blood flow at rest (0.6±0.2 ml/100g/min), the combined blockade reduced BBF during exercise by ~21%, to 5.0±1.8 ml/100g/min (p...

  2. Computational Analysis of Human Blood Flow

    Science.gov (United States)

    Panta, Yogendra; Marie, Hazel; Harvey, Mark

    2009-11-01

    Fluid flow modeling with commercially available computational fluid dynamics (CFD) software is widely used to visualize and predict physical phenomena related to various biological systems. In this presentation, a typical human aorta model was analyzed assuming the blood flow as laminar with complaint cardiac muscle wall boundaries. FLUENT, a commercially available finite volume software, coupled with Solidworks, a modeling software, was employed for the preprocessing, simulation and postprocessing of all the models.The analysis mainly consists of a fluid-dynamics analysis including a calculation of the velocity field and pressure distribution in the blood and a mechanical analysis of the deformation of the tissue and artery in terms of wall shear stress. A number of other models e.g. T branches, angle shaped were previously analyzed and compared their results for consistency for similar boundary conditions. The velocities, pressures and wall shear stress distributions achieved in all models were as expected given the similar boundary conditions. The three dimensional time dependent analysis of blood flow accounting the effect of body forces with a complaint boundary was also performed.

  3. Loneliness, eudaimonia, and the human conserved transcriptional response to adversity.

    Science.gov (United States)

    Cole, Steven W; Levine, Morgan E; Arevalo, Jesusa M G; Ma, Jeffrey; Weir, David R; Crimmins, Eileen M

    2015-12-01

    Chronic social adversity activates a conserved transcriptional response to adversity (CTRA) marked by increased expression of pro-inflammatory genes and decreased expression of antiviral- and antibody-related genes. Recent findings suggest that some psychological resilience factors may help buffer CTRA activation, but the relative impact of resilience and adversity factors remains poorly understood. Here we examined the relative strength of CTRA association for the two best-established psychological correlates of CTRA gene expression-the risk factor of perceived social isolation (loneliness) and the resilience factor of eudaimonic well-being (purpose and meaning in life). Peripheral blood samples and validated measures of loneliness and eudaimonic well-being were analyzed in 108 community-dwelling older adults participating in the longitudinal US Health and Retirement Study (56% female, mean age 73). Mixed effect linear model analyses quantified the strength of association between CTRA gene expression and measures of loneliness and eudaimonic well-being in separate and joint analyses. As in previous studies, separate analyses found CTRA gene expression to be up-regulated in association with loneliness and down-regulated in association with eudaimonic well-being. In joint analyses, effects of loneliness were completely abrogated whereas eudaimonic well-being continued to associate with CTRA down-regulation. Similar eudaimonia-dominant effects were observed for positive and negative affect, optimism and pessimism, and anxiety symptoms. All results were independent of demographic and behavioral health risk factors. Eudaimonic well-being may have the potential to compensate for the adverse impact of loneliness on CTRA gene expression. Findings suggest a novel approach to targeting the health risks associated with social isolation by promoting purpose and meaning in life. Copyright © 2015 Elsevier Ltd. All rights reserved.

  4. Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

    Science.gov (United States)

    In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A; Rubin, P; Kemp, J; Israel, E; Busse, W; Ledford, D; Murray, J J; Segal, A; Tinkleman, D; Drazen, J M

    1997-03-01

    Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, deletion of two, or addition of one zinc finger (Sp1/Egr-1) binding sites in the region 176 to 147 bp upstream from the ATG translation start site where there are normally 5 Sp1 binding motifs in tandem. Reporter gene activity directed by any of the mutant forms of the transcription factor binding region was significantly (P < 0.05) less effective than the activity driven by the wild type transcription factor binding region. Electrophoretic mobility shift assays (EMSAs) demonstrated the capacity of wild type and mutant transcription factor binding regions to bind nuclear extracts from human umbilical vein endothelial cells (HUVECs). These data are consistent with a family of mutations in the 5-LO gene that can modify reporter gene transcription possibly through differences in Sp1 and Egr-1 transactivation.

  5. Intergenic and repeat transcription in human, chimpanzee and macaque brains measured by RNA-Seq.

    Directory of Open Access Journals (Sweden)

    Augix Guohua Xu

    Full Text Available Transcription is the first step connecting genetic information with an organism's phenotype. While expression of annotated genes in the human brain has been characterized extensively, our knowledge about the scope and the conservation of transcripts located outside of the known genes' boundaries is limited. Here, we use high-throughput transcriptome sequencing (RNA-Seq to characterize the total non-ribosomal transcriptome of human, chimpanzee, and rhesus macaque brain. In all species, only 20-28% of non-ribosomal transcripts correspond to annotated exons and 20-23% to introns. By contrast, transcripts originating within intronic and intergenic repetitive sequences constitute 40-48% of the total brain transcriptome. Notably, some repeat families show elevated transcription. In non-repetitive intergenic regions, we identify and characterize 1,093 distinct regions highly expressed in the human brain. These regions are conserved at the RNA expression level across primates studied and at the DNA sequence level across mammals. A large proportion of these transcripts (20% represents 3'UTR extensions of known genes and may play roles in alternative microRNA-directed regulation. Finally, we show that while transcriptome divergence between species increases with evolutionary time, intergenic transcripts show more expression differences among species and exons show less. Our results show that many yet uncharacterized evolutionary conserved transcripts exist in the human brain. Some of these transcripts may play roles in transcriptional regulation and contribute to evolution of human-specific phenotypic traits.

  6. Accurate Gene Expression-Based Biodosimetry Using a Minimal Set of Human Gene Transcripts

    Energy Technology Data Exchange (ETDEWEB)

    Tucker, James D., E-mail: jtucker@biology.biosci.wayne.edu [Department of Biological Sciences, Wayne State University, Detroit, Michigan (United States); Joiner, Michael C. [Department of Radiation Oncology, Wayne State University, Detroit, Michigan (United States); Thomas, Robert A.; Grever, William E.; Bakhmutsky, Marina V. [Department of Biological Sciences, Wayne State University, Detroit, Michigan (United States); Chinkhota, Chantelle N.; Smolinski, Joseph M. [Department of Electrical and Computer Engineering, Wayne State University, Detroit, Michigan (United States); Divine, George W. [Department of Public Health Sciences, Henry Ford Hospital, Detroit, Michigan (United States); Auner, Gregory W. [Department of Electrical and Computer Engineering, Wayne State University, Detroit, Michigan (United States)

    2014-03-15

    Purpose: Rapid and reliable methods for conducting biological dosimetry are a necessity in the event of a large-scale nuclear event. Conventional biodosimetry methods lack the speed, portability, ease of use, and low cost required for triaging numerous victims. Here we address this need by showing that polymerase chain reaction (PCR) on a small number of gene transcripts can provide accurate and rapid dosimetry. The low cost and relative ease of PCR compared with existing dosimetry methods suggest that this approach may be useful in mass-casualty triage situations. Methods and Materials: Human peripheral blood from 60 adult donors was acutely exposed to cobalt-60 gamma rays at doses of 0 (control) to 10 Gy. mRNA expression levels of 121 selected genes were obtained 0.5, 1, and 2 days after exposure by reverse-transcriptase real-time PCR. Optimal dosimetry at each time point was obtained by stepwise regression of dose received against individual gene transcript expression levels. Results: Only 3 to 4 different gene transcripts, ASTN2, CDKN1A, GDF15, and ATM, are needed to explain ≥0.87 of the variance (R{sup 2}). Receiver-operator characteristics, a measure of sensitivity and specificity, of 0.98 for these statistical models were achieved at each time point. Conclusions: The actual and predicted radiation doses agree very closely up to 6 Gy. Dosimetry at 8 and 10 Gy shows some effect of saturation, thereby slightly diminishing the ability to quantify higher exposures. Analyses of these gene transcripts may be advantageous for use in a field-portable device designed to assess exposures in mass casualty situations or in clinical radiation emergencies.

  7. Distribution of the DAZ gene transcripts in human testis.

    Directory of Open Access Journals (Sweden)

    J B Warchoł

    2004-07-01

    Full Text Available Involvement of variety of genes, especially located on Y chromosome, is critical for the regulation of spermatogenesis. In particular, fertility candidate genes such as deleted in azoospermia (DAZ are believed to have important function in sperm production, since DAZ is frequently deleted in azoospermic and severy oligozoospermic men. The role of the DAZ gene is supported by its exclusive expression in the testis and by its deletion in about 10% of azoospermic and severely oligozoospermic patients. The distribution of DAZ transcripts in seminiferous epithelium of human testis is reported in the present study. The use of Adobe Photoshop and Scion Image softwares allowed for semi-quantitative analysis of in situ RT-PCR (ISRT-PCR results. The intensity of ISRT-PCR product's fluorescence was different within individual seminiferous tubules. It was clearly shown by using the pseudocolour scale and transforming the intensity of the fluorescence into levels of greyscale images. The more intense fluorescence characterised single spermatogonia and those organized in small groups inside separate tubules. The most intense accumulation of DAZ mRNA was observed in spermatogonia.

  8. Insight of Human Stroke from blood flow and blood pressure

    Science.gov (United States)

    Chen, Zhi; Ivanov, Plamen Ch.; Hu, Kun; Stanley, H. Eugene

    2003-03-01

    Stroke is is one of the leading cause of death and disability in the world. It is well believed that stroke is caused by the disturbance of cerebrovascular autoregulation. We investigate the blood flow on the left and right middle cerebral artery and beat-to-beat blood pressure simultaneously measured from the finger, for both subjects with stroke and healthy subjects. Synchronization technique is used to distinguish the difference between these two groups.

  9. Distinct Transcriptional and Alternative Splicing Signatures of Decidual CD4+ T Cells in Early Human Pregnancy

    Directory of Open Access Journals (Sweden)

    Weihong Zeng

    2017-06-01

    Full Text Available Decidual CD4+ T (dCD4 T cells are crucial for the maternal-fetal immune tolerance required for a healthy pregnancy outcome. However, their molecular and functional characteristics are not well elucidated. In this study, we performed the first analysis of transcriptional and alternative splicing (AS landscapes for paired decidual and peripheral blood CD4+ T (pCD4 T cells in human early pregnancy using high throughput mRNA sequencing. Our data showed that dCD4 T cells are endowed with a unique transcriptional signature when compared to pCD4 T cells: dCD4 T cells upregulate 1,695 genes enriched in immune system process whereas downregulate 1,011 genes mainly related to mRNA catabolic process and the ribosome. Moreover, dCD4 T cells were observed to be at M phase, and show increased activation, proliferation, and cytokine production, as well as display an effector-memory phenotype and a heterogenous nature containing Th1, Th17, and Treg cell subsets. However, dCD4 T cells undergo a comparable number of upregulated and downregulated AS events, both of which are enriched in the genes related to cellular metabolic process. And the changes at the AS event level do not reflect measurable differences at the gene expression level in dCD4 T cells. Collectively, our findings provide a comprehensive portrait of the unique transcriptional signature and AS profile of CD4+ T cells in human decidua and help us gain more understanding of the functional characteristic of these cells during early pregnancy.

  10. An integrative 'omics' solution to the detection of recombinant human erythropoietin and blood doping.

    Science.gov (United States)

    Pitsiladis, Yannis P; Durussel, Jérôme; Rabin, Olivier

    2014-05-01

    Administration of recombinant human erythropoietin (rHumanEPO) improves sporting performance and hence is frequently subject to abuse by athletes, although rHumanEPO is prohibited by the WADA. Approaches to detect rHumanEPO doping have improved significantly in recent years but remain imperfect. A new transcriptomic-based longitudinal screening approach is being developed that has the potential to improve the analytical performance of current detection methods. In particular, studies are being funded by WADA to identify a 'molecular signature' of rHumanEPO doping and preliminary results are promising. In the first systematic study to be conducted, the expression of hundreds of genes were found to be altered by rHumanEPO with numerous gene transcripts being differentially expressed after the first injection and further transcripts profoundly upregulated during and subsequently downregulated up to 4 weeks postadministration of the drug; with the same transcriptomic pattern observed in all participants. The identification of a blood 'molecular signature' of rHumanEPO administration is the strongest evidence to date that gene biomarkers have the potential to substantially improve the analytical performance of current antidoping methods such as the Athlete Biological Passport for rHumanEPO detection. Given the early promise of transcriptomics, research using an 'omics'-based approach involving genomics, transcriptomics, proteomics and metabolomics should be intensified in order to achieve improved detection of rHumanEPO and other doping substances and methods difficult to detect such a recombinant human growth hormone and blood transfusions.

  11. Circulating Human Eosinophils Share a Similar Transcriptional Profile in Asthma and Other Hypereosinophilic Disorders.

    Science.gov (United States)

    Barnig, Cindy; Alsaleh, Ghada; Jung, Nicolas; Dembélé, Doulaye; Paul, Nicodème; Poirot, Anh; Uring-Lambert, Béatrice; Georgel, Philippe; de Blay, Fréderic; Bahram, Seiamak

    2015-01-01

    Eosinophils are leukocytes that are released into the peripheral blood in a phenotypically mature state and are capable of being recruited into tissues in response to appropriate stimuli. Eosinophils, traditionally considered cytotoxic effector cells, are leukocytes recruited into the airways of asthma patients where they are believed to contribute to the development of many features of the disease. This perception, however, has been challenged by recent findings suggesting that eosinophils have also immunomodulatory functions and may be involved in tissue homeostasis and wound healing. Here we describe a transcriptome-based approach-in a limited number of patients and controls-to investigate the activation state of circulating human eosinophils isolated by flow cytometry. We provide an overview of the global expression pattern in eosinophils in various relevant conditions, e.g., eosinophilic asthma, hypereosinophilic dermatological diseases, parasitosis and pulmonary aspergillosis. Compared to healthy subjects, circulating eosinophils isolated from asthma patients differed in their gene expression profile which is marked by downregulation of transcripts involved in antigen presentation, pathogen recognition and mucosal innate immunity, whereas up-regulated genes were involved in response to non-specific stimulation, wounding and maintenance of homeostasis. Eosinophils from other hypereosinophilic disorders displayed a very similar transcriptional profile. Taken together, these observations seem to indicate that eosinophils exhibit non-specific immunomodulatory functions important for tissue repair and homeostasis and suggest new roles for these cells in asthma immunobiology.

  12. The effects of Ankaferd® Blood Stopper on transcription factors in HUVEC and the erythrocyte protein profile

    Directory of Open Access Journals (Sweden)

    Erkan Yılmaz

    2011-12-01

    Full Text Available Objective: Ankaferd® Blood Stopper (ABS is an herbal extract that has historically been used as a hemostatic agent in traditional Turkish medicine. ABS is comprised of a standardized herbal mixture of T. vulgaris, G. glabra, V. vinifera, A. officinarum, and U. dioica. ABS’s basic mechanism of action is the formation of an encapsulated protein web, which represents the focal point for vital erythrocyte masses. The hemostatic effects of ABS have been observed in vitro and in vivo. ABS was registered as a hemostatic agent for external hemorrhages and dental bleeding following phase I randomized, double-blind crossover placebo-controlled clinical research, and safety and efficacy reports. In terms of the potential use of ABS, transcription factors may be novel factors that play a role in the hemostatic and other pleiotropic effects of ABS. Materials and Methods: Hence, the present study aimed to investigate the effects of ABS on endothelium, and possible transcription factor changes in HUVEC (human umbilical vein endothelial cells and the erythrocyte membrane profile. ABS (5 μL and 50 μL was administered to HUVEC (in 75 cm2; ~75% fullness for 5 min and 15 min. Results: ABS caused significant increases in the level of activation of the following transcription factors; AP2, AR, CRE/ATF1, CREB, E2F1-5, E2F6, EGR, GATA, HNF-1, ISRE, Myc-Max, NF-1, NFkB, p53, PPAR, SMAD 2/3, SP1, TRE/AP1, and YY1. Following erythrocyte membrane isolation, protein complexes were undissolved, but denatured. The protein complex formed was resistant to heat and detergent. Trypsin and sonication were used in order to break this complex; the complex dissolved and erythrocyte membrane proteins were released in SDS-PAGE.Conclusion: ABS established a very fast and solid protein web, and increased the level of transcription factor activation. Therefore the cellular effects of ABS could be related to different intracellular biological pathways.

  13. Geographical, environmental and pathophysiological influences on the human blood transcriptome.

    Science.gov (United States)

    Tabassum, Rubina; Nath, Artika; Preininger, Marcela; Gibson, Greg

    2013-12-01

    Gene expression variation provides a read-out of both genetic and environmental influences on gene activity. Geographical, genomic and sociogenomic studies have highlighted how life circumstances of an individual modify the expression of hundreds and in some cases thousands of genes in a co-ordinated manner. This review places such results in the context of a conserved set of 90 transcripts known as Blood Informative Transcripts (BIT) that capture the major conserved components of variation in the peripheral blood transcriptome. Pathophysiological states are also shown to associate with the perturbation of transcript abundance along the major axes. Discussion of false negative rates leads us to argue that simple significance thresholds provide a biased perspective on assessment of differential expression that may cloud the interpretation of studies with small sample sizes.

  14. Expression profiling feline peripheral blood monocytes identifies a transcriptional signature associated with type two diabetes mellitus.

    Science.gov (United States)

    O'Leary, Caroline A; Sedhom, Mamdouh; Reeve-Johnson, Mia; Mallyon, John; Irvine, Katharine M

    2017-04-01

    Diabetes mellitus is a common disease of cats and is similar to type 2 diabetes (T2D) in humans, especially with respect to the role of obesity-induced insulin resistance, glucose toxicity, decreased number of pancreatic β-cells and pancreatic amyloid deposition. Cats have thus been proposed as a valuable translational model of T2D. In humans, inflammation associated with adipose tissue is believed to be central to T2D development, and peripheral blood monocytes (PBM) are important in the inflammatory cascade which leads to insulin resistance and β-cell failure. PBM may thus provide a useful window to study the pathogenesis of diabetes mellitus in cats, however feline monocytes are poorly characterised. In this study, we used the Affymetrix Feline 1.0ST array to profile peripheral blood monocytes from 3 domestic cats with T2D and 3 cats with normal glucose tolerance. Feline monocytes were enriched for genes expressed in human monocytes, and, despite heterogeneous gene expression, we identified a T2D-associated expression signature associated with cell cycle perturbations, DNA repair and the unfolded protein response, oxidative phosphorylation and inflammatory responses. Our data provide novel insights into the feline monocyte transcriptome, and support the hypothesis that inflammatory monocytes contribute to T2D pathogenesis in cats as well as in humans. Copyright © 2017 Elsevier B.V. All rights reserved.

  15. Whole genome transcript profiling from fingerstick blood samples: a comparison and feasibility study

    Directory of Open Access Journals (Sweden)

    Williams Adam R

    2009-12-01

    Full Text Available Abstract Background Whole genome gene expression profiling has revolutionized research in the past decade especially with the advent of microarrays. Recently, there have been significant improvements in whole blood RNA isolation techniques which, through stabilization of RNA at the time of sample collection, avoid bias and artifacts introduced during sample handling. Despite these improvements, current human whole blood RNA stabilization/isolation kits are limited by the requirement of a venous blood sample of at least 2.5 mL. While fingerstick blood collection has been used for many different assays, there has yet to be a kit developed to isolate high quality RNA for use in gene expression studies from such small human samples. The clinical and field testing advantages of obtaining reliable and reproducible gene expression data from a fingerstick are many; it is less invasive, time saving, more mobile, and eliminates the need of a trained phlebotomist. Furthermore, this method could also be employed in small animal studies, i.e. mice, where larger sample collections often require sacrificing the animal. In this study, we offer a rapid and simple method to extract sufficient amounts of high quality total RNA from approximately 70 μl of whole blood collected via a fingerstick using a modified protocol of the commercially available Qiagen PAXgene RNA Blood Kit. Results From two sets of fingerstick collections, about 70 uL whole blood collected via finger lancet and capillary tube, we recovered an average of 252.6 ng total RNA with an average RIN of 9.3. The post-amplification yields for 50 ng of total RNA averaged at 7.0 ug cDNA. The cDNA hybridized to Affymetrix HG-U133 Plus 2.0 GeneChips had an average % Present call of 52.5%. Both fingerstick collections were highly correlated with r2 values ranging from 0.94 to 0.97. Similarly both fingerstick collections were highly correlated to the venous collection with r2 values ranging from 0.88 to 0

  16. Characterization of transcription factor networks involved in umbilical cord blood CD34+ stem cells-derived erythropoiesis.

    Directory of Open Access Journals (Sweden)

    Biaoru Li

    Full Text Available Fetal stem cells isolated from umbilical cord blood (UCB possess a great capacity for proliferation and differentiation and serve as a valuable model system to study gene regulation. Expanded knowledge of the molecular control of hemoglobin synthesis will provide a basis for rational design of therapies for β-hemoglobinopathies. Transcriptome data are available for erythroid progenitors derived from adult stem cells, however studies to define molecular mechanisms controlling globin gene regulation during fetal erythropoiesis are limited. Here, we utilize UCB-CD34+ stem cells induced to undergo erythroid differentiation to characterize the transcriptome and transcription factor networks (TFNs associated with the γ/β-globin switch during fetal erythropoiesis. UCB-CD34+ stem cells grown in the one-phase liquid culture system displayed a higher proliferative capacity than adult CD34+ stem cells. The γ/β-globin switch was observed after day 42 during fetal erythropoiesis in contrast to adult progenitors where the switch occurred around day 21. To gain insights into transcription factors involved in globin gene regulation, microarray analysis was performed on RNA isolated from UCB-CD34+ cell-derived erythroid progenitors harvested on day 21, 42, 49 and 56 using the HumanHT-12 Expression BeadChip. After data normalization, Gene Set Enrichment Analysis identified transcription factors (TFs with significant changes in expression during the γ/β-globin switch. Forty-five TFs were silenced by day 56 (Profile-1 and 30 TFs were activated by day 56 (Profile-2. Both GSEA datasets were analyzed using the MIMI Cytoscape platform, which discovered TFNs centered on KLF4 and GATA2 (Profile-1 and KLF1 and GATA1 for Profile-2 genes. Subsequent shRNA studies in KU812 leukemia cells and human erythroid progenitors generated from UCB-CD34+ cells supported a negative role of MAFB in γ-globin regulation. The characteristics of erythroblasts derived from UCB-CD34

  17. Human blood serum analysis using TRXRF

    International Nuclear Information System (INIS)

    Zarkadas, C.; Karydas, A.G.; Paradellis, T.

    2000-01-01

    Total reflection x-ray fluorescence was applied in the analysis of a pool human blood serum sample, which was collected out of 100 healthy individuals during an ordinary day at a hospital in Athens. Direct measurements of 4 1 quantities were performed in a standard TRXRF module, but with the addition of a Mo filter after the cut-off reflector. In this way the exciting beam was further monochromatized leading to an improved peak to background ratio. The elements S, Cl, K, Ca, Fe, Cu, Zn, Se, Br, Rb were detected, with detection limits in the low ppb region for the elements of interest. The determined trace elements concentrations were found to be in very good agreement with values already reported in literature. For intercomparison a quantity of the same sample was freeze dried and measured in a secondary target assembly, in the form of pellets, giving almost identical results. The trace elements concentrations obtained can be considered as representative values for healthy population of Athens and therefore can be used as a monitor in order to associate the lack or excess of certain trace elements with pathological cases. (author)

  18. A transcript finishing initiative for closing gaps in the human transcriptome

    DEFF Research Database (Denmark)

    Sogayar, Mari Cleide; Camargo, Anamaria A; Bettoni, Fabiana

    2004-01-01

    We report the results of a transcript finishing initiative, undertaken for the purpose of identifying and characterizing novel human transcripts, in which RT-PCR was used to bridge gaps between paired EST clusters, mapped against the genomic sequence. Each pair of EST clusters selected...

  19. Genomic structure and cloning of two transcript isoforms of human Sp8.

    NARCIS (Netherlands)

    M.A. Milona (Maria-athina); J.E. Gough (Julie); A.J. Edgar (Alasdair)

    2004-01-01

    textabstractBACKGROUND: The Specificity proteins (Sp) are a family of transcription factors that have three highly conserved zinc-fingers located towards the carboxy-terminal that bind GC-boxes and assist in the initiation of gene transcription. Human Sp1-7 genes have been

  20. Isoforms of purified methyltransferase from human blood platelets ...

    African Journals Online (AJOL)

    ... purification from normal human blood platelets have not been investigated, hence, the aim of this study was to purify, characterise the enzyme from human blood platelets and determine its possible role in phospholipid transmethylation. The plasma membranes were purified by velocity and sucrose gradient centrifugation ...

  1. Prevalence of malaria and human blood factors among patients in ...

    African Journals Online (AJOL)

    Background: Malaria has been and is still a major protozoan disease affecting the human population. Erythrocyte polymorphisms (mainly in blood groups and genotypes) influence the susceptibility to severe malaria. Aim: This study is aimed at assessing the prevalence malaria in relation to human blood factor and to ...

  2. Regulation of human protein S gene (PROS1) transcription

    NARCIS (Netherlands)

    Wolf, Cornelia de

    2006-01-01

    This thesis describes the investigation of the transcriptional regulation of the gene for anticoagulant plasma Protein S, PROS1. Protein S is a cofactor for Protein C in the Protein C anticoagulant pathway. The coagulation cascade is negatively regulated by this pathway through inactivation of

  3. Uncovering layers of human RNA polymerase II transcription

    DEFF Research Database (Denmark)

    Jensen, Torben Heick

    In recent years DNA microarray and high-throughput sequencing technologies have challenged the “gene-centric” view that pre-mRNA is the only RNA species transcribed off protein-coding genes. Instead unorthodox transcription from within genic- and intergenic regions has been demonstrated to occur...

  4. Characteristics of functional enrichment and gene expression level of human putative transcriptional target genes.

    Science.gov (United States)

    Osato, Naoki

    2018-01-19

    Transcriptional target genes show functional enrichment of genes. However, how many and how significantly transcriptional target genes include functional enrichments are still unclear. To address these issues, I predicted human transcriptional target genes using open chromatin regions, ChIP-seq data and DNA binding sequences of transcription factors in databases, and examined functional enrichment and gene expression level of putative transcriptional target genes. Gene Ontology annotations showed four times larger numbers of functional enrichments in putative transcriptional target genes than gene expression information alone, independent of transcriptional target genes. To compare the number of functional enrichments of putative transcriptional target genes between cells or search conditions, I normalized the number of functional enrichment by calculating its ratios in the total number of transcriptional target genes. With this analysis, native putative transcriptional target genes showed the largest normalized number of functional enrichments, compared with target genes including 5-60% of randomly selected genes. The normalized number of functional enrichments was changed according to the criteria of enhancer-promoter interactions such as distance from transcriptional start sites and orientation of CTCF-binding sites. Forward-reverse orientation of CTCF-binding sites showed significantly higher normalized number of functional enrichments than the other orientations. Journal papers showed that the top five frequent functional enrichments were related to the cellular functions in the three cell types. The median expression level of transcriptional target genes changed according to the criteria of enhancer-promoter assignments (i.e. interactions) and was correlated with the changes of the normalized number of functional enrichments of transcriptional target genes. Human putative transcriptional target genes showed significant functional enrichments. Functional

  5. Data in support of transcriptional regulation and function of Fas-antisense long noncoding RNA during human erythropoiesis

    Directory of Open Access Journals (Sweden)

    Olga Villamizar

    2016-06-01

    Full Text Available This paper describes data related to a research article titled, “Fas-antisense long noncoding RNA is differentially expressed during maturation of human erythrocytes and confers resistance to Fas-mediated cell death” [1]. Long noncoding RNAs (lncRNAs are increasingly appreciated for their capacity to regulate many steps of gene expression. While recent studies suggest that many lncRNAs are functional, the scope of their actions throughout human biology is largely undefined including human red blood cell development (erythropoiesis. Here we include expression data for 82 lncRNAs during early, intermediate and late stages of human erythropoiesis using a commercial qPCR Array. From these data, we identified lncRNA Fas-antisense 1 (Fas-AS1 or Saf described in the research article. Also included are 5′ untranslated sequences (UTR for lncRNA Saf with transcription factor target sequences identified. Quantitative RT-PCR data demonstrate relative levels of critical erythroid transcription factors, GATA-1 and KLF1, in K562 human erythroleukemia cells and maturing erythroblasts derived from human CD34+ cells. End point and quantitative RT-PCR data for cDNA prepared using random hexamers versus oligo(dT18 revealed that lncRNA Saf is not effectively polyadenylated. Finally, we include flow cytometry histograms demonstrating Fas levels on maturing erythroblasts derived from human CD34+ cells transduced using mock conditions or with lentivirus particles encoding for Saf.

  6. Characterization of DNA binding, transcriptional activation, and regulated nuclear association of recombinant human NFATp

    Directory of Open Access Journals (Sweden)

    Seto Anita G

    2000-11-01

    Full Text Available Abstract Background NFATp is one member of a family of transcriptional activators whose nuclear accumulation and hence transcriptional activity is regulated in mammalian cells. Human NFATp exists as a phosphoprotein in the cytoplasm of naive T cells. Upon antigen stimulation, NFATp is dephosphorylated, accumulates in nuclei, and functions to regulate transcription of genes including those encoding cytokines. While the properties of the DNA binding domain of NFATp have been investigated in detail, biochemical studies of the transcriptional activation and regulated association with nuclei have remained unexplored because of a lack of full length, purified recombinant NFATp. Results We developed methods for expressing and purifying full length recombinant human NFATp that has all of the properties known to be associated with native NFATp. The recombinant NFATp binds DNA on its own and cooperatively with AP-1 proteins, activates transcription in vitro, is phosphorylated, can be dephosphorylated by calcineurin, and exhibits regulated association with nuclei in vitro. Importantly, activation by recombinant NFATp in a reconstituted transcription system required regions of the protein outside of the central DNA binding domain. Conclusions We conclude that NFATp is a bona fide transcriptional activator. Moreover, the reagents and methods that we developed will facilitate future studies on the mechanisms of transcriptional activation and nuclear accumulation by NFATp, a member of an important family of transcriptional regulatory proteins.

  7. Effects of butyltin exposures on MAP kinase dependent transcription regulators in human natural killer cells

    Science.gov (United States)

    Person, Rachel J.; Whalen, Margaret M.

    2010-01-01

    Natural Killer (NK) cells are a major immune defense mechanism against cancer development and viral infection. The butyltins (BTs), tributyltin (TBT) and dibutyltin (DBT) have been widely used in industrial and other applications and significantly contaminate the environment. Both TBT and DBT have been detected in human blood. These compounds inhibit the lytic and binding function of human NK cells and thus could increase the incidence of cancer and viral infections. Butyltin (BT)-induced loss of NK function is accompanied by activation of mitogen activated protein kinases (MAPKs) and decreases in expression of cell-surface and cytolytic proteins. MAPKs activate components of the transcription regulator AP-1 and activate the transcription regulator Elk-1. Based on the fact that BTs activate MAPKs and alter protein expression, the current study examined the effect of BT exposures on the levels and phosphorylation states of the components of AP-1 and the phosphorylation state of Elk-1. Exposure to 300 nM TBT for 10 min increased the phosphorylation of c-Jun in NK cells. 1 h exposures to 300 nM and 200 nM TBT increased the phosphorylation and overall level of c-Jun. During a 300 nM treatment with TBT for 1 h the binding activity of AP-1 was significantly decreased. There were no significant alterations of AP-1 components or of Elk-1 with DBT exposures. Thus, it appears that TBT-induced alterations on phosphorylation, total levels and binding activity of c-Jun might contribute to, but are not fully responsible for, TBT-induced alterations of NK protein expression. PMID:20370538

  8. A Role for the NF-kb/Rel Transcription Factors in Human Breast Cancer

    National Research Council Canada - National Science Library

    Baldwin, Albert

    1998-01-01

    Human breast cancer is characterized by the inappropriate expression of growth factors, kinases and possibly certain transcription factors Our project has focused on the regulation of the NF-kB family...

  9. E6-associated transcription patterns in human papilloma virus 16-positive cervical tissues.

    Science.gov (United States)

    Lin, Kezhi; Lu, Xulian; Chen, Jun; Zou, Ruanmin; Zhang, Lifang; Xue, Xiangyang

    2015-01-01

    The change in transcription pattern induced by post-transcriptional RNA splicing is an important mechanism in the regulation of the early gene expression of human papilloma virus (HPV). The present study was conducted to establish a method to specifically amplify HPV-16 E6-associated transcripts. The E6-related transcripts from 63 HPV-16-positive cervical tumor tissue samples were amplified, consisting of eight cases of low-risk intraepithelial lesions, 38 cases of high-risk intraepithelial lesions and 17 cases of cervical cancer (CxCa). The appropriate amplified segments were recovered following agarose gel electrophoresis, and subjected to further sequencing and sequence alignment analysis. Six groups of E6 transcription patterns were identified from HPV-16-positive cervical tumor tissue, including five newly-discovered transcripts. Different HPV-16 E6-associated transcription patterns were detected during the development of CxCa. Over the course of the progression of the low-grade squamous intraepithelial lesions to CxCa, the specific HPV-16 E6-associated transcription patterns and the dominant transcripts were all different. As indicated by this study, the transcription pattern of the E6 early gene of HPV-16 was closely associated with the stages of cervical carcinogenesis, and may also be involved in the development of CxCa.

  10. Numerical simulation of human blood flow in microvessels

    International Nuclear Information System (INIS)

    Attaullah; Chughtai, I.R.; Nadeem, M.

    2009-01-01

    In this research, steady state flow of human blood in vascular system has been studied. Computational fluid dynamics has been used to predict pressure drop in human arteriole, artery, capillary, venule and vein. Viscosity of human blood has been treated in different ways by employing Newtonian, Power law and Herschel-Bulkley models. It has been observed that the Herschel-Bulkley model predicts the pressure gradients in all diameters reasonably whereas Newtonian and Power laws have their limitations. (author)

  11. Tracking blood vessels in human forearms using visual servoing

    DEFF Research Database (Denmark)

    Savarimuthu, Thiusius Rajeeth; Ellekilde, Lars-Peter; Hansen, Morten

    compensation. By using images taken with near-infrared light to locate the blood vessels in a human forearm and using the same images to detects movements of the arm, this paper shows that it is possible make a robot arm, potentially equipped with a needle for drawing the blood, compensate for the movements......Drawing an average of more than 2 blood sample per Danish citizen per year increases the demand for an automatic blood sampling method. This paper presents a proof of concept to one of the main challenges in making a fully automated blood sampling procedure, namely: the patient movement...

  12. New insight into transcription of human endogenous retroviral elements

    Czech Academy of Sciences Publication Activity Database

    Pačes, Jan; Huang, Y.T.; Pačes, Václav; Rídl, Jakub; Chang, C.M.

    2013-01-01

    Roč. 30, č. 3 (2013), s. 314-318 ISSN 1871-6784 R&D Projects: GA ČR GCP305/10/J052 Grant - others:Chang Gung Memorial Hospital(TW) CMRPD170233; NSCNHRI Taiwan(TW) NSC 97-2314-B-182-001-MY2; NSCNHRI Taiwan(TW) NSC 99-2314-B-182-024-MY3; NSCNHRI Taiwan(TW) NSC 99-2923-B-182-001-MY3 Institutional support: RVO:68378050 Keywords : HERV * transcription * tissue-specific Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.106, year: 2013

  13. [Production of human proteins in the blood of transgenic animals

    NARCIS (Netherlands)

    Massoud, M.; Bischoff, Rainer; Dalemans, W.; Pointu, H.; Attal, J.; Schultz, H.; Clesse, D.; Stinnakre, M.G.; Pavirani, A.; Houdebine, L.M.

    1990-01-01

    The human alpha 1-antitrypsin gene has been microinjected into rabbit embryos. A line of transgenic rabbits has thus been established. Human alpha 1-antitrypsin was found in the blood of transgenic animals at the concentration of 1 mg/ml plasma. The human protein was active and separable from its

  14. Transcriptional dysregulation in NIPBL and cohesin mutant human cells.

    Directory of Open Access Journals (Sweden)

    Jinglan Liu

    2009-05-01

    Full Text Available Cohesin regulates sister chromatid cohesion during the mitotic cell cycle with Nipped-B-Like (NIPBL facilitating its loading and unloading. In addition to this canonical role, cohesin has also been demonstrated to play a critical role in regulation of gene expression in nondividing cells. Heterozygous mutations in the cohesin regulator NIPBL or cohesin structural components SMC1A and SMC3 result in the multisystem developmental disorder Cornelia de Lange Syndrome (CdLS. Genome-wide assessment of transcription in 16 mutant cell lines from severely affected CdLS probands has identified a unique profile of dysregulated gene expression that was validated in an additional 101 samples and correlates with phenotypic severity. This profile could serve as a diagnostic and classification tool. Cohesin binding analysis demonstrates a preference for intergenic regions suggesting a cis-regulatory function mimicking that of a boundary/insulator interacting protein. However, the binding sites are enriched within the promoter regions of the dysregulated genes and are significantly decreased in CdLS proband, indicating an alternative role of cohesin as a transcription factor.

  15. The transcriptional network that controls growth arrest and differentiation in a human myeloid leukemia cell line

    DEFF Research Database (Denmark)

    Suzuki, Harukazu; Forrest, Alistair R R; van Nimwegen, Erik

    2009-01-01

    , we identified the key transcription regulators, their time-dependent activities and target genes. Systematic siRNA knockdown of 52 transcription factors confirmed the roles of individual factors in the regulatory network. Our results indicate that cellular states are constrained by complex networks......Using deep sequencing (deepCAGE), the FANTOM4 study measured the genome-wide dynamics of transcription-start-site usage in the human monocytic cell line THP-1 throughout a time course of growth arrest and differentiation. Modeling the expression dynamics in terms of predicted cis-regulatory sites...... involving both positive and negative regulatory interactions among substantial numbers of transcription factors and that no single transcription factor is both necessary and sufficient to drive the differentiation process....

  16. Involvement of DNA topoisomerase I in transcription of human ribosomal RNA genes

    International Nuclear Information System (INIS)

    Zhang, H.; Wang, J.C.; Liu, L.F.

    1988-01-01

    Treatment of HeLa cells with a DNA topoisomerase I-specific inhibitor, camptothecin, results in rapid cessation of the synthesis of the 45S rRNA precursor. The inhibition of rRNA synthesis is reversible following drug removal and correlates with the presence of camptothecin-trapped topoisomerase I-DNA abortive complexes, which can be detected as topoisomerase I-linked DNA breaks upon lysis with sodium dodecyl sulfate. These breaks were found to be concentrated within the transcribed region of human rRNA genes. No such sites can be detected in the inactive human rRNA genes in mouse-human hybrid cells, suggesting a preferential association of topoisomerase I with actively transcribed genes. The distribution of RNA polymerase molecules along the transcription unit of human rRNA genes in camptothecin-treated HeLa cells, as assayed by nuclear run-on transcription, shows a graded decrease of the RNA polymerase density toward the 3' end of the transcription unit; the density is minimally affected near the 5' start of the transcription unit. These results suggest that DNA topoisomerase I is normally involved in the elongation step of transcription, especially when the transcripts are long, and that camptothecin interferes with this role

  17. Salt, Blood Pressure and Cardiovascular Changes in Human and ...

    African Journals Online (AJOL)

    Salt, Blood Pressure and Cardiovascular Changes in Human and Experimental Studies – A Review. ... Some of the pathophysiological changes include cardiac hypertrophy and enhanced cardiac contractility, enhanced contraction of blood vessels and veins in response to constrictor agonists and diminished relaxation of ...

  18. Human growth hormone alters carbohydrate storage in blood and ...

    African Journals Online (AJOL)

    MJP

    2015-06-02

    Jun 2, 2015 ... is the key hormone to maintain the glucose ... homeostasis is tissue-specific.[3] ... Key words: Human growth hormone, blood glucose, hepatic glycogen, hypoglycaemia, ..... diabetic and glycogenolytic effect, which help.

  19. Degradation of blood in the human digestive tract

    International Nuclear Information System (INIS)

    Moerup, S.; Johansen, C.

    1986-01-01

    Samples of human faeces from patients suffering from intestinal bleeding have been studied by use of Moessbauer spectroscopy. It is shown that it is possible to follow the degradation of blood in the digestive tract. (Auth.)

  20. A transcriptional profile of aging in the human kidney.

    Directory of Open Access Journals (Sweden)

    Graham E J Rodwell

    2004-12-01

    Full Text Available In this study, we found 985 genes that change expression in the cortex and the medulla of the kidney with age. Some of the genes whose transcripts increase in abundance with age are known to be specifically expressed in immune cells, suggesting that immune surveillance or inflammation increases with age. The age-regulated genes show a similar aging profile in the cortex and the medulla, suggesting a common underlying mechanism for aging. Expression profiles of these age-regulated genes mark not only age, but also the relative health and physiology of the kidney in older individuals. Finally, the set of aging-regulated kidney genes suggests specific mechanisms and pathways that may play a role in kidney degeneration with age.

  1. Bystander apoptosis in human cells mediated by irradiated blood plasma

    Energy Technology Data Exchange (ETDEWEB)

    Vinnikov, Volodymyr, E-mail: vlad.vinnikov@mail.ru [Grigoriev Institute for Medical Radiology of the National Academy of Medical Science of Ukraine (Ukraine); Lloyd, David; Finnon, Paul [Centre for Radiation, Chemical and Environmental Hazards of the Health Protection Agency of the United Kingdom (United Kingdom)

    2012-03-01

    Following exposure to high doses of ionizing radiation, due to an accident or during radiotherapy, bystander signalling poses a potential hazard to unirradiated cells and tissues. This process can be mediated by factors circulating in blood plasma. Thus, we assessed the ability of plasma taken from in vitro irradiated human blood to produce a direct cytotoxic effect, by inducing apoptosis in primary human peripheral blood mononuclear cells (PBM), which mainly comprised G{sub 0}-stage lymphocytes. Plasma was collected from healthy donors' blood irradiated in vitro to 0-40 Gy acute {gamma}-rays. Reporter PBM were separated from unirradiated blood with Histopaque and held in medium with the test plasma for 24 h at 37 Degree-Sign C. Additionally, plasma from in vitro irradiated and unirradiated blood was tested against PBM collected from blood given 4 Gy. Apoptosis in reporter PBM was measured by the Annexin V test using flow cytometry. Plasma collected from unirradiated and irradiated blood did not produce any apoptotic response above the control level in unirradiated reporter PBM. Surprisingly, plasma from irradiated blood caused a dose-dependent reduction of apoptosis in irradiated reporter PBM. The yields of radiation-induced cell death in irradiated reporter PBM (after subtracting the respective values in unirradiated reporter PBM) were 22.2 {+-} 1.8% in plasma-free cultures, 21.6 {+-} 1.1% in cultures treated with plasma from unirradiated blood, 20.2 {+-} 1.4% in cultures with plasma from blood given 2-4 Gy and 16.7 {+-} 3.2% in cultures with plasma from blood given 6-10 Gy. These results suggested that irradiated blood plasma did not cause a radiation-induced bystander cell-killing effect. Instead, a reduction of apoptosis in irradiated reporter cells cultured with irradiated blood plasma has implications concerning oncogenic risk from mutated cells surviving after high dose in vivo irradiation (e.g. radiotherapy) and requires further study.

  2. Bystander apoptosis in human cells mediated by irradiated blood plasma

    International Nuclear Information System (INIS)

    Vinnikov, Volodymyr; Lloyd, David; Finnon, Paul

    2012-01-01

    Following exposure to high doses of ionizing radiation, due to an accident or during radiotherapy, bystander signalling poses a potential hazard to unirradiated cells and tissues. This process can be mediated by factors circulating in blood plasma. Thus, we assessed the ability of plasma taken from in vitro irradiated human blood to produce a direct cytotoxic effect, by inducing apoptosis in primary human peripheral blood mononuclear cells (PBM), which mainly comprised G 0 -stage lymphocytes. Plasma was collected from healthy donors’ blood irradiated in vitro to 0–40 Gy acute γ-rays. Reporter PBM were separated from unirradiated blood with Histopaque and held in medium with the test plasma for 24 h at 37 °C. Additionally, plasma from in vitro irradiated and unirradiated blood was tested against PBM collected from blood given 4 Gy. Apoptosis in reporter PBM was measured by the Annexin V test using flow cytometry. Plasma collected from unirradiated and irradiated blood did not produce any apoptotic response above the control level in unirradiated reporter PBM. Surprisingly, plasma from irradiated blood caused a dose-dependent reduction of apoptosis in irradiated reporter PBM. The yields of radiation-induced cell death in irradiated reporter PBM (after subtracting the respective values in unirradiated reporter PBM) were 22.2 ± 1.8% in plasma-free cultures, 21.6 ± 1.1% in cultures treated with plasma from unirradiated blood, 20.2 ± 1.4% in cultures with plasma from blood given 2–4 Gy and 16.7 ± 3.2% in cultures with plasma from blood given 6–10 Gy. These results suggested that irradiated blood plasma did not cause a radiation-induced bystander cell-killing effect. Instead, a reduction of apoptosis in irradiated reporter cells cultured with irradiated blood plasma has implications concerning oncogenic risk from mutated cells surviving after high dose in vivo irradiation (e.g. radiotherapy) and requires further study.

  3. Transcriptome analysis of Neisseria meningitidis in human whole blood and mutagenesis studies identify virulence factors involved in blood survival.

    Directory of Open Access Journals (Sweden)

    Hebert Echenique-Rivera

    2011-05-01

    Full Text Available During infection Neisseria meningitidis (Nm encounters multiple environments within the host, which makes rapid adaptation a crucial factor for meningococcal survival. Despite the importance of invasion into the bloodstream in the meningococcal disease process, little is known about how Nm adapts to permit survival and growth in blood. To address this, we performed a time-course transcriptome analysis using an ex vivo model of human whole blood infection. We observed that Nm alters the expression of ≈30% of ORFs of the genome and major dynamic changes were observed in the expression of transcriptional regulators, transport and binding proteins, energy metabolism, and surface-exposed virulence factors. In particular, we found that the gene encoding the regulator Fur, as well as all genes encoding iron uptake systems, were significantly up-regulated. Analysis of regulated genes encoding for surface-exposed proteins involved in Nm pathogenesis allowed us to better understand mechanisms used to circumvent host defenses. During blood infection, Nm activates genes encoding for the factor H binding proteins, fHbp and NspA, genes encoding for detoxifying enzymes such as SodC, Kat and AniA, as well as several less characterized surface-exposed proteins that might have a role in blood survival. Through mutagenesis studies of a subset of up-regulated genes we were able to identify new proteins important for survival in human blood and also to identify additional roles of previously known virulence factors in aiding survival in blood. Nm mutant strains lacking the genes encoding the hypothetical protein NMB1483 and the surface-exposed proteins NalP, Mip and NspA, the Fur regulator, the transferrin binding protein TbpB, and the L-lactate permease LctP were sensitive to killing by human blood. This increased knowledge of how Nm responds to adaptation in blood could also be helpful to develop diagnostic and therapeutic strategies to control the devastating

  4. The transcription factor SOX18 regulates the expression of matrix metalloproteinase 7 and guidance molecules in human endothelial cells.

    Directory of Open Access Journals (Sweden)

    Martina Hoeth

    Full Text Available Mutations in the transcription factor SOX18 are responsible for specific cardiovascular defects in humans and mice. In order to gain insight into the molecular basis of its action, we identified target genes of SOX18 and analyzed one, MMP7, in detail.SOX18 was expressed in HUVEC using a recombinant adenoviral vector and the altered gene expression profile was analyzed using microarrays. Expression of several regulated candidate SOX18 target genes was verified by real-time PCR. Knock-down of SOX18 using RNA interference was then used to confirm the effect of the transcription factor on selected genes that included the guidance molecules ephrin B2 and semaphorin 3G. One gene, MMP7, was chosen for further analysis, including detailed promoter studies using reporter gene assays, electrophoretic mobility shift analysis and chromatin-immunoprecipitation, revealing that it responds directly to SOX18. Immunohistochemical analysis demonstrated the co-expression of SOX18 and MMP7 in blood vessels of human skin.The identification of MMP7 as a direct SOX18 target gene as well as other potential candidates including guidance molecules provides a molecular basis for the proposed function of this transcription factor in the regulation of vessel formation.

  5. DOT/FAA Human Factors Workshop on Aviation (5th). Transcript.

    Science.gov (United States)

    1982-01-01

    This document is a verbatim transcript of the proceedings of the Fifth Human Factors Workshop held at the Mike Monroney Aeronautical Center in Oklahoma City, Oklahoma, on July 7-9, 1981. The Sixth Human Factors Workshop was held at the same facility ...

  6. Role of natural antisense transcripts pertaining to tumor suppressor genes in human carcinomas

    International Nuclear Information System (INIS)

    Pelicci, G.; Pierotti, M.

    2009-01-01

    Overlapping transcripts in opposite orientations can potentially form perfect sense-antisense duplex RNA. Recently, several studies have revealed the extent of natural antisense transcripts (NATs) and their role in important biological phenomena also in higher organisms. In order to test the hypothesis that the function of NATs in man might represent an essential element in the regulation of gene expression, especially at transcriptional level, in this study we planned to look for, systematically examine, and characterize NATs belonging in the human genome to the tumour suppressor class of genes, so to identify physiological (and potentially pathological) modulators in this gene class

  7. Evaluation of genome damage and transcription profile of DNA damage/repair response genes in peripheral blood mononuclear cells exposed to low dose radiation

    International Nuclear Information System (INIS)

    Soren, D.C.; Saini, Divyalakshmi; Das, Birajalaxmi

    2016-01-01

    Humans are exposed to various physical and chemical mutagens in their life time. Physical mutagens, like ionizing radiation (IR), may induce adverse effect at high acute dose exposures in human cells. However, there are inconsistent results on the effect of low dose radiation exposure in human cells. There are a variety of DNA damage endpoints to evaluate the effect of low dose radiation in human cells. DNA damage response (DDR) may lead to changes in expression profile of many genes. In the present study, an attempt has been made to evaluate genome damage at low dose IR exposure in human blood lymphocytes. Cytochalasin blocked micronuclei (CBMN) assay has been used to determine the frequency of micronuclei in binucleated cells in PBMCs exposed to IR. Transcription profile of ATM, P53, GADD45A, CDKN1A, TRF1 and TRF2 genes was studied using real time quantitative PCR. Venous blood samples collected from 10 random healthy donors were irradiated with different doses of γ-radiation ( 137 Cs) along with sham irradiated control. Whole blood culture was set up using microculture technique. Blood samples were stimulated with phytohemagglutinin, and CBMN assay was performed. An average of 2,500 binucleated cells was scored for each dose point. For gene expression analysis, total RNA was isolated, cDNA was prepared, and gene expression analysis for ATM, P53, CDKN1A, GADD45A, TRF1 and TRF2 was done using real time PCR. Our results revealed no significant increase in the frequency of MN up to 100 mGy as compared to control. However, no significant alteration in gene expression profile was observed. In conclusion, no significant dose response was observed at the frequency of MN as well as the expression profile of DDR/repair genes, suggesting low dose radiation did not induce significant DNA damage at these acute dose exposures. (author)

  8. Direct RNA-based detection of CTX-M β-lactamases in human blood samples.

    Science.gov (United States)

    Stein, Claudia; Makarewicz, Oliwia; Pfeifer, Yvonne; Brandt, Christian; Pletz, Mathias W

    2015-05-01

    Bloodstream infections with ESBL-producers are associated with increased mortality, which is due to delayed appropriate treatment resulting in clinical failure. Current routine diagnostics for detection of bloodstream infections consists of blood culture followed by species identification and susceptibility testing. In attempts to improve and accelerate diagnostic procedures, PCR-based methods have been developed. These methods focus on species identification covering only a limited number of ESBL coding genes. Therefore, they fail to cover the steadily further evolving genetic diversity of clinically relevant β-lactamases. We have recently designed a fast and novel RNA targeting method to detect and specify CTX-M alleles from bacterial cultures, based on an amplification-pyrosequencing approach. We further developed this assay towards a diagnostic tool for clinical use and evaluated its sensitivity and specificity when applied directly to human blood samples. An optimized protocol for mRNA isolation allows detection of specific CTX-M groups from as little as 100 CFU/mL blood via reverse transcription, amplification, and pyrosequencing directly from human EDTA blood samples as well as from pre-incubated human blood cultures with a turnaround time for test results of <7 h. Copyright © 2015 Elsevier GmbH. All rights reserved.

  9. Skin Blood Perfusion and Oxygenation Colour Affect Perceived Human Health

    Science.gov (United States)

    Stephen, Ian D.; Coetzee, Vinet; Law Smith, Miriam; Perrett, David I.

    2009-01-01

    Skin blood perfusion and oxygenation depends upon cardiovascular, hormonal and circulatory health in humans and provides socio-sexual signals of underlying physiology, dominance and reproductive status in some primates. We allowed participants to manipulate colour calibrated facial photographs along empirically-measured oxygenated and deoxygenated blood colour axes both separately and simultaneously, to optimise healthy appearance. Participants increased skin blood colour, particularly oxygenated, above basal levels to optimise healthy appearance. We show, therefore, that skin blood perfusion and oxygenation influence perceived health in a way that may be important to mate choice. PMID:19337378

  10. Effect of fluocinolone acetonide cream on human skin blood flow

    International Nuclear Information System (INIS)

    Chimoskey, J.E.; Holloway, A. Jr.; Flanagan, W.J.

    1975-01-01

    Blood flow rate was measured in the forearm skin of human subjects exposed to ultraviolet irradiation. Blood flow was determined by the 133 Xe disappearance technique 18 hr after ultraviolet (UV) irradiation with a Westinghouse RS sunlamp held 10 inches from the skin for 10 min. Ultraviolet irradiation caused skin blood flow to increase. Application of fluocinolone acetonide cream, 0.025 percent, 4 times in the 16 hr following UV irradiation had no effect on either control skin blood flow or the UV-induced hyperemia

  11. Nocturnal variations in peripheral blood flow, systemic blood pressure, and heart rate in humans

    DEFF Research Database (Denmark)

    Sindrup, J H; Kastrup, J; Christensen, H

    1991-01-01

    Subcutaneous adipose tissue blood flow rate, together with systemic arterial blood pressure and heart rate under ambulatory conditions, was measured in the lower legs of 15 normal human subjects for 12-20 h. The 133Xe-washout technique, portable CdTe(Cl) detectors, and a portable data storage uni.......0001). The synchronism of the nocturnal subcutaneous hyperemia and the decrease in systemic mean arterial blood pressure point to a common, possibly central nervous or humoral, eliciting mechanism.......Subcutaneous adipose tissue blood flow rate, together with systemic arterial blood pressure and heart rate under ambulatory conditions, was measured in the lower legs of 15 normal human subjects for 12-20 h. The 133Xe-washout technique, portable CdTe(Cl) detectors, and a portable data storage unit...

  12. GAD2 Alternative Transcripts in the Human Prefrontal Cortex, and in Schizophrenia and Affective Disorders.

    Directory of Open Access Journals (Sweden)

    Kasey N Davis

    Full Text Available Genetic variation and early adverse environmental events work together to increase risk for schizophrenia. γ-aminobutyric acid (GABA, the major inhibitory neurotransmitter in adult mammalian brain, plays a major role in normal brain development, and has been strongly implicated in the pathobiology of schizophrenia. GABA synthesis is controlled by two glutamic acid decarboxylase (GAD genes, GAD1 and GAD2, both of which produce a number of alternative transcripts. Genetic variants in the GAD1 gene are associated with increased risk for schizophrenia, and reduced expression of its major transcript in the human dorsolateral prefrontal cortex (DLPFC. No consistent changes in GAD2 expression have been found in brains from patients with schizophrenia. In this work, with the use of RNA sequencing and PCR technologies, we confirmed and tracked the expression of an alternative truncated transcript of GAD2 (ENST00000428517 in human control DLPFC homogenates across lifespan besides the well-known full length transcript of GAD2. In addition, using quantitative RT-PCR, expression of GAD2 full length and truncated transcripts were measured in the DLPFC of patients with schizophrenia, bipolar disorder and major depression. The expression of GAD2 full length transcript is decreased in the DLPFC of schizophrenia and bipolar disorder patients, while GAD2 truncated transcript is increased in bipolar disorder patients but decreased in schizophrenia patients. Moreover, the patients with schizophrenia with completed suicide or positive nicotine exposure showed significantly higher expression of GAD2 full length transcript. Alternative transcripts of GAD2 may be important in the growth and development of GABA-synthesizing neurons as well as abnormal GABA signaling in the DLPFC of patients with schizophrenia and affective disorders.

  13. Human macrophages support persistent transcription from unintegrated HIV-1 DNA

    International Nuclear Information System (INIS)

    Kelly, Jeremy; Beddall, Margaret H.; Yu Dongyang; Iyer, Subashini R.; Marsh, Jon W.; Wu Yuntao

    2008-01-01

    Retroviruses require integration of their RNA genomes for both stability and productive viral replication. In HIV infection of non-dividing, resting CD4 T cells, where integration is greatly impeded, the reverse transcribed HIV DNA has limited biological activity and a short half-life. In metabolically active and proliferating T cells, unintegrated DNA rapidly diminishes with cell division. HIV also infects the non-dividing but metabolically active macrophage population. In an in vitro examination of HIV infection of macrophages, we find that unintegrated viral DNA not only has an unusual stability, but also maintains biological activity. The unintegrated linear DNA, 1-LTR, and 2-LTR circles are stable for at least 30 days. Additionally, there is persistent viral gene transcription, which is selective and skewed towards viral early genes such as nef and tat with highly diminished rev and vif. One viral early gene product Nef was measurably synthesized. We also find that independent of integration, the HIV infection process in macrophages leads to generation of numerous chemokines

  14. The determination of chromium-50 in human blood and its utilization for blood volume measurements

    International Nuclear Information System (INIS)

    Zeisler, R.; Young, I.

    1986-01-01

    Possible relationships between insufficient blood volume increases during pregnancy and infant mortality could be established with an adequate measurement procedure. An accurate and precise technique for blood volume measurements has been found in the isotope dilution technique using chromium-51 as a label for red blood cells. However, in a study involving pregnant women, only stable isotopes can be used for labeling. Stable chromium-50 can be determined in total blood samples before and after dilution experiments by neutron activation analysis (NAA) or mass spectrometry. However, both techniques may be affected by insufficient sensitivity and contamination problems at the inherently low natural chromium concentrations to be measured in the blood. NAA procedures involving irradiations with highly thermalized neutrons at a fluence rate of 2x10 13 n/cm 2 xs and low background gamma spectrometry are applied to the analysis of total blood. Natural levels of chromium-50 in human and animal blood have been found to be <0.1 ng/mL; i.e., total chromium levels of <3 ng/mL. Based on the NAA procedure, a new approach to the blood volume measurement via chromium-50 isotope dilution has been developed which utilizes the ratio of the induced activities of chromium-51 to the iron-59 in three blood samples taken from each individual, namely blank, labeled and diluted labeled blood. (author)

  15. A simple method for human peripheral blood monocyte Isolation

    Directory of Open Access Journals (Sweden)

    Marcos C de Almeida

    2000-04-01

    Full Text Available We describe a simple method using percoll gradient for isolation of highly enriched human monocytes. High numbers of fully functional cells are obtained from whole blood or buffy coat cells. The use of simple laboratory equipment and a relatively cheap reagent makes the described method a convenient approach to obtaining human monocytes.

  16. effects of septrin administration on blood cells parameters in humans

    African Journals Online (AJOL)

    honey

    2014-03-31

    Mar 31, 2014 ... RESEARCH PAPER. EFFECTS OF SEPTRIN ADMINISTRATION ON BLOOD CELLS PARAMETERS IN. HUMANS. *1Onyebuagu P.C., 2Kiridi K. and 1Pughikumo D.T.. 1Department of Human Physiology, Niger Delta University, Bayelsa, Nigeria. 2Department of Radiology, Niger. Delta University, Bayelsa ...

  17. Nucleotide sequences of cDNAs for human papillomavirus type 18 transcripts in HeLa cells

    International Nuclear Information System (INIS)

    Inagaki, Yutaka; Tsunokawa, Youko; Takebe, Naoko; Terada, Masaaki; Sugimura, Takashi; Nawa, Hiroyuki; Nakanishi, Shigetada

    1988-01-01

    HeLa cells expressed 3.4- and 1.6-kilobase (kb) transcripts of the integrated human papillomavirus (HPV) type 18 genome. Two types of cDNA clones representing each size of HPV type 18 transcript were isolated. Sequence analysis of these two types of cDNA clones revealed that the 3.4-kb transcript contained E6, E7, the 5' portion of E1, and human sequence and that the 1.6-kb transcript contained spliced and frameshifted E6 (E6 * ), E7, and human sequence. There was a common human sequence containing a poly(A) addition signal in the 3' end portions of both transcripts, indicating that they were transcribed from the HPV genome at the same integration site with different splicing. Furthermore, the 1.6-kb transcript contained both of the two viral TATA boxes upstream of E6, strongly indicating that a cellular promoter was used for its transcription

  18. Alterations in transcription factor binding in radioresistant human melanoma cells after ionizing radiation

    International Nuclear Information System (INIS)

    Sahijdak, W.M.; Yang, Chin-Rang; Zuckerman, J.S.; Meyers, M.; Boothman, D.A.

    1994-01-01

    We analyzed alterations in transcription factor binding to specific, known promoter DNA consensus sequences between irradiated and unirradiated radioresistant human melanoma (U1-Mel) cells. The goal of this study was to begin to investigate which transcription factors and DNA-binding sites are responsible for the induction of specific transcripts and proteins after ionizing radiation. Transcription factor binding was observed using DNA band-shift assays and oligonucleotide competition analyses. Confluence-arrested U1-Mel cells were irradiated (4.5 Gy) and harvested at 4 h. Double-stranded oligonucleotides containing known DNA-binding consensus sites for specific transcription factors were used. Increased DNA binding activity after ionizing radiation was noted with oligonucleotides containing the CREB, NF-kB and Sp1 consensus sites. No changes in protein binding to AP-1, AP-2, AP-3, or CTF/NF1, GRE or Oct-1 consensus sequences were noted. X-ray activation of select transcription factors, which bind certain consensus sites in promoters, may cause specific induction or repression of gene transcription. 22 refs., 2 figs

  19. The predictive nature of transcript expression levels on protein expression in adult human brain.

    Science.gov (United States)

    Bauernfeind, Amy L; Babbitt, Courtney C

    2017-04-24

    Next generation sequencing methods are the gold standard for evaluating expression of the transcriptome. When determining the biological implications of such studies, the assumption is often made that transcript expression levels correspond to protein levels in a meaningful way. However, the strength of the overall correlation between transcript and protein expression is inconsistent, particularly in brain samples. Following high-throughput transcriptomic (RNA-Seq) and proteomic (liquid chromatography coupled with tandem mass spectrometry) analyses of adult human brain samples, we compared the correlation in the expression of transcripts and proteins that support various biological processes, molecular functions, and that are located in different areas of the cell. Although most categories of transcripts have extremely weak predictive value for the expression of their associated proteins (R 2 values of < 10%), transcripts coding for protein kinases and membrane-associated proteins, including those that are part of receptors or ion transporters, are among those that are most predictive of downstream protein expression levels. The predictive value of transcript expression for corresponding proteins is variable in human brain samples, reflecting the complex regulation of protein expression. However, we found that transcriptomic analyses are appropriate for assessing the expression levels of certain classes of proteins, including those that modify proteins, such as kinases and phosphatases, regulate metabolic and synaptic activity, or are associated with a cellular membrane. These findings can be used to guide the interpretation of gene expression results from primate brain samples.

  20. Relationship between human cytomegalovirus transcription and symptomatic apical periodontitis in Iran.

    Science.gov (United States)

    Yazdi, K A; Sabeti, M; Jabalameli, F; Eman eini, M; Kolahdouzan, S A; Slots, J

    2008-12-01

    Apical periodontitis of endodontic origin may develop as a result of cooperative interactions among herpesviruses, specific pathogenic bacteria and tissue-destructive inflammatory mediators. This study sought to identify the presence of Epstein-Barr virus (EBV) and human cytomegalovirus (HCMV) transcripts in symptomatic and asymptomatic periapical lesions of individuals living in Iran. Fifty endodontic patients (28 with symptomatic periapical lesions and 22 with asymptomatic periapical lesions) were included in the study. In each study subject, a microbiological periapical sample was collected using a curette in conjunction with periapical surgery. A reverse transcription-polymerase chain reaction assay was used to identify transcripts of EBV and HCMV. Human cytomegalovirus transcript was detected in 15 of the 28 (53.6%) symptomatic and in six of the 22 (27.3%) asymptomatic periapical study lesions (significant difference between symptomatic and asymptomatic lesions; P = 0.03, chi-square test). Epstein-Barr virus transcript was identified in one symptomatic and in two asymptomatic periapical lesions. This study establishes that HCMV transcription is common in apical periodontitis and is most frequent in symptomatic lesions. The high frequency of active herpesvirus infections in severe apical periodontitis changes the pathogenic paradigm of the disease and may also have preventive and therapeutic implications.

  1. Controlling transcription in human pluripotent stem cells using CRISPR-effectors.

    Science.gov (United States)

    Genga, Ryan M; Kearns, Nicola A; Maehr, René

    2016-05-15

    The ability to manipulate transcription in human pluripotent stem cells (hPSCs) is fundamental for the discovery of key genes and mechanisms governing cellular state and differentiation. Recently developed CRISPR-effector systems provide a systematic approach to rapidly test gene function in mammalian cells, including hPSCs. In this review, we discuss recent advances in CRISPR-effector technologies that have been employed to control transcription through gene activation, gene repression, and epigenome engineering. We describe an application of CRISPR-effector mediated transcriptional regulation in hPSCs by targeting a synthetic promoter driving a GFP transgene, demonstrating the ease and effectiveness of CRISPR-effector mediated transcriptional regulation in hPSCs. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. Application of dried blood spot cards to determine olive oil phenols (hydroxytyrosol metabolites) in human blood.

    Science.gov (United States)

    de Las Hazas, María Carmen López; Motilva, Maria José; Piñol, Carme; Macià, Alba

    2016-10-01

    In this study, a fast and simple blood sampling and sample pre-treatment method based on the use of the dried blood spot (DBS) cards and ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) for the quantification of olive oil phenolic metabolites in human blood was developed and validated. After validation, the method was applied to determine hydroxytyrosol metabolites in human blood samples after the acute intake of an olive oil phenolic extract. Using the FTA DMPK-A DBS card under optimum conditions, with 20µL as the blood solution volume, 100µL of methanol/Milli-Q water (50/50, v/v) as the extraction solvent and 7 disks punched out from the card, the main hydroxytyrosol metabolites (hydroxytyrosol-3-O-sulphate and hydroxytyrosol acetate sulphate) were identified and quantified. The developed methodology allowed detecting and quantifying the generated metabolites at low μM levels. The proposed method is a significant improvement over existing methods to determine phenolic metabolites circulating in blood and plasma samples, thus making blood sampling possible with the volunteer pricking their own finger, and the subsequent storage of the blood in the DBS cards prior to chromatographic analysis. Copyright © 2016 Elsevier B.V. All rights reserved.

  3. Spatial and Single-Cell Transcriptional Profiling Identifies Functionally Distinct Human Dermal Fibroblast Subpopulations.

    Science.gov (United States)

    Philippeos, Christina; Telerman, Stephanie B; Oulès, Bénédicte; Pisco, Angela O; Shaw, Tanya J; Elgueta, Raul; Lombardi, Giovanna; Driskell, Ryan R; Soldin, Mark; Lynch, Magnus D; Watt, Fiona M

    2018-04-01

    Previous studies have shown that mouse dermis is composed of functionally distinct fibroblast lineages. To explore the extent of fibroblast heterogeneity in human skin, we used a combination of comparative spatial transcriptional profiling of human and mouse dermis and single-cell transcriptional profiling of human dermal fibroblasts. We show that there are at least four distinct fibroblast populations in adult human skin, not all of which are spatially segregated. We define markers permitting their isolation and show that although marker expression is lost in culture, different fibroblast subpopulations retain distinct functionality in terms of Wnt signaling, responsiveness to IFN-γ, and ability to support human epidermal reconstitution when introduced into decellularized dermis. These findings suggest that ex vivo expansion or in vivo ablation of specific fibroblast subpopulations may have therapeutic applications in wound healing and diseases characterized by excessive fibrosis. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

  4. O-GlcNAc transferase regulates transcriptional activity of human Oct4.

    Science.gov (United States)

    Constable, Sandii; Lim, Jae-Min; Vaidyanathan, Krithika; Wells, Lance

    2017-10-01

    O-linked β-N-acetylglucosamine (O-GlcNAc) is a single sugar modification found on many different classes of nuclear and cytoplasmic proteins. Addition of this modification, by the enzyme O-linked N-acetylglucosamine transferase (OGT), is dynamic and inducible. One major class of proteins modified by O-GlcNAc is transcription factors. O-GlcNAc regulates transcription factor properties through a variety of different mechanisms including localization, stability and transcriptional activation. Maintenance of embryonic stem (ES) cell pluripotency requires tight regulation of several key transcription factors, many of which are modified by O-GlcNAc. Octamer-binding protein 4 (Oct4) is one of the key transcription factors required for pluripotency of ES cells and more recently, the generation of induced pluripotent stem (iPS) cells. The action of Oct4 is modulated by the addition of several post-translational modifications, including O-GlcNAc. Previous studies in mice found a single site of O-GlcNAc addition responsible for transcriptional regulation. This study was designed to determine if this mechanism is conserved in humans. We mapped 10 novel sites of O-GlcNAc attachment on human Oct4, and confirmed a role for OGT in transcriptional activation of Oct4 at a site distinct from that found in mouse that allows distinction between different Oct4 target promoters. Additionally, we uncovered a potential new role for OGT that does not include its catalytic function. These results confirm that human Oct4 activity is being regulated by OGT by a mechanism that is distinct from mouse Oct4. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  5. High resolution analysis of the human transcriptome: detection of extensive alternative splicing independent of transcriptional activity

    Directory of Open Access Journals (Sweden)

    Rouet Fabien

    2009-10-01

    Full Text Available Abstract Background Commercially available microarrays have been used in many settings to generate expression profiles for a variety of applications, including target selection for disease detection, classification, profiling for pharmacogenomic response to therapeutics, and potential disease staging. However, many commercially available microarray platforms fail to capture transcript diversity produced by alternative splicing, a major mechanism for driving proteomic diversity through transcript heterogeneity. Results The human Genome-Wide SpliceArray™ (GWSA, a novel microarray platform, utilizes an existing probe design concept to monitor such transcript diversity on a genome scale. The human GWSA allows the detection of alternatively spliced events within the human genome through the use of exon body and exon junction probes to provide a direct measure of each transcript, through simple calculations derived from expression data. This report focuses on the performance and validation of the array when measured against standards recently published by the Microarray Quality Control (MAQC Project. The array was shown to be highly quantitative, and displayed greater than 85% correlation with the HG-U133 Plus 2.0 array at the gene level while providing more extensive coverage of each gene. Almost 60% of splice events among genes demonstrating differential expression of greater than 3 fold also contained extensive splicing alterations. Importantly, almost 10% of splice events within the gene set displaying constant overall expression values had evidence of transcript diversity. Two examples illustrate the types of events identified: LIM domain 7 showed no differential expression at the gene level, but demonstrated deregulation of an exon skip event, while erythrocyte membrane protein band 4.1 -like 3 was differentially expressed and also displayed deregulation of a skipped exon isoform. Conclusion Significant changes were detected independent of

  6. Shape Memory of Human Red Blood Cells

    OpenAIRE

    Fischer, Thomas M.

    2004-01-01

    The human red cell can be deformed by external forces but returns to the biconcave resting shape after removal of the forces. If after such shape excursions the rim is always formed by the same part of the membrane, the cell is said to have a memory of its biconcave shape. If the rim can form anywhere on the membrane, the cell would have no shape memory. The shape memory was probed by an experiment called go-and-stop. Locations on the membrane were marked by spontaneously adhering latex spher...

  7. Regulation of the skeletal muscle blood flow in humans

    DEFF Research Database (Denmark)

    Mortensen, Stefan; Saltin, Bengt

    2014-01-01

    In humans, skeletal muscle blood flow is regulated by an interaction between several locally formed vasodilators including nitric oxide (NO) and prostaglandins. In plasma, ATP is a potent vasodilator that stimulates the formation of NO and prostaglandins and very importantly can offset local...... concentration does not increase during exercise. In the skeletal muscle interstitium, there is a marked increase in the concentration of ATP and adenosine and this increase is tightly coupled to the increase in blood flow. The sources of interstitial ATP and adenosine are thought to be skeletal muscle cells...... hyperaemia whereas the role of ATP remains uncertain due to lack of specific purinergic receptor blockers for human use. The purpose of this review is to address the interaction between vasodilator systems and to discuss the multiple proposed roles of ATP in human skeletal muscle blood flow regulation...

  8. Radioprotective effect of antioxidants on human blood lymphocytes

    International Nuclear Information System (INIS)

    Wang Mingsuo; Gu Xuandi; Zhu Genbo; Feng Jixing; Su Liaoyuan

    1991-09-01

    By using an improved fluorometric method with 2-thiobarbituric acid (TBA) as fluorometric agent, the antiradiation effects of four kinds of antioxidants on 60 Co γ-ray irradiation inducing final products of lipid peroxides (LPO), i.e. malodialdehyde (MDA) content changes in human blood lymphocytes, were investigated with LPO value as an indicator. The results of the experiment were as following: (1)The radioprotective effect of exogenous antioxidants added to human blood lymphocytes on radiation-induced LPO damage of cellular membrane were remarkable; (2)The radioprotective beneficial sequences of four kinds of antioxidants were arranged like this: SOD > VE >VC, Se 4+ ; (3)Radioprotective effects of antioxidants on radiation-induced damage varied especially with the property of antioxidants, drug concentration, and pretreatment and monitoring time, etc., as well as irradiated dosage and various kinds of incubated cells. In addition, the mechanism of these antioxidants as radioprotectants on human blood lymphocytes is discussed in connection with LPO damage and radioprotection

  9. Pancreas developing markers expressed on human mononucleated umbilical cord blood cells

    International Nuclear Information System (INIS)

    Pessina, A.; Eletti, B.; Croera, C.; Savalli, N.; Diodovich, C.; Gribaldo, L.

    2004-01-01

    Haematopoietic system represents the main source of haematopoietic stem cells and probably of multipotential adult progenitor cells and mesenchimal stem cells at first described as colony forming unit-fibroblast. Whereas there are many studies on the gene expression profile of the different precursors along their haematopoietic differentiation, few data (sometimes conflicting) have been reported about the phenotype of the cells (present in bone marrow and possibly in cord blood) able to differentiate into non-haematopoietic cells. As both postnatal bone marrow and umbilical cord blood contain nestin positive cells able to proliferate and differentiate into the main neural phenotype (neuron, astroglia and oligodendroglia) many authors considered nestin a neuroepithelial precursor marker that seems to be essential also in multipotential progenitor cells of pancreas present both in rat and in human pancreatic islets (called nestin positive islet derived progenitors). Although the importance of nestin in these cells appears to be evident, it remains yet to clarify the number and the sequential expression of the genes coding all the transcription factors essential for beta cells differentiation and therefore the conditions able to induce the expression of many important transcription factors genes such as isl-1, pax-4, pdx-1 and ngn-3. Among them pdx-1 is a gene essential for pancreas development which is able to control ngn-3 in activating the expression of other differentiation factors for endocrine cells. Here, we describe for the first time in human umbilical cord blood cells (UCB) the pattern of expression of a panel of markers (nestin, CK-8, CK-18) and transcription factors (Isl-1, Pdx-1, Pax-4, Ngn-3) considered important for beta cells differentiation. Our data demonstrate that UCB contains a cell population having a phenotype very similar to endocrine cell precursors in transition to beta cells

  10. Transcriptional dynamics during human adipogenesis and its link to adipose morphology and distribution

    DEFF Research Database (Denmark)

    Ehrlund, Anna; Mejhert, Niklas; Björk, Christel

    2017-01-01

    White adipose tissue (WAT) can develop into several phenotypes with different pathophysiological impact on type 2 diabetes. To better understand the adipogenic process, the transcriptional events that occur during in vitro differentiation of human adipocytes were investigated and the findings lin...

  11. DOT/FAA Human Factors Workshop on Aviation (6th). Transcript.

    Science.gov (United States)

    1982-05-01

    This document is a verbatim transcript of the proceedings of the DOT/FAA Sixth Human Factors Workshop on Aviation held at the Mike Monroney Aeronautical Center, Oklahoma City, Oklahoma on July 7-8, 1981. The subject of the workshop was aviation maint...

  12. Associating transcription factors and conserved RNA structures with gene regulation in the human brain

    DEFF Research Database (Denmark)

    Hecker, Nikolai; Seemann, Stefan E.; Silahtaroglu, Asli

    2017-01-01

    Anatomical subdivisions of the human brain can be associated with different neuronal functions. This functional diversification is reflected by differences in gene expression. By analyzing post-mortem gene expression data from the Allen Brain Atlas, we investigated the impact of transcription fac...

  13. Transcriptional regulation of human RANK ligand gene expression by E2F1

    International Nuclear Information System (INIS)

    Hu Yan; Sun Meng; Nadiminty, Nagalakshmi; Lou Wei; Pinder, Elaine; Gao, Allen C.

    2008-01-01

    Receptor activator of nuclear factor kappa B ligand (RANKL) is a critical osteoclastogenic factor involved in the regulation of bone resorption, immune function, the development of mammary gland and cardiovascular system. To understand the transcriptional regulation of RANKL, we amplified and characterized a 1890 bp 5'-flanking sequence of human RANKL gene (-1782 bp to +108 bp relative to the transcription start site). Using a series of deletion mutations of the 1890 bp RANKL promoter, we identified a 72 bp region (-172 to -100 bp) mediating RANKL basal transcriptional activity. Sequence analysis revealed a putative E2F binding site within this 72 bp region in the human RANKL promoter. Overexpression of E2F1 increased RANKL promoter activity, while down-regulation of E2F1 expression by small interfering RNA decreased RANKL promoter activity. RT-PCR and enzyme linked immunosorbent assays (ELISA) further demonstrated that E2F1 induced the expression of RANKL. Electrophoretic gel mobility shift assays (EMSA) and antibody competition assays confirmed that E2F1 proteins bind to the consensus E2F binding site in the RANKL promoter. Mutation of the E2F consensus binding site in the RANKL promoter profoundly reduced the basal promoter activity and abolished the transcriptional modulation of RANKL by E2F1. These results suggest that E2F1 plays an important role in regulating RANKL transcription through binding to the E2F consensus binding site

  14. 1-D blood flow modelling in a running human body.

    Science.gov (United States)

    Szabó, Viktor; Halász, Gábor

    2017-07-01

    In this paper an attempt was made to simulate blood flow in a mobile human arterial network, specifically, in a running human subject. In order to simulate the effect of motion, a previously published immobile 1-D model was modified by including an inertial force term into the momentum equation. To calculate inertial force, gait analysis was performed at different levels of speed. Our results show that motion has a significant effect on the amplitudes of the blood pressure and flow rate but the average values are not effected significantly.

  15. Natural blood feeding and temperature shift modulate the global transcriptional profile of Rickettsia rickettsii infecting its tick vector.

    Directory of Open Access Journals (Sweden)

    Maria Fernanda B M Galletti

    Full Text Available Rickettsia rickettsii is an obligate intracellular tick-borne bacterium that causes Rocky Mountain Spotted Fever (RMSF, the most lethal spotted fever rickettsiosis. When an infected starving tick begins blood feeding from a vertebrate host, R. rickettsii is exposed to a temperature elevation and to components in the blood meal. These two environmental stimuli have been previously associated with the reactivation of rickettsial virulence in ticks, but the factors responsible for this phenotype conversion have not been completely elucidated. Using customized oligonucleotide microarrays and high-throughput microfluidic qRT-PCR, we analyzed the effects of a 10°C temperature elevation and of a blood meal on the transcriptional profile of R. rickettsii infecting the tick Amblyomma aureolatum. This is the first study of the transcriptome of a bacterium in the genus Rickettsia infecting a natural tick vector. Although both stimuli significantly increased bacterial load, blood feeding had a greater effect, modulating five-fold more genes than the temperature upshift. Certain components of the Type IV Secretion System (T4SS were up-regulated by blood feeding. This suggests that this important bacterial transport system may be utilized to secrete effectors during the tick vector's blood meal. Blood feeding also up-regulated the expression of antioxidant enzymes, which might correspond to an attempt by R. rickettsii to protect itself against the deleterious effects of free radicals produced by fed ticks. The modulated genes identified in this study, including those encoding hypothetical proteins, require further functional analysis and may have potential as future targets for vaccine development.

  16. The blood DNA virome in 8,000 humans.

    Directory of Open Access Journals (Sweden)

    Ahmed Moustafa

    2017-03-01

    Full Text Available The characterization of the blood virome is important for the safety of blood-derived transfusion products, and for the identification of emerging pathogens. We explored non-human sequence data from whole-genome sequencing of blood from 8,240 individuals, none of whom were ascertained for any infectious disease. Viral sequences were extracted from the pool of sequence reads that did not map to the human reference genome. Analyses sifted through close to 1 Petabyte of sequence data and performed 0.5 trillion similarity searches. With a lower bound for identification of 2 viral genomes/100,000 cells, we mapped sequences to 94 different viruses, including sequences from 19 human DNA viruses, proviruses and RNA viruses (herpesviruses, anelloviruses, papillomaviruses, three polyomaviruses, adenovirus, HIV, HTLV, hepatitis B, hepatitis C, parvovirus B19, and influenza virus in 42% of the study participants. Of possible relevance to transfusion medicine, we identified Merkel cell polyomavirus in 49 individuals, papillomavirus in blood of 13 individuals, parvovirus B19 in 6 individuals, and the presence of herpesvirus 8 in 3 individuals. The presence of DNA sequences from two RNA viruses was unexpected: Hepatitis C virus is revealing of an integration event, while the influenza virus sequence resulted from immunization with a DNA vaccine. Age, sex and ancestry contributed significantly to the prevalence of infection. The remaining 75 viruses mostly reflect extensive contamination of commercial reagents and from the environment. These technical problems represent a major challenge for the identification of novel human pathogens. Increasing availability of human whole-genome sequences will contribute substantial amounts of data on the composition of the normal and pathogenic human blood virome. Distinguishing contaminants from real human viruses is challenging.

  17. Shape memory of human red blood cells.

    Science.gov (United States)

    Fischer, Thomas M

    2004-05-01

    The human red cell can be deformed by external forces but returns to the biconcave resting shape after removal of the forces. If after such shape excursions the rim is always formed by the same part of the membrane, the cell is said to have a memory of its biconcave shape. If the rim can form anywhere on the membrane, the cell would have no shape memory. The shape memory was probed by an experiment called go-and-stop. Locations on the membrane were marked by spontaneously adhering latex spheres. Shape excursions were induced by shear flow. In virtually all red cells, a shape memory was found. After stop of flow and during the return of the latex spheres to the original location, the red cell shape was biconcave. The return occurred by a tank-tread motion of the membrane. The memory could not be eliminated by deforming the red cells in shear flow up to 4 h at room temperature as well as at 37 degrees C. It is suggested that 1). the characteristic time of stress relaxation is >80 min and 2). red cells in vivo also have a shape memory.

  18. Simulated night shift work induces circadian misalignment of the human peripheral blood mononuclear cell transcriptome.

    Science.gov (United States)

    Kervezee, Laura; Cuesta, Marc; Cermakian, Nicolas; Boivin, Diane B

    2018-05-22

    Misalignment of the endogenous circadian timing system leads to disruption of physiological rhythms and may contribute to the development of the deleterious health effects associated with night shift work. However, the molecular underpinnings remain to be elucidated. Here, we investigated the effect of a 4-day simulated night shift work protocol on the circadian regulation of the human transcriptome. Repeated blood samples were collected over two 24-hour measurement periods from eight healthy subjects under highly controlled laboratory conditions before and 4 days after a 10-hour delay of their habitual sleep period. RNA was extracted from peripheral blood mononuclear cells to obtain transcriptomic data. Cosinor analysis revealed a marked reduction of significantly rhythmic transcripts in the night shift condition compared with baseline at group and individual levels. Subsequent analysis using a mixed-effects model selection approach indicated that this decrease is mainly due to dampened rhythms rather than to a complete loss of rhythmicity: 73% of transcripts rhythmically expressed at baseline remained rhythmic during the night shift condition with a similar phase relative to habitual bedtimes, but with lower amplitudes. Functional analysis revealed that key biological processes are affected by the night shift protocol, most notably the natural killer cell-mediated immune response and Jun/AP1 and STAT pathways. These results show that 4 days of simulated night shifts leads to a loss in temporal coordination between the human circadian transcriptome and the external environment and impacts biological processes related to the adverse health effects associated to night shift work.

  19. Muscle blood flow at onset of dynamic exercise in humans.

    Science.gov (United States)

    Rådegran, G; Saltin, B

    1998-01-01

    To evaluate the temporal relationship between blood flow, blood pressure, and muscle contractions, we continuously measured femoral arterial inflow with ultrasound Doppler at onset of passive exercise and voluntary, one-legged, dynamic knee-extensor exercise in humans. Blood velocity and inflow increased (P dicrotic and diastolic blood pressure notches, respectively. Mechanical hindrance occurred (P dicrotic notch. The increase in blood flow (Q) was characterized by a one-component (approximately 15% of peak power output), two-component (approximately 40-70% of peak power output), or three-component exponential model (> or = 75% of peak power output), where Q(t) = Qpassive + delta Q1.[1 - e-(t - TD1/tau 1)]+ delta Q2.[1 - e-(t - TD2/tau 2)]+ delta Q3.[1 - e-(t - TD3/tau 3)]; Qpassive, the blood flow during passive leg movement, equals 1.17 +/- 0.11 l/min; TD is the onset latency; tau is the time constant; delta Q is the magnitude of blood flow rise; and subscripts 1-3 refer to the first, second, and third components of the exponential model, respectively. The time to reach 50% of the difference between passive and voluntary asymptotic blood flow was approximately 2.2-8.9 s. The blood flow leveled off after approximately 10-150 s, related to the power outputs. It is concluded that the elevation in blood flow with the first duty cycle(s) is due to muscle mechanical factors, but vasodilators initiate a more potent amplification within the second to fourth contraction.

  20. Smartphone-Imaged HIV-1 Reverse-Transcription Loop-Mediated Isothermal Amplification (RT-LAMP on a Chip from Whole Blood

    Directory of Open Access Journals (Sweden)

    Gregory L. Damhorst

    2015-09-01

    Full Text Available Viral load measurements are an essential tool for the long-term clinical care of human immunodeficiency virus (HIV-positive individuals. The gold standards in viral load instrumentation, however, are still too limited by their size, cost, and sophisticated operation for these measurements to be ubiquitous in remote settings with poor healthcare infrastructure, including parts of the world that are disproportionately affected by HIV infection. The challenge of developing a point-of-care platform capable of making viral load more accessible has been frequently approached but no solution has yet emerged that meets the practical requirements of low cost, portability, and ease-of-use. In this paper, we perform reverse-transcription loop-mediated isothermal amplification (RT-LAMP on minimally processed HIV-spiked whole blood samples with a microfluidic and silicon microchip platform, and perform fluorescence measurements with a consumer smartphone. Our integrated assay shows amplification from as few as three viruses in a ~ 60 nL RT-LAMP droplet, corresponding to a whole blood concentration of 670 viruses per μL of whole blood. The technology contains greater power in a digital RT-LAMP approach that could be scaled up for the determination of viral load from a finger prick of blood in the clinical care of HIV-positive individuals. We demonstrate that all aspects of this viral load approach, from a drop of blood to imaging the RT-LAMP reaction, are compatible with lab-on-a-chip components and mobile instrumentation.

  1. Two distinct promoters drive transcription of the human D1A dopamine receptor gene.

    Science.gov (United States)

    Lee, S H; Minowa, M T; Mouradian, M M

    1996-10-11

    The human D1A dopamine receptor gene has a GC-rich, TATA-less promoter located upstream of a small, noncoding exon 1, which is separated from the coding exon 2 by a 116-base pair (bp)-long intron. Serial 3'-deletions of the 5'-noncoding region of this gene, including the intron and 5'-end of exon 2, resulted in 80 and 40% decrease in transcriptional activity of the upstream promoter in two D1A-expressing neuroblastoma cell lines, SK-N-MC and NS20Y, respectively. To investigate the function of this region, the intron and 245 bp at the 5'-end of exon 2 were investigated. Transient expression analyses using various chloramphenicol acetyltransferase constructs showed that the transcriptional activity of the intron is higher than that of the upstream promoter by 12-fold in SK-N-MC cells and by 5.5-fold in NS20Y cells in an orientation-dependent manner, indicating that the D1A intron is a strong promoter. Primer extension and ribonuclease protection assays revealed that transcription driven by the intron promoter is initiated at the junction of intron and exon 2 and at a cluster of nucleotides located 50 bp downstream from this junction. The same transcription start sites are utilized by the chloramphenicol acetyltransferase constructs employed in transfections as well as by the D1A gene expressed within the human caudate. The relative abundance of D1A transcripts originating from the upstream promoter compared with those transcribed from the intron promoter is 1.5-2.9 times in SK-N-MC cells and 2 times in the human caudate. Transcript stability studies in SK-N-MC cells revealed that longer D1A mRNA molecules containing exon 1 are degraded 1.8 times faster than shorter transcripts lacking exon 1. Although gel mobility shift assay could not detect DNA-protein interaction at the D1A intron, competitive co-transfection using the intron as competitor confirmed the presence of trans-acting factors at the intron. These data taken together indicate that the human D1A gene has

  2. Blood-group-Ii-active gangliosides of human erythrocyte membranes

    International Nuclear Information System (INIS)

    Feizi, T.; Childs, R.A.; Hakomori, S.-I.; Powell, M.E.

    1978-01-01

    More than ten new types of gangliosides, in addition to haematoside and sialosylparagloboside, were isolated from human erythrocyte membranes. These were separated by successive chromatographies on DAEA-Sephadex, on porous silica-gel columns and on thin-layer silica gel as acetylated compounds. Highly potent blood-group-Ii and moderate blood-group-H activities were demonstrated in some of the ganglioside fractions. The gangliosides incorporated into chlolesterol/phosphatidylcholine liposomes stoicheiometrically inhibited binding of anti-(blood-group-I and i) antibodies to a radioiodinated blood-group-Ii-active glycoprotein. The fraction with the highest blood-group-I activity, I(g) fraction, behaved like sialosyl-deca- to dodeca-glycosylceramides on t.l.c. Certain blood-group-I and most of the i-determinants were in partially or completely cryptic form and could be unmasked by sialidase treatment. Thus the I and i antigens, which are known to occur on internal structures of blood-group-ABH-active glycoproteins in secretions, also occur in the interior of the carbohydrate chains of erythrocyte gangliosides. (author)

  3. Regulation of endogenous human gene expression by ligand-inducible TALE transcription factors.

    Science.gov (United States)

    Mercer, Andrew C; Gaj, Thomas; Sirk, Shannon J; Lamb, Brian M; Barbas, Carlos F

    2014-10-17

    The construction of increasingly sophisticated synthetic biological circuits is dependent on the development of extensible tools capable of providing specific control of gene expression in eukaryotic cells. Here, we describe a new class of synthetic transcription factors that activate gene expression in response to extracellular chemical stimuli. These inducible activators consist of customizable transcription activator-like effector (TALE) proteins combined with steroid hormone receptor ligand-binding domains. We demonstrate that these ligand-responsive TALE transcription factors allow for tunable and conditional control of gene activation and can be used to regulate the expression of endogenous genes in human cells. Since TALEs can be designed to recognize any contiguous DNA sequence, the conditional gene regulatory system described herein will enable the design of advanced synthetic gene networks.

  4. Discovery of a Regulatory Motif for Human Satellite DNA Transcription in Response to BATF2 Overexpression.

    Science.gov (United States)

    Bai, Xuejia; Huang, Wenqiu; Zhang, Chenguang; Niu, Jing; Ding, Wei

    2016-03-01

    One of the basic leucine zipper transcription factors, BATF2, has been found to suppress cancer growth and migration. However, little is known about the genes downstream of BATF2. HeLa cells were stably transfected with BATF2, then chromatin immunoprecipitation-sequencing was employed to identify the DNA motifs responsive to BATF2. Comprehensive bioinformatics analyses indicated that the most significant motif discovered as TTCCATT[CT]GATTCCATTC[AG]AT was primarily distributed among the chromosome centromere regions and mostly within human type II satellite DNA. Such motifs were able to prime the transcription of type II satellite DNA in a directional and asymmetrical manner. Consistently, satellite II transcription was up-regulated in BATF2-overexpressing cells. The present study provides insight into understanding the role of BATF2 in tumours and the importance of satellite DNA in the maintenance of genomic stability. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  5. Genotoxic damage in cultured human peripheral blood lymphocytes ...

    African Journals Online (AJOL)

    Falaq Naz

    2012-06-29

    Jun 29, 2012 ... Genotoxic damage in cultured human peripheral blood lymphocytes of oral ... catechol estrogens and quinines, via redox reactions causes oxidative damage to .... volume was prepared for each donor. About, 0.8 ml of cell sus .... duce the adverse effects of OCs, such as the reduction in the estrogen content.

  6. Sympathetic regulation of cerebral blood flow in humans : a review

    NARCIS (Netherlands)

    ter Laan, M.; van Dijk, J. M. C.; Elting, J. W. J.; Staal, M. J.; Absalom, A. R.

    Cerebral blood flow (CBF) is regulated by vasomotor, chemical, metabolic, and neurogenic mechanisms. Even though the innervation of cerebral arteries is quite extensively described and reviewed in the literature, its role in regulation of CBF in humans remains controversial. We believe that

  7. Detecting multiple DNA human profile from a mosquito blood meal.

    Science.gov (United States)

    Rabêlo, K C N; Albuquerque, C M R; Tavares, V B; Santos, S M; Souza, C A; Oliveira, T C; Moura, R R; Brandão, L A C; Crovella, S

    2016-08-26

    Criminal traces commonly found at crime scenes may present mixtures from two or more individuals. The scene of the crime is important for the collection of various types of traces in order to find the perpetrator of the crime. Thus, we propose that hematophagous mosquitoes found at crime scenes can be used to perform genetic testing of human blood and aid in suspect investigation. The aim of the study was to obtain a single Aedes aegypti mosquito profile from a human DNA mixture containing genetic materials of four individuals. We also determined the effect of blood acquisition time by setting time intervals of 24, 48, and 72 h after the blood meal. STR loci and amelogenin were analyzed, and the results showed that human DNA profiles could be obtained from hematophagous mosquitos at 24 h following the blood meal. It is possible that hematophagous mosquitoes can be used as biological remains at the scene of the crime, and can be used to detect human DNA profiles of up to four individuals.

  8. Generating autologous hematopoietic cells from human-induced pluripotent stem cells through ectopic expression of transcription factors.

    Science.gov (United States)

    Hwang, Yongsung; Broxmeyer, Hal E; Lee, Man Ryul

    2017-07-01

    Hematopoietic cell transplantation (HCT) is a successful treatment modality for patients with malignant and nonmalignant disorders, usually when no other treatment option is available. The cells supporting long-term reconstitution after HCT are the hematopoietic stem cells (HSCs), which can be limited in numbers. Moreover, finding an appropriate human leukocyte antigen-matched donor can be problematic. If HSCs can be stably produced in large numbers from autologous or allogeneic cell sources, it would benefit HCT. Induced pluripotent stem cells (iPSCs) established from patients' own somatic cells can be differentiated into hematopoietic cells in vitro. This review will highlight recent methods for regulating human (h) iPSC production of HSCs and more mature blood cells. Advancements in transcription factor-mediated regulation of the developmental stages of in-vivo hematopoietic lineage commitment have begun to provide an understanding of the molecular mechanism of hematopoiesis. Such studies involve not only directed differentiation in which transcription factors, specifically expressed in hematopoietic lineage-specific cells, are overexpressed in iPSCs, but also direct conversion in which transcription factors are introduced into patient-derived somatic cells which are dedifferentiated to hematopoietic cells. As iPSCs derived from patients suffering from genetically mutated diseases would express the same mutated genetic information, CRISPR-Cas9 gene editing has been utilized to differentiate genetically corrected iPSCs into normal hematopoietic cells. IPSCs provide a model for molecular understanding of disease, and also may function as a cell population for therapy. Efficient differentiation of patient-specific iPSCs into HSCs and progenitor cells is a potential means to overcome limitations of such cells for HCT, as well as for providing in-vitro drug screening templates as tissue-on-a-chip models.

  9. Assessing quality and completeness of human transcriptional regulatory pathways on a genome-wide scale

    Directory of Open Access Journals (Sweden)

    Aifantis Iannis

    2011-02-01

    Full Text Available Abstract Background Pathway databases are becoming increasingly important and almost omnipresent in most types of biological and translational research. However, little is known about the quality and completeness of pathways stored in these databases. The present study conducts a comprehensive assessment of transcriptional regulatory pathways in humans for seven well-studied transcription factors: MYC, NOTCH1, BCL6, TP53, AR, STAT1, and RELA. The employed benchmarking methodology first involves integrating genome-wide binding with functional gene expression data to derive direct targets of transcription factors. Then the lists of experimentally obtained direct targets are compared with relevant lists of transcriptional targets from 10 commonly used pathway databases. Results The results of this study show that for the majority of pathway databases, the overlap between experimentally obtained target genes and targets reported in transcriptional regulatory pathway databases is surprisingly small and often is not statistically significant. The only exception is MetaCore pathway database which yields statistically significant intersection with experimental results in 84% cases. Additionally, we suggest that the lists of experimentally derived direct targets obtained in this study can be used to reveal new biological insight in transcriptional regulation and suggest novel putative therapeutic targets in cancer. Conclusions Our study opens a debate on validity of using many popular pathway databases to obtain transcriptional regulatory targets. We conclude that the choice of pathway databases should be informed by solid scientific evidence and rigorous empirical evaluation. Reviewers This article was reviewed by Prof. Wing Hung Wong, Dr. Thiago Motta Venancio (nominated by Dr. L Aravind, and Prof. Geoff J McLachlan.

  10. The Inflammatory Transcription Factors NFκB, STAT1 and STAT3 Drive Age-Associated Transcriptional Changes in the Human Kidney

    Science.gov (United States)

    O’Brown, Zach K.; Van Nostrand, Eric L.; Higgins, John P.; Kim, Stuart K.

    2015-01-01

    Human kidney function declines with age, accompanied by stereotyped changes in gene expression and histopathology, but the mechanisms underlying these changes are largely unknown. To identify potential regulators of kidney aging, we compared age-associated transcriptional changes in the human kidney with genome-wide maps of transcription factor occupancy from ChIP-seq datasets in human cells. The strongest candidates were the inflammation-associated transcription factors NFκB, STAT1 and STAT3, the activities of which increase with age in epithelial compartments of the renal cortex. Stimulation of renal tubular epithelial cells with the inflammatory cytokines IL-6 (a STAT3 activator), IFNγ (a STAT1 activator), or TNFα (an NFκB activator) recapitulated age-associated gene expression changes. We show that common DNA variants in RELA and NFKB1, the two genes encoding subunits of the NFκB transcription factor, associate with kidney function and chronic kidney disease in gene association studies, providing the first evidence that genetic variation in NFκB contributes to renal aging phenotypes. Our results suggest that NFκB, STAT1 and STAT3 underlie transcriptional changes and chronic inflammation in the aging human kidney. PMID:26678048

  11. Distinguishing the Transcription Regulation Patterns in Promoters of Human Genes with Different Function or Evolutionary Age

    KAUST Repository

    Alam, Tanvir

    2012-07-01

    Distinguishing transcription regulatory patterns of different gene groups is a common problem in various bioinformatics studies. In this work we developed a methodology to deal with such a problem based on machine learning techniques. We applied our method to two biologically important problems related to detecting a difference in transcription regulation of: a/ protein-coding and long non-coding RNAs (lncRNAs) in human, as well as b/ a difference between primate-specific and non-primate-specific long non-coding RNAs. Our method is capable to classify RNAs using various regulatory features of genes that transcribe into these RNAs, such as nucleotide frequencies, transcription factor binding sites, de novo sequence motifs, CpG islands, repetitive elements, histone modification marks, and others. Ten-fold cross-validation tests suggest that our model can distinguish protein-coding and non-coding RNAs with accuracy above 80%. Twenty-fold cross-validation tests suggest that our model can distinguish primate-specific from non-primate-specific promoters of lncRNAs with accuracy above 80%. Consequently, we can hypothesize that transcription of the groups of genes mentioned above are regulated by different mechanisms. Feature selection techniques allowed us to reduce the number of features significantly while keeping the accuracy around 80%. Consequently, we can conclude that selected features play significant role in transcription regulation of coding and non-coding genes, as well as primate-specific and non-primate-specific lncRNA genes.

  12. Human GW182 Paralogs Are the Central Organizers for RNA-Mediated Control of Transcription.

    Science.gov (United States)

    Hicks, Jessica A; Li, Liande; Matsui, Masayuki; Chu, Yongjun; Volkov, Oleg; Johnson, Krystal C; Corey, David R

    2017-08-15

    In the cytoplasm, small RNAs can control mammalian translation by regulating the stability of mRNA. In the nucleus, small RNAs can also control transcription and splicing. The mechanisms for RNA-mediated nuclear regulation are not understood and remain controversial, hindering the effective application of nuclear RNAi and investigation of its natural regulatory roles. Here, we reveal that the human GW182 paralogs TNRC6A/B/C are central organizing factors critical to RNA-mediated transcriptional activation. Mass spectrometry of purified nuclear lysates followed by experimental validation demonstrates that TNRC6A interacts with proteins involved in protein degradation, RNAi, the CCR4-NOT complex, the mediator complex, and histone-modifying complexes. Functional analysis implicates TNRC6A, NAT10, MED14, and WDR5 in RNA-mediated transcriptional activation. These findings describe protein complexes capable of bridging RNA-mediated sequence-specific recognition of noncoding RNA transcripts with the regulation of gene transcription. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  13. A network of paralogous stress response transcription factors in the human pathogen Candida glabrata.

    Directory of Open Access Journals (Sweden)

    Jawad eMerhej

    2016-05-01

    Full Text Available The yeast Candida glabrata has become the second cause of systemic candidemia in humans. However, relatively few genome-wide studies have been conducted in this organism and our knowledge of its transcriptional regulatory network is quite limited. In the present work, we combined genome-wide chromatin immunoprecipitation (ChIP-seq, transcriptome analyses and DNA binding motif predictions to describe the regulatory interactions of the seven Yap (Yeast AP1 transcription factors of C. glabrata. We described a transcriptional network containing 255 regulatory interactions and 309 potential target genes. We predicted with high confidence the preferred DNA binding sites for 5 of the 7 CgYaps and showed a strong conservation of the Yap DNA binding properties between S. cerevisiae and C. glabrata. We provided reliable functional annotation for 3 of the 7 Yaps and identified for Yap1 and Yap5 a core regulon which is conserved in S. cerevisiae, C. glabrata and C. albicans. We uncovered new roles for CgYap7 in the regulation of iron-sulfur cluster biogenesis, for CgYap1 in the regulation of heme biosynthesis and for CgYap5 in the repression of GRX4 in response to iron starvation. These transcription factors define an interconnected transcriptional network at the cross-roads between redox homeostasis, oxygen consumption and iron metabolism.

  14. Transcriptional profiling of five isolated size-matched stages of human preantral follicles

    DEFF Research Database (Denmark)

    Kristensen, Stine Gry; Ebbesen, Pernille; Andersen, Claus Yding

    2015-01-01

    Little is known of the early stages of human follicular development and the complex processes that regulate follicular growth. To identify genes of potential importance, we analysed follicle-related transcripts in five populations of isolated size-matched human preantral follicles by microarray...... factors of NOTCH signalling, IGF2, orphan nuclear receptor LRH-1, and homeobox gene HOXA7, indicating potentially important regulatory roles for these genes during early human folliculogenesis. We also found that FSHR mRNA and protein were present in the earliest stages of preantral follicles, whereas LHR...

  15. TcoF-DB v2: update of the database of human and mouse transcription co-factors and transcription factor interactions

    KAUST Repository

    Schmeier, Sebastian

    2016-10-17

    Transcription factors (TFs) play a pivotal role in transcriptional regulation, making them crucial for cell survival and important biological functions. For the regulation of transcription, interactions of different regulatory proteins known as transcription co-factors (TcoFs) and TFs are essential in forming necessary protein complexes. Although TcoFs themselves do not bind DNA directly, their influence on transcriptional regulation and initiation, although indirect, has been shown to be significant, with the functionality of TFs strongly influenced by the presence of TcoFs. In the TcoF-DB v2 database, we collect information on TcoFs. In this article, we describe updates and improvements implemented in TcoF-DB v2. TcoF-DB v2 provides several new features that enables exploration of the roles of TcoFs. The content of the database has significantly expanded, and is enriched with information from Gene Ontology, biological pathways, diseases and molecular signatures. TcoF-DB v2 now includes many more TFs; has substantially increased the number of human TcoFs to 958, and now includes information on mouse (418 new TcoFs). TcoF-DB v2 enables the exploration of information on TcoFs and allows investigations into their influence on transcriptional regulation in humans and mice. TcoF-DB v2 can be accessed at http://tcofdb.org/.

  16. TcoF-DB v2: update of the database of human and mouse transcription co-factors and transcription factor interactions

    KAUST Repository

    Schmeier, Sebastian; Alam, Tanvir; Essack, Magbubah; Bajic, Vladimir B.

    2016-01-01

    Transcription factors (TFs) play a pivotal role in transcriptional regulation, making them crucial for cell survival and important biological functions. For the regulation of transcription, interactions of different regulatory proteins known as transcription co-factors (TcoFs) and TFs are essential in forming necessary protein complexes. Although TcoFs themselves do not bind DNA directly, their influence on transcriptional regulation and initiation, although indirect, has been shown to be significant, with the functionality of TFs strongly influenced by the presence of TcoFs. In the TcoF-DB v2 database, we collect information on TcoFs. In this article, we describe updates and improvements implemented in TcoF-DB v2. TcoF-DB v2 provides several new features that enables exploration of the roles of TcoFs. The content of the database has significantly expanded, and is enriched with information from Gene Ontology, biological pathways, diseases and molecular signatures. TcoF-DB v2 now includes many more TFs; has substantially increased the number of human TcoFs to 958, and now includes information on mouse (418 new TcoFs). TcoF-DB v2 enables the exploration of information on TcoFs and allows investigations into their influence on transcriptional regulation in humans and mice. TcoF-DB v2 can be accessed at http://tcofdb.org/.

  17. BloodSpot: a database of gene expression profiles and transcriptional programs for healthy and malignant haematopoiesis

    DEFF Research Database (Denmark)

    Bagger, Frederik Otzen; Sasivarevic, Damir; Hadi Sohi, Sina

    2016-01-01

    Research on human and murine haematopoiesis has resulted in a vast number of gene-expression data sets that can potentially answer questions regarding normal and aberrant blood formation. To researchers and clinicians with limited bioinformatics experience, these data have remained available, yet...

  18. Ventricular transcriptional data provide mechanistic insights into diesel exhaust induced attenuation of cardiac contractile response and blood pressure

    Science.gov (United States)

    Human exposures to near road ambient particulate matter and its major component, diesel exhaust (DE), have been associated with cardiovascular impairments however the mechanisms and the role of hypertension are not well understood. We have shown that DE exposure reduces blood pre...

  19. Sp1 and CREB regulate basal transcription of the human SNF2L gene

    International Nuclear Information System (INIS)

    Xia Yu; Jiang Baichun; Zou Yongxin; Gao Guimin; Shang Linshan; Chen Bingxi; Liu Qiji; Gong Yaoqin

    2008-01-01

    Imitation Switch (ISWI) is a member of the SWI2/SNF2 superfamily of ATP-dependent chromatin remodelers, which are involved in multiple nuclear functions, including transcriptional regulation, replication, and chromatin assembly. Mammalian genomes encode two ISWI orthologs, SNF2H and SNF2L. In order to clarify the molecular mechanisms governing the expression of human SNF2L gene, we functionally examined the transcriptional regulation of human SNF2L promoter. Reporter gene assays demonstrated that the minimal SNF2L promoter was located between positions -152 to -86 relative to the transcription start site. In this region we have identified a cAMP-response element (CRE) located at -99 to -92 and a Sp1-binding site at -145 to -135 that play a critical role in regulating basal activity of human SNF2L gene, which were proven by deletion and mutation of specific binding sites, EMSA, and down-regulating Sp1 and CREB via RNAi. This study provides the first insight into the mechanisms that control basal expression of human SNF2L gene

  20. Bisphenol A and Bisphenol S Induce Distinct Transcriptional Profiles in Differentiating Human Primary Preadipocytes.

    Directory of Open Access Journals (Sweden)

    Jonathan G Boucher

    Full Text Available Bisphenol S (BPS is increasingly used as a replacement plasticizer for bisphenol A (BPA but its effects on human health have not been thoroughly examined. Recent evidence indicates that both BPA and BPS induce adipogenesis, although the mechanisms leading to this effect are unclear. In an effort to identify common and distinct mechanisms of action in inducing adipogenesis, transcriptional profiles of differentiating human preadipocytes exposed to BPA or BPS were compared. Human subcutaneous primary preadipocytes were differentiated in the presence of either 25 μM BPA or BPS for 2 and 4 days. Poly-A RNA-sequencing was used to identify differentially expressed genes (DEGs. Functional analysis of DEGs was undertaken in Ingenuity Pathway Analysis. BPA-treatment resulted in 472 and 176 DEGs on days 2 and 4, respectively, affecting pathways such as liver X receptor (LXR/retinoid X receptor (RXR activation, hepatic fibrosis and cholestasis. BPS-treatment resulted in 195 and 51 DEGs on days 2 and 4, respectively, revealing enrichment of genes associated with adipogenesis and lipid metabolism including the adipogenesis pathway and cholesterol biosynthesis. Interestingly, the transcription repressor N-CoR was identified as a negative upstream regulator in both BPA- and BPS-treated cells. This study presents the first comparison of BPA- and BPS-induced transcriptional profiles in human differentiating preadipocytes. While we previously showed that BPA and BPS both induce adipogenesis, the results from this study show that BPS affects adipose specific transcriptional changes earlier than BPA, and alters the expression of genes specifically related to adipogenesis and lipid metabolism. The findings provide insight into potential BPS and BPA-mediated mechanisms of action in inducing adipogenesis in human primary preadipocytes.

  1. The DNA methylome of human peripheral blood mononuclear cells

    DEFF Research Database (Denmark)

    Li, Yingrui; Zhu, Jingde; Tian, Geng

    2010-01-01

    DNA methylation plays an important role in biological processes in human health and disease. Recent technological advances allow unbiased whole-genome DNA methylation (methylome) analysis to be carried out on human cells. Using whole-genome bisulfite sequencing at 24.7-fold coverage (12.3-fold per...... strand), we report a comprehensive (92.62%) methylome and analysis of the unique sequences in human peripheral blood mononuclear cells (PBMC) from the same Asian individual whose genome was deciphered in the YH project. PBMC constitute an important source for clinical blood tests world-wide. We found...... research and confirms new sequencing technology as a paradigm for large-scale epigenomics studies....

  2. Pleomorphic Structures in Human Blood Are Red Blood Cell-Derived Microparticles, Not Bacteria.

    Science.gov (United States)

    Mitchell, Adam J; Gray, Warren D; Schroeder, Max; Yi, Hong; Taylor, Jeannette V; Dillard, Rebecca S; Ke, Zunlong; Wright, Elizabeth R; Stephens, David; Roback, John D; Searles, Charles D

    2016-01-01

    Red blood cell (RBC) transfusions are a common, life-saving therapy for many patients, but they have also been associated with poor clinical outcomes. We identified unusual, pleomorphic structures in human RBC transfusion units by negative-stain electron microscopy that appeared identical to those previously reported to be bacteria in healthy human blood samples. The presence of viable, replicating bacteria in stored blood could explain poor outcomes in transfusion recipients and have major implications for transfusion medicine. Here, we investigated the possibility that these structures were bacteria. Flow cytometry, miRNA analysis, protein analysis, and additional electron microscopy studies strongly indicated that the pleomorphic structures in the supernatant of stored RBCs were RBC-derived microparticles (RMPs). Bacterial 16S rDNA PCR amplified from these samples were sequenced and was found to be highly similar to species that are known to commonly contaminate laboratory reagents. These studies suggest that pleomorphic structures identified in human blood are RMPs and not bacteria, and they provide an example in which laboratory contaminants may can mislead investigators.

  3. Sb(V reactivity with human blood components: redox effects.

    Directory of Open Access Journals (Sweden)

    Silvana López

    Full Text Available We assessed the reactivity of Sb(V in human blood. Sb(V reactivity was determined using an HPLC-HG-AFS hyphenated system. Sb(V was partially reduced to Sb(III in blood incubation experiments; however, Sb(III was a highly unstable species. The addition of 0.1 mol L(-1 EDTA prevented Sb(III oxidation, thus enabling the detection of the reduction of Sb(V to Sb(III. The transformation of Sb(V to Sb(III in human whole blood was assessed because the reduction of Sb(V in human blood may likely generate redox side effects. Our results indicate that glutathione was the reducing agent in this reaction and that Sb(V significantly decreased the GSH/GSSG ratio from 0.32 ± 0.09 to 0.07 ± 0.03. Moreover, the presence of 200 ng mL(-1 of Sb(V increased the activity of superoxide dismutase from 4.4 ± 0.1 to 7.0 ± 0.4 U mL(-1 and decreased the activity of glutathione peroxidase from 62 ± 1 to 34 ± 2 nmol min(-1 mL(-1.

  4. Empirical modelling to predict the refractive index of human blood

    Science.gov (United States)

    Yahya, M.; Saghir, M. Z.

    2016-02-01

    Optical techniques used for the measurement of the optical properties of blood are of great interest in clinical diagnostics. Blood analysis is a routine procedure used in medical diagnostics to confirm a patient’s condition. Measuring the optical properties of blood is difficult due to the non-homogenous nature of the blood itself. In addition, there is a lot of variation in the refractive indices reported in the literature. These are the reasons that motivated the researchers to develop a mathematical model that can be used to predict the refractive index of human blood as a function of concentration, temperature and wavelength. The experimental measurements were conducted on mimicking phantom hemoglobin samples using the Abbemat Refractometer. The results analysis revealed a linear relationship between the refractive index and concentration as well as temperature, and a non-linear relationship between refractive index and wavelength. These results are in agreement with those found in the literature. In addition, a new formula was developed based on empirical modelling which suggests that temperature and wavelength coefficients be added to the Barer formula. The verification of this correlation confirmed its ability to determine refractive index and/or blood hematocrit values with appropriate clinical accuracy.

  5. Empirical modelling to predict the refractive index of human blood

    International Nuclear Information System (INIS)

    Yahya, M; Saghir, M Z

    2016-01-01

    Optical techniques used for the measurement of the optical properties of blood are of great interest in clinical diagnostics. Blood analysis is a routine procedure used in medical diagnostics to confirm a patient’s condition. Measuring the optical properties of blood is difficult due to the non-homogenous nature of the blood itself. In addition, there is a lot of variation in the refractive indices reported in the literature. These are the reasons that motivated the researchers to develop a mathematical model that can be used to predict the refractive index of human blood as a function of concentration, temperature and wavelength. The experimental measurements were conducted on mimicking phantom hemoglobin samples using the Abbemat Refractometer. The results analysis revealed a linear relationship between the refractive index and concentration as well as temperature, and a non-linear relationship between refractive index and wavelength. These results are in agreement with those found in the literature. In addition, a new formula was developed based on empirical modelling which suggests that temperature and wavelength coefficients be added to the Barer formula. The verification of this correlation confirmed its ability to determine refractive index and/or blood hematocrit values with appropriate clinical accuracy. (paper)

  6. Longitudinal peripheral blood transcriptional analysis of a patient with severe Ebola virus disease.

    Science.gov (United States)

    Kash, John C; Walters, Kathie-Anne; Kindrachuk, Jason; Baxter, David; Scherler, Kelsey; Janosko, Krisztina B; Adams, Rick D; Herbert, Andrew S; James, Rebekah M; Stonier, Spencer W; Memoli, Matthew J; Dye, John M; Davey, Richard T; Chertow, Daniel S; Taubenberger, Jeffery K

    2017-04-12

    The 2013-2015 outbreak of Ebola virus disease in Guinea, Liberia, and Sierra Leone was unprecedented in the number of documented cases, but there have been few published reports on immune responses in clinical cases and their relationships with the course of illness and severity of Ebola virus disease. Symptoms of Ebola virus disease can include severe headache, myalgia, asthenia, fever, fatigue, diarrhea, vomiting, abdominal pain, and hemorrhage. Although experimental treatments are in development, there are no current U.S. Food and Drug Administration-approved vaccines or therapies. We report a detailed study of host gene expression as measured by microarray in daily peripheral blood samples collected from a patient with severe Ebola virus disease. This individual was provided with supportive care without experimental therapies at the National Institutes of Health Clinical Center from before onset of critical illness to recovery. Pearson analysis of daily gene expression signatures revealed marked gene expression changes in peripheral blood leukocytes that correlated with changes in serum and peripheral blood leukocytes, viral load, antibody responses, coagulopathy, multiple organ dysfunction, and then recovery. This study revealed marked shifts in immune and antiviral responses that preceded changes in medical condition, indicating that clearance of replicating Ebola virus from peripheral blood leukocytes is likely important for systemic viral clearance. Copyright © 2017, American Association for the Advancement of Science.

  7. Profiling of exercise-induced transcripts in the peripheral blood cells of Thoroughbred horses.

    Science.gov (United States)

    Tozaki, Teruaki; Kikuchi, Mio; Kakoi, Hironaga; Hirota, Kei-Ichi; Mukai, Kazutaka; Aida, Hiroko; Nakamura, Seiji; Nagata, Shun-Ichi

    2016-01-01

    Transcriptome analyses based on DNA microarray technology have been used to investigate gene expression profiles in horses. In this study, we aimed to identify exercise-induced changes in the expression profiles of genes in the peripheral blood of Thoroughbred horses using DNA microarray technology (15,429 genes on 43,603 probes). Blood samples from the jugular vein were collected from six horses before and 1 min, 4 hr, and 24 hr after all-out running on a treadmill. After the normalization of microarray data, a total of 26,830 probes were clustered into four groups and 11 subgroups showing similar expression changes based on k-mean clustering. The expression level of inflammation-related genes, including interleukin-1 receptor type II (IL-1R2), matrix metallopeptidase 8 (MMP8), protein S100-A8 (S100-A8), and serum amyloid A (SAA), increased at 4 hr after exercise, whereas that of c-Fos (FOS) increased at 1 min after exercise. These results indicated that the inflammatory response increased in the peripheral blood cells after exercise. Our study also revealed the presence of genes that may not be affected by all-out exercise. In conclusion, transcriptome analysis of peripheral blood cells could be used to monitor physiological changes induced by various external stress factors, including exercise, in Thoroughbred racehorses.

  8. Acrolein generation stimulates hypercontraction in isolated human blood vessels

    International Nuclear Information System (INIS)

    Conklin, D.J.; Bhatnagar, A.; Cowley, H.R.; Johnson, G.H.; Wiechmann, R.J.; Sayre, L.M.; Trent, M.B.; Boor, P.J.

    2006-01-01

    Increased risk of vasospasm, a spontaneous hyperconstriction, is associated with atherosclerosis, cigarette smoking, and hypertension-all conditions involving oxidative stress, lipid peroxidation, and inflammation. To test the role of the lipid peroxidation- and inflammation-derived aldehyde, acrolein, in human vasospasm, we developed an ex vivo model using human coronary artery bypass graft (CABG) blood vessels and a demonstrated acrolein precursor, allylamine. Allylamine induces hypercontraction in isolated rat coronary artery in a semicarbazide-sensitive amine oxidase activity (SSAO) dependent manner. Isolated human CABG blood vessels (internal mammary artery, radial artery, saphenous vein) were used to determine: (1) vessel responses and sensitivity to acrolein, allylamine, and H 2 O 2 exposure (1 μM-1 mM), (2) SSAO dependence of allylamine-induced effects using SSAO inhibitors (semicarbazide, 1 mM; MDL 72274-E, active isomer; MDL 72274-Z, inactive isomer; 100 μM), (3) the vasoactive effects of two other SSAO amine substrates, benzylamine and methylamine, and (4) the contribution of extracellular Ca 2+ to hypercontraction. Acrolein or allylamine but not H 2 O 2 , benzylamine, or methylamine stimulated spontaneous and pharmacologically intractable hypercontraction in CABG blood vessels that was similar to clinical vasospasm. Allylamine-induced hypercontraction and blood vessel SSAO activity were abolished by pretreatment with semicarbazide or MDL 72274-E but not by MDL 72274-Z. Allylamine-induced hypercontraction also was significantly attenuated in Ca 2+ -free buffer. In isolated aorta of spontaneously hypertensive rat, allylamine-induced an SSAO-dependent contraction and enhanced norepinephrine sensitivity but not in Sprague-Dawley rat aorta. We conclude that acrolein generation in the blood vessel wall increases human susceptibility to vasospasm, an event that is enhanced in hypertension

  9. Selective activation of human heat shock gene transcription by nitrosourea antitumor drugs mediated by isocyanate-induced damage and activation of heat shock transcription factor.

    Science.gov (United States)

    Kroes, R A; Abravaya, K; Seidenfeld, J; Morimoto, R I

    1991-01-01

    Treatment of cultured human tumor cells with the chloroethylnitrosourea antitumor drug 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) selectively induces transcription and protein synthesis of a subset of the human heat shock or stress-induced genes (HSP90 and HSP70) with little effect on other stress genes or on expression of the c-fos, c-myc, or beta-actin genes. The active component of BCNU and related compounds appears to be the isocyanate moiety that causes carbamoylation of proteins and nucleic acids. Transcriptional activation of the human HSP70 gene by BCNU is dependent on the heat shock element and correlates with the level of heat shock transcription factor and its binding to the heat shock element in vivo. Unlike activation by heat or heavy metals, BCNU-mediated activation is strongly dependent upon new protein synthesis. This suggests that BCNU-induced, isocyanate-mediated damage to newly synthesized protein(s) may be responsible for activation of the heat shock transcription factor and increased transcription of the HSP90 and HSP70 genes. Images PMID:2052560

  10. Elevated expression and potential roles of human Sp5, a member of Sp transcription factor family, in human cancers

    International Nuclear Information System (INIS)

    Chen Yongxin; Guo Yingqiu; Ge Xijin; Itoh, Hirotaka; Watanabe, Akira; Fujiwara, Takeshi; Kodama, Tatsuhiko; Aburatani, Hiroyuki

    2006-01-01

    In this report, we describe the expression and function of human Sp5, a member of the Sp family of zinc finger transcription factors. Like other family members, the Sp5 protein contains a Cys2His2 zinc finger DNA binding domain at the C-terminus. Our experiments employing Gal4-Sp5 fusion proteins reveal multiple transcriptional domains, including a N-terminal activity domain, an intrinsic repressive element, and a C-terminal synergistic domain. Elevated expression of Sp5 was noted in several human tumors including hepatocellular carcinoma, gastric cancer, and colon cancer. To study the effects of the Sp5 protein on growth properties of human cancer cells and facilitate the identification of its downstream genes, we combined an inducible gene expression system with microarray analysis to screen for its transcriptional targets. Transfer of Sp5 into MCF-7 cells that expressed no detectable endogenous Sp5 protein elicited significant growth promotion activity. Several of the constitutively deregulated genes have been associated with tumorigenesis (CDC25C, CEACAM6, TMPRSS2, XBP1, MYBL1, ABHD2, and CXCL12) and Wnt/β-Catenin signaling pathways (BAMBI, SIX1, IGFBP5, AES, and p21 WAF1 ). This information could be utilized for further mechanistic research and for devising optimized therapeutic strategies against human cancers

  11. Transcriptional and post-transcriptional upregulation of p27 mediates growth inhibition of isorhapontigenin (ISO) on human bladder cancer cells.

    Science.gov (United States)

    Jiang, Guosong; Huang, Chao; Li, Jingxia; Huang, Haishan; Wang, Jingjing; Li, Yawei; Xie, Fei; Jin, Honglei; Zhu, Junlan; Huang, Chuanshu

    2018-03-08

    There are few approved drugs available for the treatment of muscle-invasive bladder cancer (MIBC). Recently, we have demonstrated that isorhapontigenin (ISO), a new derivative isolated from the Chinese herb Gnetum cleistostachyum, effectively induces cell-cycle arrest at the G0/G1 phase and inhibits anchorage-independent cell growth through the miR-137/Sp1/cyclin D1 axis in human MIBC cells. Herein, we found that treatment of bladder cancer (BC) cells with ISO resulted in a significant upregulation of p27, which was also observed in ISO-treated mouse BCs that were induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). Importantly, knockdown of p27 caused a decline in the ISO-induced G0-G1 growth arrest and reversed ISO suppression of anchorage-independent growth in BC cells. Mechanistic studies revealed that ISO promoted p27 expression at mRNA transcription level through increasing direct binding of forkhead box class O1 (FOXO1) to its promoter, while knockdown of FOXO1 attenuated ISO inhibition of BC cell growth. On the other hand, ISO upregulated the 3'-untranslated region (3'-UTR) activity of p27, which was accompanied by a reduction of miR-182 expression. In line with these observations, ectopic expression of miR-182 significantly blocked p27 3'-UTR activity, whereas mutation of the miR-182-binding site at p27 mRNA 3'-UTR effectively reversed this inhibition. Accordingly, ectopic expression of miR-182 also attenuated ISO upregulation of p27 expression and impaired ISO inhibition of BC cell growth. Our results not only provide novel insight into understanding of the underlying mechanism related to regulation of MIBC cell growth but also identify new roles and mechanisms underlying ISO inhibition of BC cell growth.

  12. Human acrocentric chromosomes with transcriptionally silent nucleolar organizer regions associate with nucleoli

    OpenAIRE

    Sullivan, Gareth J.; Bridger, Joanna M.; Cuthbert, Andrew P.; Newbold, Robert F.; Bickmore, Wendy A.; McStay, Brian

    2001-01-01

    Human ribosomal gene repeats are distributed among five nucleolar organizer regions (NORs) on the p arms of acrocentric chromosomes. On exit from mitosis, nucleoli form around individual active NORs. As cells progress through the cycle, these mini-nucleoli fuse to form large nucleoli incorporating multiple NORs. It is generally assumed that nucleolar incorporation of individual NORs is dependent on ribosomal gene transcription. To test this assumption, we determined the nuclear location of in...

  13. Silencing of human T-cell leukemia virus type I gene transcription by epigenetic mechanisms

    Directory of Open Access Journals (Sweden)

    Mueller Nancy

    2005-10-01

    Full Text Available Abstract Background Human T-cell leukemia virus type I (HTLV-I causes adult T-cell leukemia (ATL after a long latent period. Among accessory genes encoded by HTLV-I, the tax gene is thought to play a central role in oncogenesis. However, Tax expression is disrupted by several mechanims including genetic changes of the tax gene, deletion/hypermethylation of 5'-LTR. To clarify the role of epigenetic changes, we analyzed DNA methylation and histone modification in the whole HTLV-I provirus genome. Results The gag, pol and env genes of HTLV-I provirus were more methylated than pX region, whereas methylation of 5'-LTR was variable and 3'-LTR was not methylated at all. In ATL cell lines, complete DNA methylation of 5'-LTR was associated with transcriptional silencing of viral genes. HTLV-I provirus was more methylated in primary ATL cells than in carrier state, indicating the association with disease progression. In seroconvertors, DNA methylation was already observed in internal sequences of provirus just after seroconversion. Taken together, it is speculated that DNA methylation first occurs in the gag, pol and env regions and then extends in the 5' and 3' directions in vivo, and when 5'-LTR becomes methylated, viral transcription is silenced. Analysis of histone modification in the HTLV-I provirus showed that the methylated provirus was associated with hypoacetylation. However, the tax gene transcript could not be detected in fresh ATL cells regardless of hyperacetylated histone H3 in 5'-LTR. The transcription rapidly recovered after in vitro culture in such ATL cells. Conclusion These results showed that epigenetic changes of provirus facilitated ATL cells to evade host immune system by suppressing viral gene transcription. In addition, this study shows the presence of another reversible mechanism that suppresses the tax gene transcription without DNA methylation and hypoacetylated histone.

  14. Expression, processing and transcriptional regulation of granulysin in short-term activated human lymphocytes

    Directory of Open Access Journals (Sweden)

    Groscurth Peter

    2007-06-01

    Full Text Available Abstract Background Granulysin, a cytotoxic protein expressed in human natural killer cells and activated T lymphocytes, exhibits cytolytic activity against a variety of intracellular microbes. Expression and transcription have been partially characterised in vitro and four transcripts (NKG5, 519, 520, and 522 were identified. However, only a single protein product of 15 kDa was found, which is subsequently processed to an active 9 kDa protein. Results In this study we investigated generation of granulysin in lymphokine activated killer (LAK cells and antigen (Listeria specific T-cells. Semiquantitative RT-PCR revealed NKG5 to be the most prominent transcript. It was found to be up-regulated in a time-dependent manner in LAK cells and antigen specific T-cells and their subsets. Two isoforms of 519 mRNA were up-regulated under IL-2 and antigen stimulation. Moreover, two novel transcripts, without any known function, comprising solely parts of the 5 prime region of the primary transcript, were detected. A significant increase of granulysin expressing LAK cells as well as antigen specific T-cells was shown by fluorescence microscopy. On the subset level, increase in CD4+ granulysin expressing cells was found only under antigen stimulation. Immunoblotting showed the 15 kDa form of granulysin to be present in the first week of stimulation either with IL-2 or with bacterial antigen. Substantial processing to the 9 kDa form was detected during the first week in LAK cells and in the second week in antigen specific T-cells. Conclusion This first comprehensive study of granulysin gene regulation in primary cultured human lymphocytes shows that the regulation of granulysin synthesis in response to IL-2 or bacterial antigen stimulation occurs at several levels: RNA expression, extensive alternative splicing and posttranslational processing.

  15. Commensal Streptococcus salivarius Modulates PPARγ Transcriptional Activity in Human Intestinal Epithelial Cells.

    Directory of Open Access Journals (Sweden)

    Benoît Couvigny

    Full Text Available The impact of commensal bacteria in eukaryotic transcriptional regulation has increasingly been demonstrated over the last decades. A multitude of studies have shown direct effects of commensal bacteria from local transcriptional activity to systemic impact. The commensal bacterium Streptococcus salivarius is one of the early bacteria colonizing the oral and gut mucosal surfaces. It has been shown to down-regulate nuclear transcription factor (NF-кB in human intestinal cells, a central regulator of the host mucosal immune system response to the microbiota. In order to evaluate its impact on a further important transcription factor shown to link metabolism and inflammation in the intestine, namely PPARγ (peroxisome proliferator-activated receptor, we used human intestinal epithelial cell-lines engineered to monitor PPARγ transcriptional activity in response to a wide range of S. salivarius strains. We demonstrated that different strains from this bacterial group share the property to inhibit PPARγ activation independently of the ligand used. First attempts to identify the nature of the active compounds showed that it is a low-molecular-weight, DNase-, proteases- and heat-resistant metabolite secreted by S. salivarius strains. Among PPARγ-targeted metabolic genes, I-FABP and Angptl4 expression levels were dramatically reduced in intestinal epithelial cells exposed to S. salivarius supernatant. Both gene products modulate lipid accumulation in cells and down-regulating their expression might consequently affect host health. Our study shows that species belonging to the salivarius group of streptococci impact both host inflammatory and metabolic regulation suggesting a possible role in the host homeostasis and health.

  16. The Impact of Endurance Training on Human Skeletal Muscle Memory, Global Isoform Expression and Novel Transcripts.

    Directory of Open Access Journals (Sweden)

    Maléne E Lindholm

    2016-09-01

    Full Text Available Regularly performed endurance training has many beneficial effects on health and skeletal muscle function, and can be used to prevent and treat common diseases e.g. cardiovascular disease, type II diabetes and obesity. The molecular adaptation mechanisms regulating these effects are incompletely understood. To date, global transcriptome changes in skeletal muscles have been studied at the gene level only. Therefore, global isoform expression changes following exercise training in humans are unknown. Also, the effects of repeated interventions on transcriptional memory or training response have not been studied before. In this study, 23 individuals trained one leg for three months. Nine months later, 12 of the same subjects trained both legs in a second training period. Skeletal muscle biopsies were obtained from both legs before and after both training periods. RNA sequencing analysis of all 119 skeletal muscle biopsies showed that training altered the expression of 3,404 gene isoforms, mainly associated with oxidative ATP production. Fifty-four genes had isoforms that changed in opposite directions. Training altered expression of 34 novel transcripts, all with protein-coding potential. After nine months of detraining, no training-induced transcriptome differences were detected between the previously trained and untrained legs. Although there were several differences in the physiological and transcriptional responses to repeated training, no coherent evidence of an endurance training induced transcriptional skeletal muscle memory was found. This human lifestyle intervention induced differential expression of thousands of isoforms and several transcripts from unannotated regions of the genome. It is likely that the observed isoform expression changes reflect adaptational mechanisms and processes that provide the functional and health benefits of regular physical activity.

  17. Human Metapneumovirus Induces Formation of Inclusion Bodies for Efficient Genome Replication and Transcription.

    Science.gov (United States)

    Cifuentes-Muñoz, Nicolás; Branttie, Jean; Slaughter, Kerri Beth; Dutch, Rebecca Ellis

    2017-12-15

    Human metapneumovirus (HMPV) causes significant upper and lower respiratory disease in all age groups worldwide. The virus possesses a negative-sense single-stranded RNA genome of approximately 13.3 kb encapsidated by multiple copies of the nucleoprotein (N), giving rise to helical nucleocapsids. In addition, copies of the phosphoprotein (P) and the large RNA polymerase (L) decorate the viral nucleocapsids. After viral attachment, endocytosis, and fusion mediated by the viral glycoproteins, HMPV nucleocapsids are released into the cell cytoplasm. To visualize the subsequent steps of genome transcription and replication, a fluorescence in situ hybridization (FISH) protocol was established to detect different viral RNA subpopulations in infected cells. The FISH probes were specific for detection of HMPV positive-sense RNA (+RNA) and viral genomic RNA (vRNA). Time course analysis of human bronchial epithelial BEAS-2B cells infected with HMPV revealed the formation of inclusion bodies (IBs) from early times postinfection. HMPV IBs were shown to be cytoplasmic sites of active transcription and replication, with the translation of viral proteins being closely associated. Inclusion body formation was consistent with an actin-dependent coalescence of multiple early replicative sites. Time course quantitative reverse transcription-PCR analysis suggested that the coalescence of inclusion bodies is a strategy to efficiently replicate and transcribe the viral genome. These results provide a better understanding of the steps following HMPV entry and have important clinical implications. IMPORTANCE Human metapneumovirus (HMPV) is a recently discovered pathogen that affects human populations of all ages worldwide. Reinfections are common throughout life, but no vaccines or antiviral treatments are currently available. In this work, a spatiotemporal analysis of HMPV replication and transcription in bronchial epithelial cell-derived immortal cells was performed. HMPV was shown to

  18. The transcription of MGAT4A glycosyl transferase is increased in white cells of peripheral blood of Type 2 Diabetes patients

    Directory of Open Access Journals (Sweden)

    Cruz Miguel

    2007-10-01

    Full Text Available Abstract Background Human glycosylase IV is involved in GLUT2 transporter regulation in pancreatic β cells. A KO of this gene along with a high fat diet in a mice model has been associated with the development of type 2 diabetes (T2D. The aims of this study were to measure and compare the MGAT4A mRNA levels in white blood cells (WBC from T2D subjects and healthy subjects (T2NB, and to measure the half-life of the MGAT4A mRNA. Results We studied a sample of 73 individuals, 40 T2D subjects and 33 T2NB subjects. Anthropometrical and biochemical profiles were registered. The MGAT4A mRNA levels in WBC and the transcript half-life in Jurkat T cells were determined by Real-Time PCR. A blood differential cell counting was made for each individual. Cell counting showed T2D subjects exhibited an increased number of WBC compared to T2NB subjects (P = 0.0001. Biochemical parameters such as fasting glucose (P = 0.0001, and triglycerides (P = 0.002 were statistically significant. T2D subjects had 4.2-fold more MGAT4A transcript compared to T2NB subjects (P = 0.002. The MGAT4A mRNA had a half-life of 2.04 h in Jurkat T cells. Conclusion The results of this work suggest that in T2D subjects, high levels of glucose and triglycerides are accompanied by an increase on MGAT4A mRNA levels and WBC count; condition that suggests a pro-inflammatory state due to a chronic metabolic stress.

  19. Inducible, tunable and multiplex human gene regulation using CRISPR-Cpf1-based transcription factors | Office of Cancer Genomics

    Science.gov (United States)

    Targeted and inducible regulation of mammalian gene expression is a broadly important research capability that may also enable development of novel therapeutics for treating human diseases. Here we demonstrate that a catalytically inactive RNA-guided CRISPR-Cpf1 nuclease fused to transcriptional activation domains can up-regulate endogenous human gene expression. We engineered drug-inducible Cpf1-based activators and show how this system can be used to tune the regulation of endogenous gene transcription in human cells.

  20. Chloride Channel 3 Channels in the Activation and Migration of Human Blood Eosinophils in Allergic Asthma.

    Science.gov (United States)

    Gaurav, Rohit; Bewtra, Againdra K; Agrawal, Devendra K

    2015-08-01

    Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase is responsible for respiratory burst in immune cells. Chloride channel 3 (CLC3) has been linked to the respiratory burst in eosinophils and neutrophils. The effect of cytokines and the involvement of CLC3 in the regulation of NADPH-dependent oxidative stress and on cytokine-mediated migration of eosinophils are not known. Human peripheral blood eosinophils were isolated from healthy individuals and from individuals with asthma by negative selection. Real-time PCR was used to detect the expression of NADPH oxidases in eosinophils. Intracellular reactive oxygen species (ROS) measurement was done with flow cytometry. Superoxide generation was measured with transforming growth factor (TGF)-β, eotaxin, and CLC3 blockers. CLC3 dependence of eosinophils in TGF-β- and eotaxin-induced migration was also examined. The messenger RNA (mRNA) transcripts of NADPH oxidase (NOX) 2, dual oxidase (DUOX) 1, and DUOX2 were detected in blood eosinophils, with very low expression of NOX1, NOX3, and NOX5 and no NOX4 mRNA. The level of NOX2 mRNA transcripts increased with disease severity in the eosinophils of subjects with asthma compared with healthy nonatopic volunteers. Change in granularity and size in eosinophils, but no change in intracellular ROS, was observed with phorbol myristate acetate (PMA). PMA, TGF-β, and eotaxin used the CLC3-dependent pathway to increase superoxide radicals. TGF-β and eotaxin induced CLC3-dependent chemotaxis of eosinophils. These findings support the requirement of CLC3 in the activation and migration of human blood eosinophils and may provide a potential novel therapeutic target to regulate eosinophil hyperactivity in allergic airway inflammation in asthma.

  1. Egg beater as centrifuge: isolating human blood plasma from whole blood in resource-poor settings.

    Science.gov (United States)

    Wong, Amy P; Gupta, Malancha; Shevkoplyas, Sergey S; Whitesides, George M

    2008-12-01

    This paper demonstrates that a hand-powered egg beater can be modified to serve as a centrifuge for separating plasma from human whole blood. Immunoassays used to diagnose infectious diseases often require plasma from whole blood, and obtaining plasma typically requires electrically-powered centrifuges, which are not widely available in resource-limited settings. Human whole blood was loaded into polyethylene (PE) tubing, and the tubing was attached to the paddle of an egg beater. Spinning the paddle pelleted the blood cells to the distal end of the PE tubing; the plasma remained as the supernatant. A cholesterol assay (run on patterned paper) demonstrated the suitability of this plasma for use in diagnostic assays. The physics of the system was also analyzed as a guide for the selection of other rotating systems for use in centrifugation. Egg beaters, polyethylene tubing, and paper are readily available devices and supplies that can facilitate the use of point-of-care diagnostics at sites far from centralized laboratory facilities.

  2. Splanchnic blood flow and hepatic glucose production in exercising humans

    DEFF Research Database (Denmark)

    Bergeron, R; Kjaer, M; Simonsen, L

    2001-01-01

    The study examined the implication of the renin-angiotensin system (RAS) in regulation of splanchnic blood flow and glucose production in exercising humans. Subjects cycled for 40 min at 50% maximal O(2) consumption (VO(2 max)) followed by 30 min at 70% VO(2 max) either with [angiotensin-converti......The study examined the implication of the renin-angiotensin system (RAS) in regulation of splanchnic blood flow and glucose production in exercising humans. Subjects cycled for 40 min at 50% maximal O(2) consumption (VO(2 max)) followed by 30 min at 70% VO(2 max) either with [angiotensin......-converting enzyme (ACE) blockade] or without (control) administration of the ACE inhibitor enalapril (10 mg iv). Splanchnic blood flow was estimated by indocyanine green, and splanchnic substrate exchange was determined by the arteriohepatic venous difference. Exercise led to an approximately 20-fold increase (P ...-blockade group vs. the control group, hormones, metabolites, VO(2), and RER followed the same pattern of changes in ACE-blockade and control groups during exercise. Splanchnic blood flow (at rest: 1.67 +/- 0.12, ACE blockade; 1.59 +/- 0.18 l/min, control) decreased during moderate exercise (0.78 +/- 0.07, ACE...

  3. Sympathetic reflex control of blood flow in human peripheral tissues

    DEFF Research Database (Denmark)

    Henriksen, O

    1991-01-01

    Sympathetic vasoconstrictor reflexes are essential for the maintenance of arterial blood pressure in upright position. It has been generally believed that supraspinal sympathetic vasoconstrictor reflexes elicited by changes in baroreceptor activity play an important role. Recent studies on human...... sympathetic vasoconstrictor reflexes are blocked. Blood flow has been measure by the local 133Xe-technique. The results indicate the presence of spinal as well as supraspinal sympathetic vasoconstrictor reflexes to human peripheral tissues. Especially is emphasized the presence of a local sympathetic veno...... skeletal muscle, cutaneous and subcutaneous tissues of the limbs indicate that the situation is more complex. Measurements have been carried out during acute as well as chronic sympathetic denervation. Spinal sympathetic reflex mechanisms have been evaluated in tetraplegic patients, where supraspinal...

  4. Comprehensive analysis of the transcriptional profile of the Mediator complex across human cancer types.

    Science.gov (United States)

    Syring, Isabella; Klümper, Niklas; Offermann, Anne; Braun, Martin; Deng, Mario; Boehm, Diana; Queisser, Angela; von Mässenhausen, Anne; Brägelmann, Johannes; Vogel, Wenzel; Schmidt, Doris; Majores, Michael; Schindler, Anne; Kristiansen, Glen; Müller, Stefan C; Ellinger, Jörg; Shaikhibrahim, Zaki; Perner, Sven

    2016-04-26

    The Mediator complex is a key regulator of gene transcription and several studies demonstrated altered expressions of particular subunits in diverse human diseases, especially cancer. However a systematic study deciphering the transcriptional expression of the Mediator across different cancer entities is still lacking.We therefore performed a comprehensive in silico cancer vs. benign analysis of the Mediator complex subunits (MEDs) for 20 tumor entities using Oncomine datasets. The transcriptional expression profiles across almost all cancer entities showed differentially expressed MEDs as compared to benign tissue. Differential expression of MED8 in renal cell carcinoma (RCC) and MED12 in lung cancer (LCa) were validated and further investigated by immunohistochemical staining on tissue microarrays containing large numbers of specimen. MED8 in clear cell RCC (ccRCC) associated with shorter survival and advanced TNM stage and showed higher expression in metastatic than primary tumors. In vitro, siRNA mediated MED8 knockdown significantly impaired proliferation and motility in ccRCC cell lines, hinting at a role for MED8 to serve as a novel therapeutic target in ccRCC. Taken together, our Mediator complex transcriptome proved to be a valid tool for identifying cancer-related shifts in Mediator complex composition, revealing that MEDs do exhibit cancer specific transcriptional expression profiles.

  5. High DNA melting temperature predicts transcription start site location in human and mouse.

    LENUS (Irish Health Repository)

    Dineen, David G

    2009-12-01

    The accurate computational prediction of transcription start sites (TSS) in vertebrate genomes is a difficult problem. The physicochemical properties of DNA can be computed in various ways and a many combinations of DNA features have been tested in the past for use as predictors of transcription. We looked in detail at melting temperature, which measures the temperature, at which two strands of DNA separate, considering the cooperative nature of this process. We find that peaks in melting temperature correspond closely to experimentally determined transcription start sites in human and mouse chromosomes. Using melting temperature alone, and with simple thresholding, we can predict TSS with accuracy that is competitive with the most accurate state-of-the-art TSS prediction methods. Accuracy is measured using both experimentally and manually determined TSS. The method works especially well with CpG island containing promoters, but also works when CpG islands are absent. This result is clear evidence of the important role of the physical properties of DNA in the process of transcription. It also points to the importance for TSS prediction methods to include melting temperature as prior information.

  6. Molecular detection of peripheral blood breast cancer mRNA transcripts as a surrogate biomarker for circulating tumor cells.

    Directory of Open Access Journals (Sweden)

    Adriana Lasa

    Full Text Available Circulating tumor cells (CTCs are becoming a scientifically recognized indicator of primary tumors and/or metastasis. These cells can now be accurately detected and characterized as the result of technological advances. We analyzed the presence of CTCs in the peripheral blood of patients with metastatic breast cancer by real-time reverse-transcription PCR (RT-qPCR using a panel of selected genes. The analysis of a single marker, without an EpCAM based enrichment approach, allowed the positive identification of 35% of the metastatic breast cancer patients. The analysis of five genes (SCGB2, TFF1, TFF3, Muc1, KRT20 performed in all the samples increased the detection to 61%. We describe a sensitive, reproducible and easy to implement approach to characterize CTC in patients with metastasic breast cancer.

  7. Regulation of tumor necrosis factor gene expression by ionizing radiation in human myeloid leukemia cells and peripheral blood monocytes

    International Nuclear Information System (INIS)

    Sherman, M.L.; Datta, R.; Hallahan, D.E.; Weichselbaum, R.R.; Kufe, D.W.

    1991-01-01

    Previous studies have demonstrated that ionizing radiation induces the expression of certain cytokines, such as TNF alpha/cachectin. However, there is presently no available information regarding the molecular mechanisms responsible for the regulation of cytokine gene expression by ionizing radiation. In this report, we describe the regulation of the TNF gene by ionizing radiation in human myeloid leukemia cells. The increase in TNF transcripts by x rays was both time- and dose-dependent as determined by Northern blot analysis. Similar findings were obtained in human peripheral blood monocytes. Transcriptional run-on analyses have demonstrated that ionizing radiation stimulates the rate of TNF gene transcription. Furthermore, induction of TNF mRNA was increased in the absence of protein synthesis. In contrast, ionizing radiation had little effect on the half-life of TNF transcripts. These findings indicate that the increase in TNF mRNA observed after irradiation is regulated by transcriptional mechanisms and suggest that production of this cytokine by myeloid cells may play a role in the pathophysiologic effects of ionizing radiation

  8. Implementation of good manufacturing practices (GMP) on human blood irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Boghi, Claudio; Napolitano, Celia M.; Ferreira, Danilo C.; Rela, Paulo Roberto [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)]. E-mails: cboghi@uol.com.br; cmnapoli@ipen.br; dancarde@ig.com.br; prela@ipen.br; Zarate, Herman S. [Comission Chilena de Energia Nuclear, Santiago (Chile)]. E-mail: hzarate@cchen.cl

    2007-07-01

    The irradiation of human blood is used to avoid the TA-GVHD (transfusion-associated graft-versus-host-disease), a rare but devastating adverse effect of leukocytes present in blood components for a immuno-competent transfusion recipients. Usually this irradiation practice is performed to a physical elimination of lymphocytes. The implementation of the GMP will assure that the properly dose in a range of 25 Gy to 50 Gy will be delivered to the blood in the bag collected in a blood tissue bank. The studies to establish the GMP were developed under the guidelines of the standard ISO 11137 - Sterilization of health care products - Requirements for validation and routine control - Radiation sterilization. In this work, two dosimetric systems were used for dose mapping during the studies of irradiator qualification, loading pattern, irradiation process validation and auditing. The CaSO{sub 4}: Dy dosimeter presented difficulties concerning to uncertainty on dose measurement, stability, trace ability and calibration system. The PMMA and gafchromic dosimetric systems have shown a better performance and were adopted on establishment of GMP procedures. The irradiation tests have been done using a Gammacell 220 Irradiator. The developed GMP can be adapted for different types of gamma irradiators, allowing to set up a quality assurance program for blood irradiation. (author)

  9. Laser-photophoretic migration and fractionation of human blood cells

    Energy Technology Data Exchange (ETDEWEB)

    Monjushiro, Hideaki; Tanahashi, Yuko; Watarai, Hitoshi, E-mail: watarai@chem.sci.osaka-u.ac.jp

    2013-05-13

    Graphical abstract: -- Highlights: •RBCs were migrated faster than WBCs and blood pellets by laser photophoresis. •Photophoretic efficiency of RBC and WBC was simulated by the Mie scattering theory. •Spontaneous orientation of RBC parallel to the migration direction was elucidated. •Laser photophoretic separation of RBC and WBC was possible in a tip flow system. -- Abstract: Laser photophoretic migration behavior of human blood cells in saline solution was investigated under the irradiation of Nd:YAG laser beam (532 nm) in the absence and the presence of the flow in a fused silica capillary. Red blood cells (RBC) were migrated faster than white blood cells (WBC) and blood pellets to the direction of propagation of laser light. The observed photophoretic velocity of RBC was about 11 times faster than those of others. This was understood from the larger photophoretic efficiency of RBC than that of WBC, which was simulated based on the Mie scattering theory. Furthermore, it was found that, during the photophoretic migration, RBCs spontaneously orientated parallel to the migration direction so as to reduce the drag force. Finally, it was demonstrated that RBC and WBC were separated in a micro-channel flow system by the laser photophoresis.

  10. Laser-photophoretic migration and fractionation of human blood cells

    International Nuclear Information System (INIS)

    Monjushiro, Hideaki; Tanahashi, Yuko; Watarai, Hitoshi

    2013-01-01

    Graphical abstract: -- Highlights: •RBCs were migrated faster than WBCs and blood pellets by laser photophoresis. •Photophoretic efficiency of RBC and WBC was simulated by the Mie scattering theory. •Spontaneous orientation of RBC parallel to the migration direction was elucidated. •Laser photophoretic separation of RBC and WBC was possible in a tip flow system. -- Abstract: Laser photophoretic migration behavior of human blood cells in saline solution was investigated under the irradiation of Nd:YAG laser beam (532 nm) in the absence and the presence of the flow in a fused silica capillary. Red blood cells (RBC) were migrated faster than white blood cells (WBC) and blood pellets to the direction of propagation of laser light. The observed photophoretic velocity of RBC was about 11 times faster than those of others. This was understood from the larger photophoretic efficiency of RBC than that of WBC, which was simulated based on the Mie scattering theory. Furthermore, it was found that, during the photophoretic migration, RBCs spontaneously orientated parallel to the migration direction so as to reduce the drag force. Finally, it was demonstrated that RBC and WBC were separated in a micro-channel flow system by the laser photophoresis

  11. Implementation of good manufacturing practices (GMP) on human blood irradiation

    International Nuclear Information System (INIS)

    Boghi, Claudio; Napolitano, Celia M.; Ferreira, Danilo C.; Rela, Paulo Roberto; Zarate, Herman S.

    2007-01-01

    The irradiation of human blood is used to avoid the TA-GVHD (transfusion-associated graft-versus-host-disease), a rare but devastating adverse effect of leukocytes present in blood components for a immuno-competent transfusion recipients. Usually this irradiation practice is performed to a physical elimination of lymphocytes. The implementation of the GMP will assure that the properly dose in a range of 25 Gy to 50 Gy will be delivered to the blood in the bag collected in a blood tissue bank. The studies to establish the GMP were developed under the guidelines of the standard ISO 11137 - Sterilization of health care products - Requirements for validation and routine control - Radiation sterilization. In this work, two dosimetric systems were used for dose mapping during the studies of irradiator qualification, loading pattern, irradiation process validation and auditing. The CaSO 4 : Dy dosimeter presented difficulties concerning to uncertainty on dose measurement, stability, trace ability and calibration system. The PMMA and gafchromic dosimetric systems have shown a better performance and were adopted on establishment of GMP procedures. The irradiation tests have been done using a Gammacell 220 Irradiator. The developed GMP can be adapted for different types of gamma irradiators, allowing to set up a quality assurance program for blood irradiation. (author)

  12. Human erythrovirus B19 and blood transfusion - an update.

    Science.gov (United States)

    Parsyan, A; Candotti, D

    2007-08-01

    Erythrovirus (parvovirus) B19 (B19) is a common human pathogen. It is a non-enveloped single-strand DNA virus packaging its genome in small tight capsids consisting of viral VP1 and VP2 proteins. It is now accepted that B19 is a relatively quickly evolving virus having diverged in several genetic variants recently identified. The main route of B19 transmission is respiratory, with a majority of infections occurring during childhood and manifesting as erythema infectiousum. B19 can also be transmitted vertically and via blood transfusion and organ transplantation. The majority of adult populations show immunological evidence of previous exposure to B19. Although the immune response is able to clear infection and provide life-long protection against B19, recent data suggest that in some, if not the majority, of individuals the acute phase of infection is followed by viral persistence in the blood or other tissues regardless of the host's immunocompetence. Transmission of B19 by blood and blood products and its resistance to common viral inactivation methods raises several blood safety questions, still unanswered. The diversity of B19 strains and the ability of the virus to persist in the presence of specific antibodies raise the issue of transmissibility by transfusion not so much to immunocompetent recipients but rather to the large proportion of recipients in whom there is some degree of immunodeficiency. The ability of the virus to reactivate in immunodeficient recipients may create difficulties in differentiating between transfusion transmission and reactivation.

  13. Peridural anesthesia and the distribution of blood in supine humans

    International Nuclear Information System (INIS)

    Arndt, J.O.; Hoeck, A.S.; Stanton-Hicks, M.; Stuehmeier, K.D.

    1985-01-01

    To determine the effects of vasomotor tone on intrathoracic and splanchnic blood volume, the distribution of radioactively (/sup 99m/Tc) labeled erythrocytes was recorded by whole body scintigraphy before and during peridural anesthesia (PDA) in eight supine men. The radioactivity was recorded with a gamma camera and its distribution determined in the thorax, abdomen, and limbs. Arterial and central venous pressure, heart rate, and calf volume and flow also were measured. During PDA with a sensory block up to T4/5, radioactivity increased only in the denervated legs, whereas it decreased in all other regions, i.e., in the thorax, the innervated upper limbs, and in the splanchnic vasculature. However, in two of the subjects, after an initial decrease, splanchnic blood content increased while intrathoracic blood volume decreased further. The effects of PDA on thoracic and splanchnic filling could be duplicated by the sequestration of about 500-600 ml of blood in both legs. In supine humans high peridural anesthesia evokes the same decrease in intrathoracic blood volume as orthostasis. Potential circulatory collapse may ensue when the vasoconstrictor response fails in the splanchnic circulation

  14. Good manufacturing practices (GMP utilized on human blood irradiation process

    Directory of Open Access Journals (Sweden)

    Cláudio Boghi

    2008-01-01

    Full Text Available Irradiation of human blood is used to avoid the TA-GVHD (transfusion-associated graft-versus-host-disease, a rare but devastating adverse effect of leukocytes present in blood components for immunocompetent transfusion recipients. Usually this irradiation practice is performed to a physical elimination of lymphocytes. The implementation of the GMP will assure that the properly dose in a range of 25Gy to 50Gy will be delivered to the blood in the bag collected in a blood tissue bank. The studies to establish the GMP were developed under the guidelines of the standard ISO 11137 - Sterilization of health care products - Requirements for validation and routine control - Radiation sterilization. In this work, two dosimetric systems were used for dose mapping during the studies of irradiator qualification, loading pattern, irradiation process validation and auditing. The CaSO4: Dy dosimeter presented difficulties concerning to uncertainty on dose measurement, stability, trace ability and calibration system. The PMMA and gafchromic dosimetric systems have shown a better performance and were adopted on establishment of GMP procedures. The irradiation tests have been done using a Gammacell 220 Irradiator. The developed GMP can be adapted for different types of gamma irradiators, allowing to set up a quality assurance program for blood irradiation.

  15. Detection of endotoxins and other pyrogens using human whole blood.

    Science.gov (United States)

    Fennrich, S; Fischer, M; Hartung, T; Lexa, P; Montag-Lessing, T; Sonntag, H G; Weigandt, M; Wendel, A

    1999-01-01

    When cells of the immune system, i.e. primarily blood monocytes and macrophages, come into contact with pyrogens (fever-inducing contaminations) they release mediators transmitting the fever reaction through the organism to the thermoregulatory centres of the brain. The new test discussed here exploits this reaction for the detection of pyrogens: human whole blood taken from healthy volunteers is incubated in the presence of the test sample. If there is pyrogen contamination, the endogenous pyrogen interleukin-1 is released, which is then determined by ELISA. According to the pharmacopoeia, the rabbit pyrogen test determines the fever reaction following injection of a test sample. In comparison, the new whole blood assay is more sensitive, less expensive and determines the reaction of the targeted species. Compared to the well established in vitro alternative, i.e. the limulus amebocyte lysate assay (LAL), the new blood assay is not restricted to endotoxins of gram-negative bacteria, it is not affected by endotoxin-binding blood proteins and it reflects the potency of different endotoxin preparations in mammals. Here, interim results of the ongoing optimization and pre-validation are reported and the present state of the evaluation for biological and pharmaceutical drugs are presented.

  16. Deep coverage mouse red blood cell proteome: a first comparison with the human red blood cell

    DEFF Research Database (Denmark)

    Pasini, Erica M; Kirkegaard, Morten; Salerno, Doris

    2008-01-01

    Mice have close genetic/physiological relationships to humans, breed rapidly, and can be genetically modified, making them the most used mammal in biomedical research. Because the red blood cell (RBC) is the sole gas transporter in vertebrates, diseases of the RBC are frequently severe; much...... proteome have been confirmed here. This comparison sheds light on several open issues in RBC biology and provides a departure point for more comprehensive understanding of RBC function....

  17. Low nucleosome occupancy is encoded around functional human transcription factor binding sites

    Directory of Open Access Journals (Sweden)

    Daenen Floris

    2008-07-01

    Full Text Available Abstract Background Transcriptional regulation of genes in eukaryotes is achieved by the interactions of multiple transcription factors with arrays of transcription factor binding sites (TFBSs on DNA and with each other. Identification of these TFBSs is an essential step in our understanding of gene regulatory networks, but computational prediction of TFBSs with either consensus or commonly used stochastic models such as Position-Specific Scoring Matrices (PSSMs results in an unacceptably high number of hits consisting of a few true functional binding sites and numerous false non-functional binding sites. This is due to the inability of the models to incorporate higher order properties of sequences including sequences surrounding TFBSs and influencing the positioning of nucleosomes and/or the interactions that might occur between transcription factors. Results Significant improvement can be expected through the development of a new framework for the modeling and prediction of TFBSs that considers explicitly these higher order sequence properties. It would be particularly interesting to include in the new modeling framework the information present in the nucleosome positioning sequences (NPSs surrounding TFBSs, as it can be hypothesized that genomes use this information to encode the formation of stable nucleosomes over non-functional sites, while functional sites have a more open chromatin configuration. In this report we evaluate the usefulness of the latter feature by comparing the nucleosome occupancy probabilities around experimentally verified human TFBSs with the nucleosome occupancy probabilities around false positive TFBSs and in random sequences. Conclusion We present evidence that nucleosome occupancy is remarkably lower around true functional human TFBSs as compared to non-functional human TFBSs, which supports the use of this feature to improve current TFBS prediction approaches in higher eukaryotes.

  18. Metabolic control of muscle blood flow during exercise in humans

    DEFF Research Database (Denmark)

    Boushel, Robert Christopher

    2003-01-01

    that combined blockade of NOS and PGI2, and NOS and cytochrome P450, both attenuate exercise-induced hyperemia in humans. Combined vasodilator blockade studies offer the potential to uncover important interactions and compensatory vasodilator responses. The signaling pathways that link metabolic events evoked...... to exert control of muscle vasodilation. Adenosine, nitric oxide (NO), prostacyclin (PGI2), and endothelial-derived hyperpolarization factor (EDHF) are possible mediators of muscle vasodilation during exercise. In humans, adenosine has been shown to contribute to functional hyperemia as blood flow...... by muscle contraction to vasodilatory signals in the local vascular bed remains an important area of study....

  19. Transcription factor fos-related antigen-2 induces progressive peripheral vasculopathy in mice closely resembling human systemic sclerosis.

    Science.gov (United States)

    Maurer, Britta; Busch, Nicole; Jüngel, Astrid; Pileckyte, Margarita; Gay, Renate E; Michel, Beat A; Schett, Georg; Gay, Steffen; Distler, Jörg; Distler, Oliver

    2009-12-08

    Microvascular damage is one of the first pathological changes in systemic sclerosis. In this study, we investigated the role of Fos-related antigen-2 (Fra-2), a transcription factor of the activator protein-1 family, in the peripheral vasculopathy of systemic sclerosis and examined the underlying mechanisms. Expression of Fra-2 protein was significantly increased in skin biopsies of systemic sclerosis patients compared with healthy controls, especially in endothelial and vascular smooth muscle cells. Fra-2 transgenic mice developed a severe loss of small blood vessels in the skin that was paralleled by progressive skin fibrosis at 12 weeks of age. The reduction in capillary density was preceded by a significant increase in apoptosis in endothelial cells at week 9 as detected by immunohistochemistry. Similarly, suppression of Fra-2 by small interfering RNA prevented human microvascular endothelial cells from staurosporine-induced apoptosis and improved both the number of tubes and the cumulative tube lengths in the tube formation assay. In addition, cell migration in the scratch assay and vascular endothelial growth factor-dependent chemotaxis in a modified Boyden chamber assay were increased after transfection of human microvascular endothelial cells with Fra-2 small interfering RNA, whereas proliferation was not affected. Fra-2 is present in human systemic sclerosis and may contribute to the development of microvasculopathy by inducing endothelial cell apoptosis and by reducing endothelial cell migration and chemotaxis. Fra-2 transgenic mice are a promising preclinical model to study the mechanisms and therapeutic approaches of the peripheral vasculopathy in systemic sclerosis.

  20. BLR1 and FCGR1A transcripts in peripheral blood associate with the extent of intrathoracic tuberculosis in children and predict treatment outcome

    DEFF Research Database (Denmark)

    Jenum, Synne; Bakken, Rasmus; Dhanasekaran, S

    2016-01-01

    children with intrathoracic tuberculosis (TB), we performed blood transcriptome kinetic analysis during ATT to explore 1) the association between transcriptional biomarkers in whole blood (WB) and the extent of TB disease at diagnosis and treatment outcomes at 2 and 6 months, and 2) the potential...... of the biomarkers to predict treatment response at 2 and 6 months. We present the first data on the association between transcriptional biomarkers and the extent of TB disease as well as outcome of ATT in children: Expression of three genes down-regulated on ATT (FCGR1A, FPR1 and MMP9) exhibited a positive...

  1. Human BLCAP transcript: new editing events in normal and cancerous tissues.

    Science.gov (United States)

    Galeano, Federica; Leroy, Anne; Rossetti, Claudia; Gromova, Irina; Gautier, Philippe; Keegan, Liam P; Massimi, Luca; Di Rocco, Concezio; O'Connell, Mary A; Gallo, Angela

    2010-07-01

    Bladder cancer-associated protein (BLCAP) is a highly conserved protein among species, and it is considered a novel candidate tumor suppressor gene originally identified from human bladder carcinoma. However, little is known about the regulation or the function of this protein. Here, we show that the human BLCAP transcript undergoes multiple A-to-I editing events. Some of the new editing events alter the highly conserved amino terminus of the protein creating alternative protein isoforms by changing the genetically coded amino acids. We found that both ADAR1 and ADAR2-editing enzymes cooperate to edit this transcript and that different tissues displayed distinctive ratios of edited and unedited BLCAP transcripts. Moreover, we observed a general decrease in BLCAP-editing level in astrocytomas, bladder cancer and colorectal cancer when compared with the related normal tissues. The newly identified editing events, found to be downregulated in cancers, could be useful for future studies as a diagnostic tool to distinguish malignancies or epigenetic changes in different tumors.

  2. Ionizing radiation activates vascular endothelial growth factor-A transcription in human umbilical vein endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Hyounji; Kim, Kwang Seok; Jeong, Jae Hoon; Lim, Young Bin [Radiation Cancer Biology Team, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2016-12-15

    Vascular endothelial growth factor (VEGF) is an essential paracrine factor for developmental and pathological angiogenesis. VEGF also exerts its effects in an autocrine manner in VEGF-producing cells. For instance, autocrine VEGF signaling occurs in tumor cells and contributes to key aspects of tumorigenesis, such as in the function of cancer stem cells and tumor initiation, which are independent of angiogenesis. In addition to tumors cells, non-transformed cells also express VEGF. For example, a VEGF dependent intracellular autocrine mechanism is crucial for the survival of hematopoietic stem cells and hematopoiesis. Stereotactic body radiation therapy (SBRT) is a novel treatment modality for early primary cancer and oligometastatic disease. SBRT delivers high-dose hypofractionated radiation, such as 20-60 Gy, to tumors in a single fraction or 2-5 fractions. As VEGF is a critical regulator of functional integrity and viability of vascular endothelial cells, we examined whether high-dose irradiation alters VEGF signaling by measuring the expression levels of VEGFA transcript. It is generally believed that endothelial cells do not produce VEGF in response to radiation. In present study, however, we provide the first demonstration of transcriptional regulation of VEGFA in human vascular endothelial cells by IR treatment. Irradiation with doses higher than 10 Gy in a single exposure triggers up-regulation of VEGFA transcription within 2 hours in HUVECs, whereas irradiation with 10 Gy does not alter VEGFA levels. Our data have shown that high-dose irradiation triggers immediate transactivation of VEGFA in human vascular endothelial cells.

  3. Optimizing the method for generation of integration-free induced pluripotent stem cells from human peripheral blood.

    Science.gov (United States)

    Gu, Haihui; Huang, Xia; Xu, Jing; Song, Lili; Liu, Shuping; Zhang, Xiao-Bing; Yuan, Weiping; Li, Yanxin

    2018-06-15

    Generation of induced pluripotent stem cells (iPSCs) from human peripheral blood provides a convenient and low-invasive way to obtain patient-specific iPSCs. The episomal vector is one of the best approaches for reprogramming somatic cells to pluripotent status because of its simplicity and affordability. However, the efficiency of episomal vector reprogramming of adult peripheral blood cells is relatively low compared with cord blood and bone marrow cells. In the present study, integration-free human iPSCs derived from peripheral blood were established via episomal technology. We optimized mononuclear cell isolation and cultivation, episomal vector promoters, and a combination of transcriptional factors to improve reprogramming efficiency. Here, we improved the generation efficiency of integration-free iPSCs from human peripheral blood mononuclear cells by optimizing the method of isolating mononuclear cells from peripheral blood, by modifying the integration of culture medium, and by adjusting the duration of culture time and the combination of different episomal vectors. With this optimized protocol, a valuable asset for banking patient-specific iPSCs has been established.

  4. Transcriptional profiles of supragranular-enriched genes associate with corticocortical network architecture in the human brain.

    Science.gov (United States)

    Krienen, Fenna M; Yeo, B T Thomas; Ge, Tian; Buckner, Randy L; Sherwood, Chet C

    2016-01-26

    The human brain is patterned with disproportionately large, distributed cerebral networks that connect multiple association zones in the frontal, temporal, and parietal lobes. The expansion of the cortical surface, along with the emergence of long-range connectivity networks, may be reflected in changes to the underlying molecular architecture. Using the Allen Institute's human brain transcriptional atlas, we demonstrate that genes particularly enriched in supragranular layers of the human cerebral cortex relative to mouse distinguish major cortical classes. The topography of transcriptional expression reflects large-scale brain network organization consistent with estimates from functional connectivity MRI and anatomical tracing in nonhuman primates. Microarray expression data for genes preferentially expressed in human upper layers (II/III), but enriched only in lower layers (V/VI) of mouse, were cross-correlated to identify molecular profiles across the cerebral cortex of postmortem human brains (n = 6). Unimodal sensory and motor zones have similar molecular profiles, despite being distributed across the cortical mantle. Sensory/motor profiles were anticorrelated with paralimbic and certain distributed association network profiles. Tests of alternative gene sets did not consistently distinguish sensory and motor regions from paralimbic and association regions: (i) genes enriched in supragranular layers in both humans and mice, (ii) genes cortically enriched in humans relative to nonhuman primates, (iii) genes related to connectivity in rodents, (iv) genes associated with human and mouse connectivity, and (v) 1,454 gene sets curated from known gene ontologies. Molecular innovations of upper cortical layers may be an important component in the evolution of long-range corticocortical projections.

  5. Deep Surveying of the Transcriptional and Alternative Splicing Signatures for Decidual CD8+ T Cells at the First Trimester of Human Healthy Pregnancy

    Directory of Open Access Journals (Sweden)

    Weihong Zeng

    2018-05-01

    Full Text Available Decidual CD8+ (dCD8 T cells have been proposed to play important roles in immune protection against the invading pathogens and in tolerance toward the growing semi-allogeneic fetus during early pregnancy. However, their phenotypic and functional characteristics remain poorly defined. Here, we performed the first analysis of the transcriptional and alternative splicing (AS signatures for human first-trimester dCD8 T cells using high-throughput mRNA sequencing. Our data revealed that dCD8 T cells have distinct transcriptional and AS landscapes when compared with their autologous peripheral blood CD8+ (pCD8 T counterparts. Furthermore, human dCD8 T cells were observed to contain CD8-Treg and effector-memory T-cell subsets, and display enhanced functionality in terms of degranulation and cytokine production on a per-cell basis. Additionally, we have identified the novel splice junctions that use a high ratio of the non-canonical splicing motif GC-AG and found that AS is not a major contributor to the gene expression-level changes between paired pCD8 and dCD8 T cells. Together, our findings not only provide a comprehensive framework of the transcriptional and AS landscapes but also reveal the functional feature of human dCD8 T cells, which are of great importance in understanding the biology of these cells and the physiology of human healthy pregnancy.

  6. Naturally occurring mutations in the human 5-lipoxygenase gene promoter that modify transcription factor binding and reporter gene transcription.

    OpenAIRE

    In, K H; Asano, K; Beier, D; Grobholz, J; Finn, P W; Silverman, E K; Silverman, E S; Collins, T; Fischer, A R; Keith, T P; Serino, K; Kim, S W; De Sanctis, G T; Yandava, C; Pillari, A

    1997-01-01

    Five lipoxygenase (5-LO) is the first committed enzyme in the metabolic pathway leading to the synthesis of the leukotrienes. We examined genomic DNA isolated from 25 normal subjects and 31 patients with asthma (6 of whom had aspirin-sensitive asthma) for mutations in the known transcription factor binding regions and the protein encoding region of the 5-LO gene. A family of mutations in the G + C-rich transcription factor binding region was identified consisting of the deletion of one, delet...

  7. Substrate availability and transcriptional regulation of metabolic genes in human skeletal muscle during recovery from exercise

    DEFF Research Database (Denmark)

    Pilegaard, Henriette; Osada, Takuya; Andersen, Lisbeth Tingsted

    2005-01-01

    before exercise and 2, 5, 8, and 24 hours after exercise. Muscle glycogen was restored to near resting levels within 5 hours in the HC trial, but remained depressed through 24 hours in the LC trial. During the 2- to 8-hour recovery period, leg glucose uptake was 5- to 15-fold higher with HC ingestion......In skeletal muscle of humans, transcription of several metabolic genes is transiently induced during recovery from exercise when no food is consumed. To determine the potential influence of substrate availability on the transcriptional regulation of metabolic genes during recovery from exercise, 9...... male subjects (aged 22-27) completed 75 minutes of cycling exercise at 75% V¿o2max on 2 occasions, consuming either a high-carbohydrate (HC) or low-carbohydrate (LC) diet during the subsequent 24 hours of recovery. Nuclei were isolated and tissue frozen from vastus lateralis muscle biopsies obtained...

  8. Genome-wide transcriptional profiling of human glioblastoma cells in response to ITE treatment.

    Science.gov (United States)

    Kang, Bo; Zhou, Yanwen; Zheng, Min; Wang, Ying-Jie

    2015-09-01

    A ligand-activated transcription factor aryl hydrocarbon receptor (AhR) is recently revealed to play a key role in embryogenesis and tumorigenesis (Feng et al. [1], Safe et al. [2]) and 2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) (Song et al. [3]) is an endogenous AhR ligand that possesses anti-tumor activity. In order to gain insights into how ITE acts via the AhR in embryogenesis and tumorigenesis, we analyzed the genome-wide transcriptional profiles of the following three groups of cells: the human glioblastoma U87 parental cells, U87 tumor sphere cells treated with vehicle (DMSO) and U87 tumor sphere cells treated with ITE. Here, we provide the details of the sample gathering strategy and show the quality controls and the analyses associated with our gene array data deposited into the Gene Expression Omnibus (GEO) under the accession code of GSE67986.

  9. Atomic force microscope observation of branching in single transcript molecules derived from human cardiac muscle

    International Nuclear Information System (INIS)

    Reed, Jason; Hsueh, Carlin; Gimzewski, James K; Mishra, Bud

    2008-01-01

    We have used an atomic force microscope to examine a clinically derived sample of single-molecule gene transcripts, in the form of double-stranded cDNA, (c: complementary) obtained from human cardiac muscle without the use of polymerase chain reaction (PCR) amplification. We observed a log-normal distribution of transcript sizes, with most molecules being in the range of 0.4-7.0 kilobase pairs (kb) or 130-2300 nm in contour length, in accordance with the expected distribution of mRNA (m: messenger) sizes in mammalian cells. We observed novel branching structures not previously known to exist in cDNA, and which could have profound negative effects on traditional analysis of cDNA samples through cloning, PCR and DNA sequencing

  10. Expression of the Transcription Factor E4BP4 in Human Basophils

    DEFF Research Database (Denmark)

    Jensen, Bettina Margrethe; Gohr, Maria; Poulsen, Lars Kærgaard

    2014-01-01

    Rationale The cytokine IL-3 plays an important role for human basophil development, function and survival. IL-3 is also reported to induce the expression of the transcription factor E4BP4, but it is not known whether E4BP4 is expressed in basophils and influences basophil responsiveness. The aim...... by Alcian blue. RNA was extracted (0.005-0.02 µg RNA from 0.5 - 1 x 106 cells), and the corresponding cDNA analyzed by real-time PCR where E4BP4 expression was calculated as 2-(CT(E4BP4) - CT(β-actin)). E4BP4 protein expression was visualized in basophil lysates (107 cells/ml) by Western blot followed...... the transcription factor E4BP4 which might have an impact on basophil histamine release....

  11. Plastic downregulation of the transcriptional repressor BCL6 during maturation of human dendritic cells

    International Nuclear Information System (INIS)

    Pantano, Serafino; Jarrossay, David; Saccani, Simona; Bosisio, Daniela; Natoli, Gioacchino

    2006-01-01

    Dendritic cell (DC) maturation links peripheral events initiated by the encounter with pathogens to the activation and expansion of antigen-specific T lymphocytes in secondary lymphoid organs. Here, we describe an as yet unrecognized modulator of human DC maturation, the transcriptional repressor BCL6. We found that both myeloid and plasmacytoid DCs constitutively express BCL6, which is rapidly downregulated following maturation triggered by selected stimuli. Both in unstimulated and maturing DCs, control of BCL6 protein levels reflects the convergence of several mechanisms regulating BCL6 stability, mRNA transcription and nuclear export. By regulating the induction of several genes implicated in the immune response, including inflammatory cytokines, chemokines and survival genes, BCL6 may represent a pivotal modulator of the afferent branch of the immune response

  12. Transcriptionally Driven DNA Replication Program of the Human Parasite Leishmania major

    Directory of Open Access Journals (Sweden)

    Rodrigo Lombraña

    2016-08-01

    Full Text Available Faithful inheritance of eukaryotic genomes requires the orchestrated activation of multiple DNA replication origins (ORIs. Although origin firing is mechanistically conserved, how origins are specified and selected for activation varies across different model systems. Here, we provide a complete analysis of the nucleosomal landscape and replication program of the human parasite Leishmania major, building on a better evolutionary understanding of replication organization in Eukarya. We found that active transcription is a driving force for the nucleosomal organization of the L. major genome and that both the spatial and the temporal program of DNA replication can be explained as associated to RNA polymerase kinetics. This simple scenario likely provides flexibility and robustness to deal with the environmental changes that impose alterations in the genetic programs during parasitic life cycle stages. Our findings also suggest that coupling replication initiation to transcription elongation could be an ancient solution used by eukaryotic cells for origin maintenance.

  13. In vitro transdifferentiation of human peripheral blood mononuclear cells to photoreceptor-like cells

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    Yukari Komuta

    2016-06-01

    Full Text Available Direct reprogramming is a promising, simple and low-cost approach to generate target cells from somatic cells without using induced pluripotent stem cells. Recently, peripheral blood mononuclear cells (PBMCs have attracted considerable attention as a somatic cell source for reprogramming. As a cell source, PBMCs have an advantage over dermal fibroblasts with respect to the ease of collecting tissues. Based on our studies involving generation of photosensitive photoreceptor cells from human iris cells and human dermal fibroblasts by transduction of photoreceptor-related transcription factors via retrovirus vectors, we transduced these transcription factors into PBMCs via Sendai virus vectors. We found that retinal disease-related genes were efficiently detected in CRX-transduced cells, most of which are crucial to photoreceptor functions. In functional studies, a light-induced inward current was detected in some CRX-transduced cells. Moreover, by modification of the culture conditions including additional transduction of RAX1 and NEUROD1, we found a greater variety of retinal disease-related genes than that observed in CRX-transduced PBMCs. These data suggest that CRX acts as a master control gene for reprogramming PBMCs into photoreceptor-like cells and that our induced photoreceptor-like cells might contribute to individualized drug screening and disease modeling of inherited retinal degeneration.

  14. Analysis of transcript and protein overlap in a human osteosarcoma cell line

    Directory of Open Access Journals (Sweden)

    Emanuelsson Olof

    2010-12-01

    Full Text Available Abstract Background An interesting field of research in genomics and proteomics is to compare the overlap between the transcriptome and the proteome. Recently, the tools to analyse gene and protein expression on a whole-genome scale have been improved, including the availability of the new generation sequencing instruments and high-throughput antibody-based methods to analyze the presence and localization of proteins. In this study, we used massive transcriptome sequencing (RNA-seq to investigate the transcriptome of a human osteosarcoma cell line and compared the expression levels with in situ protein data obtained in-situ from antibody-based immunohistochemistry (IHC and immunofluorescence microscopy (IF. Results A large-scale analysis based on 2749 genes was performed, corresponding to approximately 13% of the protein coding genes in the human genome. We found the presence of both RNA and proteins to a large fraction of the analyzed genes with 60% of the analyzed human genes detected by all three methods. Only 34 genes (1.2% were not detected on the transcriptional or protein level with any method. Our data suggest that the majority of the human genes are expressed at detectable transcript or protein levels in this cell line. Since the reliability of antibodies depends on possible cross-reactivity, we compared the RNA and protein data using antibodies with different reliability scores based on various criteria, including Western blot analysis. Gene products detected in all three platforms generally have good antibody validation scores, while those detected only by antibodies, but not by RNA sequencing, generally consist of more low-scoring antibodies. Conclusion This suggests that some antibodies are staining the cells in an unspecific manner, and that assessment of transcript presence by RNA-seq can provide guidance for validation of the corresponding antibodies.

  15. The DNA methylome of human peripheral blood mononuclear cells.

    Directory of Open Access Journals (Sweden)

    Yingrui Li

    2010-11-01

    Full Text Available DNA methylation plays an important role in biological processes in human health and disease. Recent technological advances allow unbiased whole-genome DNA methylation (methylome analysis to be carried out on human cells. Using whole-genome bisulfite sequencing at 24.7-fold coverage (12.3-fold per strand, we report a comprehensive (92.62% methylome and analysis of the unique sequences in human peripheral blood mononuclear cells (PBMC from the same Asian individual whose genome was deciphered in the YH project. PBMC constitute an important source for clinical blood tests world-wide. We found that 68.4% of CpG sites and 80% displayed allele-specific expression (ASE. These data demonstrate that ASM is a recurrent phenomenon and is highly correlated with ASE in human PBMCs. Together with recently reported similar studies, our study provides a comprehensive resource for future epigenomic research and confirms new sequencing technology as a paradigm for large-scale epigenomics studies.

  16. Identification of the UBP1 locus as a critical blood pressure determinant using a combination of mouse and human genetics

    DEFF Research Database (Denmark)

    Koutnikova, Hana; Laakso, Markku; Lu, Lu

    2009-01-01

    complementarities of mouse and human genetic approaches, identifies the UBP1 locus as a critical blood pressure determinant. UBP1 plays a role in cholesterol and steroid metabolism via the transcriptional activation of CYP11A, the rate-limiting enzyme in pregnenolone and aldosterone biosynthesis. We suggest......Hypertension is a major health problem of largely unknown genetic origins. To identify new genes responsible for hypertension, genetic analysis of recombinant inbred strains of mice followed by human association studies might prove powerful and was exploited in our current study. Using a set of 27...... recombinant BXD strains of mice we identified a quantitative trait locus (QTL) for blood pressure (BP) on distal chromosome 9. The association analysis of markers encompassing the syntenic region on human chromosome 3 gave in an additive genetic model the strongest association for rs17030583 C/T and rs2291897...

  17. Transcriptional and epigenetic regulation of KIAA1199 gene expression in human breast cancer.

    Directory of Open Access Journals (Sweden)

    Cem Kuscu

    Full Text Available Emerging evidence has demonstrated that upregulated expression of KIAA1199 in human cancer bodes for poor survival. The regulatory mechanism controlling KIAA1199 expression in cancer remains to be characterized. In the present study, we have isolated and characterized the human KIAA1199 promoter in terms of regulation of KIAA1199 gene expression. A 3.3 kb fragment of human genomic DNA containing the 5'-flanking sequence of the KIAA1199 gene possesses both suppressive and activating elements. Employing a deletion mutagenesis approach, a 1.4 kb proximal region was defined as the basic KIAA1199 promoter containing a TATA-box close to the transcription start site. A combination of 5'-primer extension study with 5'RACE DNA sequencing analysis revealed one major transcription start site that is utilized in the human KIAA1199 gene. Bioinformatics analysis suggested that the 1.4 kb KIAA1199 promoter contains putative activating regulatory elements, including activator protein-1(AP-1, Twist-1, and NF-κB sites. Sequential deletion and site-direct mutagenesis analysis demonstrated that the AP-1 and distal NF-κB sites are required for KIAA1199 gene expression. Further analyses using an electrophoretic mobility-shift assay and chromatin immunoprecipitation confirmed the requirement of these cis- and trans-acting elements in controlling KIAA1199 gene expression. Finally, we found that upregulated KIAA1199 expression in human breast cancer specimens correlated with hypomethylation of the regulatory region. Involvement of DNA methylation in regulation of KIAA1199 expression was recapitulated in human breast cancer cell lines. Taken together, our study unraveled the regulatory mechanisms controlling KIAA1199 gene expression in human cancer.

  18. Aluminum induces lipid peroxidation and aggregation of human blood platelets

    Directory of Open Access Journals (Sweden)

    T.J.C. Neiva

    1997-05-01

    Full Text Available Aluminum (Al3+ intoxication is thought to play a major role in the development of Alzheimer's disease and in certain pathologic manifestations arising from long-term hemodialysis. Although the metal does not present redox capacity, it can stimulate tissue lipid peroxidation in animal models. Furthermore, in vitro studies have revealed that the fluoroaluminate complex induces diacylglycerol formation, 43-kDa protein phosphorylation and aggregation. Based on these observations, we postulated that Al3+-induced blood platelet aggregation was mediated by lipid peroxidation. Using chemiluminescence (CL of luminol as an index of total lipid peroxidation capacity, we established a correlation between lipid peroxidation capacity and platelet aggregation. Al3+ (20-100 µM stimulated CL production by human blood platelets as well as their aggregation. Incubation of the platelets with the antioxidants nor-dihydroguaiaretic acid (NDGA (100 µM and n-propyl gallate (NPG (100 µM, inhibitors of the lipoxygenase pathway, completely prevented CL and platelet aggregation. Acetyl salicylic acid (ASA (100 µM, an inhibitor of the cyclooxygenase pathway, was a weaker inhibitor of both events. These findings suggest that Al3+ stimulates lipid peroxidation and the lipoxygenase pathway in human blood platelets thereby causing their aggregation

  19. Complete genes may pass from food to human blood

    DEFF Research Database (Denmark)

    Spisák, Sándor; Solymosi, Norbert; Ittzés, Péter

    2013-01-01

    Our bloodstream is considered to be an environment well separated from the outside world and the digestive tract. According to the standard paradigm large macromolecules consumed with food cannot pass directly to the circulatory system. During digestion proteins and DNA are thought to be degraded...... into small constituents, amino acids and nucleic acids, respectively, and then absorbed by a complex active process and distributed to various parts of the body through the circulation system. Here, based on the analysis of over 1000 human samples from four independent studies, we report evidence that meal......-derived DNA fragments which are large enough to carry complete genes can avoid degradation and through an unknown mechanism enter the human circulation system. In one of the blood samples the relative concentration of plant DNA is higher than the human DNA. The plant DNA concentration shows a surprisingly...

  20. Sialotranscriptomics of Rhipicephalus zambeziensis reveals intricate expression profiles of secretory proteins and suggests tight temporal transcriptional regulation during blood-feeding.

    Science.gov (United States)

    de Castro, Minique Hilda; de Klerk, Daniel; Pienaar, Ronel; Rees, D Jasper G; Mans, Ben J

    2017-08-10

    Ticks secrete a diverse mixture of secretory proteins into the host to evade its immune response and facilitate blood-feeding, making secretory proteins attractive targets for the production of recombinant anti-tick vaccines. The largely neglected tick species, Rhipicephalus zambeziensis, is an efficient vector of Theileria parva in southern Africa but its available sequence information is limited. Next generation sequencing has advanced sequence availability for ticks in recent years and has assisted the characterisation of secretory proteins. This study focused on the de novo assembly and annotation of the salivary gland transcriptome of R. zambeziensis and the temporal expression of secretory protein transcripts in female and male ticks, before the onset of feeding and during early and late feeding. The sialotranscriptome of R. zambeziensis yielded 23,631 transcripts from which 13,584 non-redundant proteins were predicted. Eighty-six percent of these contained a predicted start and stop codon and were estimated to be putatively full-length proteins. A fifth (2569) of the predicted proteins were annotated as putative secretory proteins and explained 52% of the expression in the transcriptome. Expression analyses revealed that 2832 transcripts were differentially expressed among feeding time points and 1209 between the tick sexes. The expression analyses further indicated that 57% of the annotated secretory protein transcripts were differentially expressed. Dynamic expression profiles of secretory protein transcripts were observed during feeding of female ticks. Whereby a number of transcripts were upregulated during early feeding, presumably for feeding site establishment and then during late feeding, 52% of these were downregulated, indicating that transcripts were required at specific feeding stages. This suggested that secretory proteins are under stringent transcriptional regulation that fine-tunes their expression in salivary glands during feeding. No open

  1. Transcriptional profiling of rat skeletal muscle hypertrophy under restriction of blood flow.

    Science.gov (United States)

    Xu, Shouyu; Liu, Xueyun; Chen, Zhenhuang; Li, Gaoquan; Chen, Qin; Zhou, Guoqing; Ma, Ruijie; Yao, Xinmiao; Huang, Xiao

    2016-12-15

    Blood flow restriction (BFR) under low-intensity resistance training (LIRT) can produce similar effects upon muscles to that of high-intensity resistance training (HIRT) while overcoming many of the restrictions to HIRT that occurs in a clinical setting. However, the potential molecular mechanisms of BFR induced muscle hypertrophy remain largely unknown. Here, using a BFR rat model, we aim to better elucidate the mechanisms regulating muscle hypertrophy as induced by BFR and reveal possible clinical therapeutic targets for atrophy cases. We performed genome wide screening with microarray analysis to identify unique differentially expressed genes during rat muscle hypertrophy. We then successfully separated the differentially expressed genes from BRF treated soleus samples by comparing the Affymetrix rat Genome U34 2.0 array with the control. Using qRT-PCR and immunohistochemistry (IHC) we also analyzed other related differentially expressed genes. Results suggested that muscle hypertrophy induced by BFR is essentially regulated by the rate of protein turnover. Specifically, PI3K/AKT and MAPK pathways act as positive regulators in controlling protein synthesis where ubiquitin-proteasome acts as a negative regulator. This represents the first general genome wide level investigation of the gene expression profile in the rat soleus after BFR treatment. This may aid our understanding of the molecular mechanisms regulating and controlling muscle hypertrophy and provide support to the BFR strategies aiming to prevent muscle atrophy in a clinical setting. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Exploring the Limitations of Peripheral Blood Transcriptional Biomarkers in Predicting Influenza Vaccine Responsiveness

    Directory of Open Access Journals (Sweden)

    Luca Marchetti

    2017-01-01

    Full Text Available Systems biology has been recently applied to vaccinology to better understand immunological responses to the influenza vaccine. Particular attention has been paid to the identification of early signatures capable of predicting vaccine immunogenicity. Building from previous studies, we employed a recently established algorithm for signature-based clustering of expression profiles, SCUDO, to provide new insights into why blood-derived transcriptome biomarkers often fail to predict the seroresponse to the influenza virus vaccination. Specifically, preexisting immunity against one or more vaccine antigens, which was found to negatively affect the seroresponse, was identified as a confounding factor able to decouple early transcriptome from later antibody responses, resulting in the degradation of a biomarker predictive power. Finally, the broadly accepted definition of seroresponse to influenza virus vaccine, represented by the maximum response across the vaccine-targeted strains, was compared to a composite measure integrating the responses against all strains. This analysis revealed that composite measures provide a more accurate assessment of the seroresponse to multicomponent influenza vaccines.

  3. Aging affects the transcriptional regulation of human skeletal muscle disuse atrophy

    DEFF Research Database (Denmark)

    Suetta, Charlotte Arneboe; Frandsen, Ulrik; Jensen, Line

    2012-01-01

    Important insights concerning the molecular basis of skeletal muscle disuse-atrophy and aging related muscle loss have been obtained in cell culture and animal models, but these regulatory signaling pathways have not previously been studied in aging human muscle. In the present study, muscle...... atrophy was induced by immobilization in healthy old and young individuals to study the time-course and transcriptional factors underlying human skeletal muscle atrophy. The results reveal that irrespectively of age, mRNA expression levels of MuRF-1 and Atrogin-1 increased in the very initial phase (2......-4 days) of human disuse-muscle atrophy along with a marked reduction in PGC-1α and PGC-1β (1-4 days) and a ∼10% decrease in myofiber size (4 days). Further, an age-specific decrease in Akt and S6 phosphorylation was observed in young muscle within the first days (1-4 days) of immobilization. In contrast...

  4. Study on differential transcriptional profile in human hepatocyte exposed to different doses γ ray

    International Nuclear Information System (INIS)

    Li Jianguo; Wen Jianhua; Duan Zhikai; Tian Yu; Wang Fang; Zuo Yahui

    2009-01-01

    The study analyzed the differential transcriptional profile of normal human hepatic cell and human hepatic cell radiated with three different doses (0.5 Gy, 2 Gy, 4 Gy γ ray) by gene chip technique. The results showed that the whole differentially expressed genes of three different doses have 284 in 14112 human genes analyzed, in which 261 genes were up-regulated and 23 genes were down-regulated. These genes are mainly associated with interferon receptor, mitochondrial regulation, homo sapiens hepatitis A virus cellular receptor, cell cycle regulation, kinase and zinc finger protein etc. RT-PCR results indicated that up-regulated expression of gene HAVcr-1, HAVcr-2, MFTC, MOAP1 and down-regulated expression of gene TRIP12, DCN are consistent with gene chip data. (authors)

  5. Optoelectronic investigation of nanodiamond interactions with human blood

    Science.gov (United States)

    Ficek, M.; Wróbel, M. S.; Wasowicz, M.; Jedrzejewska-Szczerska, M.

    2016-03-01

    We present optoelectronic investigation of in vitro interactions of whole human blood with different nanodiamond biomarkers. Plasmo-chemical modifications of detonation nanodiamond particles gives the possibility for controlling their surface for biological applications. Optical investigations reveal the biological activity of nanodiamonds in blood dependent on its surface termination. We compare different types of nanodiamonds: commercial non-modified detonation nanodiamonds, and nanodiamonds modified by MW PACVD method with H2-termination, and chemically modified nanodiamond with O2-termination. The absorption spectra, and optical microscope investigations were conducted. The results indicate haemocompatibility of non-modified detonation nanodiamond as well as modified nanodiamonds, which enables their application for drug delivery, as well as sensing applications.

  6. Analysis of cytotoxic effects of nickel on human blood lymphocytes.

    Science.gov (United States)

    Zarei, Mohammad Hadi; Hosseini Shirazi, Seyed Farshad; Aghvami, Marjan; Salimi, Ahmad; Pourahmad, Jalal

    2018-02-01

    Nickel compounds possess many applications in different industrial processes. Human beings are exposed to nickel commonly through occupational exposure and food. Although a few studies so far have investigated the effects of nickel compounds on human lymphocytes, the complete mechanism of cytotoxicity of this metal on human lymphocytes is yet to be determined. The intention of this paper was to determine the cytotoxicity mechanism of water soluble NiCl 2 toward human lymphocytes using the accelerated cytotoxicity mechanisms screening (ACMS) technique. Human lymphocytes were isolated from the blood of healthy subjects based on Ficoll-Paque PLUS standard method. For the assessment of cell viability, lymphocytes were incubated with 0.05-1 mM NiCl 2 for 12 h. Determination of mechanistic parameters was performed 2, 4 and 6 h after treatment of cells with ½ EC50 12h , EC50 12h and 2EC50 12h of NiCl 2 . Our results demonstrate that cytotoxicity of NiCl 2 on human lymphocytes is associated with increased ROS formation, mitochondrial membrane potential collapse, glutathione depletion, lysosomal membrane damage, cellular proteolysis and activation of caspase-3 before cytotoxicity ensued.

  7. A repertoire of the dominant transcripts from the salivary glands of the blood-sucking bug, Triatoma dimidiata, a vector of Chagas disease

    Science.gov (United States)

    Kato, Hirotomo; Jochim, Ryan C.; Gomez, Eduardo A.; Sakoda, Ryo; Iwata, Hiroyuki; Valenzuela, Jesus G.; Hashiguchi, Yoshihisa

    2010-01-01

    Triatoma (T.) dimidiata is a hematophagous Hemiptera and a main vector of Chagas disease. The saliva of this and other blood-sucking insects contains potent pharmacologically active components that assist them in counteracting the host hemostatic and inflammatory systems during blood feeding. To describe the repertoire of potential bioactive salivary molecules from this insect, a number of randomly selected transcripts from the salivary gland cDNA library of T. dimidiata were sequenced and analyzed. This analysis showed that 77.5% of the isolated transcripts coded for putative secreted proteins, and 89.9% of these coded for variants of the lipocalin family proteins. The most abundant transcript was a homologue of procalin, the major allergen of T. protracta saliva, and contributed more than 50% of the transcripts coding for putative secreted proteins, suggesting that it may play an important role in the blood-feeding process. Other salivary transcripts encoding lipocalin family proteins had homology to triabin (a thrombin inhibitor), triafestin (an inhibitor of kallikrein–kinin system), pallidipin (an inhibitor of collagen-induced platelet aggregation) and others with unknown function. PMID:19900580

  8. Preclinical evaluation of transcriptional targeting strategy for human hepatocellular carcinoma in an orthotopic xenograft mouse model.

    Science.gov (United States)

    Sia, Kian Chuan; Huynh, Hung; Chung, Alexander Yaw Fui; Ooi, London Lucien Peng Jin; Lim, Kiat Hon; Hui, Kam Man; Lam, Paula Yeng Po

    2013-08-01

    Gene regulation of many key cell-cycle players in S-, G(2) phase, and mitosis results from transcriptional repression in their respective promoter regions during the G(0) and G(1) phases of cell cycle. Within these promoter regions are phylogenetically conserved sequences known as the cell-cycle-dependent element (CDE) and cell-cycle genes homology regions (CHR) sites. Thus, we hypothesize that transcriptional regulation of cell-cycle regulation via the CDE/CHR region together with liver-specific apolipoprotein E (apoE)-hAAT promoter could bring about a selective transgene expression in proliferating human hepatocellular carcinoma. We show that the newly generated vector AH-6CC-L2C could mediate hepatocyte-targeted luciferase gene expression in tumor cells and freshly isolated short-term hepatocellular carcinoma cultures from patient biopsy. In contrast, normal murine and human hepatocytes infected with AH-6CC-L2C expressed minimal or low luciferase activities. In the presence of prodrug 5-fluorocytosine (5-FC), AH-6CC-L2C effectively suppressed the growth of orthotopic hepatocellular carcinoma patient-derived xenograft mouse model via the expression of yeast cytosine deaminase (yCD) that converts 5-FC to anticancer metabolite 5-fluoruracil. More importantly, we show that combination treatment of AH-6CC-L2C with an EZH2 inhibitor, DZNep, that targets EpCAM-positive hepatocellular carcinoma, can bring about a greater therapeutic efficacy compared with a single treatment of virus or inhibitor. Our study showed that targeting proliferating human hepatocellular carcinoma cells through the transcriptional control of therapeutic gene could represent a feasible approach against hepatocellular carcinoma.

  9. Transcriptional factor PU.1 regulates decidual C1q expression in early pregnancy in human

    Directory of Open Access Journals (Sweden)

    Priyaa Madhukaran Raj

    2015-02-01

    Full Text Available C1q is the first recognition subcomponent of the complement classical pathway, which in addition to being synthesized in the liver, is also expressed by macrophages and dendritic cells. Trophoblast invasion during early placentation results in accumulation of debris that triggers the complement system. Hence, both early and late components of the classical pathway are widely distributed in the placenta and decidua. In addition, C1q has recently been shown to significantly contribute to feto-maternal tolerance, trophoblast migration, and spiral artery remodeling, although the exact mechanism remains unknown. Pregnancy in mice, genetically deficient in C1q, mirrors symptoms similar to that of human preeclampsia. Thus, regulated complement activation has been proposed as an essential requirement for normal successful pregnancy. Little is known about the molecular pathways that regulate C1q expression in pregnancy. PU.1, an Ets-family transcription factor, is required for the development of hematopoietic myeloid lineage immune cells, and its expression is tissue- specific. Recently, PU.1 has been shown to regulate C1q gene expression in dendritic cells and macrophages. Here, we have examined if PU.1 transcription factor regulates decidual C1q expression. We used immune-histochemical analysis, PCR and immunostaining to localize and study the gene expression of PU.1 transcription factor in early human decidua. PU.1 was highly expressed at gene and protein level in early human decidual cells including trophoblast and stromal cells. Surprisingly, nuclear as well as cytoplasmic PU.1 expression was observed. Decidual cells with predominantly nuclear PU.1 expression had higher C1q expression. It is likely that nuclear and cytoplasmic PU.1 localization has a role to play in early pregnancy via regulating C1q expression in the decidua during implantation.

  10. Candidate gene biodosimeters of mice and human exposure to ionizing radiation by quantitative reverse transcription polymerase chain reaction

    Directory of Open Access Journals (Sweden)

    Hamed Rezaeejam

    2015-01-01

    Full Text Available Understanding of cellular responses to ionizing radiation (IR is essential for the development of predictive markers useful for assessing human exposure. Biological markers of exposure to IR in human populations are of great interest for assessing normal tissue injury in radiation oncology and for biodosimetry in nuclear incidents and accidental radiation exposures. Traditional radiation exposure biomarkers based on cytogenetic assays (biodosimetry, are time-consuming and do not provide results fast enough and requires highly trained personnel for scoring. Hence, the development of rapid biodosimetry methods is one of the highest priorities. Exposure of cells to IR activates multiple signal transduction pathways, which result in complex alterations in gene-expression. Real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR has become the benchmark for the detection and quantification of RNA targets and is being utilized increasingly in monitoring the specific genes with more accurately and sensitively. This review evaluates the RT-qPCR as a biodosimetry method and we investigated the papers from 2000 up to now, which identified the genes-expression related the DNA repair, cell cycle checkpoint, and apoptosis induced by ionization radiation in peripheral blood and determined as biodosimeters. In conclusion, it could be say that RT-qPCR technique for determining the specific genes as biodosimeters could be a fully quantitative reliable and sensitive method. Furthermore, the results of the current review will help the researchers to recognize the most expressed genes induced by ionization radiation.

  11. Single-Cell Analysis of Human Pancreas Reveals Transcriptional Signatures of Aging and Somatic Mutation Patterns.

    Science.gov (United States)

    Enge, Martin; Arda, H Efsun; Mignardi, Marco; Beausang, John; Bottino, Rita; Kim, Seung K; Quake, Stephen R

    2017-10-05

    As organisms age, cells accumulate genetic and epigenetic errors that eventually lead to impaired organ function or catastrophic transformation such as cancer. Because aging reflects a stochastic process of increasing disorder, cells in an organ will be individually affected in different ways, thus rendering bulk analyses of postmitotic adult cells difficult to interpret. Here, we directly measure the effects of aging in human tissue by performing single-cell transcriptome analysis of 2,544 human pancreas cells from eight donors spanning six decades of life. We find that islet endocrine cells from older donors display increased levels of transcriptional noise and potential fate drift. By determining the mutational history of individual cells, we uncover a novel mutational signature in healthy aging endocrine cells. Our results demonstrate the feasibility of using single-cell RNA sequencing (RNA-seq) data from primary cells to derive insights into genetic and transcriptional processes that operate on aging human tissue. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Seroprevalence of human parvovirus B19 in healthy blood donors.

    Science.gov (United States)

    Kumar, Satish; Gupta, R M; Sen, Sourav; Sarkar, R S; Philip, J; Kotwal, Atul; Sumathi, S H

    2013-07-01

    Human parvovirus B19 is an emerging transfusion transmitted infection. Although parvovirus B19 infection is connected with severe complications in some recipients, donor screening is not yet mandatory. To reduce the risk of contamination, plasma-pool screening and exclusion of highly viraemic donations are recommended. In this study the prevalence of parvovirus B19 in healthy blood donors was detected by ELISA. A total of 1633 samples were screened for IgM and IgG antibodies against parvovirus B19 by ELISA. The initial 540 samples were screened for both IgM and IgG class antibodies and remaining 1093 samples were screened for only IgM class antibodies by ELISA. Net prevalence of IgM antibodies to human parvovirus B19 in our study was 7.53% and prevalence of IgG antibodies was 27.96%. Dual positivity (IgG and IgM) was 2.40%. The seroprevalence of human parvovirus B19 among blood donor population in our study is high, and poses an adverse transfusion risk especially in high-risk group of patients who have no detectable antibodies to B19. Studies with large sample size are needed to validate these results.

  13. Genomic localization, sequence analysis, and transcription of the putative human cytomegalovirus DNA polymerase gene

    International Nuclear Information System (INIS)

    Heilbronn, T.; Jahn, G.; Buerkle, A.; Freese, U.K.; Fleckenstein, B.; Zur Hausen, H.

    1987-01-01

    The human cytomegalovirus (HCMV)-induced DNA polymerase has been well characterized biochemically and functionally, but its genomic location has not yet been assigned. To identify the coding sequence, cross-hybridization with the herpes simplex virus type 1 (HSV-1) polymerase gene was used, as suggested by the close similarity of the herpes group virus-induced DNA polymerases to the HCMV DNA polymerase. A cosmid and plasmid library of the entire HCMV genome was screened with the BamHI Q fragment of HSF-1 at different stringency conditions. One PstI-HincII restriction fragment of 850 base pairs mapping within the EcoRI M fragment of HCMV cross-hybridized at T/sub m/ - 25/degrees/C. Sequence analysis revealed one open reading frame spanning the entire sequence. The amino acid sequence showed a highly conserved domain of 133 amino acids shared with the HSV and putative Esptein-Barr virus polymerase sequences. This domain maps within the C-terminal part of the HSV polymerase gene, which has been suggested to contain part of the catalytic center of the enzyme. Transcription analysis revealed one 5.4-kilobase early transcript in the sense orientation with respect to the open reading frame identified. This transcript appears to code for the 140-kilodalton HCMV polymerase protein

  14. Low prevalence of transcriptionally active human papilloma virus in Indian patients with HNSCC and leukoplakia.

    Science.gov (United States)

    Bhosale, Priyanka G; Pandey, Manishkumar; Desai, Rajiv S; Patil, Asawari; Kane, Shubhada; Prabhash, Kumar; Mahimkar, Manoj B

    2016-11-01

    In the present study, we comprehensively analyzed the prevalence of transcriptionally active human papilloma virus (HPV) in tissue samples of Indian patients with leukoplakia, predominantly hyperplastic lesions and head and neck squamous cell carcinoma (HNSCC). In addition, saliva samples from patients with HNSCC were screened for HPV detection. P16 overexpression was analyzed by immunohistochemistry. Tissue samples of leukoplakia (n = 121) and HNSCC (n = 427) and saliva from patients with HNSCC (n = 215) were tested for HPV using nested polymerase chain reaction. Positive samples were sequenced for subtyping. The presence of HPV E6/E7 mRNA was confirmed by RNA in situ hybridization. P16 expression and HPV DNA were not detected in any of the leukoplakia specimens. Of the 427 HNSCC tumors, 9 showed p16 overexpression and 7/427 cases were positive for HPV16 DNA, in saliva or tissue. E6/E7 mRNA positivity was observed in 8 HNSCC samples, primarily from patients with no habit of tobacco consumption. The prevalence of high-risk HPV was restricted to oropharynx and larynx, with very little concordance between p16 overexpression and HPV positivity. All patients with HPV-positive saliva samples had transcriptionally active HPV present in their tumors. The presence of HPV DNA does not necessarily reflect transcriptionally active virus in tumors; hence, it is important to consider this fact while categorizing HPV-associated tumors. Copyright © 2016 Elsevier Inc. All rights reserved.

  15. Mycobacterium leprae induces NF-κB-dependent transcription repression in human Schwann cells

    International Nuclear Information System (INIS)

    Pereira, Renata M.S.; Calegari-Silva, Teresa Cristina; Hernandez, Maristela O.; Saliba, Alessandra M.; Redner, Paulo; Pessolani, Maria Cristina V.; Sarno, Euzenir N.; Sampaio, Elizabeth P.; Lopes, Ulisses G.

    2005-01-01

    Mycobacterium leprae, the causative agent of leprosy, invades peripheral nerve Schwann cells, resulting in deformities associated with this disease. NF-κB is an important transcription factor involved in the regulation of host immune antimicrobial responses. We aimed in this work to investigate NF-κB signaling pathways in the human ST88-14 Schwannoma cell line infected with M. leprae. Gel shift and supershift assays indicate that two NF-κB dimers, p65/p50 and p50/p50, translocate to the nucleus in Schwann cells treated with lethally irradiated M. leprae. Consistent with p65/p50 and p50/p50 activation, we observed IκB-α degradation and reduction of p105 levels. The nuclear translocation of p50/p50 complex due to M. leprae treatment correlated with repression of NF-κB-driven transcription induced by TNF-α. Moreover, thalidomide inhibited p50 homodimer nuclear translocation induced by M. leprae and consequently rescues Schwann cells from NF-κB-dependent transcriptional repression. Here, we report for the first time that M. leprae induces NF-κB activation in Schwann cells and thalidomide is able to modulate this activation

  16. Transcriptional interactions suggest niche segregation among microorganisms in the human gut

    DEFF Research Database (Denmark)

    Plichta, Damian Rafal; Juncker, Agnieszka; dos Santos, Marcelo Bertalan Quintanilha

    2016-01-01

    The human gastrointestinal (GI) tract is the habitat for hundreds of microbial species, of which many cannot be cultivated readily, presumably because of the dependencies between species 1. Studies of microbial co-occurrence in the gut have indicated community substructures that may reflect...... functional and metabolic interactions between cohabiting species 2,3. To move beyond species co-occurrence networks, we systematically identified transcriptional interactions between pairs of coexisting gut microbes using metagenomics and microarray-based metatranscriptomics data from 233 stool samples from...

  17. Outbreak of hepatitis E virus infection in Darfur, Sudan: effectiveness of real-time reverse transcription-PCR analysis of dried blood spots.

    Science.gov (United States)

    Mérens, Audrey; Guérin, Philippe Jean; Guthmann, Jean-Paul; Nicand, Elisabeth

    2009-06-01

    Biological samples collected in refugee camps during an outbreak of hepatitis E were used to compare the accuracy of hepatitis E virus RNA amplification by real-time reverse transcription-PCR (RT-PCR) for sera and dried blood spots (concordance of 90.6%). Biological profiles (RT-PCR and serology) of asymptomatic individuals were also analyzed.

  18. Paper-based device for rapid typing of secondary human blood groups.

    Science.gov (United States)

    Li, Miaosi; Then, Whui Lyn; Li, Lizi; Shen, Wei

    2014-01-01

    We report the use of bioactive paper for typing of secondary human blood groups. Our recent work on using bioactive paper for human blood typing has led to the discovery of a new method for identifying haemagglutination of red blood cells. The primary human blood groups, i.e., ABO and RhD groups, have been successfully typed with this method. Clinically, however, many secondary blood groups can also cause fatal blood transfusion accidents, despite the fact that the haemagglutination reactions of secondary blood groups are generally weaker than those of the primary blood groups. We describe the design of a user-friendly sensor for rapid typing of secondary blood groups using bioactive paper. We also present mechanistic insights into interactions between secondary blood group antibodies and red blood cells obtained using confocal microscopy. Haemagglutination patterns under different conditions are revealed for optimization of the assay conditions.

  19. DDX11L: a novel transcript family emerging from human subtelomeric regions

    Directory of Open Access Journals (Sweden)

    D'Urso Michele

    2009-05-01

    Full Text Available Abstract Background The subtelomeric regions of human chromosomes exhibit an extraordinary plasticity. To date, due to the high GC content and to the presence of telomeric repeats, the subtelomeric sequences are underrepresented in the genomic libraries and consequently their sequences are incomplete in the finished human genome sequence, and still much remains to be learned about subtelomere organization, evolution and function. Indeed, only in recent years, several studies have disclosed, within human subtelomeres, novel gene family members. Results During a project aimed to analyze genes located in the telomeric region of the long arm of the human X chromosome, we have identified a novel transcript family, DDX11L, members of which map to 1pter, 2q13/14.1, 2qter, 3qter, 6pter, 9pter/9qter, 11pter, 12pter, 15qter, 16pter, 17pter, 19pter, 20pter/20qter, Xpter/Xqter and Yqter. Furthermore, we partially sequenced the underrepresented subtelomeres of human chromosomes showing a common evolutionary origin. Conclusion Our data indicate that an ancestral gene, originated as a rearranged portion of the primate DDX11 gene, and propagated along many subtelomeric locations, is emerging within subtelomeres of human chromosomes, defining a novel gene family. These findings support the possibility that the high plasticity of these regions, sites of DNA exchange among different chromosomes, could trigger the emergence of new genes.

  20. Whole genome transcription profiling of Anaplasma phagocytophilum in human and tick host cells by tiling array analysis

    Directory of Open Access Journals (Sweden)

    Chavez Adela

    2008-07-01

    Full Text Available Abstract Background Anaplasma phagocytophilum (Ap is an obligate intracellular bacterium and the agent of human granulocytic anaplasmosis, an emerging tick-borne disease. Ap alternately infects ticks and mammals and a variety of cell types within each. Understanding the biology behind such versatile cellular parasitism may be derived through the use of tiling microarrays to establish high resolution, genome-wide transcription profiles of the organism as it infects cell lines representative of its life cycle (tick; ISE6 and pathogenesis (human; HL-60 and HMEC-1. Results Detailed, host cell specific transcriptional behavior was revealed. There was extensive differential Ap gene transcription between the tick (ISE6 and the human (HL-60 and HMEC-1 cell lines, with far fewer differentially transcribed genes between the human cell lines, and all disproportionately represented by membrane or surface proteins. There were Ap genes exclusively transcribed in each cell line, apparent human- and tick-specific operons and paralogs, and anti-sense transcripts that suggest novel expression regulation processes. Seven virB2 paralogs (of the bacterial type IV secretion system showed human or tick cell dependent transcription. Previously unrecognized genes and coding sequences were identified, as were the expressed p44/msp2 (major surface proteins paralogs (of 114 total, through elevated signal produced to the unique hypervariable region of each – 2/114 in HL-60, 3/114 in HMEC-1, and none in ISE6. Conclusion Using these methods, whole genome transcription profiles can likely be generated for Ap, as well as other obligate intracellular organisms, in any host cells and for all stages of the cell infection process. Visual representation of comprehensive transcription data alongside an annotated map of the genome renders complex transcription into discernable patterns.

  1. Immunosuppressive activity of human cord-blood lipoproteins

    International Nuclear Information System (INIS)

    Forte, T.M.; Davis, P.A.; Curtiss, L.K.

    1985-01-01

    It is now known that the role of plasma lipoproteins is multifunctional. More recently it has been shown that lipoproteins may regulate immune responses as well. Low-density lipoproteins carrying apolipoprotein B (apoB) are known to suppress phytohemagglutinin (PHA) activated lymphocytes by inhibiting DNA synthesis. More recently, an immunoregulatory role has been described for another apolipoprotein, apoE, which is found in low quantities in normal plasma. In these studies with human umbilical-cord blood the authors were intrigued by two factors: the low level of LDL and hence apoB, and the elevated quantity of apoE. This study examines the hypothesis that apoE may regulate lymphocyte function in the human fetus

  2. Simple, accurate equations for human blood O2 dissociation computations.

    Science.gov (United States)

    Severinghaus, J W

    1979-03-01

    Hill's equation can be slightly modified to fit the standard human blood O2 dissociation curve to within plus or minus 0.0055 fractional saturation (S) from O less than S less than 1. Other modifications of Hill's equation may be used to compute Po2 (Torr) from S (Eq. 2), and the temperature coefficient of Po2 (Eq. 3). Variations of the Bohr coefficient with Po2 are given by Eq. 4. S = (((Po2(3) + 150 Po2)(-1) x 23,400) + 1)(-1) (1) In Po2 = 0.385 In (S-1 - 1)(-1) + 3.32 - (72 S)(-1) - 0.17(S6) (2) DELTA In Po2/delta T = 0.058 ((0.243 X Po2/100)(3.88) + 1)(-1) + 0.013 (3) delta In Po2/delta pH = (Po2/26.6)(0.184) - 2.2 (4) Procedures are described to determine Po2 and S of blood iteratively after extraction or addition of a defined amount of O2 and to compute P50 of blood from a single sample after measuring Po2, pH, and S.

  3. Transcriptional Inhibition of the Human Papilloma Virus Reactivates Tumor Suppressor p53 in Cervical Carcinoma Cells

    Science.gov (United States)

    Kochetkov, D. V.; Ilyinskaya, G. V.; Komarov, P. G.; Strom, E.; Agapova, L. S.; Ivanov, A. V.; Budanov, A. V.; Frolova, E. I.; Chumakov, P. M.

    2009-01-01

    Inactivation of tumor suppressor p53 accompanies the majority of human malignancies. Restoration of p53 function causes death of tumor cells and is potentially suitable for gene therapy of cancer. In cervical carcinoma, human papilloma virus (HPV) E6 facilitates proteasomal degradation of p53. Hence, a possible approach to p53 reactivation is the use of small molecules suppressing the function of viral proteins. HeLa cervical carcinoma cells (HPV-18) with a reporter construct containing the b-galactosidase gene under the control of a p53-responsive promoter were used as a test system to screen a library of small molecules for restoration of the transcriptional activity of p53. The effect of the two most active compounds was studied with cell lines differing in the state of p53-dependent signaling pathways. The compounds each specifically activated p53 in cells expressing HPV-18 and, to a lesser extent, HPV-16 and exerted no effect on control p53-negative cells or cells with the intact p53-dependent pathways. Activation of p53 in cervical carcinoma cells was accompanied by induction of p53-dependent CDKN1 (p21), inhibition of cell proliferation, and induction of apoptosis. In addition, the two compounds dramatically decreased transcription of the HPV genome, which was assumed to cause p53 reactivation. The compounds were low-toxic for normal cells and can be considered as prototypes of new anticancer drugs. PMID:17685229

  4. Distinct Contributions of Replication and Transcription to Mutation Rate Variation of Human Genomes

    KAUST Repository

    Cui, Peng; Ding, Feng; Lin, Qiang; Zhang, Lingfang; Li, Ang; Zhang, Zhang; Hu, Songnian; Yu, Jun

    2012-01-01

    Here, we evaluate the contribution of two major biological processes—DNA replication and transcription—to mutation rate variation in human genomes. Based on analysis of the public human tissue transcriptomics data, high-resolution replicating map of Hela cells and dbSNP data, we present significant correlations between expression breadth, replication time in local regions and SNP density. SNP density of tissue-specific (TS) genes is significantly higher than that of housekeeping (HK) genes. TS genes tend to locate in late-replicating genomic regions and genes in such regions have a higher SNP density compared to those in early-replication regions. In addition, SNP density is found to be positively correlated with expression level among HK genes. We conclude that the process of DNA replication generates stronger mutational pressure than transcription-associated biological processes do, resulting in an increase of mutation rate in TS genes while having weaker effects on HK genes. In contrast, transcription-associated processes are mainly responsible for the accumulation of mutations in highly-expressed HK genes.

  5. Transcription factors and stress response gene alterations in human keratinocytes following Solar Simulated Ultra Violet Radiation.

    Science.gov (United States)

    Marais, Thomas L Des; Kluz, Thomas; Xu, Dazhong; Zhang, Xiaoru; Gesumaria, Lisa; Matsui, Mary S; Costa, Max; Sun, Hong

    2017-10-19

    Ultraviolet radiation (UVR) from sunlight is the major effector for skin aging and carcinogenesis. However, genes and pathways altered by solar-simulated UVR (ssUVR), a mixture of UVA and UVB, are not well characterized. Here we report global changes in gene expression as well as associated pathways and upstream transcription factors in human keratinocytes exposed to ssUVR. Human HaCaT keratinocytes were exposed to either a single dose or 5 repetitive doses of ssUVR. Comprehensive analyses of gene expression profiles as well as functional annotation were performed at 24 hours post irradiation. Our results revealed that ssUVR modulated genes with diverse cellular functions changed in a dose-dependent manner. Gene expression in cells exposed to a single dose of ssUVR differed significantly from those that underwent repetitive exposures. While single ssUVR caused a significant inhibition in genes involved in cell cycle progression, especially G2/M checkpoint and mitotic regulation, repetitive ssUVR led to extensive changes in genes related to cell signaling and metabolism. We have also identified a panel of ssUVR target genes that exhibited persistent changes in gene expression even at 1 week after irradiation. These results revealed a complex network of transcriptional regulators and pathways that orchestrate the cellular response to ssUVR.

  6. Distinct Contributions of Replication and Transcription to Mutation Rate Variation of Human Genomes

    KAUST Repository

    Cui, Peng

    2012-03-23

    Here, we evaluate the contribution of two major biological processes—DNA replication and transcription—to mutation rate variation in human genomes. Based on analysis of the public human tissue transcriptomics data, high-resolution replicating map of Hela cells and dbSNP data, we present significant correlations between expression breadth, replication time in local regions and SNP density. SNP density of tissue-specific (TS) genes is significantly higher than that of housekeeping (HK) genes. TS genes tend to locate in late-replicating genomic regions and genes in such regions have a higher SNP density compared to those in early-replication regions. In addition, SNP density is found to be positively correlated with expression level among HK genes. We conclude that the process of DNA replication generates stronger mutational pressure than transcription-associated biological processes do, resulting in an increase of mutation rate in TS genes while having weaker effects on HK genes. In contrast, transcription-associated processes are mainly responsible for the accumulation of mutations in highly-expressed HK genes.

  7. UVB induces IL-12 transcription in human keratinocytes in vivo and in vitro

    International Nuclear Information System (INIS)

    Enk, C.D.; Blauvet, A.; Katz, S.I.; Mahanty, S.

    1996-01-01

    Human epidermal cells produce a wide range of cytokines, including those characteristic of Th2-like responses such as interleukin (IL)-4 and IL-10. As well, keratinocytes have recently been shown to produce Th1-like cytokines such as IL-12. Exposure to UVB has profound effects on the skin and systemic immune system, which is in part mediated by secretion of tumor necrosis factor (TNF)-α by epidermal cells. Because IL-12 induces production of TNF-α by certain cells of the immune system, we sought to determine whether UVB is an inducer of IL-12 gene expression in epidermal cells. Human epidermal cells were exposed to UVB radiation in vivo, isolated by suction blister technique and trypsinization and transcription of the IL-12 p35 and p40 chains was examined by RT-PCR. (Author)

  8. Genomic binding profiles of functionally distinct RNA polymerase III transcription complexes in human cells.

    Science.gov (United States)

    Moqtaderi, Zarmik; Wang, Jie; Raha, Debasish; White, Robert J; Snyder, Michael; Weng, Zhiping; Struhl, Kevin

    2010-05-01

    Genome-wide occupancy profiles of five components of the RNA polymerase III (Pol III) machinery in human cells identified the expected tRNA and noncoding RNA targets and revealed many additional Pol III-associated loci, mostly near short interspersed elements (SINEs). Several genes are targets of an alternative transcription factor IIIB (TFIIIB) containing Brf2 instead of Brf1 and have extremely low levels of TFIIIC. Strikingly, expressed Pol III genes, unlike nonexpressed Pol III genes, are situated in regions with a pattern of histone modifications associated with functional Pol II promoters. TFIIIC alone associates with numerous ETC loci, via the B box or a novel motif. ETCs are often near CTCF binding sites, suggesting a potential role in chromosome organization. Our results suggest that human Pol III complexes associate preferentially with regions near functional Pol II promoters and that TFIIIC-mediated recruitment of TFIIIB is regulated in a locus-specific manner.

  9. Infrared spectra in monitoring biochemical parameters of human blood

    International Nuclear Information System (INIS)

    Prabhakar, S; Singh, R A; Jain, N

    2012-01-01

    Infrared spectroscopy is gaining recognition as a promising method. The infrared spectra of selected regions (2000-400cm -1 ) of blood tissue samples are reported. Present study related to the role of spectral peak fitting in the study of human blood and quantitative interpretations of infrared spectra based on chemometrics. The spectral variations are interpreted in terms of the biochemical and pathological processes involved. The mean RNA/DNA ratio of fitted intensities and analytical area as calculated from the transmittance peaks at 1121cm -1 /1020cm -1 is found to be 0.911A.U and 2.00A.U. respectively. The ratio of 1659cm -1 /1544cm -1 (amide-I/amide-II) bands is found to shed light on the change in the DNA content. The ratio of amide-I/amide-II is almost unity (≅1.054) for blood spectra. The deviation from unity is an indication of DNA absorption from the RBC cells. The total phosphate content has found to be 25.09A.U. The level for glycogen/phosphate ratio (areas under peaks 1030cm -1 /1082cm -1 ) is found to be 0.286A.U. The ratio of unsaturated and saturated carbonyl compounds (C=O) in blood samples is in form of esters and the analytical areas under the spectral peaks at 1740cm -1 and 1731cm -1 for unsaturated esters and saturated esters respectively found to be 0.618A.U.

  10. The in-vitro study of human blood leukemic cells by pulsed NMR

    International Nuclear Information System (INIS)

    Zulkarnaen, M.; Munawir; Wibowo, Tono; Suyitno, Gogot

    1983-01-01

    The diagram of leukemic cells in human blood has been studied by using the NMR longitudinal relaxation technique. The observation was treated in whole blood, serum and blood cell. Every result was compared with previous observation and show that the values of the proton longitudinal relaxation in the leukemic whole blood almost twice or more that of normal blood, while in the serum and the blood cell, the values are nearly the same. (author)

  11. Friends-enemies: endogenous retroviruses are major transcriptional regulators of human DNA

    Science.gov (United States)

    Buzdin, Anton A.; Prassolov, Vladimir; Garazha, Andrew V.

    2017-06-01

    Endogenous retroviruses are mobile genetic elements hardly distinguishable from infectious, or “exogenous”, retroviruses at the time of insertion in the host DNA. Human endogenous retroviruses (HERVs) are not rare. They gave rise to multiple families of closely related mobile elements that occupy 8% of the human genome. Together, they shape genomic regulatory landscape by providing at least 320,000 human transcription factor binding sites (TFBS) located on 110,000 individual HERV elements. The HERVs host as many as 155,000 mapped DNaseI hypersensitivity sites, which denote loci active in the regulation of gene expression or chromatin structure. The contemporary view of the HERVs evolutionary dynamics suggests that at the early stages after insertion, the HERV is treated by the host cells as a foreign genetic element, and is likely to be suppressed by the targeted methylation and mutations. However, at the later stages, when significant number of mutations has been already accumulated and when the retroviral genes are broken, the regulatory potential of a HERV may be released and recruited to modify the genomic balance of transcription factor binding sites. This process goes together with further accumulation and selection of mutations, which reshape the regulatory landscape of the human DNA. However, developmental reprogramming, stress or pathological conditions like cancer, inflammation and infectious diseases, can remove the blocks limiting expression and HERV-mediated host gene regulation. This, in turn, can dramatically alter the gene expression equilibrium and shift it to a newer state, thus further amplifying instability and exacerbating the stressful situation.

  12. Deciphering the transcriptional circuitry of microRNA genes expressed during human monocytic differentiation

    KAUST Repository

    Schmeier, Sebastian; MacPherson, Cameron R; Essack, Magbubah; Kaur, Mandeep; Schaefer, Ulf; Suzuki, Harukazu; Hayashizaki, Yoshihide; Bajic, Vladimir B.

    2009-01-01

    Background: Macrophages are immune cells involved in various biological processes including host defence, homeostasis, differentiation, and organogenesis. Disruption of macrophage biology has been linked to increased pathogen infection, inflammation and malignant diseases. Differential gene expression observed in monocytic differentiation is primarily regulated by interacting transcription factors (TFs). Current research suggests that microRNAs (miRNAs) degrade and repress translation of mRNA, but also may target genes involved in differentiation. We focus on getting insights into the transcriptional circuitry regulating miRNA genes expressed during monocytic differentiation. Results: We computationally analysed the transcriptional circuitry of miRNA genes during monocytic differentiation using in vitro time-course expression data for TFs and miRNAs. A set of TF?miRNA associations was derived from predicted TF binding sites in promoter regions of miRNA genes. Time-lagged expression correlation analysis was utilised to evaluate the TF?miRNA associations. Our analysis identified 12 TFs that potentially play a central role in regulating miRNAs throughout the differentiation process. Six of these 12 TFs (ATF2, E2F3, HOXA4, NFE2L1, SP3, and YY1) have not previously been described to be important for monocytic differentiation. The remaining six TFs are CEBPB, CREB1, ELK1, NFE2L2, RUNX1, and USF2. For several miRNAs (miR-21, miR-155, miR-424, and miR-17-92), we show how their inferred transcriptional regulation impacts monocytic differentiation. Conclusions: The study demonstrates that miRNAs and their transcriptional regulatory control are integral molecular mechanisms during differentiation. Furthermore, it is the first study to decipher on a large-scale, how miRNAs are controlled by TFs during human monocytic differentiation. Subsequently, we have identified 12 candidate key controllers of miRNAs during this differentiation process. 2009 Schmeier et al; licensee Bio

  13. Deciphering the transcriptional circuitry of microRNA genes expressed during human monocytic differentiation

    KAUST Repository

    Schmeier, Sebastian

    2009-12-10

    Background: Macrophages are immune cells involved in various biological processes including host defence, homeostasis, differentiation, and organogenesis. Disruption of macrophage biology has been linked to increased pathogen infection, inflammation and malignant diseases. Differential gene expression observed in monocytic differentiation is primarily regulated by interacting transcription factors (TFs). Current research suggests that microRNAs (miRNAs) degrade and repress translation of mRNA, but also may target genes involved in differentiation. We focus on getting insights into the transcriptional circuitry regulating miRNA genes expressed during monocytic differentiation. Results: We computationally analysed the transcriptional circuitry of miRNA genes during monocytic differentiation using in vitro time-course expression data for TFs and miRNAs. A set of TF?miRNA associations was derived from predicted TF binding sites in promoter regions of miRNA genes. Time-lagged expression correlation analysis was utilised to evaluate the TF?miRNA associations. Our analysis identified 12 TFs that potentially play a central role in regulating miRNAs throughout the differentiation process. Six of these 12 TFs (ATF2, E2F3, HOXA4, NFE2L1, SP3, and YY1) have not previously been described to be important for monocytic differentiation. The remaining six TFs are CEBPB, CREB1, ELK1, NFE2L2, RUNX1, and USF2. For several miRNAs (miR-21, miR-155, miR-424, and miR-17-92), we show how their inferred transcriptional regulation impacts monocytic differentiation. Conclusions: The study demonstrates that miRNAs and their transcriptional regulatory control are integral molecular mechanisms during differentiation. Furthermore, it is the first study to decipher on a large-scale, how miRNAs are controlled by TFs during human monocytic differentiation. Subsequently, we have identified 12 candidate key controllers of miRNAs during this differentiation process. 2009 Schmeier et al; licensee Bio

  14. Human Galectin-9 Is a Potent Mediator of HIV Transcription and Reactivation.

    Directory of Open Access Journals (Sweden)

    Mohamed Abdel-Mohsen

    2016-06-01

    Full Text Available Identifying host immune determinants governing HIV transcription, latency and infectivity in vivo is critical to developing an HIV cure. Based on our recent finding that the host factor p21 regulates HIV transcription during antiretroviral therapy (ART, and published data demonstrating that the human carbohydrate-binding immunomodulatory protein galectin-9 regulates p21, we hypothesized that galectin-9 modulates HIV transcription. We report that the administration of a recombinant, stable form of galectin-9 (rGal-9 potently reverses HIV latency in vitro in the J-Lat HIV latency model. Furthermore, rGal-9 reverses HIV latency ex vivo in primary CD4+ T cells from HIV-infected, ART-suppressed individuals (p = 0.002, more potently than vorinostat (p = 0.02. rGal-9 co-administration with the latency reversal agent "JQ1", a bromodomain inhibitor, exhibits synergistic activity (p<0.05. rGal-9 signals through N-linked oligosaccharides and O-linked hexasaccharides on the T cell surface, modulating the gene expression levels of key transcription initiation, promoter proximal-pausing, and chromatin remodeling factors that regulate HIV latency. Beyond latent viral reactivation, rGal-9 induces robust expression of the host antiviral deaminase APOBEC3G in vitro and ex vivo (FDR<0.006 and significantly reduces infectivity of progeny virus, decreasing the probability that the HIV reservoir will be replenished when latency is reversed therapeutically. Lastly, endogenous levels of soluble galectin-9 in the plasma of 72 HIV-infected ART-suppressed individuals were associated with levels of HIV RNA in CD4+ T cells (p<0.02 and with the quantity and binding avidity of circulating anti-HIV antibodies (p<0.009, suggesting a role of galectin-9 in regulating HIV transcription and viral production in vivo during therapy. Our data suggest that galectin-9 and the host glycosylation machinery should be explored as foundations for novel HIV cure strategies.

  15. Transient Genome-Wide Transcriptional Response to Low-Dose Ionizing Radiation In Vivo in Humans

    International Nuclear Information System (INIS)

    Berglund, Susanne R.; Rocke, David M.; Dai Jian; Schwietert, Chad W.; Santana, Alison; Stern, Robin L.; Lehmann, Joerg; Hartmann Siantar, Christine L.; Goldberg, Zelanna

    2008-01-01

    Purpose: The in vivo effects of low-dose low linear energy transfer ionizing radiation on healthy human skin are largely unknown. Using a patient-based tissue acquisition protocol, we have performed a series of genomic analyses on the temporal dynamics over a 24-hour period to determine the radiation response after a single exposure of 10 cGy. Methods and Materials: RNA from each patient tissue sample was hybridized to an Affymetrix Human Genome U133 Plus 2.0 array. Data analysis was performed on selected gene groups and pathways. Results: Nineteen gene groups and seven gene pathways that had been shown to be radiation responsive were analyzed. Of these, nine gene groups showed significant transient transcriptional changes in the human tissue samples, which returned to baseline by 24 hours postexposure. Conclusions: Low doses of ionizing radiation on full-thickness human skin produce a definable temporal response out to 24 hours postexposure. Genes involved in DNA and tissue remodeling, cell cycle transition, and inflammation show statistically significant changes in expression, despite variability between patients. These data serve as a reference for the temporal dynamics of ionizing radiation response following low-dose exposure in healthy full-thickness human skin

  16. Increased abundance of ADAM9 transcripts in the blood is associated with tissue damage [version 2; referees: 2 approved, 1 approved with reservations

    Directory of Open Access Journals (Sweden)

    Darawan Rinchai

    2016-10-01

    Full Text Available Background: Members of the ADAM (a disintegrin and metalloprotease domain family have emerged as critical regulators of cell-cell signaling during development and homeostasis. ADAM9 is consistently overexpressed in various human cancers, and has been shown to play an important role in tumorigenesis. However, little is known about the involvement of ADAM9 during immune-mediated processes. Results: Mining of an extensive compendium of transcriptomic datasets identified important gaps in knowledge regarding the possible role of ADAM9 in immunological homeostasis and inflammation: 1 The abundance of ADAM9 transcripts in the blood was increased in patients with acute infection but, 2 changed very little after in vitro exposure to a wide range of pathogen-associated molecular patterns (PAMPs. 3 Furthermore it was found to increase significantly in subjects as a result of tissue injury or tissue remodeling, in absence of infectious processes. Conclusions: Our findings indicate that ADAM9 may constitute a valuable biomarker for the assessment of tissue damage, especially in clinical situations where other inflammatory markers are confounded by infectious processes.

  17. DNA repair and cell cycle biomarkers of radiation exposure and inflammation stress in human blood.

    Directory of Open Access Journals (Sweden)

    Helen Budworth

    Full Text Available DNA damage and repair are hallmarks of cellular responses to ionizing radiation. We hypothesized that monitoring the expression of DNA repair-associated genes would enhance the detection of individuals exposed to radiation versus other forms of physiological stress. We employed the human blood ex vivo radiation model to investigate the expression responses of DNA repair genes in repeated blood samples from healthy, non-smoking men and women exposed to 2 Gy of X-rays in the context of inflammation stress mimicked by the bacterial endotoxin lipopolysaccharide (LPS. Radiation exposure significantly modulated the transcript expression of 12 genes of 40 tested (2.2E-06human blood ex vivo dataset, and 100% accuracy for discriminating patients who received total body radiation. Three genes of this panel (CDKN1A, FDXR and BBC3 were also highly sensitive to LPS treatment in the absence of radiation exposure, and LPS co-treatment significantly affected their radiation responses. At the protein level, BAX and pCHK2-thr68 were elevated after radiation exposure, but the pCHK2-thr68 response was significantly decreased in the presence of LPS. Our combined panel yields an estimated 4-group accuracy of ∼90% to discriminate between radiation alone, inflammation alone, or combined exposures. Our findings suggest that DNA repair gene expression may be helpful to identify biodosimeters of exposure to radiation, especially within high-complexity exposure scenarios.

  18. Identification and complete sequencing of novel human transcripts through the use of mouse orthologs and testis cDNA sequences

    DEFF Research Database (Denmark)

    Ferreira, Elisa N; Pires, Lilian C; Parmigiani, Raphael B

    2004-01-01

    The correct identification of all human genes, and their derived transcripts, has not yet been achieved, and it remains one of the major aims of the worldwide genomics community. Computational programs suggest the existence of 30,000 to 40,000 human genes. However, definitive gene identification ...

  19. SMAD4 regulates cell motility through transcription of N-cadherin in human pancreatic ductal epithelium.

    Directory of Open Access Journals (Sweden)

    Ya'an Kang

    Full Text Available Expression of the cellular adhesion protein N-cadherin is a critical event during epithelial-mesenchymal transition (EMT. The SMAD4 protein has been identified as a mediator of transforming growth factor-β (TGF-β superfamily signaling, which regulates EMT, but the mechanisms linking TGF-β signaling to N-cadherin expression remain unclear. When the TGF-β pathway is activated, SMAD proteins, including the common mediator SMAD4, are subsequently translocated into the nucleus, where they influence gene transcription via SMAD binding elements (SBEs. Here we describe a mechanism for control of CDH2, the gene encoding N-cadherin, through the canonical TGFβ-SMAD4 pathway. We first identified four previously undescribed SBEs within the CDH2 promoter. Using telomerase immortalized human pancreatic ductal epithelium, we found that TGF-β stimulation prompted specific SMAD4 binding to all four SBEs. Luciferase reporter and SMAD4-knockdown experiments demonstrated that specific SMAD4 binding to the SBE located at -3790 bp to -3795 bp within the promoter region of CDH2 was necessary for TGF-β-stimulated transcription. Expression of N-cadherin on the surface of epithelial cells facilitates motility and invasion, and we demonstrated that knockdown of SMAD4 causes decreased N-cadherin expression, which results in diminished migration and invasion of human pancreatic ductal epithelial cells. Similar reduction of cell motility was produced after CDH2 knockdown. Together, these findings suggest that SMAD4 is critical for the TGF-β-driven upregulation of N-cadherin and the resultant invasive phenotype of human pancreatic ductal epithelial cells during EMT.

  20. Responses of human cells to ZnO nanoparticles: a gene transcription study†

    Science.gov (United States)

    Moos, Philip J.; Olszewski, Kyle; Honeggar, Matthew; Cassidy, Pamela; Leachman, Sancy; Woessner, David; Cutler, N. Shane; Veranth, John M.

    2013-01-01

    The gene transcript profile responses to metal oxide nanoparticles was studied using human cell lines derived from the colon and skin tumors. Much of the research on nanoparticle toxicology has focused on models of inhalation and intact skin exposure, and effects of ingestion exposure and application to diseased skin are relatively unknown. Powders of nominally nanosized SiO2, TiO2, ZnO and Fe2O3 were chosen because these substances are widely used in consumer products. The four oxides were evaluated using colon-derived cell lines, RKO and CaCo-2, and ZnO and TiO2 were evaluated further using skin-derived cell lines HaCaT and SK Mel-28. ZnO induced the most notable gene transcription changes, even though this material was applied at the lowest concentration. Nano-sized and conventional ZnO induced similar responses suggesting common mechanisms of action. The results showed neither a non-specific response pattern common to all substances nor synergy of the particles with TNF-α cotreatment. The response to ZnO was not consistent with a pronounced proinflammatory signature, but involved changes in metal metabolism, chaperonin proteins, and protein folding genes. This response was observed in all cell lines when ZnO was in contact with the human cells. When the cells were exposed to soluble Zn, the genes involved in metal metabolism were induced but the genes involved in protein refoldling were unaffected. This provides some of the first data on the effects of commercial metal oxide nanoparticles on human colon-derived and skin-derived cells. PMID:21769377

  1. HOXB5 cooperates with NKX2-1 in the transcription of human RET.

    Directory of Open Access Journals (Sweden)

    Jiang Zhu

    Full Text Available The enteric nervous system (ENS regulates peristaltic movement of the gut, and abnormal ENS causes Hirschsprung's disease (HSCR in newborns. HSCR is a congenital complex genetic disorder characterised by a lack of enteric ganglia along a variable length of the intestine. The receptor tyrosine kinase gene (RET is the major HSCR gene and its expression is crucial for ENS development. We have previously reported that (i HOXB5 transcription factor mediates RET expression, and (ii mouse with defective HOXB5 activity develop HSCR phenotype. In this study, we (i elucidate the underlying mechanisms that HOXB5 mediate RET expression, and (ii examine the interactions between HOXB5 and other transcription factors implicated in RET expression. We show that human HOXB5 binds to the promoter region 5' upstream of the binding site of NKX2-1 and regulates RET expression. HOXB5 and NKX2-1 form a protein complex and mediate RET expression in a synergistic manner. HSCR associated SNPs at the NKX2-1 binding site (-5G>A rs10900296; -1A>C rs10900297, which reduce NKX2-1 binding, abolish the synergistic trans-activation of RET by HOXB5 and NKX2-1. In contrast to the synergistic activation of RET with NKX2-1, HOXB5 cooperates in an additive manner with SOX10, PAX3 and PHOX2B in trans-activation of RET promoter. Taken together, our data suggests that HOXB5 in coordination with other transcription factors mediates RET expression. Therefore, defects in cis- or trans-regulation of RET by HOXB5 could lead to reduction of RET expression and contribute to the manifestation of the HSCR phenotype.

  2. Enteroviruses in blood of patients with type 1 diabetes detected by integrated cell culture and reverse transcription quantitative real-time PCR.

    Science.gov (United States)

    Alidjinou, Enagnon Kazali; Sane, Famara; Lefevre, Christine; Baras, Agathe; Moumna, Ilham; Engelmann, Ilka; Vantyghem, Marie-Christine; Hober, Didier

    2017-11-01

    Enteroviruses (EV) have been associated with type 1 diabetes (T1D), but EV RNA detection has been reported in only a small proportion of T1D patients. We studied whether integrated cell culture and reverse transcription real-time PCR could improve EV detection in blood samples from patients with T1D. Blood was collected from 13 patients with T1D. The presence of EV RNA in blood was investigated by using real-time RT-PCR. In addition, plasma and white blood cells (WBC) were inoculated to BGM and Vero cell line cultures. Culture supernatants and cells collected on day 7 and day 14 were tested for EV RNA by real-time RT-PCR. Enterovirus identification was performed through sequencing of the VP4/VP2 region. Enterovirus RNA was detected in blood by using real-time RT-PCR in only one out of 13 patients. The detection of EV RNA in cultures inoculated with clinical samples (plasma and/or WBC) gave positive results in five other patients. The viral loads were low, ranging from 45 to 4420 copies/ng of total RNA. One isolate was successfully identified as coxsackievirus B1. Integrated cell culture and reverse transcription real-time PCR can improve the detection rate of EV in blood samples of patients with T1D and can be useful to investigate further the relationship between EV and the disease.

  3. Amyloid β levels in human red blood cells.

    Directory of Open Access Journals (Sweden)

    Takehiro Kiko

    Full Text Available UNLABELLED: Amyloid β-peptide (Aβ is hypothesized to play a key role by oxidatively impairing the capacity of red blood cells (RBCs to deliver oxygen to the brain. These processes are implicated in the pathogenesis of Alzheimer's disease (AD. Although plasma Aβ has been investigated thoroughly, the presence and distribution of Aβ in human RBCs are still unclear. In this study, we quantitated Aβ40 and Aβ42 in human RBCs with ELISA assays, and provided evidence that significant amounts of Aβ could be detected in RBCs and that the RBC Aβ levels increased with aging. The RBC Aβ levels increased with aging. On the other hand, providing an antioxidant supplement (astaxanthin, a polar carotenoid to humans was found to decrease RBC Aβ as well as oxidative stress marker levels. These results suggest that plasma Aβ40 and Aβ42 bind to RBCs (possibly with aging, implying a pathogenic role of RBC Aβ. Moreover, the data indicate that RBC Aβ40 and Aβ42 may constitute biomarkers of AD. As a preventive strategy, therapeutic application of astaxanthin as an Aβ-lowering agent in RBCs could be considered as a possible anti-dementia agent. TRIAL REGISTRATION: Controlled-Trials.com ISRCTN42483402.

  4. Reprogramming of human fibroblasts to pluripotent stem cells using mRNA of four transcription factors

    Energy Technology Data Exchange (ETDEWEB)

    Yakubov, Eduard [Department of Molecular Cell Biology, Weizmann Institute of Science, 76100 Rehovot (Israel); Rechavi, Gidi [Cancer Research Center, Chaim Sheba Medical Center, Tel-Hashomer and Sackler School of Medicine, Tel-Aviv University, Tel-Aviv (Israel); Rozenblatt, Shmuel [Department of Molecular Microbiology and Biotechnology, Tel-Aviv University, Tel-Aviv (Israel); Givol, David, E-mail: david.givol@weizmann.ac.il [Department of Molecular Cell Biology, Weizmann Institute of Science, 76100 Rehovot (Israel)

    2010-03-26

    Reprogramming of differentiated cells into induced pluripotent cells (iPS) was accomplished in 2006 by expressing four, or less, embryonic stem cell (ESC)-specific transcription factors. Due to the possible danger of DNA damage and the potential tumorigenicity associated with such DNA damage, attempts were made to minimize DNA integration by the vectors involved in this process without complete success. Here we present a method of using RNA transfection as a tool for reprogramming human fibroblasts to iPS. We used RNA synthesized in vitro from cDNA of the same reprogramming four transcription factors. After transfection of the RNA, we show intracellular expression and nuclear localization of the respective proteins in at least 70% of the cells. We used five consecutive transfections to support continuous protein expression resulting in the formation of iPS colonies that express alkaline phosphatase and several ESC markers and that can be expanded. This method completely avoids DNA integration and may be developed to replace the use of DNA vectors in the formation of iPS.

  5. HOCOMOCO: a comprehensive collection of human transcription factor binding sites models

    Science.gov (United States)

    Kulakovskiy, Ivan V.; Medvedeva, Yulia A.; Schaefer, Ulf; Kasianov, Artem S.; Vorontsov, Ilya E.; Bajic, Vladimir B.; Makeev, Vsevolod J.

    2013-01-01

    Transcription factor (TF) binding site (TFBS) models are crucial for computational reconstruction of transcription regulatory networks. In existing repositories, a TF often has several models (also called binding profiles or motifs), obtained from different experimental data. Having a single TFBS model for a TF is more pragmatic for practical applications. We show that integration of TFBS data from various types of experiments into a single model typically results in the improved model quality probably due to partial correction of source specific technique bias. We present the Homo sapiens comprehensive model collection (HOCOMOCO, http://autosome.ru/HOCOMOCO/, http://cbrc.kaust.edu.sa/hocomoco/) containing carefully hand-curated TFBS models constructed by integration of binding sequences obtained by both low- and high-throughput methods. To construct position weight matrices to represent these TFBS models, we used ChIPMunk software in four computational modes, including newly developed periodic positional prior mode associated with DNA helix pitch. We selected only one TFBS model per TF, unless there was a clear experimental evidence for two rather distinct TFBS models. We assigned a quality rating to each model. HOCOMOCO contains 426 systematically curated TFBS models for 401 human TFs, where 172 models are based on more than one data source. PMID:23175603

  6. HOCOMOCO: A comprehensive collection of human transcription factor binding sites models

    KAUST Repository

    Kulakovskiy, Ivan V.; Medvedeva, Yulia A.; Schaefer, Ulf; Kasianov, Artem S.; Vorontsov, Ilya E.; Bajic, Vladimir B.; Makeev, Vsevolod J.

    2012-01-01

    Transcription factor (TF) binding site (TFBS) models are crucial for computational reconstruction of transcription regulatory networks. In existing repositories, a TF often has several models (also called binding profiles or motifs), obtained from different experimental data. Having a single TFBS model for a TF is more pragmatic for practical applications. We show that integration of TFBS data from various types of experiments into a single model typically results in the improved model quality probably due to partial correction of source specific technique bias. We present the Homo sapiens comprehensive model collection (HOCOMOCO, http://autosome.ru/HOCOMOCO/, http://cbrc.kaust.edu.sa/ hocomoco/) containing carefully hand-curated TFBS models constructed by integration of binding sequences obtained by both low- and high-throughput methods. To construct position weight matrices to represent these TFBS models, we used ChIPMunk software in four computational modes, including newly developed periodic positional prior mode associated with DNA helix pitch. We selected only one TFBS model per TF, unless there was a clear experimental evidence for two rather distinct TFBS models. We assigned a quality rating to each model. HOCOMOCO contains 426 systematically curated TFBS models for 401 human TFs, where 172 models are based on more than one data source. The Author(s) 2012.

  7. Regulation of the human ADAMTS-4 promoter by transcription factors and cytokines

    International Nuclear Information System (INIS)

    Thirunavukkarasu, Kannan; Pei, Yong; Moore, Terry L.; Wang, He; Yu, Xiao-peng; Geiser, Andrew G.; Chandrasekhar, Srinivasan

    2006-01-01

    ADAMTS-4 (aggrecanase-1) is a metalloprotease that plays a role in aggrecan degradation in the cartilage extracellular matrix. In order to understand the regulation of ADAMTS-4 gene expression we have cloned and characterized a functional 4.5 kb human ADAMTS-4 promoter. Sequence analysis of the promoter revealed the presence of putative binding sites for nuclear factor of activated T cells (NFAT) and Runx family of transcription factors that are known to regulate chondrocyte maturation and differentiation. Using promoter-reporter assays and mRNA analysis we have analyzed the role of chondrocyte-expressed transcription factors NFATp and Runx2 and have shown that ADAMTS-4 is a potential downstream target of these two factors. Our results suggest that inhibition of the expression/function of NFATp and/or Runx2 may enable us to modulate aggrecan degradation in normal physiology and/or in degenerative joint diseases. The ADAMTS-4 promoter would serve as a valuable mechanistic tool to better understand the regulation of ADAMTS-4 expression by signaling pathways that modulate cartilage matrix breakdown

  8. Transcriptionally Driven DNA Replication Program of the Human Parasite Leishmania major.

    Science.gov (United States)

    Lombraña, Rodrigo; Álvarez, Alba; Fernández-Justel, José Miguel; Almeida, Ricardo; Poza-Carrión, César; Gomes, Fábia; Calzada, Arturo; Requena, José María; Gómez, María

    2016-08-09

    Faithful inheritance of eukaryotic genomes requires the orchestrated activation of multiple DNA replication origins (ORIs). Although origin firing is mechanistically conserved, how origins are specified and selected for activation varies across different model systems. Here, we provide a complete analysis of the nucleosomal landscape and replication program of the human parasite Leishmania major, building on a better evolutionary understanding of replication organization in Eukarya. We found that active transcription is a driving force for the nucleosomal organization of the L. major genome and that both the spatial and the temporal program of DNA replication can be explained as associated to RNA polymerase kinetics. This simple scenario likely provides flexibility and robustness to deal with the environmental changes that impose alterations in the genetic programs during parasitic life cycle stages. Our findings also suggest that coupling replication initiation to transcription elongation could be an ancient solution used by eukaryotic cells for origin maintenance. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  9. Transcriptional regulatory elements in the noncoding region of human papillomavirus type 6

    International Nuclear Information System (INIS)

    Wu, Tzyy-Choou.

    1989-01-01

    The structure and function of the transcriptional regulatory region of human papillomavirus type 6 (HPV-6) has been investigated. To investigate tissue specific gene expression, a sensitive method to detect and localize HPV-6 viral DNA, mRNA and protein in plastic-embedded tissue sections of genital and respiratory tract papillomata by using in situ hybridization and immunoperoxidase assays has been developed. This method, using ultrathin sections and strand-specific 3 H labeled riboprobes, offers the advantages of superior morphological preservation and detection of viral genomes at low copy number with good resolution, and the modified immunocytochemistry provides better sensitivity. The results suggest that genital tract epithelium is more permissive for HPV-6 replication than respiratory tract epithelium. To study the tissue tropism of HPV-6 at the level of regulation of viral gene expression, the polymerase chain reaction was used to isolate the noncoding region (NCR) of HPV-6 in independent isolates. Nucleotide sequence analysis of molecularly cloned DNA identified base substitutions, deletions/insertions and tandem duplications. Transcriptional regulatory elements in the NCR were assayed in recombinant plasmids containing the bacterial gene for chloramphenicol acetyl transferase

  10. HOCOMOCO: A comprehensive collection of human transcription factor binding sites models

    KAUST Repository

    Kulakovskiy, Ivan V.

    2012-11-21

    Transcription factor (TF) binding site (TFBS) models are crucial for computational reconstruction of transcription regulatory networks. In existing repositories, a TF often has several models (also called binding profiles or motifs), obtained from different experimental data. Having a single TFBS model for a TF is more pragmatic for practical applications. We show that integration of TFBS data from various types of experiments into a single model typically results in the improved model quality probably due to partial correction of source specific technique bias. We present the Homo sapiens comprehensive model collection (HOCOMOCO, http://autosome.ru/HOCOMOCO/, http://cbrc.kaust.edu.sa/ hocomoco/) containing carefully hand-curated TFBS models constructed by integration of binding sequences obtained by both low- and high-throughput methods. To construct position weight matrices to represent these TFBS models, we used ChIPMunk software in four computational modes, including newly developed periodic positional prior mode associated with DNA helix pitch. We selected only one TFBS model per TF, unless there was a clear experimental evidence for two rather distinct TFBS models. We assigned a quality rating to each model. HOCOMOCO contains 426 systematically curated TFBS models for 401 human TFs, where 172 models are based on more than one data source. The Author(s) 2012.

  11. Distinction of Fly Artifacts from Human Blood using Immunodetection.

    Science.gov (United States)

    Rivers, David B; Acca, Gillian; Fink, Marc; Brogan, Rebecca; Chen, Dorothy; Schoeffield, Andrew

    2018-02-21

    Insect stains produced by necrophagous flies are indistinguishable morphologically from human bloodstains. At present, no diagnostic tests exist to overcome this deficiency. As the first step toward developing a chemical test to recognize fly artifacts, polyclonal antisera were generated in rats against three distinct antigenic sequences of fly cathepsin D-like proteinase, an enzyme that is structurally distinct in cyclorrhaphous Diptera from other animals. The resulting rat antisera bound to artifacts produced by Protophormia terraenovae and synthetic peptides used to generate the polyclonal antisera, but not with any type of mammalian blood tested in immunoassays. Among the three antisera, anti-md3 serum displayed the highest reactivity for fly stains, demonstrated cross-reactivity for all synthetic peptides representing antigenic sequences of the mature fly enzyme, and bound artifacts originating from the fly digestive tract. Further work is needed to determine whether the antisera are suitable for non-laboratory conditions. © 2018 American Academy of Forensic Sciences.

  12. Trace-element measurement in human blood samples

    International Nuclear Information System (INIS)

    Hamidian, M.R.; Ebrahimi-Fakhar, F.

    1992-01-01

    It is conceivable that some essential elements such as zinc, iron, calcium, copper, phosphorus, selenium, etc., have a major impact on biological and metabolical functions in the human body. The concentration of these elements is normally very minute and changes within a naturally set tolerance. The accurate measurement of these elements in biological samples, such as in blood, is one of the objectives of medical physics in diagnosis. There are many sophisticated methods to measure the accurate amount of each element in biological samples. The methods used in this project are a combination of proton-induced X-ray emission (PIXE) and neutron activation analysis (NAA). The PIXE and NAA are fast and reliable techniques for multielement analysis at the level of parts per million and less

  13. Low level dose induced chromosome aberrations in human blood lymphocytes

    International Nuclear Information System (INIS)

    Pohl-Rueling, J.

    1992-01-01

    Unstable structural aberrations in chromosomes of human blood lymphocytes cannot be used as biological dosemeters in the low dose range, when extrapolating from high doses using a linear dose response, as required by the original formula of the dual radiation action theory. A survey is given of experimental dose-response curves of chromosome aberrations, obtained in investigations not only by this institute, in cooperation with many other laboratories, but also by various authors in different areas of the world. The results are not compatible with the predicted linear dose relationships at in vivo dose ranges up to 30 mGy.y -1 . The aberration frequencies rise sharply with dose within the normal environmental exposure up to about twice that level. At higher doses, aberration frequencies increase less rapidly and reach a plateau. Some in vitro experiments of various authors with higher doses of low LET radiations, up to about 400 mGy have found dose responses with steps. (author)

  14. Calculation of the uptake of CO into the human blood

    Energy Technology Data Exchange (ETDEWEB)

    Zankl, J.G.

    1981-01-01

    CO is a toxic substance mainly because, due to its high hemoglobin affinity, it inhibits oxygen transport in the human blood. This process must be quantified in order to establish limiting CO concentrations for garages, road tunnels, places of work, and for purpose of accident analysis. The medical information given is based on literature and is considered only as an introduction for non-experts. The emphasis of the work was on the development of a computer program by which the influence also of variable air CO concentrations and sea levels can be calculated. The program is available at the author's institute along with detailed calculations and results not considered in this publication.

  15. Human-Specific Histone Methylation Signatures at Transcription Start Sites in Prefrontal Neurons

    Science.gov (United States)

    Cheung, Iris; Bharadwaj, Rahul; Chou, Hsin-Jung; Houston, Isaac B.; Peter, Cyril J.; Mitchell, Amanda C.; Yao, Wei-Dong; Myers, Richard H.; Chen, Jiang-fan; Preuss, Todd M.; Rogaev, Evgeny I.; Jensen, Jeffrey D.; Weng, Zhiping; Akbarian, Schahram

    2012-01-01

    Cognitive abilities and disorders unique to humans are thought to result from adaptively driven changes in brain transcriptomes, but little is known about the role of cis-regulatory changes affecting transcription start sites (TSS). Here, we mapped in human, chimpanzee, and macaque prefrontal cortex the genome-wide distribution of histone H3 trimethylated at lysine 4 (H3K4me3), an epigenetic mark sharply regulated at TSS, and identified 471 sequences with human-specific enrichment or depletion. Among these were 33 loci selectively methylated in neuronal but not non-neuronal chromatin from children and adults, including TSS at DPP10 (2q14.1), CNTN4 and CHL1 (3p26.3), and other neuropsychiatric susceptibility genes. Regulatory sequences at DPP10 and additional loci carried a strong footprint of hominid adaptation, including elevated nucleotide substitution rates and regulatory motifs absent in other primates (including archaic hominins), with evidence for selective pressures during more recent evolution and adaptive fixations in modern populations. Chromosome conformation capture at two neurodevelopmental disease loci, 2q14.1 and 16p11.2, revealed higher order chromatin structures resulting in physical contact of multiple human-specific H3K4me3 peaks spaced 0.5–1 Mb apart, in conjunction with a novel cis-bound antisense RNA linked to Polycomb repressor proteins and downregulated DPP10 expression. Therefore, coordinated epigenetic regulation via newly derived TSS chromatin could play an important role in the emergence of human-specific gene expression networks in brain that contribute to cognitive functions and neurological disease susceptibility in modern day humans. PMID:23185133

  16. PCR amplification of Bartonella koehlerae from human blood and enrichment blood cultures

    Directory of Open Access Journals (Sweden)

    Breitschwerdt Edward B

    2010-08-01

    Full Text Available Abstract Background Cats appear to be the primary reservoir host for Bartonella koehlerae, an alpha Proteobacteria that is most likely transmitted among cat populations by fleas (Ctenocephalides felis. Bartonella koehlerae has caused endocarditis in a dog and in one human patient from Israel, but other clinically relevant reports involving this bacterium are lacking. Despite publication of numerous, worldwide epidemiological studies designed to determine the prevalence of Bartonella spp. bacteremia in cats, B. koehlerae has never been isolated using conventional blood agar plates. To date, successful isolation of B. koehlerae from cats and from the one human endocarditis patient has consistently required the use of chocolate agar plates. Results In this study, Bartonella koehlerae bacteremia was documented in eight immunocompetent patients by PCR amplification and DNA sequencing, either prior to or after enrichment blood culture using Bartonella alpha Proteobacteria growth medium. Presenting symptoms most often included fatigue, insomnia, joint pain, headache, memory loss, and muscle pain. Four patients were also infected with Bartonella vinsonii subsp. berkhoffii genotype II. After molecular documentation of B. koehlerae infection in these patients, a serological test was developed and serum samples were tested retrospectively. Bartonella koehlerae antibodies were not detected (titers B. koehlerae antibody titers of 1:64 or greater. Conclusions Although biased by a study population consisting of individuals with extensive arthropod and animal exposure, the results of this study suggest that B. koehlerae bacteremia is more common in immunocompetent people than has been previously suspected. Future studies should more thoroughly define modes of transmission and risk factors for acquiring infection with B. koehlerae. In addition, studies are needed to determine if B. koehlerae is a cause or cofactor in the development of arthritis, peripheral

  17. Type 2 diabetes and obesity induce similar transcriptional reprogramming in human myocytes

    DEFF Research Database (Denmark)

    Väremo, Leif; Henriksen, Tora Ida; Scheele, Camilla

    2017-01-01

    BACKGROUND: Skeletal muscle is one of the primary tissues involved in the development of type 2 diabetes (T2D). The close association between obesity and T2D makes it difficult to isolate specific effects attributed to the disease alone. Therefore, here we set out to identify and characterize...... in sphingolipid metabolism was transcriptionally regulated. CONCLUSIONS: Our findings identify inherent characteristics in myocytes, as a memory of the in vivo phenotype, without the influence from a diabetic or obese extracellular environment, highlighting their importance in the development of T2D....... intrinsic properties of myocytes, associated independently with T2D or obesity. METHODS: We generated and analyzed RNA-seq data from primary differentiated myotubes from 24 human subjects, using a factorial design (healthy/T2D and non-obese/obese), to determine the influence of each specific factor...

  18. (Some) Cellular Mechanisms Influencing the Transcription of Human Endogenous Retrovirus, HERV-Fc1

    DEFF Research Database (Denmark)

    Laska, Magdalena Janina; Nissen, Kari Konstantin; Nexø, Bjørn Andersen

    2013-01-01

    DNA methylation and histone acetylation are epigenetic modifications that act as regulators of gene expression. DNA methylation is considered an important mechanism for silencing of retroelements in the mammalian genome. However, the methylation of human endogenous retroviruses (HERVs) is not well...... investigated. The aim of this study was to investigate the transcriptional potential of HERV-Fc1 proviral 5'LTR in more detail, and examined the specific influence of CpG methylation on this LTR in number of cell lines. Specifically, the role of demethylating chemicals e.g. 5-aza-2' deoxycytidine...... and Trichostatin-A, in inducing or reactivating expression of HERV-Fc1 specific sequences and the mechanisms were investigated. In our present study, 5-aza-dC is shown to be a powerful inducer of HERV-Fc1, and at the same time it strongly inhibits methylation of DNA. Treatment with this demethylating agent 5-aza...

  19. Exercise induces transient transcriptional activation of the PGC-1a gene in human skeletal muscle

    DEFF Research Database (Denmark)

    Pilegaard, Henriette; Saltin, Bengt; Neufer, P. Darrell

    2003-01-01

    Endurance exercise training induces mitochondrial biogenesis in skeletal muscle. The peroxisome proliferator activated receptor co-activator 1a (PGC-1a) has recently been identified as a nuclear factor critical for coordinating the activation of genes required for mitochondrial biogenesis in cell...... culture and rodent skeletal muscle. To determine whether PGC-1a transcription is regulated by acute exercise and exercise training in human skeletal muscle, seven male subjects performed 4 weeks of one-legged knee extensor exercise training. At the end of training, subjects completed 3 h of two......-legged knee extensor exercise. Biopsies were obtained from the vastus lateralis muscle of both the untrained and trained legs before exercise and after 0, 2, 6 and 24 h of recovery. Time to exhaustion (2 min maximum resistance), as well as hexokinase II (HKII), citrate synthase and 3-hydroxyacyl...

  20. Methylation and Transcripts Expression at the Imprinted GNAS Locus in Human Embryonic and Induced Pluripotent Stem Cells and Their Derivatives

    Directory of Open Access Journals (Sweden)

    Virginie Grybek

    2014-09-01

    Full Text Available Data from the literature indicate that genomic imprint marks are disturbed in human pluripotent stem cells (PSCs. GNAS is an imprinted locus that produces one biallelic (Gsα and four monoallelic (NESP55, GNAS-AS1, XLsα, and A/B transcripts due to differential methylation of their promoters (DMR. To document imprinting at the GNAS locus in PSCs, we studied GNAS locus DMR methylation and transcript (NESP55, XLsα, and A/B expression in human embryonic stem cells (hESCs and human induced pluripotent stem cells (hiPSCs derived from two human fibroblasts and their progenies. Results showed that (1 methylation at the GNAS locus DMRs is DMR and cell line specific, (2 changes in allelic transcript expression can be independent of a change in allele-specific DNA methylation, and (3 interestingly, methylation at A/B DMR is correlated with A/B transcript expression. These results indicate that these models are valuable to study the mechanisms controlling GNAS methylation, factors involved in transcript expression, and possibly mechanisms involved in the pathophysiology of pseudohypoparathyroidism type 1B.

  1. ZNF328, a novel human zinc-finger protein, suppresses transcriptional activities of SRE and AP-1

    International Nuclear Information System (INIS)

    Ou Ying; Wang Shenqiu; Cai Zhenyu; Wang Yuequn; Wang Canding; Li Yongqing; Li Fang; Yuan Wuzhou; Liu Bisheng; Wu Xiushan; Liu Mingyao

    2005-01-01

    The zinc finger proteins containing the Kruppel-associated box domain (KRAB-ZFPs) are the single largest class of transcription factors in human genome. Many of the KRAB-ZFPs are involved in cardiac development or cardiovascular diseases. Here, we have identified a novel human KRAB zinc finger gene, named ZNF328, from the human fetal heart cDNA library. The complete sequence of ZNF328 cDNA contains a 2376-bp open reading frame (ORF) and encodes a 792 amino acid protein with an N-terminal KRAB domain and classical zinc finger C 2 H 2 motifs in the C-terminus. Northern blot analysis indicates that the protein is expressed in most of the examined human adult and embryonic tissues. ZNF328 is a transcription suppressor when fused to Gal-4 DNA-binding domain and cotransfected with VP-16. Overexpression of ZNF328 in COS-7 cells inhibits the transcriptional activities of SRE and AP-1. Deletion analysis with a series of truncated fusion proteins indicates that the KRAB motif is a basal repression domain when cotransfected with VP-16. Similar results were obtained when the truncated fusion proteins were assayed for the transcriptional activities of SRE and AP-1. These results suggest that ZNF328 protein may act as a transcriptional repressor in mitogen-activated protein kinase (MAPK) signaling pathway to mediate cellular functions

  2. Transcriptional Repressor HIC1 Contributes to Suppressive Function of Human Induced Regulatory T Cells

    Directory of Open Access Journals (Sweden)

    Ubaid Ullah

    2018-02-01

    Full Text Available Regulatory T (Treg cells are critical in regulating the immune response. In vitro induced Treg (iTreg cells have significant potential in clinical medicine. However, applying iTreg cells as therapeutics is complicated by the poor stability of human iTreg cells and their variable suppressive activity. Therefore, it is important to understand the molecular mechanisms of human iTreg cell specification. We identified hypermethylated in cancer 1 (HIC1 as a transcription factor upregulated early during the differentiation of human iTreg cells. Although FOXP3 expression was unaffected, HIC1 deficiency led to a considerable loss of suppression by iTreg cells with a concomitant increase in the expression of effector T cell associated genes. SNPs linked to several immune-mediated disorders were enriched around HIC1 binding sites, and in vitro binding assays indicated that these SNPs may alter the binding of HIC1. Our results suggest that HIC1 is an important contributor to iTreg cell development and function.

  3. A paradox of transcriptional and functional innate interferon responses of human intestinal enteroids to enteric virus infection

    OpenAIRE

    Saxena, Kapil; Simon, Lukas M.; Zeng, Xi-Lei; Blutt, Sarah E.; Crawford, Sue E.; Sastri, Narayan P.; Karandikar, Umesh C.; Ajami, Nadim J.; Zachos, Nicholas C.; Kovbasnjuk, Olga; Donowitz, Mark; Conner, Margaret E.; Shaw, Chad A.; Estes, Mary K.

    2017-01-01

    Understanding host?enteric virus interactions has been limited by the inability to culture nontransformed small intestinal epithelial cells and to infect animal models with human viruses. We report epithelial responses in human small intestinal enteroid cultures from different individuals following infection with human rotavirus (HRV), a model enteric pathogen. RNA-sequencing and functional assays revealed type III IFN as the dominant transcriptional response that activates interferon-stimula...

  4. Repression of hTERT transcription by the introduction of chromosome 3 into human oral squamous cell carcinoma

    International Nuclear Information System (INIS)

    Nishio, Sachiyo; Ohira, Takahito; Sunamura, Naohiro; Oshimura, Mitsuo; Ryoke, Kazuo; Kugoh, Hiroyuki

    2015-01-01

    Telomerase is a ribonucleoprotein enzyme that maintains telomere length. Telomerase activity is primarily attributed to the expression of telomerase reverse transcriptase (TERT). It has been reported that introduction of an intact human chromosome 3 into the human oral squamous cell carcinoma cell line HSC3 suppresses the tumorigenicity of these cells. However, the mechanisms that regulate tumorigenicity have not been elucidated. To determine whether this reduction in tumorigenicity was accompanied by a reduction in telomerase activity, we investigated the transcriptional activation of TERT in HSC3 microcell hybrid clones with an introduced human chromosome 3 (HSC3#3). HSC#3 cells showed inhibition of hTERT transcription compared to that of the parental HSC3 cells. Furthermore, cell fusion experiments showed that hybrids of HSC3 cells and cells of the RCC23 renal carcinoma cell line, which also exhibits suppression of TERT transcription by the introduction of human chromosome 3, also displayed suppressed TERT transcription. These results suggested that human chromosome 3 may carry functionally distinct, additional TERT repressor genes. - Highlights: • hTERT mRNA expression level decreased in the chromosome 3 introduced HSC3 clones. • hTERT mRNA expression level was tend to suppressed in HSC3 and RCC23 hybrid cells. • We provide evidence that human chromosome 3 carries at least two distinct hTERT regulatory factors.

  5. Repression of hTERT transcription by the introduction of chromosome 3 into human oral squamous cell carcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Nishio, Sachiyo [Division of Oral and Maxillofacial Biopathological Surgery, Faculty of Medicine, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503 (Japan); Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, Yonago, Tottori, 683-8503 (Japan); Ohira, Takahito; Sunamura, Naohiro [Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, Yonago, Tottori, 683-8503 (Japan); Oshimura, Mitsuo [Chromosome Engineering Research Center, Tottori University, Yonago, Tottori, 683-8503 (Japan); Ryoke, Kazuo [Division of Oral and Maxillofacial Biopathological Surgery, Faculty of Medicine, Tottori University, 86 Nishi-cho, Yonago, Tottori, 683-8503 (Japan); Kugoh, Hiroyuki, E-mail: kugoh@med.tottori-u.ac.jp [Department of Biomedical Science, Institute of Regenerative Medicine and Biofunction, Graduate School of Medical Science, Tottori University, Yonago, Tottori, 683-8503 (Japan); Chromosome Engineering Research Center, Tottori University, Yonago, Tottori, 683-8503 (Japan)

    2015-10-30

    Telomerase is a ribonucleoprotein enzyme that maintains telomere length. Telomerase activity is primarily attributed to the expression of telomerase reverse transcriptase (TERT). It has been reported that introduction of an intact human chromosome 3 into the human oral squamous cell carcinoma cell line HSC3 suppresses the tumorigenicity of these cells. However, the mechanisms that regulate tumorigenicity have not been elucidated. To determine whether this reduction in tumorigenicity was accompanied by a reduction in telomerase activity, we investigated the transcriptional activation of TERT in HSC3 microcell hybrid clones with an introduced human chromosome 3 (HSC3#3). HSC#3 cells showed inhibition of hTERT transcription compared to that of the parental HSC3 cells. Furthermore, cell fusion experiments showed that hybrids of HSC3 cells and cells of the RCC23 renal carcinoma cell line, which also exhibits suppression of TERT transcription by the introduction of human chromosome 3, also displayed suppressed TERT transcription. These results suggested that human chromosome 3 may carry functionally distinct, additional TERT repressor genes. - Highlights: • hTERT mRNA expression level decreased in the chromosome 3 introduced HSC3 clones. • hTERT mRNA expression level was tend to suppressed in HSC3 and RCC23 hybrid cells. • We provide evidence that human chromosome 3 carries at least two distinct hTERT regulatory factors.

  6. Genome-wide analysis of differential transcriptional and epigenetic variability across human immune cell types

    DEFF Research Database (Denmark)

    Ecker, Simone; Chen, Lu; Pancaldi, Vera

    2017-01-01

    Background: A healthy immune system requires immune cells that adapt rapidly to environmental challenges. This phenotypic plasticity can be mediated by transcriptional and epigenetic variability. Results: We apply a novel analytical approach to measure and compare transcriptional and epigenetic v...

  7. Transcription of human resistin gene involves an interaction of Sp1 with peroxisome proliferator-activating receptor gamma (PPARgamma.

    Directory of Open Access Journals (Sweden)

    Anil K Singh

    2010-03-01

    Full Text Available Resistin is a cysteine rich protein, mainly expressed and secreted by circulating human mononuclear cells. While several factors responsible for transcription of mouse resistin gene have been identified, not much is known about the factors responsible for the differential expression of human resistin.We show that the minimal promoter of human resistin lies within approximately 80 bp sequence upstream of the transcriptional start site (-240 whereas binding sites for cRel, CCAAT enhancer binding protein alpha (C/EBP-alpha, activating transcription factor 2 (ATF-2 and activator protein 1 (AP-1 transcription factors, important for induced expression, are present within sequences up to -619. Specificity Protein 1(Sp1 binding site (-276 to -295 is also present and an interaction of Sp1 with peroxisome proliferator activating receptor gamma (PPARgamma is necessary for constitutive expression in U937 cells. Indeed co-immunoprecipitation assay demonstrated a direct physical interaction of Sp1 with PPARgamma in whole cell extracts of U937 cells. Phorbol myristate acetate (PMA upregulated the expression of resistin mRNA in U937 cells by increasing the recruitment of Sp1, ATF-2 and PPARgamma on the resistin gene promoter. Furthermore, PMA stimulation of U937 cells resulted in the disruption of Sp1 and PPARgamma interaction. Chromatin immunoprecipitation (ChIP assay confirmed the recruitment of transcription factors phospho ATF-2, Sp1, Sp3, PPARgamma, chromatin modifier histone deacetylase 1 (HDAC1 and the acetylated form of histone H3 but not cRel, C/EBP-alpha and phospho c-Jun during resistin gene transcription.Our findings suggest a complex interplay of Sp1 and PPARgamma along with other transcription factors that drives the expression of resistin in human monocytic U937 cells.

  8. Microphthalmia-associated transcription factor as the molecular target of cadmium toxicity in human melanocytes

    International Nuclear Information System (INIS)

    Chantarawong, Wipa; Takeda, Kazuhisa; Sangartit, Weerapon; Yoshizawa, Miki; Pradermwong, Kantimanee; Shibahara, Shigeki

    2014-01-01

    Highlights: • In human melanocytes, cadmium decreases the expression of MITF-M and tyrosinase and their mRNAs. • In human melanoma cells, cadmium decreases the expression of MITF-M protein and tyrosinase mRNA. • Expression of MITF-H is less sensitive to cadmium toxicity in melanocyte-linage cells. • Cadmium does not decrease the expression of MITF-H in retinal pigment epithelial cells. • MITF-M is the molecular target of cadmium toxicity in melanocytes. - Abstract: Dietary intake of cadmium is inevitable, causing age-related increase in cadmium accumulation in many organs, including hair, choroid and retinal pigment epithelium (RPE). Cadmium has been implicated in the pathogenesis of hearing loss and macular degeneration. The functions of cochlea and retina are maintained by melanocytes and RPE, respectively, and the differentiation of these pigment cells is regulated by microphthalmia-associated transcription factor (MITF). In the present study, we explored the potential toxicity of cadmium in the cochlea and retina by using cultured human melanocytes and human RPE cell lines. MITF consists of multiple isoforms, including melanocyte-specific MITF-M and widely expressed MITF-H. Levels of MITF-M protein and its mRNA in human epidermal melanocytes and HMV-II melanoma cells were decreased significantly by cadmium. In parallel with the MITF reduction, mRNA levels of tyrosinase, the key enzyme of melanin biosynthesis that is regulated by MITF-M, were also decreased. In RPE cells, however, the levels of total MITF protein, constituting mainly MITF-H, were not decreased by cadmium. We thus identify MITF-M as the molecular target of cadmium toxicity in melanocytes, thereby accounting for the increased risk of disability from melanocyte malfunction, such as hearing and vision loss among people with elevated cadmium exposure

  9. Microphthalmia-associated transcription factor as the molecular target of cadmium toxicity in human melanocytes

    Energy Technology Data Exchange (ETDEWEB)

    Chantarawong, Wipa [Department of Molecular Biology and Applied Physiology, Tohoku University School of Medicine, Sendai (Japan); Inter Departmental Multidisciplinary Graduate Program in Bioscience, Faculty of Science, Kasetsart University, Bangkok (Thailand); Takeda, Kazuhisa; Sangartit, Weerapon; Yoshizawa, Miki [Department of Molecular Biology and Applied Physiology, Tohoku University School of Medicine, Sendai (Japan); Pradermwong, Kantimanee [Department of Zoology, Faculty of Science, Kasetsart University, Bangkok (Thailand); Shibahara, Shigeki, E-mail: shibahar@med.tohoku.ac.jp [Department of Molecular Biology and Applied Physiology, Tohoku University School of Medicine, Sendai (Japan)

    2014-11-28

    Highlights: • In human melanocytes, cadmium decreases the expression of MITF-M and tyrosinase and their mRNAs. • In human melanoma cells, cadmium decreases the expression of MITF-M protein and tyrosinase mRNA. • Expression of MITF-H is less sensitive to cadmium toxicity in melanocyte-linage cells. • Cadmium does not decrease the expression of MITF-H in retinal pigment epithelial cells. • MITF-M is the molecular target of cadmium toxicity in melanocytes. - Abstract: Dietary intake of cadmium is inevitable, causing age-related increase in cadmium accumulation in many organs, including hair, choroid and retinal pigment epithelium (RPE). Cadmium has been implicated in the pathogenesis of hearing loss and macular degeneration. The functions of cochlea and retina are maintained by melanocytes and RPE, respectively, and the differentiation of these pigment cells is regulated by microphthalmia-associated transcription factor (MITF). In the present study, we explored the potential toxicity of cadmium in the cochlea and retina by using cultured human melanocytes and human RPE cell lines. MITF consists of multiple isoforms, including melanocyte-specific MITF-M and widely expressed MITF-H. Levels of MITF-M protein and its mRNA in human epidermal melanocytes and HMV-II melanoma cells were decreased significantly by cadmium. In parallel with the MITF reduction, mRNA levels of tyrosinase, the key enzyme of melanin biosynthesis that is regulated by MITF-M, were also decreased. In RPE cells, however, the levels of total MITF protein, constituting mainly MITF-H, were not decreased by cadmium. We thus identify MITF-M as the molecular target of cadmium toxicity in melanocytes, thereby accounting for the increased risk of disability from melanocyte malfunction, such as hearing and vision loss among people with elevated cadmium exposure.

  10. Overexpression of octamer transcription factors 1 or 2 alone has no effect on HIV-1 transcription in primary human CD4 T cells

    International Nuclear Information System (INIS)

    Zhang Mingce; Genin, Anna; Cron, Randy Q.

    2004-01-01

    We explored the binding of octamer (Oct) transcription factors to the HIV-1 long terminal repeat (LTR) by gel shift assays and showed none of the previously identified four potential Oct binding sites bound Oct-1 or Oct-2. Overexpression of Oct-1 or Oct-2 had no effect on HIV-1 LTR activity in transiently transfected primary human CD4 T cells. Next, primary human CD4 T cells were co-transfected with a green fluorescent protein (GFP)-expression vector and an Oct-1 or Oct-2 expression plasmid. The transfected cells were stimulated for 2 days and then infected with the NL4-3 strain of HIV-1. After 3 days of infection, there were no differences in HIV-1 p24 supernatant levels. Apoptosis of infected or bystander cells overexpressing Oct-1 or Oct-2 compared to control was also unaffected. Our studies demonstrate that Oct-1 and Oct-2 fail to bind to the HIV-1 LTR and have no effect on HIV-1 transcription in primary human CD4 T cells

  11. In vitro activation of transcription by the human T-cell leukemia virus type I Tax protein.

    Science.gov (United States)

    Matthews, M A; Markowitz, R B; Dynan, W S

    1992-05-01

    The human T-cell leukemia virus type I (HTLV-I) regulatory protein Tax activates transcription of the proviral long terminal repeats and a number of cellular promoters. We have developed an in vitro system to characterize the mechanism by which Tax interacts with the host cell transcription machinery. Tax was purified from cells infected with a baculovirus expression vector. Addition of these Tax preparations to nuclear extracts from uninfected human T lymphocytes activated transcription of the HTLV-I long terminal repeat approximately 10-fold. Transcription-stimulatory activity copurified with the immunoreactive 40-kDa Tax polypeptide on gel filtration chromatography, and, as expected, the effect of recombinant Tax was diminished in HTLV-I-infected T-lymphocyte extracts containing endogenous Tax. Tax-mediated transactivation in vivo has been previously shown to require 21-bp-repeat Tax-responsive elements (TxREs) in the promoter DNA. Stimulation of transcription in vitro was also strongly dependent on these sequences. To investigate the mechanism of Tax transactivation, cellular proteins that bind the 21-bp-repeat TxREs were prepared by DNA affinity chromatography. Recombinant Tax markedly increased the formation of a specific host protein-DNA complex detected in an electrophoretic mobility shift assay. These data suggest that Tax activates transcription through a direct interaction with cellular proteins that bind to the 21-bp-repeat TxREs.

  12. Expression of a splice variant of the platelet-activating factor receptor transcript 2 in various human cancer cell lines

    Directory of Open Access Journals (Sweden)

    Ibtissam Youlyouz

    2002-01-01

    Full Text Available Platelet-activating factor receptor (PAF-R transcripts were analysed by reverse transcriptase-polymerase chain reaction in five human cancer cell lines derived from the breast (BT20, SKBR3 and T47D cells, the pancreas (Miapaca cells and the bladder (5637 cells in order to confirm the existence of a splice variant of the PAF-R transcript 2. After cloning and sequencing, we confirmed its existence in all cell lines. It consisted of the PAF-R transcript 2 lengthening with 82 nucleotides from the 3' end of exon 1 of the PAF-R gene. The role of this elongated form of the tissue-type PAF-R transcript in cell physiology remains to be elucidated.

  13. Generation of human induced pluripotent stem cells from a Bombay individual: Moving towards 'universal-donor' red blood cells

    International Nuclear Information System (INIS)

    Seifinejad, Ali; Taei, Adeleh; Totonchi, Mehdi; Vazirinasab, Hamed; Hassani, Seideh Nafiseh; Aghdami, Nasser; Shahbazi, Ebrahim; Yazdi, Reza Salman; Salekdeh, Ghasem Hosseini; Baharvand, Hossein

    2010-01-01

    Bombay phenotype is one of the rare phenotypes in the ABO blood group system that fails to express ABH antigens on red blood cells. Nonsense or missense mutations in fucosyltransfrase1 (FUT1) and fucosyltransfrase2 (FUT2) genes are known to create this phenotype. This blood group is compatible with all other blood groups as a donor, as it does not express the H antigen on the red blood cells. In this study, we describe the establishment of human induced pluripotent stem cells (iPSCs) from the dermal fibroblasts of a Bombay blood-type individual by the ectopic expression of established transcription factors Klf4, Oct4, Sox2, and c-Myc. Sequence analyses of fibroblasts and iPSCs revealed a nonsense mutation 826C to T (276 Gln to Ter) in the FUT1 gene and a missense mutation 739G to A (247 Gly to Ser) in the FUT2 gene in the Bombay phenotype under study. The established iPSCs resemble human embryonic stem cells in morphology, passaging, surface and pluripotency markers, normal karyotype, gene expression, DNA methylation of critical pluripotency genes, and in-vitro differentiation. The directed differentiation of the iPSCs into hematopoietic lineage cells displayed increased expression of the hematopoietic lineage markers such as CD34, CD133, RUNX1, KDR, α-globulin, and γ-globulin. Such specific stem cells provide an unprecedented opportunity to produce a universal blood group donor, in-vitro, thus enabling cellular replacement therapies, once the safety issue is resolved.

  14. Direct activation of human and mouse Oct4 genes using engineered TALE and Cas9 transcription factors.

    Science.gov (United States)

    Hu, Jiabiao; Lei, Yong; Wong, Wing-Ki; Liu, Senquan; Lee, Kai-Chuen; He, Xiangjun; You, Wenxing; Zhou, Rui; Guo, Jun-Tao; Chen, Xiongfong; Peng, Xianlu; Sun, Hao; Huang, He; Zhao, Hui; Feng, Bo

    2014-04-01

    The newly developed transcription activator-like effector protein (TALE) and clustered regularly interspaced short palindromic repeats/Cas9 transcription factors (TF) offered a powerful and precise approach for modulating gene expression. In this article, we systematically investigated the potential of these new tools in activating the stringently silenced pluripotency gene Oct4 (Pou5f1) in mouse and human somatic cells. First, with a number of TALEs and sgRNAs targeting various regions in the mouse and human Oct4 promoters, we found that the most efficient TALE-VP64s bound around -120 to -80 bp, while highly effective sgRNAs targeted from -147 to -89-bp upstream of the transcription start sites to induce high activity of luciferase reporters. In addition, we observed significant transcriptional synergy when multiple TFs were applied simultaneously. Although individual TFs exhibited marginal activity to up-regulate endogenous gene expression, optimized combinations of TALE-VP64s could enhance endogenous Oct4 transcription up to 30-fold in mouse NIH3T3 cells and 20-fold in human HEK293T cells. More importantly, the enhancement of OCT4 transcription ultimately generated OCT4 proteins. Furthermore, examination of different epigenetic modifiers showed that histone acetyltransferase p300 could enhance both TALE-VP64 and sgRNA/dCas9-VP64 induced transcription of endogenous OCT4. Taken together, our study suggested that engineered TALE-TF and dCas9-TF are useful tools for modulating gene expression in mammalian cells.

  15. Papillomavirus genomes in human cervical carcinoma: Analysis of their integration and transcriptional activity

    International Nuclear Information System (INIS)

    Matulic, M.; Soric, J.

    1994-01-01

    Eighty-four biopsies derived from cervical tissues were analyzed for the presence of human papillomavirus (HPV) DNA types 6, 16 and 18 using Southern blot hybridization. HPV 6 was found in none of the cervical biopsies, and HPV types 16 and 18 were found in 44% of them. The rate of HPV 16/18 positive samples increased proportionally to the severity of the lesion. In normal tissue there were no positive samples, in mild and moderate dysplasia HPV 16/18 was present in 20% and in severe dysplasia and invasive carcinomas in 37 and 50%, respectively. In biopsies from 13 cases with squamous cell carcinoma of the uterine cervix and CIN III lesions HPV 16 was integrated within the host genome. It was concluded that the virus could be integrated at variable, presumably randomly selected chromosomal loci and with different number of copies. Transcription of HPV 16 and 18 was detected in one cervical cancer in HeLa cells, respectively. These results imply that HPV types 16 and 18 play an etiological role in the carcinogenesis of human cervical epithelial cells. (author)

  16. Architecture of the human and yeast general transcription and DNA repair factor TFIIH

    Science.gov (United States)

    Luo, Jie; Cimermancic, Peter; Viswanath, Shruthi; Ebmeier, Christopher C.; Kim, Bong; Dehecq, Marine; Raman, Vishnu; Greenberg, Charles H.; Pellarin, Riccardo; Sali, Andrej; Taatjes, Dylan J.; Hahn, Steven; Ranish, Jeff

    2015-01-01

    Summary TFIIH is essential for both RNA polymerase II transcription and DNA repair, and mutations in TFIIH can result in human disease. Here, we determine the molecular architecture of human and yeast TFIIH by an integrative approach using chemical crosslinking/mass spectrometry (CXMS) data, biochemical analyses, and previously published electron microscopy maps. We identified four new conserved “topological regions” that function as hubs for TFIIH assembly and more than 35 conserved topological features within TFIIH, illuminating a network of interactions involved in TFIIH assembly and regulation of its activities. We show that one of these conserved regions, the p62/Tfb1 Anchor region, directly interacts with the DNA helicase subunit XPD/Rad3 in native TFIIH and is required for the integrity and function of TFIIH. We also reveal the structural basis for defects in patients with Xeroderma pigmentosum and Trichothiodystrophy, with mutations found at the interface between the p62 Anchor region and the XPD subunit. PMID:26340423

  17. Comprehensive human transcription factor binding site map for combinatory binding motifs discovery.

    Directory of Open Access Journals (Sweden)

    Arnoldo J Müller-Molina

    Full Text Available To know the map between transcription factors (TFs and their binding sites is essential to reverse engineer the regulation process. Only about 10%-20% of the transcription factor binding motifs (TFBMs have been reported. This lack of data hinders understanding gene regulation. To address this drawback, we propose a computational method that exploits never used TF properties to discover the missing TFBMs and their sites in all human gene promoters. The method starts by predicting a dictionary of regulatory "DNA words." From this dictionary, it distills 4098 novel predictions. To disclose the crosstalk between motifs, an additional algorithm extracts TF combinatorial binding patterns creating a collection of TF regulatory syntactic rules. Using these rules, we narrowed down a list of 504 novel motifs that appear frequently in syntax patterns. We tested the predictions against 509 known motifs confirming that our system can reliably predict ab initio motifs with an accuracy of 81%-far higher than previous approaches. We found that on average, 90% of the discovered combinatorial binding patterns target at least 10 genes, suggesting that to control in an independent manner smaller gene sets, supplementary regulatory mechanisms are required. Additionally, we discovered that the new TFBMs and their combinatorial patterns convey biological meaning, targeting TFs and genes related to developmental functions. Thus, among all the possible available targets in the genome, the TFs tend to regulate other TFs and genes involved in developmental functions. We provide a comprehensive resource for regulation analysis that includes a dictionary of "DNA words," newly predicted motifs and their corresponding combinatorial patterns. Combinatorial patterns are a useful filter to discover TFBMs that play a major role in orchestrating other factors and thus, are likely to lock/unlock cellular functional clusters.

  18. Transcriptional Regulation of Frizzled-1 in Human Osteoblasts by Sp1.

    Directory of Open Access Journals (Sweden)

    Shibing Yu

    Full Text Available The wingless pathway has a powerful influence on bone metabolism and is a therapeutic target in skeletal disorders. Wingless signaling is mediated in part through the Frizzled (FZD receptor family. FZD transcriptional regulation is poorly understood. Herein we tested the hypothesis that Sp1 plays an important role in the transcriptional regulation of FZD1 expression in osteoblasts and osteoblast mineralization. To test this hypothesis, we conducted FZD1 promoter assays in Saos2 cells with and without Sp1 overexpression. We found that Sp1 significantly up-regulates FZD1 promoter activity in Saos2 cells. Chromatin immunoprecipitation (ChIP and electrophoretic mobility shift (EMSA assays identified a novel and functional Sp1 binding site at -44 to -40 from the translation start site in the FZD1 promoter. The Sp1-dependent activation of the FZD1 promoter was abolished by mithramycin A (MMA, an antibiotic affecting both Sp1 binding and Sp1 protein levels in Saos2 cells. Similarly, down-regulation of Sp1 in hFOB cells resulted in less FZD1 expression and lower alkaline phosphatase activity. Moreover, over-expression of Sp1 increased FZD1 expression and Saos2 cell mineralization while MMA decreased Sp1 and FZD1 expression and Saos2 cell mineralization. Knockdown of FZD1 prior to Sp1 overexpression partially abolished Sp1 stimulation of osteoblast differentiation markers. Taken together, our results suggest that Sp1 plays a role in human osteoblast differentiation and mineralization, which is at least partially mediated by Sp1-dependent transactivation of FZD1.

  19. Comprehensive prediction in 78 human cell lines reveals rigidity and compactness of transcription factor dimers

    Science.gov (United States)

    Jankowski, Aleksander; Szczurek, Ewa; Jauch, Ralf; Tiuryn, Jerzy; Prabhakar, Shyam

    2013-01-01

    The binding of transcription factors (TFs) to their specific motifs in genomic regulatory regions is commonly studied in isolation. However, in order to elucidate the mechanisms of transcriptional regulation, it is essential to determine which TFs bind DNA cooperatively as dimers and to infer the precise nature of these interactions. So far, only a small number of such dimeric complexes are known. Here, we present an algorithm for predicting cell-type–specific TF–TF dimerization on DNA on a large scale, using DNase I hypersensitivity data from 78 human cell lines. We represented the universe of possible TF complexes by their corresponding motif complexes, and analyzed their occurrence at cell-type–specific DNase I hypersensitive sites. Based on ∼1.4 billion tests for motif complex enrichment, we predicted 603 highly significant cell-type–specific TF dimers, the vast majority of which are novel. Our predictions included 76% (19/25) of the known dimeric complexes and showed significant overlap with an experimental database of protein–protein interactions. They were also independently supported by evolutionary conservation, as well as quantitative variation in DNase I digestion patterns. Notably, the known and predicted TF dimers were almost always highly compact and rigidly spaced, suggesting that TFs dimerize in close proximity to their partners, which results in strict constraints on the structure of the DNA-bound complex. Overall, our results indicate that chromatin openness profiles are highly predictive of cell-type–specific TF–TF interactions. Moreover, cooperative TF dimerization seems to be a widespread phenomenon, with multiple TF complexes predicted in most cell types. PMID:23554463

  20. Profiling ethanol-targeted transcription factors in human carcinoma cell-derived embryoid bodies.

    Science.gov (United States)

    Mandal, Chanchal; Halder, Debasish; Chai, Jin Choul; Lee, Young Seek; Jung, Kyoung Hwa; Chai, Young Gyu

    2016-01-15

    Fetal alcohol spectrum disorder is a collective term that represents fetal abnormalities associated with maternal alcohol consumption. Prenatal alcohol exposure and related anomalies are well characterized, but the molecular mechanism behind this phenomenon is not yet understood. Few insights have been gained from genetic and epigenetic studies of fetal alcohol spectrum disorder. Our aim was to profile the important molecular regulators of ethanol-related alterations of the genome. For this purpose, we have analyzed the gene expression pattern of human carcinoma cell-derived embryoid bodies in the absence or presence of ethanol. A cDNA microarray analysis was used to profile mRNA expression in embryoid bodies at day 7 with or without ethanol treatment. A total of 493 differentially expressed genes were identified in response to 50 mM ethanol exposure. Of these, 111 genes were up-regulated, and 382 were down-regulated. Gene ontology term enrichment analysis revealed that these genes are involved in important biological processes: neurological system processes, cognition, behavior, sensory perception of smell, taste and chemical stimuli and synaptic transmission. Similarly, the enrichment of disease-related genes included relevant categories such as neurological diseases, developmental disorders, skeletal and muscular disorders, and connective tissue disorders. Furthermore, we have identified a group of 26 genes that encode transcription factors. We validated the relative gene expression of several transcription factors using quantitative real time PCR. We hope that our study substantially contributes to the understanding of the molecular mechanisms underlying the pathology of alcohol-mediated anomalies and facilitates further research. Copyright © 2015 Elsevier B.V. All rights reserved.

  1. Variation in Bluetongue virus real-time reverse transcription polymerase chain reaction assay results in blood samples of sheep, cattle, and alpaca.

    Science.gov (United States)

    Brito, Barbara P; Gardner, Ian A; Hietala, Sharon K; Crossley, Beate M

    2011-07-01

    Bluetongue is a vector-borne viral disease that affects domestic and wild ruminants. The epidemiology of this disease has recently changed, with occurrence in new geographic areas. Various real-time quantitative reverse transcription polymerase chain reaction (real-time qRT-PCR) assays are used to detect Bluetongue virus (BTV); however, the impact of biologic differences between New World camelids and domestic ruminant samples on PCR efficiency, for which the BTV real-time qRT-PCR was initially validated are unknown. New world camelids are known to have important biologic differences in whole blood composition, including hemoglobin concentration, which can alter PCR performance. In the present study, sheep, cattle, and alpaca blood were spiked with BTV serotypes 10, 11, 13, and 17 and analyzed in 10-fold dilutions by real-time qRT-PCR to determine if species affected nucleic acid recovery and assay performance. A separate experiment was performed using spiked alpaca blood subsequently diluted in 10-fold series in sheep blood to assess the influence of alpaca blood on performance efficiency of the BTV real-time qRT-PCR assay. Results showed that BTV-specific nucleic acid detection from alpaca blood was consistently 1-2 logs lower than from sheep and cattle blood, and results were similar for each of the 4 BTV serotypes analyzed.

  2. Genes Involved in Human Ribosome Biogenesis areTranscriptionally Upregulated in Colorectal Cancer

    DEFF Research Database (Denmark)

    Mansilla, Francisco; Lamy, Philippe; Ørntoft, Torben Falck

    2009-01-01

    Microarray gene expression profiling comprising 168 colorectal adenocarcinomas and 10 normal mucosas showed that over 79% of the genes involved in human ribosome biogenesis are significantly upregulated (log2>0.5, p<10-3) when compared to normal mucosa. Overexpression was independent of microsate......Microarray gene expression profiling comprising 168 colorectal adenocarcinomas and 10 normal mucosas showed that over 79% of the genes involved in human ribosome biogenesis are significantly upregulated (log2>0.5, p... of microsatellite status. The promoters of the genes studied showed a significant enrichment for several transcription factor binding sites. There was a significant correlation between the number of binding site targets for these transcription factors and the observed gene transcript upregulation. The upregulation...

  3. Genome-wide profiling of H3K56 acetylation and transcription factor binding sites in human adipocytes.

    Directory of Open Access Journals (Sweden)

    Kinyui Alice Lo

    Full Text Available The growing epidemic of obesity and metabolic diseases calls for a better understanding of adipocyte biology. The regulation of transcription in adipocytes is particularly important, as it is a target for several therapeutic approaches. Transcriptional outcomes are influenced by both histone modifications and transcription factor binding. Although the epigenetic states and binding sites of several important transcription factors have been profiled in the mouse 3T3-L1 cell line, such data are lacking in human adipocytes. In this study, we identified H3K56 acetylation sites in human adipocytes derived from mesenchymal stem cells. H3K56 is acetylated by CBP and p300, and deacetylated by SIRT1, all are proteins with important roles in diabetes and insulin signaling. We found that while almost half of the genome shows signs of H3K56 acetylation, the highest level of H3K56 acetylation is associated with transcription factors and proteins in the adipokine signaling and Type II Diabetes pathways. In order to discover the transcription factors that recruit acetyltransferases and deacetylases to sites of H3K56 acetylation, we analyzed DNA sequences near H3K56 acetylated regions and found that the E2F recognition sequence was enriched. Using chromatin immunoprecipitation followed by high-throughput sequencing, we confirmed that genes bound by E2F4, as well as those by HSF-1 and C/EBPα, have higher than expected levels of H3K56 acetylation, and that the transcription factor binding sites and acetylation sites are often adjacent but rarely overlap. We also discovered a significant difference between bound targets of C/EBPα in 3T3-L1 and human adipocytes, highlighting the need to construct species-specific epigenetic and transcription factor binding site maps. This is the first genome-wide profile of H3K56 acetylation, E2F4, C/EBPα and HSF-1 binding in human adipocytes, and will serve as an important resource for better understanding adipocyte

  4. Hypoxia, hypoxia-inducible transcription factor, and macrophages in human atherosclerotic plaques are correlated with intraplaque angiogenesis

    NARCIS (Netherlands)

    Sluimer, Judith C.; Gasc, Jean-Marie; van Wanroij, Job L.; Kisters, Natasja; Groeneweg, Mathijs; Sollewijn Gelpke, Maarten D.; Cleutjens, Jack P.; van den Akker, Luc H.; Corvol, Pierre; Wouters, Bradly G.; Daemen, Mat J.; Bijnens, Ann-Pascale J.

    2008-01-01

    We sought to examine the presence of hypoxia in human carotid atherosclerosis and its association with hypoxia-inducible transcription factor (HIF) and intraplaque angiogenesis. Atherosclerotic plaques develop intraplaque angiogenesis, which is a typical feature of hypoxic tissue and expression of

  5. Analysis of the human intestinal epithelial cell transcriptional response to Lactobacillus acidophilus, Lactobacillus salivarius, Bifidobacterium lactis and Escherichia coli

    DEFF Research Database (Denmark)

    Putaala, H; Barrangou, R; Leyer, G J

    2010-01-01

    a comparative analysis of the global in vitro transcriptional response of human intestinal epithelial cells to Lactobacillus acidophilus NCFM™, Lactobacillus salivarius Ls-33, Bifidobacterium animalis subsp. lactis 420, and enterohaemorrhagic Escherichia coli O157:H7 (EHEC). Interestingly, L. salivarius Ls-33...

  6. Noncoding RNA in the Transcriptional Landscape of Human Neural Progenitor Cell Differentiation

    Directory of Open Access Journals (Sweden)

    Patrick eHecht

    2015-10-01

    Full Text Available Increasing evidence suggests that noncoding RNAs play key roles in cellular processes, particularly in the brain. The present study used RNA sequencing to identify the transcriptional landscape of two human neural progenitor cell lines, SK-N-SH and ReNcell CX, as they differentiate into human cortical projection neurons. Protein coding genes were found to account for 54.8% and 57.0% of expressed genes, respectively, and alignment of RNA sequencing reads revealed that only 25.5-28.1% mapped to exonic regions of the genome. Differential expression analysis in the two cell lines identified altered gene expression in both protein coding and noncoding RNAs as they undergo neural differentiation with 222 differentially expressed genes observed in SK-N-SH cells and 19 differentially expressed genes in ReNcell CX. Interestingly, genes showing differential expression in SK-N-SH cells are enriched in genes implicated in autism spectrum disorder, but not in gene sets related to cancer or Alzheimer’s disease. Weighted gene co-expression network analysis (WGCNA was used to detect modules of co-expressed protein coding and noncoding RNAs in SK-N-SH cells and found four modules to be associated with neural differentiation. These modules contain varying levels of noncoding RNAs ranging from 10.7% to 49.7% with gene ontology suggesting roles in numerous cellular processes important for differentiation. These results indicate that noncoding RNAs are highly expressed in human neural progenitor cells and likely hold key regulatory roles in gene networks underlying neural differentiation and neurodevelopmental disorders.

  7. Transcription factors GATA-4 and GATA-6 in normal and neoplastic human gastrointestinal mucosa

    Directory of Open Access Journals (Sweden)

    Mäki Markku

    2008-04-01

    Full Text Available Abstract Background Human gastrointestinal mucosa regenerates vigorously throughout life, but the factors controlling cell fate in mature mucosa are poorly understood. GATA transcription factors direct cell proliferation and differentiation in many organs, and are implicated in tumorigenesis. GATA-4 and GATA-6 are considered crucial for the formation of murine gastrointestinal mucosa, but their role in human gastrointestinal tract remains unexplored. We studied in detail the expression patterns of these two GATA factors and a GATA-6 down-stream target, Indian hedgehog (Ihh, in normal human gastrointestinal mucosa. Since these factors are considered important for proliferation and differentiation, we also explored the possible alterations in their expression in gastrointestinal neoplasias. The expression of the carcinogenesis-related protein Indian hedgehog was also investigated in comparison to GATA factors. Methods Samples of normal and neoplastic gastrointestinal tract from children and adults were subjected to RNA in situ hybridization with 33P labelled probes and immunohistochemistry, using an avidin-biotin immunoperoxidase system. The pathological tissues examined included samples of chronic and atrophic gastritis as well as adenomas and adenocarcinomas of the colon and rectum. Results GATA-4 was abundant in the differentiated epithelial cells of the proximal parts of the gastrointestinal tract but was absent from the distal parts. In contrast, GATA-6 was expressed throughout the gastrointestinal epithelium, and in the distal gut its expression was most intense at the bottom of the crypts, i.e. cells with proliferative capacity. Both factors were also present in Barrett's esophagus and metaplasia of the stomach. GATA-6 expression was reduced in colon carcinoma. Ihh expression overlapped with that of GATA-6 especially in benign gastrointestinal neoplasias. Conclusion The results suggest differential but overlapping functions for GATA-4 and

  8. Chlorinated Flavonoids Modulate the Inflammatory Process in Human Blood.

    Science.gov (United States)

    Proença, Carina; Ribeiro, Daniela; Soares, Tânia; Tomé, Sara M; Silva, Artur M S; Lima, José L F C; Fernandes, Eduarda; Freitas, Marisa

    2017-08-01

    Flavonoids are known to react with neutrophil-generated hypochlorous acid (HOCl) at inflammation loci to form stable mono- and dichlorinated products. Some of these products have been shown to retain or even enhance their inflammatory potential, but further information is required in a broader approach to inflammatory mechanisms. In that sense, we performed an integrated evaluation on the anti-inflammatory potential of a panel of novel chlorinated flavonoids and their parent compounds, in several steps of the complex inflammatory cascade, namely, in the activity of cyclooxygenase (COX)-1 and COX-2, and in the production of cytokines [interleukin (IL)-6, IL-1β, tumor necrosis factor (TNF)], and the chemokine, IL-8, as well as in the production of reactive species, using human whole blood as a representative in vitro model, establishing, whenever possible, a structure-activity relationship. Although luteolin was the most active compound, chlorinated flavonoids demonstrated a remarkable pattern of activity for the resolution of the inflammatory processes. Our results demonstrated that 6-chloro-3',4',5,7-tetrahydroxyflavone deserves scientific attention due to its ability to modulate the reactive species and cytokines/chemokine production. In this regard, the therapeutic potential of flavonoids' metabolites, and in this particular case the chlorinated flavonoids, should not be neglected.

  9. Identification of transcriptional macromolecular associations in human bone using browser based in silico analysis in a giant correlation matrix.

    Science.gov (United States)

    Reppe, Sjur; Sachse, Daniel; Olstad, Ole K; Gautvik, Vigdis T; Sanderson, Paul; Datta, Harish K; Berg, Jens P; Gautvik, Kaare M

    2013-03-01

    Intracellular signaling is critically dependent on gene regulatory networks comprising physical molecular interactions. Presently, there is a lack of comprehensive databases for most human tissue types to verify such macromolecular interactions. We present a user friendly browser which helps to identify functional macromolecular interactions in human bone as significant correlations at the transcriptional level. The molecular skeletal phenotype has been characterized by transcriptome analysis of iliac crest bone biopsies from 84 postmenopausal women through quantifications of ~23,000 mRNA species. When the signal levels were inter-correlated, an array containing >260 million correlations was generated, thus recognizing the human bone interactome at the RNA level. The matrix correlation and p values were made easily accessible by a freely available online browser. We show that significant correlations within the giant matrix are reproduced in a replica set of 13 male vertebral biopsies. The identified correlations differ somewhat from transcriptional interactions identified in cell culture experiments and transgenic mice, thus demonstrating that care should be taken in extrapolating such results to the in vivo situation in human bone. The current giant matrix and web browser are a valuable tool for easy access to the human bone transcriptome and molecular interactions represented as significant correlations at the RNA-level. The browser and matrix should be a valuable hypothesis generating tool for identification of regulatory mechanisms and serve as a library of transcript relationships in human bone, a relatively inaccessible tissue. Copyright © 2012 Elsevier Inc. All rights reserved.

  10. Transcriptional profile of isoproterenol-induced cardiomyopathy and comparison to exercise-induced cardiac hypertrophy and human cardiac failure

    Directory of Open Access Journals (Sweden)

    McIver Lauren J

    2009-12-01

    Full Text Available Abstract Background Isoproterenol-induced cardiac hypertrophy in mice has been used in a number of studies to model human cardiac disease. In this study, we compared the transcriptional response of the heart in this model to other animal models of heart failure, as well as to the transcriptional response of human hearts suffering heart failure. Results We performed microarray analyses on RNA from mice with isoproterenol-induced cardiac hypertrophy and mice with exercise-induced physiological hypertrophy and identified 865 and 2,534 genes that were significantly altered in pathological and physiological cardiac hypertrophy models, respectively. We compared our results to 18 different microarray data sets (318 individual arrays representing various other animal models and four human cardiac diseases and identified a canonical set of 64 genes that are generally altered in failing hearts. We also produced a pairwise similarity matrix to illustrate relatedness of animal models with human heart disease and identified ischemia as the human condition that most resembles isoproterenol treatment. Conclusion The overall patterns of gene expression are consistent with observed structural and molecular differences between normal and maladaptive cardiac hypertrophy and support a role for the immune system (or immune cell infiltration in the pathology of stress-induced hypertrophy. Cross-study comparisons such as the results presented here provide targets for further research of cardiac disease that might generally apply to maladaptive cardiac stresses and are also a means of identifying which animal models best recapitulate human disease at the transcriptional level.

  11. Blood-brain transfer of Pittsburgh compound B in humans

    DEFF Research Database (Denmark)

    Gjedde, Albert; Aanerud, Joel; Braendgaard, Hans

    2013-01-01

    -brain barrier is held to be high but the permeability-surface area product and extraction fractions in patients or healthy volunteers are not known. We used PET to determine the clearance associated with the unidrectional blood-brain transfer of [(11)C]PiB and the corresponding cerebral blood flow rates...... with the observation that numerically, but insignificantly, unidirectional blood-brain clearances are lower and extraction fractions higher in the patients. The evidence of unchanged permeability-surface area products in the patients implies that blood flow changes can be deduced from the unidirectional blood......In the labeled form, the Pittsburgh compound B (2-(4'-{N-methyl-[(11)C]}methyl-aminophenyl)-6-hydroxy-benzothiazole, [(11)C]PiB), is used as a biomarker for positron emission tomography (PET) of brain β-amyloid deposition in Alzheimer's disease (AD). The permeability of [(11)C]PiB in the blood...

  12. Investigation of the proteins relaxation time in human blood serum; Badania relaksacyjne bialek surowicy krwi II

    Energy Technology Data Exchange (ETDEWEB)

    Blicharska, B.; Klauza, M. [Inst. Fizyki, Uniwersytet Jagiellonski, Cracow (Poland); Kuliszkiewicz-Janus, M. [Akademia Medyczna, Wroclaw (Poland)

    1994-12-31

    In this paper the results of human blood serum proteins relaxation time measurements by means of NMR method are presented. The measurements have been done for three samples of human blood: i/laudably ii/leukemia iii/granulomas. The dependences of the relaxation time on the temperature are also presented. 3 refs, 4 figs.

  13. Light-scattering properties of undiluted human blood subjected to simple shear

    NARCIS (Netherlands)

    Steenbergen, Wiendelt; Kolkman, R.G.M.; de Mul, F.F.M.

    1999-01-01

    An experimental investigation was performed into the effect of simple shear on the light-scattering properties of undiluted human blood. Undiluted human blood was enclosed between two glass plates with an adjustable separation between 30 and 120 mm and with one plate moving parallel to the other.

  14. RNA/DNA co-analysis from human menstrual blood and vaginal secretion stains

    DEFF Research Database (Denmark)

    Haas, Claus; Hanson, E; Anjos, M J

    2014-01-01

    housekeeping genes for their suitability as reference genes. Six menstrual blood and six vaginal secretion stains, two dilution series (1/4-1/64 pieces of a menstrual blood/vaginal swab) and, optionally, bona fide or mock casework samples of human or non-human origin were analyzed by 24 participating...

  15. Transient receptor potential canonical type 3 channels and blood pressure in humans

    DEFF Research Database (Denmark)

    Thilo, Florian; Baumunk, Daniel; Krause, Hans

    2009-01-01

    There is evidence that transient receptor potential canonical type 3 (TRPC3) cation channels are involved in the regulation of blood pressure, but this has not been studied using human renal tissue. We tested the hypothesis that the expression of TRPC3 in human renal tissue is associated with blood...

  16. Time-dependent histamine release from stored human blood products

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Edvardsen, L; Vangsgaard, K

    1996-01-01

    .0 (range 176.0-910.0) nmol/l in whole blood and 475.0 (range 360.0-1560.0) nmol/l in plasma-reduced whole blood, while it was undetectable in SAGM blood. Spontaneous histamine release increased in a time-dependent manner from a median of 6.7 (range 2.2-17.4) nmol/l at the time of storage to 175.0 (range 33...

  17. Transcriptional profiling of whole blood and serum protein analysis of mice exposed to the neurotoxin Pacific Ciguatoxin-1.

    Science.gov (United States)

    Ryan, James C; Bottein Dechraoui, Marie-Yasmine; Morey, Jeanine S; Rezvani, Amir; Levin, Edward D; Gordon, Christopher J; Ramsdell, John S; Van Dolah, Frances M

    2007-11-01

    Ciguatoxins (CTX) are a suite of cyclic polyether toxins produced by the marine dinoflagellate Gambierdiscus sp., are potent activators of voltage-gated sodium channels and a leading cause of human poisoning from food fish. This report characterizes the genomic and proteomic response in whole blood of adult male mice exposed i.p. to 264 ng/kg of the Pacific congener of CTX (P-CTX-1) at 1, 4 and 24h. Whole genome microarray expression data were filtered by tightness of fit between replicates, fold change (1.8) and p-value (10(-5)), resulting in 183 annotated genes used for trending analysis, K-means clustering and ontology classification. Genes involved with cytokine signaling, proteasome complex and ribosomal function were dominant. qPCR performed on 19 genes of interest had a correlation of 0.95 to array results by Pearson's correlation coefficient. Serum protein analysis showed small but significant changes in 6 of 60 proteins assayed: Ccl2, Ccl12, CD40, IL-10, leptin and M-CSF. In large part, the gene expression was consistent with a Th2 immune response with interesting similarities to expression seen in asthmatic models.

  18. Effects of Contrast Media on Blood Rheology: Comparison in Humans, Pigs, and Sheep

    International Nuclear Information System (INIS)

    Laurent, Alexandre; Durussel, Jean Jacques; Dufaux, Jacques; Penhouet, Laurence; Bailly, Anne Laure; Bonneau, Michel; Merland, Jean Jacques

    1999-01-01

    Purpose: To compare whole blood viscosity and erythrocyte aggregation in humans, pigs, and sheep, before and after adding water-soluble iodinated contrast medium (CM). Methods: Two CMs were studied: iopromide (nonionic) and ioxaglate (ionic). The blood-CM viscosity was measured with a Couette viscometer. Erythrocyte aggregation was measured with an erythroaggregometer. Results: The blood-CM viscosity was increased up to +20% (relative to pure blood) with a CM concentration of 0%-10%. At CM concentrations from 10% to 50%, the viscosity decreased. The disaggregation shear stress was increased (relative to pure blood) at low CM concentration (0%-10%). When the CM concentration increased from 10% to 20%, the disaggregation shear stress was decreased, except with the pig blood-ioxaglate mixture. Conclusion: At low CM concentration the blood viscosity was increased in pig, sheep, and humans and the disaggregation shear stress was increased in pig and humans. The aggregation of sheep blood was too low to be detected by the erythroaggregometer. This rise can be explained by the formation of poorly deformable echinocytes. At higher CM concentration, the viscosity and the disaggregation shear stress decreased in relation to the blood dilution. We conclude that pig blood and sheep blood can both be used to study the effect of CM injection on blood viscosity. Nevertheless, the rheologic behavior of pig blood in terms of erythrocyte aggregation is closer to that of human blood than is sheep blood when mixed with CM. Pigs could thus be more suitable than sheep for in vivo studies of CM miscibility with blood during selective cannulation procedures

  19. Generation of Human Induced Pluripotent Stem Cells from Peripheral Blood Mononuclear Cells Using Sendai Virus.

    Science.gov (United States)

    Soares, Filipa A C; Pedersen, Roger A; Vallier, Ludovic

    2016-01-01

    This protocol describes the efficient isolation of peripheral blood mononuclear cells from circulating blood via density gradient centrifugation and subsequent generation of integration-free human induced pluripotent stem cells. Peripheral blood mononuclear cells are cultured for 9 days to allow expansion of the erythroblast population. The erythroblasts are then used to derive human induced pluripotent stem cells using Sendai viral vectors, each expressing one of the four reprogramming factors Oct4, Sox2, Klf4, and c-Myc.

  20. Time-dependent histamine release from stored human blood products

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Edvardsen, L; Vangsgaard, K

    1996-01-01

    storage. Whole blood (six units), plasma-reduced whole blood (six units), and plasma- and buffy coat-reduced (saline-adenine-glucose-mannitol) (SAGM) blood (six units) from unpaid healthy donors were stored in the blood bank for 35 days at 4 degrees C. Plasma histamine and total cell-bound histamine......Perioperative transfusion of whole blood has been shown to amplify trauma-induced immunosuppression, which could be attenuated by perioperative administration of histamine2 receptor antagonists. Supernatants from different blood products were, therefore, analysed for histamine content during.......0 (range 176.0-910.0) nmol/l in whole blood and 475.0 (range 360.0-1560.0) nmol/l in plasma-reduced whole blood, while it was undetectable in SAGM blood. Spontaneous histamine release increased in a time-dependent manner from a median of 6.7 (range 2.2-17.4) nmol/l at the time of storage to 175.0 (range 33...

  1. Core Transcriptional Regulatory Circuit Controlled by the TAL1 Complex in Human T Cell Acute Lymphoblastic Leukemia

    OpenAIRE

    Sanda, Takaomi; Lawton, Lee N.; Barrasa, M. Inmaculada; Fan, Zi Peng; Kohlhammer, Holger; Gutierrez, Alejandro; Ma, Wenxue; Tatarek, Jessica; Ahn, Yebin; Kelliher, Michelle A.; Jamieson, Catriona H.M.; Staudt, Louis M.; Young, Richard A.; Look, A. Thomas

    2012-01-01

    The oncogenic transcription factor TAL1/SCL is aberrantly expressed in over 40% of cases of human T-cell acute lymphoblastic leukemia (T-ALL), emphasizing its importance in the molecular pathogenesis of T-ALL. Here we identify the core transcriptional regulatory circuit controlled by TAL1 and its regulatory partners HEB, E2A, LMO1/2, GATA3 and RUNX1. We show that TAL1 forms a positive interconnected auto-regulatory loop with GATA3 and RUNX1, and that the TAL1 complex directly activates the MY...

  2. Specific interactions between transcription factors and the promoter-regulatory region of the human cytomegalovirus major immediate-early gene

    International Nuclear Information System (INIS)

    Ghazal, P.; Lubon, H.; Hennighausen, L.

    1988-01-01

    Repeat sequence motifs as well as unique sequences between nucleotides -150 and -22 of the human cytomegalovirus immediate-early 1 gene interact in vitro with nuclear proteins. The authors show that a transcriptional element between nucleotides -91 and -65 stimulated promoter activity in vivo and in vitro by binding specific cellular transcription factors. Finally, a common sequence motif, (T)TGG/AC, present in 15 of the determined binding sites suggests a particular class of nuclear factors associated with the immediate-early 1 gene

  3. Whole thorax irradiation of non-human primates induces persistent nuclear damage and gene expression changes in peripheral blood cells.

    Directory of Open Access Journals (Sweden)

    Shanaz A Ghandhi

    Full Text Available We investigated the cytogenetic and gene expression responses of peripheral blood cells of non-human primates (NHP, Macaca mulatta that were whole-thorax irradiated with a single dose of 10 Gy. In this model, partial irradiation of NHPs in the thoracic region (Whole Thorax Lung Irradiation, WTLI allows the study of late radiation-induced lung injury, while avoiding acute radiation syndromes related to hematopoietic and gastrointestinal injury. A transient drop in circulating lymphocytes and platelets was seen by 9 days, followed by elevations in respiratory rate, circulating neutrophils, lymphocytes, and monocytes at 60-100 days, corresponding to computed tomography (CT and histologic evidence of pneumonitis, and elective euthanasia of four animals. To evaluate long-term DNA damage in NHP peripheral blood lymphocytes after 10 Gy WTLI, we used the cytokinesis-block micronucleus (CBMN assay to measure chromosomal aberrations as post-mitotic micronuclei in blood samples collected up to 8 months after irradiation. Regression analysis showed significant induction of micronuclei in NHP blood cells that persisted with a gradual decline over the 8-month study period, suggesting long-term DNA damage in blood lymphocytes after WTLI. We also report transcriptomic changes in blood up to 30 days after WTLI. We isolated total RNA from peripheral blood at 3 days before and then at 2, 5 and 30 days after irradiation. We identified 1187 transcripts that were significantly changed across the 30-day time course. From changes in gene expression, we identified biological processes related to immune responses, which persisted across the 30-day study. Response to oxygen-containing compounds and bacteria were implicated by gene-expression changes at the earliest day 2 and latest, day 30 time-points. Gene expression changes suggest a persistent altered state of the immune system, specifically response to infection, for at least a month after WTLI.

  4. Whole thorax irradiation of non-human primates induces persistent nuclear damage and gene expression changes in peripheral blood cells.

    Science.gov (United States)

    Ghandhi, Shanaz A; Turner, Helen C; Shuryak, Igor; Dugan, Gregory O; Bourland, J Daniel; Olson, John D; Tooze, Janet A; Morton, Shad R; Batinic-Haberle, Ines; Cline, J Mark; Amundson, Sally A

    2018-01-01

    We investigated the cytogenetic and gene expression responses of peripheral blood cells of non-human primates (NHP, Macaca mulatta) that were whole-thorax irradiated with a single dose of 10 Gy. In this model, partial irradiation of NHPs in the thoracic region (Whole Thorax Lung Irradiation, WTLI) allows the study of late radiation-induced lung injury, while avoiding acute radiation syndromes related to hematopoietic and gastrointestinal injury. A transient drop in circulating lymphocytes and platelets was seen by 9 days, followed by elevations in respiratory rate, circulating neutrophils, lymphocytes, and monocytes at 60-100 days, corresponding to computed tomography (CT) and histologic evidence of pneumonitis, and elective euthanasia of four animals. To evaluate long-term DNA damage in NHP peripheral blood lymphocytes after 10 Gy WTLI, we used the cytokinesis-block micronucleus (CBMN) assay to measure chromosomal aberrations as post-mitotic micronuclei in blood samples collected up to 8 months after irradiation. Regression analysis showed significant induction of micronuclei in NHP blood cells that persisted with a gradual decline over the 8-month study period, suggesting long-term DNA damage in blood lymphocytes after WTLI. We also report transcriptomic changes in blood up to 30 days after WTLI. We isolated total RNA from peripheral blood at 3 days before and then at 2, 5 and 30 days after irradiation. We identified 1187 transcripts that were significantly changed across the 30-day time course. From changes in gene expression, we identified biological processes related to immune responses, which persisted across the 30-day study. Response to oxygen-containing compounds and bacteria were implicated by gene-expression changes at the earliest day 2 and latest, day 30 time-points. Gene expression changes suggest a persistent altered state of the immune system, specifically response to infection, for at least a month after WTLI.

  5. Transcriptional and Cell Cycle Alterations Mark Aging of Primary Human Adipose-Derived Stem Cells.

    Science.gov (United States)

    Shan, Xiaoyin; Roberts, Cleresa; Kim, Eun Ji; Brenner, Ariana; Grant, Gregory; Percec, Ivona

    2017-05-01

    Adult stem cells play a critical role in the maintenance of tissue homeostasis and prevention of aging. While the regenerative potential of stem cells with low cellular turnover, such as adipose-derived stem cells (ASCs), is increasingly recognized, the study of chronological aging in ASCs is technically difficult and remains poorly understood. Here, we use our model of chronological aging in primary human ASCs to examine genome-wide transcriptional networks. We demonstrate first that the transcriptome of aging ASCs is distinctly more stable than that of age-matched fibroblasts, and further, that age-dependent modifications in cell cycle progression and translation initiation specifically characterize aging ASCs in conjunction with increased nascent protein synthesis and a distinctly shortened G1 phase. Our results reveal novel chronological aging mechanisms in ASCs that are inherently different from differentiated cells and that may reflect an organismal attempt to meet the increased demands of tissue and organ homeostasis during aging. Stem Cells 2017;35:1392-1401. © 2017 AlphaMed Press.

  6. Musa paradisiaca inflorescence induces human colon cancer cell death by modulating cascades of transcriptional events.

    Science.gov (United States)

    K B, Arun; Madhavan, Aravind; T R, Reshmitha; Thomas, Sithara; Nisha, P

    2018-01-24

    Colorectal cancer (CRC) is one of the leading causes of cancer death, and diet plays an important role in the etiology of CRC. Traditional medical practitioners in many South Asian countries use plantain inflorescence to treat various gastro-intestinal ailments. The aim of the present study was to investigate the anticancer effects of extracts of inflorescence of Musa paradisiaca against HT29 human colon cancer cells and elucidate the mechanism of these effects by studying the modulation of cascades of transcriptional events. In vitro assays depicted that methanol extract of Musa paradisiaca inflorescence (PIMET) was cytotoxic to HT29 cells. PIMET induced DNA damage and arrested the cell cycle at the G2/M phase. Expression studies showed that PIMET pretreatment upregulates pro-apoptotic Bcl2 and downregulates anti-apoptotic Bax proteins. Different assays showed that the deregulation of pro/antiapoptotic proteins reduces the mitochondrial membrane potential and ATP production; moreover, it enhances cytochrome c release, which triggers the apoptotic pathway, and further cleaves caspase 3 and PARP proteins, resulting in apoptosis. Changes in the protein expression profile of HT29 cells after PIMET treatment were analyzed using mass-spectrometry-based proteomics. PIMET treatment significantly altered the expression of HT29 protein; interestingly, X-linked inhibitor of apoptosis protein was also downregulated. Alteration in the expression of this protein has significant effects, leading to HT29 cell death.

  7. Sequence and transcription analysis of the human cytomegalovirus DNA polymerase gene

    International Nuclear Information System (INIS)

    Kouzarides, T.; Bankier, A.T.; Satchwell, S.C.; Weston, K.; Tomlinson, P.; Barrell, B.G.

    1987-01-01

    DNA sequence analysis has revealed that the gene coding for the human cytomegalovirus (HCMV) DNA polymerase is present within the long unique region of the virus genome. Identification is based on extensive amino acid homology between the predicted HCMV open reading frame HFLF2 and the DNA polymerase of herpes simplex virus type 1. The authors present here a 5280 base-pair DNA sequence containing the HCMV pol gene, along with the analysis of transcripts encoded within this region. Since HCMV pol also shows homology to the predicted Epstein-Barr virus pol, they were able to analyze the extent of homology between the DNA polymerases of three distantly related herpes viruses, HCMV, Epstein-Barr virus, and herpes simplex virus. The comparison shows that these DNA polymerases exhibit considerable amino acid homology and highlights a number of highly conserved regions; two such regions show homology to sequences within the adenovirus type 2 DNA polymerase. The HCMV pol gene is flanked by open reading frames with homology to those of other herpes viruses; upstream, there is a reading frame homologous to the glycoprotein B gene of herpes simplex virus type I and Epstein-Barr virus, and downstream there is a reading frame homologous to BFLF2 of Epstein-Barr virus

  8. Characterization of herpes simplex virus type 2 latency-associated transcription in human sacral ganglia and in cell culture.

    Science.gov (United States)

    Croen, K D; Ostrove, J M; Dragovic, L; Straus, S E

    1991-01-01

    The ability of herpes simplex virus type 2 (HSV-2) to establish latency in and reactivate from sacral dorsal root sensory ganglia is the basis for recurrent genital herpes. The expression of HSV-2 genes in latently infected human sacral ganglia was investigated by in situ hybridization. Hybridizations with a probe from the long repeat region of HSV-2 revealed strong nuclear signals overlying neurons in sacral ganglia from five of nine individuals. The RNA detected overlaps with the transcript for infected cell protein O but in the opposite, or "anti-sense," orientation. These observations mimic those made previously with HSV-1 in human trigeminal ganglia and confirm the recent findings during latency in HSV-2-infected mice and guinea pigs. Northern hybridization of RNA from infected Vero cells showed that an HSV-2 latency-associated transcript was similar in size to the larger (1.85 kb) latency transcript of HSV-1. Thus, HSV-1 and HSV-2 latency in human sensory ganglia are similar, if not identical, in terms of their cellular localization and pattern of transcription.

  9. Comparative transcriptional analysis of three human ligaments with distinct biomechanical properties

    Science.gov (United States)

    Lorda-Diez, Carlos I; Canga-Villegas, Ana; Cerezal, Luis; Plaza, Santiago; Hurlé, Juan M; García-Porrero, Juan A; Montero, Juan A

    2013-01-01

    One major aim of regenerative medicine targeting the musculoskeletal system is to provide complementary and/or alternative therapeutic approaches to current surgical therapies, often involving the removal and prosthetic substitution of damaged tissues such as ligaments. For these approaches to be successful, detailed information regarding the cellular and molecular composition of different musculoskeletal tissues is required. Ligaments have often been considered homogeneous tissues with common biomechanical properties. However, advances in tissue engineering research have highlighted the functional relevance of the organisational and compositional differences between ligament types, especially in those with higher risks of injury. The aim of this study was to provide information concerning the relative expression levels of a subset of key genes (including extracellular matrix components, transcription factors and growth factors) that confer functional identity to ligaments. We compared the transcriptomes of three representative human ligaments subjected to different biomechanical demands: the anterior cruciate ligament (ACL); the ligamentum teres of the hip (LT); and the iliofemoral ligament (IL). We revealed significant differences in the expression of type I collagen, elastin, fibromodulin, biglycan, transforming growth factor β1, transforming growth interacting factor 1, hypoxia-inducible factor 1-alpha and transforming growth factor β-induced gene between the IL and the other two ligaments. Thus, considerable molecular heterogeneity can exist between anatomically distinct ligaments with differing biomechanical demands. However, the LT and ACL were found to show remarkable molecular homology, suggesting common functional properties. This finding provides experimental support for the proposed role of the LT as a hip joint stabiliser in humans. PMID:24128114

  10. Transcriptional Modulation of Human Endogenous Retroviruses in Primary CD4+ T Cells Following Vorinostat Treatment

    Directory of Open Access Journals (Sweden)

    Cory H. White

    2018-04-01

    Full Text Available The greatest obstacle to a cure for HIV is the provirus that integrates into the genome of the infected cell and persists despite antiretroviral therapy. A “shock and kill” approach has been proposed as a strategy for an HIV cure whereby drugs and compounds referred to as latency-reversing agents (LRAs are used to “shock” the silent provirus into active replication to permit “killing” by virus-induced pathology or immune recognition. The LRA most utilized to date in clinical trials has been the histone deacetylase (HDAC inhibitor—vorinostat. Potentially, pathological off-target effects of vorinostat may result from the activation of human endogenous retroviruses (HERVs, which share common ancestry with exogenous retroviruses including HIV. To explore the effects of HDAC inhibition on HERV transcription, an unbiased pharmacogenomics approach (total RNA-Seq was used to evaluate HERV expression following the exposure of primary CD4+ T cells to a high dose of vorinostat. Over 2,000 individual HERV elements were found to be significantly modulated by vorinostat, whereby elements belonging to the ERVL family (e.g., LTR16C and LTR33 were predominantly downregulated, in contrast to LTR12 elements of the HERV-9 family, which exhibited the greatest signal, with the upregulation of 140 distinct elements. The modulation of three different LTR12 elements by vorinostat was confirmed by droplet digital PCR along a dose–response curve. The monitoring of LTR12 expression during clinical trials with vorinostat may be indicated to assess the impact of this HERV on the human genome and host immunity.

  11. Transcriptional profiling of human femoral mesenchymal stem cells in osteoporosis and its association with adipogenesis.

    Science.gov (United States)

    Choi, Yong Jun; Song, Insun; Jin, Yilan; Jin, Hyun-Seok; Ji, Hyung Min; Jeong, Seon-Yong; Won, Ye-Yeon; Chung, Yoon-Sok

    2017-10-20

    Genetic alterations are major contributing factors in the development of osteoporosis. Osteoblasts and adipocytes share a common origin, mesenchymal stem cells (MSCs), and their genetic determinants might be important in the relationship between osteoporosis and obesity. In the present study, we aimed to isolate differentially expressed genes (DEGs) in osteoporosis and normal controls using human MSCs, and elucidate the common pathways and genes related to osteoporosis and adipogenesis. Human MSCs were obtained from the bone marrow of femurs from postmenopausal women during orthopedic surgeries. RNA sequencing (RNA-seq) was carried out using next-generation sequencing (NGS) technology. DEGs were identified using RNA-seq data. Ingenuity pathway analysis (IPA) was used to elucidate the common pathway related to osteoporosis and adipogenesis. Candidate genes for the common pathway were validated with other independent osteoporosis and obese subjects using RT-PCR (reverse transcription-polymerase chain reaction) analysis. Fifty-three DEGs were identified between postmenopausal osteoporosis patients and normal bone mineral density (BMD) controls. Most of the genetic changes were related to the differentiation of cells. The nuclear receptor subfamily 4 group A (NR4A) family was identified as possible common genes related to osteogenesis and adipogenesis. The expression level of the mRNA of NR4A1 was significantly higher in osteoporosis patients than in controls (p=0.018). The expression level of the mRNA of NR4A2 was significantly higher in obese patients than in controls (p=0.041). Some genetic changes in MSCs are involved in the pathophysiology of osteoporosis. The NR4A family might comprise common genes related to osteoporosis and obesity. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Transcriptional regulation of the human Liver X Receptor α gene by Hepatocyte Nuclear Factor 4α

    Energy Technology Data Exchange (ETDEWEB)

    Theofilatos, Dimitris; Anestis, Aristomenis [University of Crete Medical School and Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology of Hellas, Heraklion, 71003, Crete (Greece); Hashimoto, Koshi [Department of Preemptive Medicine and Metabolism, Tokyo Medical and Dental University, 1-5-45 Yushima, Bunkyo-city, Tokyo, 113-8510 (Japan); Kardassis, Dimitris, E-mail: kardasis@imbb.forth.gr [University of Crete Medical School and Institute of Molecular Biology and Biotechnology, Foundation for Research and Technology of Hellas, Heraklion, 71003, Crete (Greece)

    2016-01-15

    Liver X Receptors (LXRs) are sterol-activated transcription factors that play major roles in cellular cholesterol homeostasis, HDL biogenesis and reverse cholesterol transport. The aim of the present study was to investigate the mechanisms that control the expression of the human LXRα gene in hepatic cells. A series of reporter plasmids containing consecutive 5′ deletions of the hLXRα promoter upstream of the luciferase gene were constructed and the activity of each construct was measured in HepG2 cells. This analysis showed that the activity of the human LXRα promoter was significantly reduced by deleting the −111 to −42 region suggesting the presence of positive regulatory elements in this short proximal fragment. Bioinformatics data including motif search and ChIP-Seq revealed the presence of a potential binding motif for Hepatocyte Nuclear Factor 4 α (HNF-4α) in this area. Overexpression of HNF-4α in HEK 293T cells increased the expression of all LXRα promoter constructs except −42/+384. In line, silencing the expression of endogenous HNF-4α in HepG2 cells was associated with reduced LXRα protein levels and reduced activity of the −111/+384 LXRα promoter but not of the −42/+384 promoter. Using ChiP assays in HepG2 cells combined with DNAP assays we mapped the novel HNF-4α specific binding motif (H4-SBM) in the −50 to −40 region of the human LXRα promoter. A triple mutation in this H4-SBM abolished HNF-4α binding and reduced the activity of the promoter to 65% relative to the wild type. Furthermore, the mutant promoter could not be transactivated by HNF-4α. In conclusion, our data indicate that HNF-4α may have a wider role in cell and plasma cholesterol homeostasis by controlling the expression of LXRα in hepatic cells. - Highlights: • The human LXRα promoter contains a HNF-4α specific binding motif in the proximal −50/−40 region. • Mutations in this motif abolished HNF4α binding and transactivation of the h

  13. Transcriptional regulation of the human Liver X Receptor α gene by Hepatocyte Nuclear Factor 4α

    International Nuclear Information System (INIS)

    Theofilatos, Dimitris; Anestis, Aristomenis; Hashimoto, Koshi; Kardassis, Dimitris

    2016-01-01

    Liver X Receptors (LXRs) are sterol-activated transcription factors that play major roles in cellular cholesterol homeostasis, HDL biogenesis and reverse cholesterol transport. The aim of the present study was to investigate the mechanisms that control the expression of the human LXRα gene in hepatic cells. A series of reporter plasmids containing consecutive 5′ deletions of the hLXRα promoter upstream of the luciferase gene were constructed and the activity of each construct was measured in HepG2 cells. This analysis showed that the activity of the human LXRα promoter was significantly reduced by deleting the −111 to −42 region suggesting the presence of positive regulatory elements in this short proximal fragment. Bioinformatics data including motif search and ChIP-Seq revealed the presence of a potential binding motif for Hepatocyte Nuclear Factor 4 α (HNF-4α) in this area. Overexpression of HNF-4α in HEK 293T cells increased the expression of all LXRα promoter constructs except −42/+384. In line, silencing the expression of endogenous HNF-4α in HepG2 cells was associated with reduced LXRα protein levels and reduced activity of the −111/+384 LXRα promoter but not of the −42/+384 promoter. Using ChiP assays in HepG2 cells combined with DNAP assays we mapped the novel HNF-4α specific binding motif (H4-SBM) in the −50 to −40 region of the human LXRα promoter. A triple mutation in this H4-SBM abolished HNF-4α binding and reduced the activity of the promoter to 65% relative to the wild type. Furthermore, the mutant promoter could not be transactivated by HNF-4α. In conclusion, our data indicate that HNF-4α may have a wider role in cell and plasma cholesterol homeostasis by controlling the expression of LXRα in hepatic cells. - Highlights: • The human LXRα promoter contains a HNF-4α specific binding motif in the proximal −50/−40 region. • Mutations in this motif abolished HNF4α binding and transactivation of the h

  14. The viral transcription group determines the HLA class I cellular immune response against human respiratory syncytial virus.

    Science.gov (United States)

    Johnstone, Carolina; Lorente, Elena; Barriga, Alejandro; Barnea, Eilon; Infantes, Susana; Lemonnier, François A; David, Chella S; Admon, Arie; López, Daniel

    2015-04-01

    The cytotoxic T-lymphocyte-mediated killing of virus-infected cells requires previous recognition of short viral antigenic peptides bound to human leukocyte antigen class I molecules that are exposed on the surface of infected cells. The cytotoxic T-lymphocyte response is critical for the clearance of human respiratory syncytial virus infection. In this study, naturally processed viral human leukocyte antigen class I ligands were identified with mass spectrometry analysis of complex human leukocyte antigen-bound peptide pools isolated from large amounts of human respiratory syncytial virus-infected cells. Acute antiviral T-cell response characterization showed that viral transcription determines both the immunoprevalence and immunodominance of the human leukocyte antigen class I response to human respiratory syncytial virus. These findings have clear implications for antiviral vaccine design. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. The Viral Transcription Group Determines the HLA Class I Cellular Immune Response Against Human Respiratory Syncytial Virus*

    Science.gov (United States)

    Johnstone, Carolina; Lorente, Elena; Barriga, Alejandro; Barnea, Eilon; Infantes, Susana; Lemonnier, François A.; David, Chella S.; Admon, Arie; López, Daniel

    2015-01-01

    The cytotoxic T-lymphocyte-mediated killing of virus-infected cells requires previous recognition of short viral antigenic peptides bound to human leukocyte antigen class I molecules that are exposed on the surface of infected cells. The cytotoxic T-lymphocyte response is critical for the clearance of human respiratory syncytial virus infection. In this study, naturally processed viral human leukocyte antigen class I ligands were identified with mass spectrometry analysis of complex human leukocyte antigen-bound peptide pools isolated from large amounts of human respiratory syncytial virus-infected cells. Acute antiviral T-cell response characterization showed that viral transcription determines both the immunoprevalence and immunodominance of the human leukocyte antigen class I response to human respiratory syncytial virus. These findings have clear implications for antiviral vaccine design. PMID:25635267

  16. Characterization of Microvesicles Released from Human Red Blood Cells

    Directory of Open Access Journals (Sweden)

    Duc Bach Nguyen

    2016-03-01

    Full Text Available Background/Aims: Extracellular vesicles (EVs are spherical fragments of cell membrane released from various cell types under physiological as well as pathological conditions. Based on their size and origin, EVs are classified as exosome, microvesicles (MVs and apoptotic bodies. Recently, the release of MVs from human red blood cells (RBCs under different conditions has been reported. MVs are released by outward budding and fission of the plasma membrane. However, the outward budding process itself, the release of MVs and the physical properties of these MVs have not been well investigated. The aim of this study is to investigate the formation process, isolation and characterization of MVs released from RBCs under conditions of stimulating Ca2+ uptake and activation of protein kinase C. Methods: Experiments were performed based on single cell fluorescence imaging, fluorescence activated cell sorter/flow cytometer (FACS, scanning electron microscopy (SEM, atomic force microscopy (AFM and dynamic light scattering (DLS. The released MVs were collected by differential centrifugation and characterized in both their size and zeta potential. Results: Treatment of RBCs with 4-bromo-A23187 (positive control, lysophosphatidic acid (LPA, or phorbol-12 myristate-13 acetate (PMA in the presence of 2 mM extracellular Ca2+ led to an alteration of cell volume and cell morphology. In stimulated RBCs, exposure of phosphatidylserine (PS and formation of MVs were observed by using annexin V-FITC. The shedding of MVs was also observed in the case of PMA treatment in the absence of Ca2+, especially under the transmitted bright field illumination. By using SEM, AFM and DLS the morphology and size of stimulated RBCs, MVs were characterized. The sizes of the two populations of MVs were 205.8 ± 51.4 nm and 125.6 ± 31.4 nm, respectively. Adhesion of stimulated RBCs and MVs was observed. The zeta potential of MVs was determined in the range from - 40 mV to - 10 m

  17. The secondary biogenic radiation of gamma-irradiated human blood

    International Nuclear Information System (INIS)

    Kuzin, A.M.; Surkenova, G.N.; Budagovskij, A.V.; Gudi, G.A.

    1997-01-01

    The sample of blood freshly taken from healthy men were gamma-irradiated with a dose of 10 Gy. It was shown that after the treatment the blood gained the capacity to emit secondary biogenic radiation. Emission lasted for some hours, passed through quartz-glass curette and was revealed by stimulating influence on biological detector (sprouting seeds)

  18. Optoacoustic measurements of human placenta and umbilical blood oxygenation

    Science.gov (United States)

    Nanovskaya, T. N.; Petrov, I. Y.; Petrov, Y.; Patrikeeva, S. L.; Ahmed, M. S.; Hankins, G. D. V.; Prough, D. S.; Esenaliev, R. O.

    2016-03-01

    Adequate oxygenation is essential for normal embryogenesis and fetal growth. Perturbations in the intrauterine oxidative environment during pregnancy are associated with several pathophysiological disorders such as pregnancy loss, preeclampsia, and intrauterine growth restriction. We proposed to use optoacoustic technology for monitoring placental and fetal umbilical blood oxygenation. In this work, we studied optoacoustic monitoring of oxygenation in placenta and umbilical cord blood ex vivo using technique of placenta perfusion. We used a medical grade, nearinfrared, tunable, optoacoustic system developed and built for oxygenation monitoring in blood vessels and in tissues. First, we calibrated the system for cord blood oxygenation measurements by using a CO-Oximeter (gold standard). Then we performed validation in cord blood circulating through the catheters localized on the fetal side of an isolated placental lobule. Finally, the oxygenation measurements were performed in the perfused placental tissue. To increase or decrease blood oxygenation, we used infusion of a gas mixture of 95% O2 + 5% CO2 and 95% N2 + 5% CO2, respectively. In placental tissue, up to four cycles of changes in oxygenation were performed. The optoacoustically measured oxygenation in circulating cord blood and in placental lobule closely correlated with the actual oxygenation data measured by CO-Oximeter. We plan to further test the placental and cord blood oxygenation monitoring with optoacoustics in animal and clinical studies.

  19. Transcription of human 7S K DNA in vitro and in vivo is exclusively controlled by an upstream promoter

    Energy Technology Data Exchange (ETDEWEB)

    Kleinert, H.; Benecke, B.J.

    1988-02-25

    The authors have analyzed the transcription of a recently isolated human 7S K RNA gene in vitro and in vivo. In contrast to hitherto characterized class III genes (genes transcribed by RNA polymerase III), the coding sequence of this gene is not required for faithful and efficient transcription by RNA polymerase III. In fact, a procaryotic vector DNA sequence was efficiently transcribed by RNA polymerase III under the control of the 7S K RNA gene upstream sequence in vitro and in vivo. S/sub 1/-nuclease protection analyses confirmed that the 7S K 5'flanking sequence was sufficient for accurate transcription initiation. These data demonstrate that 7S K DNA represents a novel class III gene, the promoter elements of which are located outside the coding sequence.

  20. Human blood RNA stabilization in samples collected and transported for a large biobank

    Science.gov (United States)

    2012-01-01

    Background The Norwegian Mother and Child Cohort Study (MoBa) is a nation-wide population-based pregnancy cohort initiated in 1999, comprising more than 108.000 pregnancies recruited between 1999 and 2008. In this study we evaluated the feasibility of integrating RNA analyses into existing MoBa protocols. We compared two different blood RNA collection tube systems – the PAXgene™ Blood RNA system and the Tempus™ Blood RNA system - and assessed the effects of suboptimal blood volumes in collection tubes and of transportation of blood samples by standard mail. Endpoints to characterize the samples were RNA quality and yield, and the RNA transcript stability of selected genes. Findings High-quality RNA could be extracted from blood samples stabilized with both PAXgene and Tempus tubes. The RNA yields obtained from the blood samples collected in Tempus tubes were consistently higher than from PAXgene tubes. Higher RNA yields were obtained from cord blood (3 – 4 times) compared to adult blood with both types of tubes. Transportation of samples by standard mail had moderate effects on RNA quality and RNA transcript stability; the overall RNA quality of the transported samples was high. Some unexplained changes in gene expression were noted, which seemed to correlate with suboptimal blood volumes collected in the tubes. Temperature variations during transportation may also be of some importance. Conclusions Our results strongly suggest that special collection tubes are necessary for RNA stabilization and they should be used for establishing new biobanks. We also show that the 50,000 samples collected in the MoBa biobank provide RNA of high quality and in sufficient amounts to allow gene expression analyses for studying the association of disease with altered patterns of gene expression. PMID:22988904

  1. Human extrahepatic portal vein obstruction correlates with decreased factor VII and protein C transcription but increased hepatocyte proliferation.

    Science.gov (United States)

    Chiu, Bill; Melin-Aldana, Hector; Superina, Riccardo A

    2007-10-01

    A 3-year-old girl developed extrahepatic portal vein obstruction (EHPVO) after a liver transplant. She had sequelae of portal hypertension that required another transplantation. The circumstances allowed for comparison of liver-dependent coagulation factor production between the second donor liver and the explanted liver with EHPVO. Liver samples from the explanted first graft and the second transplant were obtained. Fresh tissue was used to perform reverse transcription-polymerase chain reaction with primers against factors V, VII, as well as VIII, protein C, and paraffin-embedded sections for hepatocyte proliferation using Ki-67 antibody as well as for apoptosis using TUNEL assay. The transcription of factor VII and that of protein C were decreased in the explant as compared with the newly transplanted liver (factor VII, 77% of the donor; protein C, 88% of the donor). The transcription of factor V and that of factor VIII were unchanged. The explant had a greater percentage of proliferating hepatocytes than the new organ (0.85% +/- 0.75% vs 0.11% +/- 0.21%). The percentage of apoptotic cells was similar between the 2 livers (0.09% +/- 0.13% vs 0.09% +/- 0.13%). Idiopathic EHPVO is associated with a reduction in liver-dependent coagulation factor transcription and an increase in hepatocyte proliferation. Portal blood flow deprivation alters hepatic homeostasis and initiates mechanisms that attempt to restore liver-dependent coagulation factors.

  2. Usefulness of Transcriptional Blood Biomarkers as a Non-invasive Surrogate Marker of Mucosal Healing and Endoscopic Response in Ulcerative Colitis.

    Science.gov (United States)

    Planell, Núria; Masamunt, M Carme; Leal, Raquel Franco; Rodríguez, Lorena; Esteller, Miriam; Lozano, Juan J; Ramírez, Anna; Ayrizono, Maria de Lourdes Setsuko; Coy, Claudio Saddy Rodrigues; Alfaro, Ignacio; Ordás, Ingrid; Visvanathan, Sudha; Ricart, Elena; Guardiola, Jordi; Panés, Julián; Salas, Azucena

    2017-10-27

    Ulcerative colitis [UC] is a chronic inflammatory disease of the colon. Colonoscopy remains the gold standard for evaluating disease activity, as clinical symptoms are not sufficiently accurate. The aim of this study is to identify new accurate non-invasive biomarkers based on whole-blood transcriptomics that can predict mucosal lesions and response to treatment in UC patients. Whole-blood samples were collected for a total of 152 UC patients at endoscopy. Blood RNA from 25 UC individuals and 20 controls was analysed using microarrays. Genes that correlated with endoscopic activity were validated using real-time polymerase chain reaction in an independent group of 111 UC patients, and a prediction model for mucosal lesions was evaluated. Responsiveness to treatment was assessed in a longitudinal cohort of 16 UC patients who started anti-tumour necrosis factor [TNF] therapy and were followed up for 14 weeks. Microarray analysis identified 122 genes significantly altered in the blood of endoscopically active UC patients. A significant correlation with the degree of endoscopic activity was observed in several genes, including HP, CD177, GPR84, and S100A12. Using HP as a predictor of endoscopic disease activity, an accuracy of 67.3% was observed, compared with 52.4%, 45.2%, and 30.3% for C-reactive protein, erythrocyte sedimentation rate, and platelet count, respectively. Finally, at 14 weeks of treatment, response to anti-TNF therapy induced alterations in blood HP, CD177, GPR84, and S100A12 transcripts that correlated with changes in endoscopic activity. Transcriptional changes in UC patients are sensitive to endoscopic improvement and appear to be an effective tool to monitor patients over time. © European Crohn’s and Colitis Organisation (ECCO) 2017.

  3. Transcription arrest by a G quadruplex forming-trinucleotide repeat sequence from the human c-myb gene.

    Science.gov (United States)

    Broxson, Christopher; Beckett, Joshua; Tornaletti, Silvia

    2011-05-17

    Non canonical DNA structures correspond to genomic regions particularly susceptible to genetic instability. The transcription process facilitates formation of these structures and plays a major role in generating the instability associated with these genomic sites. However, little is known about how non canonical structures are processed when encountered by an elongating RNA polymerase. Here we have studied the behavior of T7 RNA polymerase (T7RNAP) when encountering a G quadruplex forming-(GGA)(4) repeat located in the human c-myb proto-oncogene. To make direct correlations between formation of the structure and effects on transcription, we have taken advantage of the ability of the T7 polymerase to transcribe single-stranded substrates and of G4 DNA to form in single-stranded G-rich sequences in the presence of potassium ions. Under physiological KCl concentrations, we found that T7 RNAP transcription was arrested at two sites that mapped to the c-myb (GGA)(4) repeat sequence. The extent of arrest did not change with time, indicating that the c-myb repeat represented an absolute block and not a transient pause to T7 RNAP. Consistent with G4 DNA formation, arrest was not observed in the absence of KCl or in the presence of LiCl. Furthermore, mutations in the c-myb (GGA)(4) repeat, expected to prevent transition to G4, also eliminated the transcription block. We show T7 RNAP arrest at the c-myb repeat in double-stranded DNA under conditions mimicking the cellular concentration of biomolecules and potassium ions, suggesting that the G4 structure formed in the c-myb repeat may represent a transcription roadblock in vivo. Our results support a mechanism of transcription-coupled DNA repair initiated by arrest of transcription at G4 structures.

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

    Directory of Open Access Journals (Sweden)

    Unger Elizabeth R

    2011-09-01

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

  5. NMR water-proton spin-lattice relaxation time of human red blood cells and red blood cell suspensions

    International Nuclear Information System (INIS)

    Sullivan, S.G.; Rosenthal, J.S.; Winston, A.; Stern, A.

    1988-01-01

    NMR water-proton spin-lattice relaxation times were studied as probes of water structure in human red blood cells and red blood cell suspensions. Normal saline had a relaxation time of about 3000 ms while packed red blood cells had a relaxation time of about 500 ms. The relaxation time of a red blood cell suspension at 50% hematocrit was about 750 ms showing that surface charges and polar groups of the red cell membrane effectively structure extracellular water. Incubation of red cells in hypotonic saline increases relaxation time whereas hypertonic saline decreases relaxation time. Relaxation times varied independently of mean corpuscular volume and mean corpuscular hemoglobin concentration in a sample population. Studies with lysates and resealed membrane ghosts show that hemoglobin is very effective in lowering water-proton relaxation time whereas resealed membrane ghosts in the absence of hemoglobin are less effective than intact red cells. 9 refs.; 3 figs.; 1 table

  6. Cloning and functional analysis of human mTERFL encoding a novel mitochondrial transcription termination factor-like protein

    International Nuclear Information System (INIS)

    Chen Yao; Zhou Guangjin; Yu Min; He Yungang; Tang Wei; Lai Jianhua; He Jie; Liu Wanguo; Tan Deyong

    2005-01-01

    Serum plays an important role in the regulation of cell cycle and cell growth. To identify novel serum-inhibitory factors and study their roles in cell cycle regulation, we performed mRNA differential display analysis of U251 cells in the presence or absence of serum and cloned a novel gene encoding the human mitochondrial transcription termination factor-like protein (mTERFL). The full-length mTERFL cDNA has been isolated and the genomic structure determined. The mTERFL gene consists of three exons and encodes 385 amino acids with 52% sequence similarity to the human mitochondrial transcription termination factor (mTERF). However, mTERFL and mTERF have an opposite expression pattern in response to serum. The expression of mTERFL is dramatically inhibited by the addition of serum in serum-starved cells while the mTERF is rather induced. Northern blot analysis detected three mTERFL transcripts of 1.7, 3.2, and 3.5 kb. Besides the 3.2 kb transcript that is unique to skeletal muscle, other two transcripts express predominant in heart, liver, pancreas, and skeletal muscle. Expression of the GFP-mTERFL fusion protein in HeLa cells localized it to the mitochondria. Furthermore, ectopic expression of mTERFL suppresses cell growth and arrests cells in the G1 stage demonstrated by MTT and flow cytometry analysis. Collectively, our data suggest that mTERFL is a novel mTERF family member and a serum-inhibitory factor probably participating in the regulation of cell growth through the modulation of mitochondrial transcription

  7. Changes in Transcriptional Output of Human Peripheral Blood Mononuclear Cells Following Resistance Exercise

    Science.gov (United States)

    2011-01-01

    immunodeficiency virus (HIV), or had been diagnosed with acquired immune deficiency syndrome (AIDS). Participants were also excluded if they were... Laron Z (1995) Mathematical formulae for the prediction of the residual beta cell function during the first two years of disease in children and

  8. Transcriptional Profiling of Human Peripheral Blood Mononuclear Cells Exposed to Bacillus anthracis in vitro

    Science.gov (United States)

    2013-03-11

    sterile screw-capped tube containing 5 ml of sterile PBS. New sporulation medium (3 grams of Tryptone, 3 grams of yeast extract, 2 grams of agar, 23...United States. Emerg Infect Dis 7: 933-944. 3. Penn CC, Klotz SA (1997) Anthrax pneumonia. Semin Respir Infect 12: 28-30. 4. Sweeney DA, Hicks CW, Cui X...Li Y, Eichacker PQ (2011) Anthrax infection. Am J Respir Crit Care Med 184: 1333-1341. 5. Roy CJ, Reed DS, Hutt JA (2010) Aerobiology and inhalation

  9. Diesel exhaust particulate extracts inhibit transcription of nuclear respiratory factor-1 and cell viability in human umbilical vein endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Mattingly, Kathleen A.; Klinge, Carolyn M. [University of Louisville School of Medicine, Department of Biochemistry and Molecular Biology, Center for Genetics and Molecular Medicine, Louisville, KY (United States)

    2012-04-15

    Endothelial dysfunction precedes cardiovascular disease and is accompanied by mitochondrial dysfunction. Here we tested the hypothesis that diesel exhaust particulate extracts (DEPEs), prepared from a truck run at different speeds and engine loads, would inhibit genomic estrogen receptor activation of nuclear respiratory factor-1 (NRF-1) transcription in human umbilical vein endothelial cells (HUVECs). Additionally, we examined how DEPEs affect NRF-1-regulated TFAM expression and, in turn, Tfam-regulated mtDNA-encoded cytochrome c oxidase subunit I (COI, MTCO1) and NADH dehydrogenase subunit I (NDI) expression as well as cell proliferation and viability. We report that 17{beta}-estradiol (E{sub 2}), 4-hydroxytamoxifen (4-OHT), and raloxifene increased NRF-1 transcription in HUVECs in an ER-dependent manner. DEPEs inhibited NRF-1 transcription, and this suppression was not ablated by concomitant treatment with E{sub 2}, 4-OHT, or raloxifene, indicating that the effect was not due to inhibition of ER activity. While E{sub 2} increased HUVEC proliferation and viability, DEPEs inhibited viability but not proliferation. Resveratrol increased NRF-1 transcription in an ER-dependent manner in HUVECs, and ablated DEPE inhibition of basal NRF-1 expression. Given that NRF-1 is a key nuclear transcription factor regulating genes involved in mitochondrial activity and biogenesis, these data suggest that DEPEs may adversely affect mitochondrial function leading to endothelial dysfunction and resveratrol may block these effects. (orig.)

  10. Exosomes derived from human umbilical cord blood mesenchymal stem cells stimulates rejuvenation of human skin.

    Science.gov (United States)

    Kim, Yoon-Jin; Yoo, Sae Mi; Park, Hwan Hee; Lim, Hye Jin; Kim, Yu-Lee; Lee, Seunghee; Seo, Kwang-Won; Kang, Kyung-Sun

    2017-11-18

    Human umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) play an important role in cutaneous wound healing, and recent studies suggested that MSC-derived exosomes activate several signaling pathways, which are conducive in wound healing and cell growth. In this study, we investigated the roles of exosomes that are derived from USC-CM (USC-CM Exos) in cutaneous collagen synthesis and permeation. We found that USC-CM has various growth factors associated with skin rejuvenation. Our in vitro results showed that USC-CM Exos integrate in Human Dermal Fibroblasts (HDFs) and consequently promote cell migration and collagen synthesis of HDFs. Moreover, we evaluated skin permeation of USC-CM Exos by using human skin tissues. Results showed that Exo-Green labeled USC-CM Exos approached the outermost layer of the epidermis after 3 h and gradually approached the epidermis after 18 h. Moreover, increased expressions of Collagen I and Elastin were found after 3 days of treatment on human skin. The results showed that USC-CM Exos is absorbed into human skin, it promotes Collagen I and Elastin synthesis in the skin, which are essential to skin rejuvenation and shows the potential of USC-CM integration with the cosmetics or therapeutics. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Inner ocular blood flow responses to an acute decrease in blood pressure in resting humans

    International Nuclear Information System (INIS)

    Ikemura, Tsukasa; Kashima, Hideaki; Yamaguchi, Yuji; Miyaji, Akane; Hayashi, Naoyuki

    2015-01-01

    Whether inner ocular vessels have an autoregulatory response to acute fluctuations in blood pressure is unclear. We tried to examine the validity of acute hypotension elicited by thigh-cuff release as to assess the dynamic autoregulation in the ocular circulation. Blood flow velocity in the superior nasal and inferior temporal retinal arterioles, and in the retinal and choroidal vasculature were measured with the aid of laser speckle flowgraphy before and immediately after an acute decrease in blood pressure in 20 healthy subjects. Acute hypotension was induced by a rapid release of bilateral thigh occlusion cuffs that had been inflated to 220 mmHg for 2 min. The ratio of the relative change in retinal and choroidal blood flow velocity to the relative change in mean arterial blood pressure (MAP) was calculated. Immediately after cuff release, the MAP and blood flows in the all ocular target vessels decreased significantly from the baseline values obtained before thigh-cuff release. The ratio of the relative change in inner ocular blood flow velocity to that in the MAP exceeded 1% / %mmHg. An explicit dynamic autoregulation in inner ocular vessels cannot be demonstrated in response to an acute hypotension induced by the thigh-cuff release technique. (paper)

  12. Gene expression profiling analysis of CRTC1-MAML2 fusion oncogene-induced transcriptional program in human mucoepidermoid carcinoma cells

    International Nuclear Information System (INIS)

    Chen, Jie; Li, Jian-Liang; Chen, Zirong; Griffin, James D.; Wu, Lizi

    2015-01-01

    Mucoepidermoid carcinoma (MEC) arises from multiple organs and accounts for the most common types of salivary gland malignancies. Currently, patients with unresectable and metastatic MEC have poor long-term clinical outcomes and no targeted therapies are available. The majority of MEC tumors contain a t(11;19) chromosomal translocation that fuses two genes, CRTC1 and MAML2, to generate the chimeric protein CRTC1-MAML2. CRTC1-MAML2 displays transforming activity in vitro and is required for human MEC cell growth and survival, partially due to its ability to constitutively activate CREB-mediated transcription. Consequently, CRTC1-MAML2 is implicated as a major etiologic molecular event and a therapeutic target for MEC. However, the molecular mechanisms underlying CRTC1-MAML2 oncogenic action in MEC have not yet been systematically analyzed. Elucidation of the CRTC1-MAML2-regulated transcriptional program and its underlying mechanisms will provide important insights into MEC pathogenesis that are essential for the development of targeted therapeutics. Transcriptional profiling was performed on human MEC cells with the depletion of endogenous CRTC1-MAML2 fusion or its interacting partner CREB via shRNA-mediated gene knockdown. A subset of target genes was validated via real-time RT-PCR assays. CRTC1-MAML2-perturbed molecular pathways in MEC were identified through pathway analyses. Finally, comparative analysis of CRTC1-MAML2-regulated and CREB-regulated transcriptional profiles was carried out to assess the contribution of CREB in mediating CRTC1-MAML2-induced transcription. A total of 808 differentially expressed genes were identified in human MEC cells after CRTC1-MAML2 knockdown and a subset of known and novel fusion target genes was confirmed by real-time RT-PCR. Pathway Analysis revealed that CRTC1-MAML2-regulated genes were associated with network functions that are important for cell growth, proliferation, survival, migration, and metabolism. Comparison of CRTC

  13. Revised genomic structure of the human ghrelin gene and identification of novel exons, alternative splice variants and natural antisense transcripts

    Directory of Open Access Journals (Sweden)

    Herington Adrian C

    2007-08-01

    Full Text Available Abstract Background Ghrelin is a multifunctional peptide hormone expressed in a range of normal tissues and pathologies. It has been reported that the human ghrelin gene consists of five exons which span 5 kb of genomic DNA on chromosome 3 and includes a 20 bp non-coding first exon (20 bp exon 0. The availability of bioinformatic tools enabling comparative analysis and the finalisation of the human genome prompted us to re-examine the genomic structure of the ghrelin locus. Results We have demonstrated the presence of an additional novel exon (exon -1 and 5' extensions to exon 0 and 1 using comparative in silico analysis and have demonstrated their existence experimentally using RT-PCR and 5' RACE. A revised exon-intron structure demonstrates that the human ghrelin gene spans 7.2 kb and consists of six rather than five exons. Several ghrelin gene-derived splice forms were detected in a range of human tissues and cell lines. We have demonstrated ghrelin gene-derived mRNA transcripts that do not code for ghrelin, but instead may encode the C-terminal region of full-length preproghrelin (C-ghrelin, which contains the coding region for obestatin and a transcript encoding obestatin-only. Splice variants that differed in their 5' untranslated regions were also found, suggesting a role of these regions in the post-transcriptional regulation of preproghrelin translation. Finally, several natural antisense transcripts, termed ghrelinOS (ghrelin opposite strand transcripts, were demonstrated via orientation-specific RT-PCR, 5' RACE and in silico analysis of ESTs and cloned amplicons. Conclusion The sense and antisense alternative transcripts demonstrated in this study may function as non-coding regulatory RNA, or code for novel protein isoforms. This is the first demonstration of putative obestatin and C-ghrelin specific transcripts and these findings suggest that these ghrelin gene-derived peptides may also be produced independently of preproghrelin

  14. TAS1R3 and UCN2 Transcript Levels in Blood Cells Are Associated With Sugary and Fatty Food Consumption in Children.

    Science.gov (United States)

    Priego, T; Sánchez, J; Picó, C; Ahrens, W; De Henauw, S; Kourides, Y; Lissner, L; Molnár, D; Moreno, L A; Russo, P; Siani, A; Veidebaum, T; Palou, A

    2015-09-01

    New types of dietary exposure biomarkers are needed to implement effective strategies for obesity prevention in children. Of special interest are biomarkers of consumption of food rich in simple sugars and fat because their intake has been associated with obesity development. Peripheral blood cells (PBCs) represent a promising new tool for identifying novel, transcript-based biomarkers. This study aimed to study potential associations between the transcripts of taste receptor type 1 member 3 (TAS1R3) and urocortin II (UCN2) genes in PBCs and the frequency of sugary and fatty food consumption in children. Four hundred sixty-three children from the IDEFICS cohort were selected to include a similar number of boys and girls, both normal-weight and overweight, belonging to eight European countries. Anthropometric parameters (measured at baseline and in a subset of 193 children after 2 years), food consumption frequency and transcript levels of TAS1R3 and UCN2 genes in PBCs were measured. Children with low-frequency consumption of sugary foods displayed higher TAS1R3 expression levels with respect to those with intermediate or high frequency. In turn, children with high-frequency consumption of fatty foods showed lower UCN2 expression levels with respect to those with low or intermediate frequency. Moreover, transcripts of TAS1R3 were related with body mass index and fat-mass changes after a 2-year follow-up period, with low expression levels of this gene being related with increased fat accumulation over time. The transcripts of TAS1R3 and UCN2 in PBCs may be considered potential biomarkers of consumption of sugary and fatty food, respectively, to complement data of food-intake questionnaires.

  15. Lin- CD34hi CD117int/hi FcεRI+ cells in human blood constitute a rare population of mast cell progenitors.

    Science.gov (United States)

    Dahlin, Joakim S; Malinovschi, Andrei; Öhrvik, Helena; Sandelin, Martin; Janson, Christer; Alving, Kjell; Hallgren, Jenny

    2016-01-28

    Mast cells are rare tissue-resident immune cells that are involved in allergic reactions, and their numbers are increased in the lungs of asthmatics. Murine lung mast cells arise from committed bone marrow-derived progenitors that enter the blood circulation, migrate through the pulmonary endothelium, and mature in the tissue. In humans, mast cells can be cultured from multipotent CD34(+) progenitor cells. However, a population of distinct precursor cells that give rise to mast cells has remained undiscovered. To our knowledge, this is the first report of human lineage-negative (Lin(-)) CD34(hi) CD117(int/hi) FcεRI(+) progenitor cells, which represented only 0.0053% of the isolated blood cells in healthy individuals. These cells expressed integrin β7 and developed a mast cell-like phenotype, although with a slow cell division capacity in vitro. Isolated Lin(-) CD34(hi) CD117(int/hi) FcεRI(+) blood cells had an immature mast cell-like appearance and expressed high levels of many mast cell-related genes as compared with human blood basophils in whole-transcriptome microarray analyses. Furthermore, serglycin, tryptase, and carboxypeptidase A messenger RNA transcripts were detected by quantitative reverse transcription-polymerase chain reaction. Altogether, we propose that the Lin(-) CD34(hi) CD117(int/hi) FcεRI(+) blood cells are closely related to human tissue mast cells and likely constitute an immediate precursor population, which can give rise to predominantly mast cells. Furthermore, asthmatics with reduced lung function had a higher frequency of Lin(-) CD34(hi) CD117(int/hi) FcεRI(+) blood mast cell progenitors than asthmatics with normal lung function. © 2016 by The American Society of Hematology.

  16. Multilayered epithelium in a rat model and human Barrett's esophagus: Similar expression patterns of transcription factors and differentiation markers

    Directory of Open Access Journals (Sweden)

    Yang Chung S

    2008-01-01

    Full Text Available Abstract Background In rats, esophagogastroduodenal anastomosis (EGDA without concomitant chemical carcinogen treatment leads to gastroesophageal reflux disease, multilayered epithelium (MLE, a presumed precursor in intestinal metaplasia, columnar-lined esophagus, dysplasia, and esophageal adenocarcinoma. Previously we have shown that columnar-lined esophagus in EGDA rats resembled human Barrett's esophagus (BE in its morphology, mucin features and expression of differentiation markers (Lab. Invest. 2004;84:753–765. The purpose of this study was to compare the phenotype of rat MLE with human MLE, in order to gain insight into the nature of MLE and its potential role in the development of BE. Methods Serial sectioning was performed on tissue samples from 32 EGDA rats and 13 patients with established BE. Tissue sections were immunohistochemically stained for a variety of transcription factors and differentiation markers of esophageal squamous epithelium and intestinal columnar epithelium. Results We detected MLE in 56.3% (18/32 of EGDA rats, and in all human samples. As expected, both rat and human squamous epithelium, but not intestinal metaplasia, expressed squamous transcription factors and differentiation markers (p63, Sox2, CK14 and CK4 in all cases. Both rat and human intestinal metaplasia, but not squamous epithelium, expressed intestinal transcription factors and differentiation markers (Cdx2, GATA4, HNF1α, villin and Muc2 in all cases. Rat MLE shared expression patterns of Sox2, CK4, Cdx2, GATA4, villin and Muc2 with human MLE. However, p63 and CK14 were expressed in a higher proportion of rat MLE compared to humans. Conclusion These data indicate that rat MLE shares similar properties to human MLE in its expression pattern of these markers, not withstanding small differences, and support the concept that MLE may be a transitional stage in the metaplastic conversion of squamous to columnar epithelium in BE.

  17. Wetting and spreading of human blood: Recent advances and applications

    OpenAIRE

    Smith , F.R. ,; Brutin , David

    2018-01-01

    International audience; Investigation of the physical phenomena involved in blood interactions with real surfaces present new exciting challenges. The fluid mechanical properties of such a fluid is singular due its non-Newtonian and complex behaviour, depending on the surrounding ambient conditions and the donor/victim's blood biological properties. The fundamental research on the topic remains fairly recent; although it finds applications in fields such as forensic science, with bloodstain p...

  18. The uptake kinetics and immunotoxic effects of microcystin-LR in human and chicken peripheral blood lymphocytes in vitro

    International Nuclear Information System (INIS)

    Lankoff, Anna; Carmichael, Wayne W.; Grasman, Keith A.; Yuan, Moucun

    2004-01-01

    Microcystin-LR is a cyanobacterial heptapeptide that presents acute and chronic hazards to animal and human health. We investigated the influence of this toxin on human and chicken immune system modulation in vitro. Peripheral blood lymphocytes were treated with microcystin-LR at environmentally relevant doses of 1, 10 and 25 μg/ml for 12, 24, 48, 72 h (for proliferation assay cells were treated for 72 h). T-cell and B-cell proliferation as well as apoptosis and necrosis were determined in human and chicken samples. IL-2 and IL-6 production by human lymphocytes also was measured. In addition, uptake kinetics of microcystin-LR into human and chicken peripheral blood lymphocytes were calculated by Liquid Chromatography (LS) /Mass Spectrometry (MS) analysis. At the highest dose microcystin-LR decreased T-cell proliferation and all doses of microcystin-LR inhibited B-cell proliferation. The frequency of apoptotic and necrotic cells increased in a dose and time-dependent manner. Human lymphocytes responded to stimulation with microcystin-LR by increased production of IL-6 and decreased production of IL-2. Human lymphocytes were able to uptake from 0.014 to 1.663 μg/ml and chicken lymphocytes from 0.035 to 1.733 μg/ml of the microcystin-LR added to the cultures, depending on the treatment time and dose. In conclusion, microcystin-LR acted as an immunomodulator in cytokine production and down-regulated lymphocyte functions by induction of apoptosis and necrosis. However, further studies dealing with the influence of microcystin-LR on expression cytokine genes and transcription factors are necessary to confirm these hypotheses

  19. Alternative splicing and extensive RNA editing of human TPH2 transcripts.

    Directory of Open Access Journals (Sweden)

    Maik Grohmann

    Full Text Available Brain serotonin (5-HT neurotransmission plays a key role in the regulation of mood and has been implicated in a variety of neuropsychiatric conditions. Tryptophan hydroxylase (TPH is the rate-limiting enzyme in the biosynthesis of 5-HT. Recently, we discovered a second TPH isoform (TPH2 in vertebrates, including man, which is predominantly expressed in brain, while the previously known TPH isoform (TPH1 is primarly a non-neuronal enzyme. Overwhelming evidence now points to TPH2 as a candidate gene for 5-HT-related psychiatric disorders. To assess the role of TPH2 gene variability in the etiology of psychiatric diseases we performed cDNA sequence analysis of TPH2 transcripts from human post mortem amygdala samples obtained from individuals with psychiatric disorders (drug abuse, schizophrenia, suicide and controls. Here we show that TPH2 exists in two alternatively spliced variants in the coding region, denoted TPH2a and TPH2b. Moreover, we found evidence that the pre-mRNAs of both splice variants are dynamically RNA-edited in a mutually exclusive manner. Kinetic studies with cell lines expressing recombinant TPH2 variants revealed a higher activity of the novel TPH2B protein compared with the previously known TPH2A, whereas RNA editing was shown to inhibit the enzymatic activity of both TPH2 splice variants. Therefore, our results strongly suggest a complex fine-tuning of central nervous system 5-HT biosynthesis by TPH2 alternative splicing and RNA editing. Finally, we present molecular and large-scale linkage data evidencing that deregulated alternative splicing and RNA editing is involved in the etiology of psychiatric diseases, such as suicidal behaviour.

  20. Preanalytical Biases in the Measurement of Human Blood Sphingolipids

    Directory of Open Access Journals (Sweden)

    Robert Brunkhorst

    2018-05-01

    Full Text Available Dysregulation of blood sphingolipids is an emerging topic in clinical science. The objective of this study was to determine preanalytical biases that typically occur in clinical and translational studies and that influence measured blood sphingolipid levels. Therefore, we collected blood samples from four healthy male volunteers to investigate the effect of storage conditions (time, temperature, long-term storage, freeze–thaw cycles, blood drawing (venous or arterial sampling, prolonged venous compression, and sample preparation (centrifugation, freezing on sphingolipid levels measured by LC-MS/MS. Our data show that sphingosine 1-phosphate (S1P and sphinganine 1-phosphate (SA1P were upregulated in whole blood samples in a time- and temperature-dependent manner. Increased centrifugation at higher speeds led to lower amounts of S1P and SA1P. All other preanalytical biases did not significantly alter the amounts of S1P and SA1P. Further, in almost all settings, we did not detect differences in (dihydroceramide levels. In summary, besides time-, temperature-, and centrifugation-dependent changes in S1P and SA1P levels, sphingolipids in blood remained stable under practically relevant preanalytical conditions.

  1. A RAPID Method for Blood Processing to Increase the Yield of Plasma Peptide Levels in Human Blood.

    Science.gov (United States)

    Teuffel, Pauline; Goebel-Stengel, Miriam; Hofmann, Tobias; Prinz, Philip; Scharner, Sophie; Körner, Jan L; Grötzinger, Carsten; Rose, Matthias; Klapp, Burghard F; Stengel, Andreas

    2016-04-28

    Research in the field of food intake regulation is gaining importance. This often includes the measurement of peptides regulating food intake. For the correct determination of a peptide's concentration, it should be stable during blood processing. However, this is not the case for several peptides which are quickly degraded by endogenous peptidases. Recently, we developed a blood processing method employing Reduced temperatures, Acidification, Protease inhibition, Isotopic exogenous controls and Dilution (RAPID) for the use in rats. Here, we have established this technique for the use in humans and investigated recovery, molecular form and circulating concentration of food intake regulatory hormones. The RAPID method significantly improved the recovery for (125)I-labeled somatostatin-28 (+39%), glucagon-like peptide-1 (+35%), acyl ghrelin and glucagon (+32%), insulin and kisspeptin (+29%), nesfatin-1 (+28%), leptin (+21%) and peptide YY3-36 (+19%) compared to standard processing (EDTA blood on ice, p processing, while after standard processing 62% of acyl ghrelin were degraded resulting in an earlier peak likely representing desacyl ghrelin. After RAPID processing the acyl/desacyl ghrelin ratio in blood of normal weight subjects was 1:3 compared to 1:23 following standard processing (p = 0.03). Also endogenous kisspeptin levels were higher after RAPID compared to standard processing (+99%, p = 0.02). The RAPID blood processing method can be used in humans, yields higher peptide levels and allows for assessment of the correct molecular form.

  2. Immunotoxicity assessment of rice-derived recombinant human serum albumin using human peripheral blood mononuclear cells.

    Directory of Open Access Journals (Sweden)

    Kai Fu

    Full Text Available Human serum albumin (HSA is extensively used in clinics to treat a variety of diseases, such as hypoproteinemia, hemorrhagic shock, serious burn injuries, cirrhotic ascites and fetal erythroblastosis. To address supply shortages and high safety risks from limited human donors, we recently developed recombinant technology to produce HSA from rice endosperm. To assess the risk potential of HSA derived from Oryza sativa (OsrHSA before a First-in-human (FIH trial, we compared OsrHSA and plasma-derived HSA (pHSA, evaluating the potential for an immune reaction and toxicity using human peripheral blood mononuclear cells (PBMCs. The results indicated that neither OsrHSA nor pHSA stimulated T cell proliferation at 1x and 5x dosages. We also found no significant differences in the profiles of the CD4(+ and CD8(+ T cell subsets between OsrHSA- and pHSA-treated cells. Furthermore, the results showed that there were no significant differences between OsrHSA and pHSA in the production of cytokines such as interferon-gamma (IFN-γ, tumor necrosis factor-alpha (TNF-α, interleukin (IL-10 and IL-4. Our results demonstrated that OsrHSA has equivalent immunotoxicity to pHSA when using the PBMC model. Moreover, this ex vivo system could provide an alternative approach to predict potential risks in novel biopharmaceutical development.

  3. Clone-specific expression, transcriptional regulation, and action of interleukin-6 in human colon carcinoma cells

    International Nuclear Information System (INIS)

    Brozek, Wolfgang; Bises, Giovanna; Fabjani, Gerhild; Cross, Heide S; Peterlik, Meinrad

    2008-01-01

    Many cancer cells produce interleukin-6 (IL-6), a cytokine that plays a role in growth stimulation, metastasis, and angiogenesis of secondary tumours in a variety of malignancies, including colorectal cancer. Effectiveness of IL-6 in this respect may depend on the quantity of basal and inducible IL-6 expressed as the tumour progresses through stages of malignancy. We therefore have evaluated the effect of IL-6 modulators, i.e. IL-1β, prostaglandin E 2 , 17β-estradiol, and 1,25-dihydroxyvitamin D 3 , on expression and synthesis of the cytokine at different stages of tumour progression. We utilized cultures of the human colon carcinoma cell clones Caco-2/AQ, COGA-1A and COGA-13, all of which expressed differentiation and proliferation markers typical of distinct stages of tumour progression. IL-6 mRNA and protein levels were assayed by RT-PCR and ELISA, respectively. DNA sequencing was utilized to detect polymorphisms in the IL-6 gene promoter. IL-6 mRNA and protein concentrations were low in well and moderately differentiated Caco-2/AQ and COGA-1A cells, but were high in poorly differentiated COGA-13 cells. Addition of IL-1β (5 ng/ml) to a COGA-13 culture raised IL-6 production approximately thousandfold via a prostaglandin-independent mechanism. Addition of 17β-estradiol (10 -7 M) reduced basal IL-6 production by one-third, but IL-1β-inducible IL-6 was unaffected. Search for polymorphisms in the IL-6 promoter revealed the presence of a single haplotype, i.e., -597A/-572G/-174C, in COGA-13 cells, which is associated with a high degree of transcriptional activity of the IL-6 gene. IL-6 blocked differentiation only in Caco-2/AQ cells and stimulated mitosis through up-regulation of c-myc proto-oncogene expression. These effects were inhibited by 10 -8 M 1,25-dihydroxyvitamin D 3 . In human colon carcinoma cells derived from well and moderately differentiated tumours, IL-6 expression is low and only marginally affected, if at all, by PGE 2 , 1,25-dihydroxyvitamin D

  4. Induction of human adiponectin gene transcription by telmisartan, angiotensin receptor blocker, independently on PPAR-γ activation

    International Nuclear Information System (INIS)

    Moriuchi, Akie; Yamasaki, Hironori; Shimamura, Mika; Kita, Atsushi; Kuwahara, Hironaga; Fujishima, Keiichiro; Satoh, Tsuyoshi; Fukushima, Keiko; Fukushima, Tetsuya; Hayakawa, Takao; Mizuguchi, Hiroyuki; Nagayama, Yuji; Abiru, Norio; Kawasaki, Eiji; Eguchi, Katsumi

    2007-01-01

    Adiponectin, an adipose tissue-specific plasma protein, has been shown to ameliorate insulin resistance and inhibit the process of atherosclerosis. Recently, several reports have stated that angiotensin type 1 receptor blockers (ARBs), increase adiponectin plasma level, and ameliorate insulin resistance. Telmisartan, a subclass of ARBs, has been shown to be a partial agonist of the peroxisome proliferator-activated receptor (PPAR)-γ, and to increase the plasma adiponectin level. However, the transcriptional regulation of the human adiponectin gene by telmisartan has not been determined yet. To elucidate the effect of telmisartan on adiponectin, the stimulatory regulation of human adiponectin gene by telmisartan was investigated in 3T3-L1 adipocytes, utilizing adenovirus-mediated luciferase reporter gene-transferring technique. This study indicates that telmisartan may stimulate adiponectin transcription independent of PPAR-γ

  5. Transcriptome-wide mega-analyses reveal joint dysregulation of immunologic genes and transcription regulators in brain and blood in schizophrenia.

    Science.gov (United States)

    Hess, Jonathan L; Tylee, Daniel S; Barve, Rahul; de Jong, Simone; Ophoff, Roel A; Kumarasinghe, Nishantha; Tooney, Paul; Schall, Ulrich; Gardiner, Erin; Beveridge, Natalie Jane; Scott, Rodney J; Yasawardene, Surangi; Perera, Antionette; Mendis, Jayan; Carr, Vaughan; Kelly, Brian; Cairns, Murray; Tsuang, Ming T; Glatt, Stephen J

    2016-10-01

    The application of microarray technology in schizophrenia research was heralded as paradigm-shifting, as it allowed for high-throughput assessment of cell and tissue function. This technology was widely adopted, initially in studies of postmortem brain tissue, and later in studies of peripheral blood. The collective body of schizophrenia microarray literature contains apparent inconsistencies between studies, with failures to replicate top hits, in part due to small sample sizes, cohort-specific effects, differences in array types, and other confounders. In an attempt to summarize existing studies of schizophrenia cases and non-related comparison subjects, we performed two mega-analyses of a combined set of microarray data from postmortem prefrontal cortices (n=315) and from ex-vivo blood tissues (n=578). We adjusted regression models per gene to remove non-significant covariates, providing best-estimates of transcripts dysregulated in schizophrenia. We also examined dysregulation of functionally related gene sets and gene co-expression modules, and assessed enrichment of cell types and genetic risk factors. The identities of the most significantly dysregulated genes were largely distinct for each tissue, but the findings indicated common emergent biological functions (e.g. immunity) and regulatory factors (e.g., predicted targets of transcription factors and miRNA species across tissues). Our network-based analyses converged upon similar patterns of heightened innate immune gene expression in both brain and blood in schizophrenia. We also constructed generalizable machine-learning classifiers using the blood-based microarray data. Our study provides an informative atlas for future pathophysiologic and biomarker studies of schizophrenia. Published by Elsevier B.V.

  6. A broad but restricted requirement for TAF-5 (human TAFII100) for embryonic transcription in Caenorhabditis elegans.

    Science.gov (United States)

    Walker, Amy K; Blackwell, T Keith

    2003-02-21

    As conserved components of the transcription factor (TF) IID- and TFTC/SAGA-related complexes, TATA-binding protein-associated factors (TAF(II)s) are important for eukaryotic mRNA transcription. In yeast, genetic analyses suggest that, although some individual TAF(II)s are required for transcription of most genes, others have highly specialized functions. Much less is known about the functions of TAF(II)s in metazoans, which have more complex genomes that include many tissue-specific genes. TAF-5 (human (h) TAF(II)100) is of particular interest because it is predicted to have an important structural role. Here we describe the first genetics-based analysis of TAF-5 in a metazoan. By performing RNA interference in Caenorhabditis elegans embryos, which can survive for several cell generations without transcription, we found that taf-5 is important for a significant fraction of transcription. However, TAF-5 is apparently not essential for the expression of multiple developmental and other metazoan-specific genes. This phenotype remarkably resembles the previously described effects of similarly depleting two C. elegans histone fold TAF(II)s, TAF-9 (hTAF(II)31/32) and TAF-10 (hTAF(II)30), but is distinct from the widespread transcription block caused by TAF-4 (hTAF(II)130) depletion. Our findings suggest that TAF-5, TAF-9, and TAF-10 are part of a functional module of TFIID- and TFTC/SAGA-related complexes that can be bypassed in many metazoan-specific genes.

  7. Interaction between C/EBPβ and Tax down-regulates human T-cell leukemia virus type I transcription

    International Nuclear Information System (INIS)

    Hivin, P.; Gaudray, G.; Devaux, C.; Mesnard, J.-M.

    2004-01-01

    The human T-cell leukemia virus type I (HTLV-I) Tax protein trans-activates viral transcription through three imperfect tandem repeats of a 21-bp sequence called Tax-responsive element (TxRE). Tax regulates transcription via direct interaction with some members of the activating transcription factor/CRE-binding protein (ATF/CREB) family including CREM, CREB, and CREB-2. By interacting with their ZIP domain, Tax stimulates the binding of these cellular factors to the CRE-like sequence present in the TxREs. Recent observations have shown that CCAAT/enhancer binding protein β (C/EBPβ) forms stable complexes on the CRE site in the presence of CREB-2. Given that C/EBPβ has also been found to interact with Tax, we analyzed the effects of C/EBPβ on viral Tax-dependent transcription. We show here that C/EBPβ represses viral transcription and that Tax is no more able to form a stable complex with CREB-2 on the TxRE site in the presence of C/EBPβ. We also analyzed the physical interactions between Tax and C/EBPβ and found that the central region of C/EBPβ, excluding its ZIP domain, is required for direct interaction with Tax. It is the first time that Tax is described to interact with a basic leucine-zipper (bZIP) factor without recognizing its ZIP domain. Although unexpected, this result explains why C/EBPβ would be unable to form a stable complex with Tax on the TxRE site and could then down-regulate viral transcription. Lastly, we found that C/EBPβ was able to inhibit Tax expression in vivo from an infectious HTLV-I molecular clone. In conclusion, we propose that during cell activation events, which stimulate the Tax synthesis, C/EBPβ may down-regulate the level of HTLV-I expression to escape the cytotoxic-T-lymphocyte response

  8. Sp1 is a transcription repressor to stanniocalcin-1 expression in TSA-treated human colon cancer cells, HT29.

    Science.gov (United States)

    Law, Alice Y S; Yeung, B H Y; Ching, L Y; Wong, Chris K C

    2011-08-01

    Our previous study demonstrated that, stanniocalcin-1 (STC1) was a target of histone deacetylase (HDAC) inhibitors and was involved in trichostatin A (TSA) induced apoptosis in the human colon cancer cells, HT29. In this study, we reported that the transcriptional factor, specificity protein 1 (Sp1) in association with retinoblastoma (Rb) repressed STC1 gene transcription in TSA-treated HT29 cells. Our data demonstrated that, a co-treatment of the cells with TSA and Sp1 inhibitor, mithramycin A (MTM) led to a marked synergistic induction of STC1 transcript levels, STC1 promoter (1 kb)-driven luciferase activity and an increase of apoptotic cell population. The knockdown of Sp1 gene expression in TSA treated cells, revealed the repressor role of Sp1 in STC1 transcription. Using a protein phosphatase inhibitor okadaic acid (OKA), an increase of Sp1 hyperphosphorylation and so a reduction of its transcriptional activity, led to a significant induction of STC1 gene expression. Chromatin immunoprecipitation (ChIP) assay revealed that Sp1 binding on STC1 proximal promoter in TSA treated cells. The binding of Sp1 to STC1 promoter was abolished by the co-treatment of MTM or OKA in TSA-treated cells. Re-ChIP assay illustrated that Sp1-mediated inhibition of STC1 transcription was associated with the recruitment of another repressor molecule, Rb. Collectively our findings identify STC1 is a downstream target of Sp1. Copyright © 2011 Wiley-Liss, Inc.

  9. The Peripheral Whole Blood Transcriptome of Acute Pyelonephritis in Human Pregnancy

    Science.gov (United States)

    Madan, Ichchha; Than, Nandor Gabor; Romero, Roberto; Chaemsaithong, Piya; Miranda, Jezid; Tarca, Adi L.; Bhatti, Gaurav; Draghici, Sorin; Yeo, Lami; Mazor, Moshe; Hassan, Sonia S.; Chaiworapongsa, Tinnakorn

    2018-01-01

    Objective Human pregnancy is characterized by activation of the innate immune response and suppression of adaptive immunity. The former is thought to provide protection against infection to the mother, and the latter, tolerance against paternal antigens expressed in fetal cells. Acute pyelonephritis is associated with an increased risk of acute respiratory distress syndrome and sepsis in pregnant (vs. nonpregnant) women. The objective of this study was to describe the gene expression profile (transcriptome) of maternal whole blood in acute pyelonephritis. Method A case-control study was conducted to include pregnant women with acute pyelonephritis (n=15) and women with a normal pregnancy (n=34). Affymetrix HG-U133 Plus 2.0 arrays (Affymetrix, Santa Clara, CA, USA) were used for gene expression profiling. A linear model was used to test the association between the presence of pyelonephritis and gene expression levels while controlling for white blood cell count and gestational age. A fold change of 1.5 was considered significant at a false discovery rate of 0.1. A subset of differentially expressed genes (n=56) was tested with real-time quantitative reverse transcription-polymerase chain reaction (qRT-PCR) (cases, n=19; controls, n=59). Gene ontology and pathway analysis were applied. Results A total of 983 genes were differentially expressed in acute pyelonephritis: 457 were up-regulated and 526 were down-regulated. Significant enrichment of 300 biological processes and 63 molecular functions was found in pyelonephritis. Significantly impacted pathways in pyelonephritis included a) cytokine-cytokine receptor interaction; b) T-cell receptor signaling; c) Jak-STAT signaling; and d) complement and coagulation cascades. Of 56 genes tested by qRT-PCR, 48 (85.7%) had confirmation of differential expression. Conclusion This is the first study of the transcriptomic signature of whole blood in pregnant women with acute pyelonephritis. Acute infection during pregnancy is

  10. Sex differences of human cortical blood flow and energy metabolism.

    Science.gov (United States)

    Aanerud, Joel; Borghammer, Per; Rodell, Anders; Jónsdottir, Kristjana Y; Gjedde, Albert

    2017-07-01

    Brain energy metabolism is held to reflect energy demanding processes in neuropil related to the density and activity of synapses. There is recent evidence that men have higher density of synapses in temporal cortex than women. One consequence of these differences would be different rates of cortical energy turnover and blood flow in men and women. To test the hypotheses that rates of oxygen consumption (CMRO 2 ) and cerebral blood flow are higher in men than in women in regions of cerebral cortex, and that the differences persist with aging, we used positron emission tomography to determine cerebral blood flow and cerebral metabolic rate of oxygen as functions of age in healthy volunteers of both sexes. Cerebral metabolic rate of oxygen did not change with age for either sex and there were no differences of mean values of cerebral metabolic rate of oxygen between men and women in cerebral cortex. Women had significant decreases of cerebral blood flow as function of age in frontal and parietal lobes. Young women had significantly higher cerebral blood flow than men in frontal and temporal lobes, but these differences had disappeared at age 65. The absent sex difference of cerebral energy turnover suggests that the known differences of synaptic density between the sexes are counteracted by opposite differences of individual synaptic activity.

  11. Transcriptional program of ciliated epithelial cells reveals new cilium and centrosome components and links to human disease.

    Directory of Open Access Journals (Sweden)

    Ramona A Hoh

    Full Text Available Defects in the centrosome and cilium are associated with a set of human diseases having diverse phenotypes. To further characterize the components that define the function of these organelles we determined the transcriptional profile of multiciliated tracheal epithelial cells. Cultures of mouse tracheal epithelial cells undergoing differentiation in vitro were derived from mice expressing GFP from the ciliated-cell specific FOXJ1 promoter (FOXJ1:GFP. The transcriptional profile of ciliating GFP+ cells from these cultures was defined at an early and a late time point during differentiation and was refined by subtraction of the profile of the non-ciliated GFP- cells. We identified 649 genes upregulated early, when most cells were forming basal bodies, and 73 genes genes upregulated late, when most cells were fully ciliated. Most, but not all, of known centrosome proteins are transcriptionally upregulated early, particularly Plk4, a master regulator of centriole formation. We found that three genes associated with human disease states, Mdm1, Mlf1, and Dyx1c1, are upregulated during ciliogenesis and localize to centrioles and cilia. This transcriptome for mammalian multiciliated epithelial cells identifies new candidate centrosome and cilia proteins, highlights similarities between components of motile and primary cilia, and identifies new links between cilia proteins and human disease.

  12. Transcription-factor occupancy at HOT regions quantitatively predicts RNA polymerase recruitment in five human cell lines.

    KAUST Repository

    Foley, Joseph W

    2013-10-20

    BACKGROUND: High-occupancy target (HOT) regions are compact genome loci occupied by many different transcription factors (TFs). HOT regions were initially defined in invertebrate model organisms, and we here show that they are a ubiquitous feature of the human gene-regulation landscape. RESULTS: We identified HOT regions by a comprehensive analysis of ChIP-seq data from 96 DNA-associated proteins in 5 human cell lines. Most HOT regions co-localize with RNA polymerase II binding sites, but many are not near the promoters of annotated genes. At HOT promoters, TF occupancy is strongly predictive of transcription preinitiation complex recruitment and moderately predictive of initiating Pol II recruitment, but only weakly predictive of elongating Pol II and RNA transcript abundance. TF occupancy varies quantitatively within human HOT regions; we used this variation to discover novel associations between TFs. The sequence motif associated with any given TF\\'s direct DNA binding is somewhat predictive of its empirical occupancy, but a great deal of occupancy occurs at sites without the TF\\'s motif, implying indirect recruitment by another TF whose motif is present. CONCLUSIONS: Mammalian HOT regions are regulatory hubs that integrate the signals from diverse regulatory pathways to quantitatively tune the promoter for RNA polymerase II recruitment.

  13. Transcription-factor occupancy at HOT regions quantitatively predicts RNA polymerase recruitment in five human cell lines.

    KAUST Repository

    Foley, Joseph W; Sidow, Arend

    2013-01-01

    BACKGROUND: High-occupancy target (HOT) regions are compact genome loci occupied by many different transcription factors (TFs). HOT regions were initially defined in invertebrate model organisms, and we here show that they are a ubiquitous feature of the human gene-regulation landscape. RESULTS: We identified HOT regions by a comprehensive analysis of ChIP-seq data from 96 DNA-associated proteins in 5 human cell lines. Most HOT regions co-localize with RNA polymerase II binding sites, but many are not near the promoters of annotated genes. At HOT promoters, TF occupancy is strongly predictive of transcription preinitiation complex recruitment and moderately predictive of initiating Pol II recruitment, but only weakly predictive of elongating Pol II and RNA transcript abundance. TF occupancy varies quantitatively within human HOT regions; we used this variation to discover novel associations between TFs. The sequence motif associated with any given TF's direct DNA binding is somewhat predictive of its empirical occupancy, but a great deal of occupancy occurs at sites without the TF's motif, implying indirect recruitment by another TF whose motif is present. CONCLUSIONS: Mammalian HOT regions are regulatory hubs that integrate the signals from diverse regulatory pathways to quantitatively tune the promoter for RNA polymerase II recruitment.

  14. Chorionic gonadotropin regulates the transcript level of VHL, p53, and HIF-2alpha in human granulosa lutein cells.

    Science.gov (United States)

    Herr, D; Keck, C; Tempfer, C; Pietrowski, Detlef

    2004-12-01

    The ovarian corpus luteum plays a critical role in reproduction being the primary source of circulating progesterone. After ovulation the corpus luteum is build by avascular granulosa lutein cells through rapid vascularization regulated by gonadotropic hormones. The present study was performed to investigate whether this process might be influenced by the human chorionic gonadotropin (hCG)-dependent expression of different tumor suppressor genes and hypoxia dependent transcription factors. RNA was isolated from cultured granulosa lutein cells, transcribed into cDNA, and the transcript level of following genes were determined: RB-1, VHL, NF-1, NF-2, Wt-1, p53, APC, and hypoxia inducible factor-1 (HIF-1), -2, and -3alpha. Additionally, the influence of hCG on the expression of VHL, p53, and HIf2alpha were investigated. We demonstrate that in human granulosa lutein cells the tumor suppressor genes RB-1, VHL, NF-1, NF-2, Wt-1, p53, and APC and the hypoxia dependent transcription factors HIF-1alpha, -2alpha, and -3alpha are expressed. In addition, we showed that hCG regulates the expression of p53, VHL, and HIF-2alpha. Our results indicate that hCG may determine the growth and development of the corpus luteum by mediating hypoxic and apoptotic pathways in human granulosa lutein cells. Copyright 2004 Wiley-Liss, Inc.

  15. Cloning and characterization of the mouse Mcoln1 gene reveals an alternatively spliced transcript not seen in humans

    Directory of Open Access Journals (Sweden)

    Stahl Stefanie

    2002-02-01

    Full Text Available Abstract Background Mucolipidosis type IV (MLIV is an autosomal recessive lysosomal storage disorder characterized by severe neurologic and ophthalmologic abnormalities. Recently the MLIV gene, MCOLN1, has been identified as a new member of the transient receptor potential (TRP cation channel superfamily. Here we report the cloning and characterization of the mouse homologue, Mcoln1, and report a novel splice variant that is not seen in humans. Results The human and mouse genes display a high degree of synteny. Mcoln1 shows 91% amino acid and 86% nucleotide identity to MCOLN1. Also, Mcoln1 maps to chromosome 8 and contains an open reading frame of 580 amino acids, with a transcript length of approximately 2 kb encoded by 14 exons, similar to its human counterpart. The transcript that results from murine specific alternative splicing encodes a 611 amino acid protein that differs at the c-terminus. Conclusions Mcoln1 is highly similar to MCOLN1, especially in the transmembrane domains and ion pore region. Also, the late endosomal/lysosomal targeting signal is conserved, supporting the hypothesis that the protein is localized to these vesicle membranes. To date, there are very few reports describing species-specific splice variants. While identification of Mcoln1 is crucial to the development of mouse models for MLIV, the fact that there are two transcripts in mice suggests an additional or alternate function of the gene that may complicate phenotypic assessment.

  16. In Vitro Anticancer Activity of Phlorofucofuroeckol A via Upregulation of Activating Transcription Factor 3 against Human Colorectal Cancer Cells

    Directory of Open Access Journals (Sweden)

    Hyun Ji Eo

    2016-03-01

    Full Text Available Phlorofucofuroeckol A (PFF-A, one of the phlorotannins found in brown algae, has been reported to exert anti-cancer property. However, the molecular mechanism for the anti-cancer effect of PFF-A has not been known. Activating transcription factor 3 (ATF3 has been reported to be associated with apoptosis in colorectal cancer. The present study was performed to investigate the molecular mechanism by which PFF-A stimulates ATF3 expression and apoptosis in human colorectal cancer cells. PFF-A decreased cell viability through apoptosis of human colorectal cancer cells. PFF-A increased ATF3 expression through regulating transcriptional activity. The responsible cis-element for ATF3 transcriptional activation by PFF-A was cAMP response element binding protein (CREB, located between positions −147 and −85 of the ATF3 promoter. Inhibition of p38, c-Jun N-terminal kinases (JNK, glycogen synthase kinase (GSK 3β, and IκB kinase (IKK-α blocked PFF-A-mediated ATF3 expression. ATF3 knockdown by ATF3 siRNA attenuated the cleavage of poly (ADP-ribose polymerase (PARP by PFF-A, while ATF3 overexpression increased PFF-A-mediated cleaved PARP. These results suggest that PFF-A may exert anti-cancer property through inducing apoptosis via the ATF3-mediated pathway in human colorectal cancer cells.

  17. Transcriptional regulation of human FE65, a ligand of Alzheimer's disease amyloid precursor protein, by Sp1.

    LENUS (Irish Health Repository)

    Yu, Hoi-Tin

    2010-03-01

    FE65 is a neuronal-enriched adaptor protein that binds to the Alzheimer\\'s disease amyloid precursor protein (APP). FE65 forms a transcriptionally active complex with the APP intracellular domain (AICD). The precise gene targets for this complex are unclear but several Alzheimer\\'s disease-linked genes have been proposed. Additionally, evidence suggests that FE65 influences APP metabolism. The mechanism by which FE65 expression is regulated is as yet unknown. To gain insight into the regulatory mechanism, we cloned a 1.6 kb fragment upstream of the human FE65 gene and found that it possesses particularly strong promoter activity in neurones. To delineate essential regions in the human FE65 promoter, a series of deletion mutants were generated. The minimal FE65 promoter was located between -100 and +5, which contains a functional Sp1 site. Overexpression of the transcription factor Sp1 potentiates the FE65 promoter activity. Conversely, suppression of the FE65 promoter was observed in cells either treated with an Sp1 inhibitor or in which Sp1 was knocked down. Furthermore, reduced levels of Sp1 resulted in downregulation of endogenous FE65 mRNA and protein. These findings reveal that Sp1 plays a crucial role in transcriptional control of the human FE65 gene.

  18. Transcriptional regulation of the human Na+/H+ exchanger NHE3 by serotonin in intestinal epithelial cells

    International Nuclear Information System (INIS)

    Amin, Md Ruhul; Ghannad, Leda; Othman, Ahmad; Gill, Ravinder K.; Dudeja, Pradeep K.; Ramaswamy, Krishnamurthy; Malakooti, Jaleh

    2009-01-01

    Serotonin (5-HT) decreases NHE2 and NHE3 activities under acute conditions in human intestinal epithelial cells. Here, we have investigated the effects of 5-HT on expression of the human NHE3 gene and the mechanisms underlying its transcriptional regulation in differentiated C2BBe1 cells. Treatment of the human intestinal epithelial cell line, C2BBe1, with 5-HT (20 μM) resulted in a significant decrease in NHE3 mRNA and protein expression. In transient transfection studies, 5-HT repressed the NHE3 promoter activity by ∼55%. The repression of the NHE3 promoter activity in response to 5-HT was accompanied by reduced DNA-binding activity of transcription factors Sp1 and Sp3 to the NHE3 promoter without alteration in their nuclear levels. Pharmacological inhibitors of protein kinase C reversed the inhibitory effect of 5-HT on the promoter activity. Our data indicate that 5-HT suppresses the transcriptional activity of the NHE3 promoter and this effect may be mediated by PKCα and modulation of DNA-binding affinities of Sp1 and Sp3.

  19. Validation of whole-blood transcriptome signature during microdose recombinant human erythropoietin (rHuEpo) administration.

    Science.gov (United States)

    Wang, Guan; Durussel, Jérôme; Shurlock, Jonathan; Mooses, Martin; Fuku, Noriyuki; Bruinvels, Georgie; Pedlar, Charles; Burden, Richard; Murray, Andrew; Yee, Brendan; Keenan, Anne; McClure, John D; Sottas, Pierre-Edouard; Pitsiladis, Yannis P

    2017-11-14

    Recombinant human erythropoietin (rHuEpo) can improve human performance and is therefore frequently abused by athletes. As a result, the World Anti-Doping Agency (WADA) introduced the Athlete Biological Passport (ABP) as an indirect method to detect blood doping. Despite this progress, challenges remain to detect blood manipulations such as the use of microdoses of rHuEpo. Forty-five whole-blood transcriptional markers of rHuEpo previously derived from a high-dose rHuEpo administration trial were used to assess whether microdoses of rHuEpo could be detected in 14 trained subjects and whether these markers may be confounded by exercise (n = 14 trained subjects) and altitude training (n = 21 elite runners and n = 4 elite rowers, respectively). Differential gene expression analysis was carried out following normalisation and significance declared following application of a 5% false discovery rate (FDR) and a 1.5 fold-change. Adaptive model analysis was also applied to incorporate these markers for the detection of rHuEpo. ALAS2, BCL2L1, DCAF12, EPB42, GMPR, SELENBP1, SLC4A1, TMOD1 and TRIM58 were differentially expressed during and throughout the post phase of microdose rHuEpo administration. The CD247 and TRIM58 genes were significantly up- and down-regulated, respectively, immediately following exercise when compared with the baseline both before and after rHuEpo/placebo. No significant gene expression changes were found 30 min after exercise in either rHuEpo or placebo groups. ALAS2, BCL2L1, DCAF12, SLC4A1, TMOD1 and TRIM58 tended to be significantly expressed in the elite runners ten days after arriving at altitude and one week after returning from altitude (FDR > 0.059, fold-change varying from 1.39 to 1.63). Following application of the adaptive model, 15 genes showed a high sensitivity (≥ 93%) and specificity (≥ 71%), with BCL2L1 and CSDA having the highest sensitivity (93%) and specificity (93%). Current results provide further evidence that

  20. Whole blood microculture assay of human lymphocyte function.

    Science.gov (United States)

    Pauly, J L; Han, T

    1976-11-01

    A whole blood microculture assay is described for measuring lymphocyte reactivity to mitogenic and antigenic stimulants. This assay employs heparinized whole blood, serum-free culture medium, microtiter plates, and a Multiple Automated Sample Harvester (MASH). When this assay is compared to other leukocyte assays, its major advantages include (1) the utilization of fewer lymphocytes per microculture, thuus reducing the amount of blood required per test while increasing the number of test agents and replicate cultures which can be employed in any given experiment; (2) the conservation of mitogens, antigens, drugs, enzymes, hormones, lymphokines, and other test agents, some of which are either expensive of difficult to prepare in large quantities; (3) the elimination of lymphocyte isolation and purification procedures which may disrupt the relative proportion of T cells, B cells and antigen-processing cells; and (4) the application of an automated harvester which simplifies and expedites procedures required for processing cells for liquid scintillation counting.

  1. Reduced blood flow to contracting skeletal muscle in ageing humans

    DEFF Research Database (Denmark)

    Nyberg, Michael Permin; Hellsten, Ylva

    2016-01-01

    The ability to sustain a given absolute submaximal workload declines with advancing age likely due to a lower level of blood flow and O2 delivery to the exercising muscles. Given that physical inactivity mimics many of the physiological changes associated with ageing, separating the physiological...... consequences of ageing and physical inactivity can be challenging; yet, observations from cross-sectional and longitudinal studies on the effects of physical activity have provided some insight. Physical activity has the potential to offset the age-related decline in blood flow to contracting skeletal muscle...... the O2 demand of the active skeletal muscle of aged individuals during conditions where systemic blood flow is not limited by cardiac output seems to a large extent to be related to the level of physical activity. This article is protected by copyright. All rights reserved....

  2. Sex differences of human cortical blood flow and energy metabolism

    DEFF Research Database (Denmark)

    Aanerud, Joel; Borghammer, Per; Rodell, Anders

    2017-01-01

    cerebral blood flow and cerebral metabolic rate of oxygen as functions of age in healthy volunteers of both sexes. Cerebral metabolic rate of oxygen did not change with age for either sex and there were no differences of mean values of cerebral metabolic rate of oxygen between men and women in cerebral...... cortex. Women had significant decreases of cerebral blood flow as function of age in frontal and parietal lobes. Young women had significantly higher cerebral blood flow than men in frontal and temporal lobes, but these differences had disappeared at age 65. The absent sex difference of cerebral energy...... turnover suggests that the known differences of synaptic density between the sexes are counteracted by opposite differences of individual synaptic activity....

  3. Effects of midazolam on cerebral blood flow in human volunteers

    International Nuclear Information System (INIS)

    Forster, A.; Juge, O.; Morel, D.

    1982-01-01

    The effects of intravenously administered midazolam on cerebral blood flow were evaluated in eight healthy volunteers using the 133 Xe inhalation technique. Six minutes after an intravenous dose of 0.15 mg/kg midazolam, the cerebral blood flow decreased significantly (P less than 0.001) from a value of 40.6 +/- 3.3 to a value of 27.0 +/- 5.0 ml . 100 g-1 . min-1. Cerebrovascular resistance (CVR) increased from 2.8 +/- 0.2 to 3.9 to 0.6 mmHg/(ml . 100 g-1 . min-1)(P less than 0.001). Mean arterial blood pressure decreased significantly (P less than 0.05) from 117 +/- 8 to 109 +/- 9 mmHg and arterial carbon dioxide tension increased from 33.9 +/- 2.3 to 38.6 +/- 3.2 mmHg (P less than 0.05). Arterial oxygen tension remained stable throughout the study, 484 +/- 95 mmHg before the administration of midazolam and 453 +/- 76 mmHg after. All the subjects slept after the injection of the drug and had anterograde amnesia of 24.5 +/- 5 min. The decrease in mean arterial blood pressure was probably not important since it remained in the physiologic range for cerebral blood flow autoregulation. The increase in arterial carbon dioxide tension observed after the midazolam injection may have partially counteracted the effect of this new benzodiazepine on cerebral blood flow. Our data suggest that midazolam might be a safe agent to use for the induction of anethesia in neurosurgical patients with intracranial hypertension

  4. Knowledge, attitude, and beliefs of young, college student blood donors about Human immunodeficiency virus

    OpenAIRE

    Dubey, Anju; Sonker, Atul; Chaudhary, Rajendra K.

    2014-01-01

    Introduction: Young people, who tend to be healthy, idealistic, and motivated, are an excellent pool of potential voluntary unpaid blood donors. Recruiting and retaining young blood donors improves the long term safety and sufficiency of a country′s blood supply. Knowledge, attitude, and beliefs about Human immunodeficiency virus (HIV) should play an important role in prevention of disease transmission. Materials and Methods: This study was a questionnaire based survey, conducted to explore t...

  5. Kub5-Hera, the human Rtt103 homolog, plays dual functional roles in transcription termination and DNA repair.

    Science.gov (United States)

    Morales, Julio C; Richard, Patricia; Rommel, Amy; Fattah, Farjana J; Motea, Edward A; Patidar, Praveen L; Xiao, Ling; Leskov, Konstantin; Wu, Shwu-Yuan; Hittelman, Walter N; Chiang, Cheng-Ming; Manley, James L; Boothman, David A

    2014-04-01

    Functions of Kub5-Hera (In Greek Mythology Hera controlled Artemis) (K-H), the human homolog of the yeast transcription termination factor Rtt103, remain undefined. Here, we show that K-H has functions in both transcription termination and DNA double-strand break (DSB) repair. K-H forms distinct protein complexes with factors that repair DSBs (e.g. Ku70, Ku86, Artemis) and terminate transcription (e.g. RNA polymerase II). K-H loss resulted in increased basal R-loop levels, DSBs, activated DNA-damage responses and enhanced genomic instability. Significantly lowered Artemis protein levels were detected in K-H knockdown cells, which were restored with specific K-H cDNA re-expression. K-H deficient cells were hypersensitive to cytotoxic agents that induce DSBs, unable to reseal complex DSB ends, and showed significantly delayed γ-H2AX and 53BP1 repair-related foci regression. Artemis re-expression in K-H-deficient cells restored DNA-repair function and resistance to DSB-inducing agents. However, R loops persisted consistent with dual roles of K-H in transcription termination and DSB repair.

  6. Insight into podocyte differentiation from the study of human genetic disease: nail-patella syndrome and transcriptional regulation in podocytes.

    Science.gov (United States)

    Morello, Roy; Lee, Brendan

    2002-05-01

    In recent years, our understanding of the molecular basis of kidney development has benefited from the study of rare genetic diseases affecting renal function. This has especially been the case with the differentiation of the highly specialized podocyte in the pathogenesis of human disorders and mouse phenotypes affecting the renal filtration barrier. This filtration barrier represents the end product of a complex series of signaling events that produce a tripartite structure consisting of interdigitating podocyte foot processes with intervening slit diaphragms, the glomerular basement membrane, and the fenestrated endothelial cell. Dysregulation of unique cytoskeletal and extracellular matrix proteins in genetic forms of nephrotic syndrome has shown how specific structural proteins contribute to podocyte function and differentiation. However, much less is known about the transcriptional determinants that both specify and maintain this differentiated cell. Our studies of a skeletal malformation syndrome, nail-patella syndrome, have shown how the LIM homeodomain transcription factor, Lmx1b, contributes to transcriptional regulation of glomerular basement membrane collagen expression by podocytes. Moreover, they raise intriguing questions about more global transcriptional regulation of podocyte morphogenesis.

  7. Bone-Like Hydroxyapatite Formation in Human Blood

    Science.gov (United States)

    Titov, Anatoly T.; Larionov, Peter M.; Ivanova, Alexandra S.; Zaikovskii, Vladimir I.; Chernyavskiy, Mikhail A.

    2016-01-01

    The purpose of this study was to prove the mechanism of mineralization, when hydroxyapatite (HAP) is formed in blood plasma. These observations were substantiated by in vitro simulation of HAP crystallization in the plasma of healthy adults in a controllable quasi-physiological environment (T = 37°C, pH = 7.4) and at concentrations of dissolved Ca…

  8. In vitro differentiation of human umbilical cord blood mesenchymal ...

    African Journals Online (AJOL)

    Mesenchymal stem cells (MSCs) were isolated by gradient density centrifugation from umbilical cord blood. Spindle-shaped adherent cells were permitted to grow to 70% confluence in primary culture media which was reached by day 12. Induction of differentiation started by culturing cells with differentiation medium ...

  9. Effect on some haematological indices of human whole blood when ...

    African Journals Online (AJOL)

    DR UGHACHUKWU

    2013-07-31

    Jul 31, 2013 ... Awka Campus, Anambra State Nigeria. 2Department of Obstetrics and Gynaecology. Anambra State University Teaching Hospital, Amaku Awka,. Anambra State Nigeria. 3Department of Hematology and Blood Group Serology. College of Health Sciences, Nnamdi Azikiwe University, Awka,. Nnewi Campus.

  10. Roughness influence on human blood drop spreading and splashing

    Science.gov (United States)

    Smith, Fiona; Buntsma, Naomi; Brutin, David

    2017-11-01

    The impact behaviour of complex fluid droplets is a topic that has been extensively studied but with much debate. The Bloodstain Pattern Analysis (BPA) community is encountering this scientific problem with daily practical cases since they use bloodstains as evidence in crime scene reconstruction. We aim to provide fundamental explanations in the study of blood drip stains by investigating the influence of surface roughness and wettability on the splashing limit of droplets of blood, a non-Newtonian colloidal fluid. Droplets of blood impacting perpendicularly different surfaces at different velocities were recorded. The recordings were analysed as well as the surfaces characteristics in order to find an empirical solution since we found that roughness plays a major role in the threshold of the splashing/non-splashing behaviour of blood compared to the wettability. Moreover it appears that roughness alters the deformation of the drip stains. These observations are key in characterising features of drip stains with the impacting conditions, which would answer some forensic issues.

  11. Effects of Septrin Administration on Blood Cells Parameters in Humans

    African Journals Online (AJOL)

    The results showed that the packed cell volume (PCV), total white blood cell count (WBC), neutrophils and platelets were significantly decreased (p<0.05), especially after 7-10 days of septrin administration, compared to the control values. On the other hand, the reticulocytes, lymphocytes, eosinophils and prothrombin time ...

  12. A Laboratory Exercise to Determine Human ABO Blood Type by Noninvasive Methods

    Science.gov (United States)

    Martin, Michael P.; Detzel, Stephen M.

    2008-01-01

    Analysis of single-nucleotide polymorphisms and their association with diseases and nondisease phenotypes is of growing importance in human biology studies. In this laboratory exercise, students determine the genetic basis for their ABO blood type; however, no blood is drawn. Students isolate genomic DNA from buccal mucosa cells that are present…

  13. Determination of Elements in Normal and Leukemic Human Whole Blood by Neutron Activation Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Brune, D; Frykberg, B; Samsahl, K; Wester, P O

    1961-11-15

    By means of gamma-spectrometry the following elements were simultaneously determined in normal and leukemic human whole blood: Cu, Mn, Zn, Sr, Na, P, Ca, Rb, Cd, Sb, Au, Cs and Fe. Chemical separations were performed according to a group separation method using ion-exchange technique. No significant difference between the concentrations of the elements in normal- and leukemic blood was observed.

  14. Determination of telmisartan in human blood plasma: Part I: Immunoassay development

    NARCIS (Netherlands)

    Hempen, C.M.; Gläsle-Schwarz, Liane; Kunz, Ulrich; Karst, U.

    2006-01-01

    Telmisartan is an angiotensin II receptor antagonist and a known drug against high blood pressure. In this report, the development of a new and rapid analytical technique, an enzyme-linked immunosorbent assay (ELISA) for the determination of telmisartan in human blood plasma is described. The

  15. Determination of Elements in Normal and Leukemic Human Whole Blood by Neutron Activation Analysis

    International Nuclear Information System (INIS)

    Brune, D.; Frykberg, B.; Samsahl, K.; Wester, P.O.

    1961-11-01

    By means of gamma-spectrometry the following elements were simultaneously determined in normal and leukemic human whole blood: Cu, Mn, Zn, Sr, Na, P, Ca, Rb, Cd, Sb, Au, Cs and Fe. Chemical separations were performed according to a group separation method using ion-exchange technique. No significant difference between the concentrations of the elements in normal- and leukemic blood was observed

  16. Gene expression in human peripheral blood 48 hours after exposure to ionizing radiation

    Data.gov (United States)

    National Aeronautics and Space Administration — Analysis of human peripheral blood 48 hours after irradiation ex vivo with graded doses of gamma rays. Results have been used in building and testing classifiers to...

  17. microRNA expression profiles in human peripheral blood lymphocytes cultured in modeled microgravity

    Data.gov (United States)

    National Aeronautics and Space Administration — In the present study we analyzed miRNA and mRNA expression profiles in human peripheral blood lymphocytes (PBLs) incubated in microgravity condition simulated by a...

  18. Gene expression profiling of human peripheral blood lymphocytes cultured in modeled microgravity

    Data.gov (United States)

    National Aeronautics and Space Administration — In the present study we analyzed miRNA and mRNA expression profiles in human peripheral blood lymphocytes (PBLs) incubated in microgravity condition simulated by a...

  19. Survey and evaluation of mutations in the human KLF1 transcription unit.

    Science.gov (United States)

    Gnanapragasam, Merlin Nithya; Crispino, John D; Ali, Abdullah M; Weinberg, Rona; Hoffman, Ronald; Raza, Azra; Bieker, James J

    2018-04-26

    Erythroid Krüppel-like Factor (EKLF/KLF1) is an erythroid-enriched transcription factor that plays a global role in all aspects of erythropoiesis, including cell cycle control and differentiation. We queried whether its mutation might play a role in red cell malignancies by genomic sequencing of the KLF1 transcription unit in cell lines, erythroid neoplasms, dysplastic disorders, and leukemia. In addition, we queried published databases from a number of varied sources. In all cases we only found changes in commonly notated SNPs. Our results suggest that if there are mutations in KLF1 associated with erythroid malignancies, they are exceedingly rare.

  20. Elastolytic activity of human blood monocytes characterized by a new monoclonal antibody against human leucocyte elastase. Relationship to rheumatoid arthritis

    DEFF Research Database (Denmark)

    Jensen, H S; Christensen, L D

    1990-01-01

    The leucocyte elastase of human blood monocytes was investigated by applying a new monoclonal antibody which did not block the enzyme activity against elastin. In a fixed population of mononuclear cells (MNC) and using fluorescence activated cell sorting (FACS), the human leucocyte elastase (HLE...

  1. DNA damage response and role of shelterin complex in human peripheral blood mononuclear cells exposed to gamma radiation

    International Nuclear Information System (INIS)

    Saini, Divyalakshmi; Das, Birajalaxmi

    2013-01-01

    Telomeres are the DNA protein structures that cap the ends of linear DNA. It consists of short repetitive DNA sequences (TTAGGG)n and specialized telomere binding proteins. There are six telomeric proteins (TRF1, TRF2, TIN2, TERF2, PTOP and POT1) called as shelterin complex/telosome which maintains telomere integrity. The function of this 'telosome' is to protect the natural ends of the chromosomes from being recognized as artificial DNA breaks, thereby preventing chromosome end-to-end fusions. DNA Damage Response (DDR) induced by radiation and its interaction with telomeric protein complex is poorly understood in human PBMCs at G 0 stage. Alterations in either telomeric DNA or telomere binding proteins can impair the function of the telosome, which may lead to senescence or apoptosis. Ionizing radiation which induces a plethora of DNA lesions in human cell may also alter the expression of telomere associated proteins. In the present study, we have made an attempt to study the DNA damage response of telomere proteins in human peripheral blood mononuclear cells exposed to gamma radiation. Venous blood samples were collected from eight random healthy volunteers and PBMCs were separated. Dose response as well as time point kinetics study was carried out at transcription as well as protein level. PBMCs were irradiated at various doses between 10 cGy to 2.0 Gy at a dose rate of 1.0 Gy/min. Total RNA was isolated for gene expression analysis at 0 hour and 4 hours respectively. cDNA was prepared and transcriptional pattern as studied using real time q-PCR where Taqman probes were used. Time point kinetics of transcriptional pattern of TRF1, TRF2, TIN2, TERF2, PTOP and POT1 was carried out at 0 min, 15 min, 30 min, 60 min, and 120 min for two different doses (1.0 Gy and 2.0 Gy). Dose response and time point kinetics of TRF2 was studied at similar doses using confocal microscopy. Our results revealed that at 2.0 Gy there was a two fold increase at the level of transcription

  2. Aspirin-triggered lipoxin A4 and lipoxin A4 up-regulate transcriptional corepressor NAB1 in human neutrophils.

    Science.gov (United States)

    Qiu, F H; Devchand, P R; Wada, K; Serhan, C N

    2001-12-01

    Aspirin-triggered 15-epi-lipoxin A4 (ATL) is an endogenous lipid mediator that mimics the actions of native lipoxin A4, a putative "stop signal" involved in regulating resolution of inflammation. A metabolically more stable analog of ATL, 15-epi-16-(para-fluoro)-phenoxy-lipoxin A4 analog (ATLa), inhibits neutrophil recruitment in vitro and in vivo and displays potent anti-inflammatory actions. ATLa binds with high affinity to the lipoxin A4 receptor, a G protein-coupled receptor on the surface of leukocytes. In this study, we used freshly isolated human neutrophils to examine ATLa's potential for initiating rapid nuclear responses. Using differential display reverse transcription polymerase chain reaction, we identified a subset of genes that was selectively up-regulated upon short exposure of polymorphonuclear leukocytes to ATLa but not to the chemoattractant leukotriene B4 or vehicle alone. We further investigated ATLa regulation of one of the genes, NAB1, a transcriptional corepressor identified previously as a glucocorticoid-responsive gene in hamster smooth muscle cells. Treatment of human neutrophils with pertussis toxin blocked ATLa up-regulation of NAB1. In addition, ATLa stimulated NAB1 gene expression in murine lung vascular smooth muscle in vivo. These findings provide evidence for rapid transcriptional induction of a cassette of genes via an ATLa-stimulated G protein-coupled receptor pathway that is potentially protective and overlaps with the anti-inflammatory glucocorticoid regulatory circuit.

  3. Transcriptional profiling of MEF2-regulated genes in human neural progenitor cells derived from embryonic stem cells

    Directory of Open Access Journals (Sweden)

    Shing Fai Chan

    2015-03-01

    Full Text Available The myocyte enhancer factor 2 (MEF2 family of transcription factors is highly expressed in the brain and constitutes a key determinant of neuronal survival, differentiation, and synaptic plasticity. However, genome-wide transcriptional profiling of MEF2-regulated genes has not yet been fully elucidated, particularly at the neural stem cell stage. Here we report the results of microarray analysis comparing mRNAs isolated from human neural progenitor/stem cells (hNPCs derived from embryonic stem cells expressing a control vector versus progenitors expressing a constitutively-active form of MEF2 (MEF2CA, which increases MEF2 activity. Microarray experiments were performed using the Illumina Human HT-12 V4.0 expression beadchip (GEO#: GSE57184. By comparing vector-control cells to MEF2CA cells, microarray analysis identified 1880 unique genes that were differentially expressed. Among these genes, 1121 genes were up-regulated and 759 genes were down-regulated. Our results provide a valuable resource for identifying transcriptional targets of MEF2 in hNPCs.

  4. An Algorithm for Generating Small RNAs Capable of Epigenetically Modulating Transcriptional Gene Silencing and Activation in Human Cells

    Directory of Open Access Journals (Sweden)

    Amanda Ackley

    2013-01-01

    Full Text Available Small noncoding antisense RNAs (sasRNAs guide epigenetic silencing complexes to target loci in human cells and modulate gene transcription. When these targeted loci are situated within a promoter, long-term, stable epigenetic silencing of transcription can occur. Recent studies suggest that there exists an endogenous form of such epigenetic regulation in human cells involving long noncoding RNAs. In this article, we present and validate an algorithm for the generation of highly effective sasRNAs that can mimic the endogenous noncoding RNAs involved in the epigenetic regulation of gene expression. We validate this algorithm by targeting several oncogenes including AKT-1, c-MYC, K-RAS, and H-RAS. We also target a long antisense RNA that mediates the epigenetic repression of the tumor suppressor gene DUSP6, silenced in pancreatic cancer. An algorithm that can efficiently design small noncoding RNAs for the epigenetic transcriptional silencing or activation of specific genes has potential therapeutic and experimental applications.

  5. A T-cell specific transcriptional enhancer element 3' of Cα in the human T-cell receptor α locus

    International Nuclear Information System (INIS)

    Ho, Icheng; Yang, Lihsuan; Morle, G.; Leiden, J.M.

    1989-01-01

    A transcriptional enhancer element has been identified 4.5 kilobases 3' of C α (constant region α chain) in the human T-cell receptor (TCR) α-chain locus. This enhancer is active on both a TCR V α (variable region α chain) promoter and the minimal simian virus 40 promoter in TCR α/β Jurkat and EL4 cells but is inactive on a V α promoter TCR γ/δ PEER and Molt-13 cells, clone 13 B cells, and HeLa fibroblasts. The enhancer has been localized to a 116-base-pair BstXI/Dra I restriction enzyme fragment, which lacks immunoglobulin octamer and κB enhancer motifs but does contain a consensus cAMP-response element (CRE). DNase I footprint analyses demonstrated that the minimal enhancer contains two binding sites for Jurkat nuclear proteins. One of these sites corresponds to the CRE, while the other does not correspond to a known transcriptional enhancer motif. These data support a model in which TCR α gene transcription is regulated by a unique set of cis-acting sequences and trans-acting factors, which are differentially active in cells of the TCR α/β lineage. In addition, the TCR α enhancer may play a role in activating oncogene expression in T-lymphoblastoid tumors that have previously been shown to display chromosomal translocations into the human TCR α locus

  6. Polymerase chain reaction amplification fails to detect aromatase cytochrome P450 transcripts in normal human endometrium or decidua.

    Science.gov (United States)

    Bulun, S E; Mahendroo, M S; Simpson, E R

    1993-06-01

    It has been proposed that the biosynthesis of estrogens by the human endometrium may be of physiological significance during the menstrual cycle. Local estrogen production was also suggested to be important in the development of endometrial cancer; however, the presence or absence of aromatase enzyme activity in normal human endometrium is controversial. To address this issue, we used a sensitive technique capable of detecting mRNA transcripts present in only very low copy number. The polymerase chain reaction linked to reverse transcription (RT-PCR) was used to evaluate the presence or absence of aromatase cytochrome P450 (P450arom) transcripts in endometrial tissues (n = 7) and endometrial stromal cells (n = 9) under various culture conditions. RNA was isolated from four proliferative and three secretory tissue samples and from cultured endometrial stromal cells isolated from seven proliferative and two secretory endometria. Five sets of cultures were treated with medroxyprogesterone acetate (MPA), estradiol (E2), and forskolin. Additionally, RNA was isolated from decidualized endometrium obtained from a patient with tubal pregnancy. A single stranded cDNA was synthesized from total RNA using Moloney murine leukemia virus reverse transcriptase and a P450arom-specific oligonucleotide. The single stranded cDNA was used as a template for PCR and was amplified for 20-35 cycles using P450arom-specific primers. RNA from adipose tissue and placenta was amplified to provide positive controls, whereas myometrial RNA was used as a negative control. In two experiments involving two endometrial tissues and three sets of cells in culture, a rat P450arom cRNA was coamplified in each sample as an internal control to demonstrate that the remote possibility of RT-PCR failures in individual test samples cannot account for our negative results. By Southern or slot blot hybridization of the amplified fragments using human and rat P450arom-specific probes, we found no evidence for

  7. Allogeneic human dermal fibroblasts are viable in peripheral blood mononuclear co-culture

    Directory of Open Access Journals (Sweden)

    Restu Syamsul Hadi

    2014-08-01

    Full Text Available Background Transplanted allogeneic dermal fibroblasts retain stem cell subpopulations, and are easily isolated, expanded and stored using standard techniques. Their potential for regenerative therapy of chronic wounds should be evaluated. The aim of this study was to determine allogeneic fibroblast viability in the presence of peripheral blood mononuclear cells (PBMC. Methods In this experimental study, fibroblasts were isolated from foreskin explants, expanded in the presence of serum, and stored using slow-freezing. We used one intervention group of allogeneic fibroblasts co-cultured with PBMC and 2 control groups of separate fibroblast and PBMC cultures.Fibroblasts were characterized by their collagen secretion and octamer-binding transcription factor 4 (OCT4 expression. Viability was evaluated using water soluble tetrazolium-1 (WST-1 proliferation assay. Absorbances were measured at 450 nm. Data analysis was performed by student’s paired t-test. Results Dermal fibroblasts were shown to secrete collagen, express OCT4, be recoverable after cryopreservation, and become attached to the culture dish in a co-culture with PBMC. Co-cultured and control fibroblasts had no significantly different cell viabilities (p>0.05. Calculated viable cell numbers increased 1.8 and 5.1-fold, respectively, at days 2 and 4 in vitro. Both groups showed comparable doubling times at days 2 and 4 in vitro. PBMC did not interfere with allogeneic fibroblast viability and proliferative capacity Conclusions Allogeneic fibroblasts remain viable and proliferate in the presence of host PBMC. Future research should evaluate allogeneic human dermal fibroblast competency in clinical settings. Dermal fibroblasts are a potential source for cell therapy in chronic wound management.

  8. Ensemble approach combining multiple methods improves human transcription start site prediction.

    LENUS (Irish Health Repository)

    Dineen, David G

    2010-01-01

    The computational prediction of transcription start sites is an important unsolved problem. Some recent progress has been made, but many promoters, particularly those not associated with CpG islands, are still difficult to locate using current methods. These methods use different features and training sets, along with a variety of machine learning techniques and result in different prediction sets.

  9. Transcription factor ZNF25 is associated with osteoblast differentiation of human skeletal stem cells

    DEFF Research Database (Denmark)

    Twine, Natalie A.; Harkness, Linda; Kassem, Moustapha

    2016-01-01

    containing G protein-coupled receptor 5 and RAN-binding protein 3-like. We also observed enrichment in extracellular matrix organization, skeletal system development and regulation of ossification in the entire upregulated set of genes. Consistent with its function as a transcription factor during osteoblast...

  10. Core transcriptional regulatory circuit controlled by the TAL1 complex in human T cell acute lymphoblastic leukemia.

    Science.gov (United States)

    Sanda, Takaomi; Lawton, Lee N; Barrasa, M Inmaculada; Fan, Zi Peng; Kohlhammer, Holger; Gutierrez, Alejandro; Ma, Wenxue; Tatarek, Jessica; Ahn, Yebin; Kelliher, Michelle A; Jamieson, Catriona H M; Staudt, Louis M; Young, Richard A; Look, A Thomas

    2012-08-14

    The oncogenic transcription factor TAL1/SCL is aberrantly expressed in over 40% of cases of human T cell acute lymphoblastic leukemia (T-ALL), emphasizing its importance in the molecular pathogenesis of T-ALL. Here we identify the core transcriptional regulatory circuit controlled by TAL1 and its regulatory partners HEB, E2A, LMO1/2, GATA3, and RUNX1. We show that TAL1 forms a positive interconnected autoregulatory loop with GATA3 and RUNX1 and that the TAL1 complex directly activates the MYB oncogene, forming a positive feed-forward regulatory loop that reinforces and stabilizes the TAL1-regulated oncogenic program. One of the critical downstream targets in this circuitry is the TRIB2 gene, which is oppositely regulated by TAL1 and E2A/HEB and is essential for the survival of T-ALL cells. Copyright © 2012 Elsevier Inc. All rights reserved.

  11. Transcriptional activation of immediate-early gene ETR101 by human T-cell leukaemia virus type I Tax

    DEFF Research Database (Denmark)

    Chen, Li; Ma, Shiliang; Li, Bo

    2003-01-01

    Human T-cell leukaemia virus type I (HTLV-I) Tax regulates viral and cellular gene expression through interactions with multiple cellular transcription pathways. This study describes the finding of immediate-early gene ETR101 expression in HTLV-I-infected cells and its regulation by Tax. ETR101...... was persistently expressed in HTLV-I-infected cells but not in HTLV-I uninfected cells. Expression of ETR101 was dependent upon Tax expression in the inducible Tax-expressing cell line JPX-9 and also in Jurkat cells transiently transfected with Tax-expressing vectors. Tax transactivated the ETR101 gene promoter......-DNA complex in HTLV-I-infected cell lines. EMSA with specific antibodies confirmed that the CREB transcription factor was responsible for formation of this specific protein-DNA complex. These results suggested that Tax directly transactivated ETR101 gene expression, mainly through a CRE sequence via the CREB...

  12. Nanoparticle-mediated transcriptional modification enhances neuronal differentiation of human neural stem cells following transplantation in rat brain.

    Science.gov (United States)

    Li, Xiaowei; Tzeng, Stephany Y; Liu, Xiaoyan; Tammia, Markus; Cheng, Yu-Hao; Rolfe, Andrew; Sun, Dong; Zhang, Ning; Green, Jordan J; Wen, Xuejun; Mao, Hai-Quan

    2016-04-01

    Strategies to enhance survival and direct the differentiation of stem cells in vivo following transplantation in tissue repair site are critical to realizing the potential of stem cell-based therapies. Here we demonstrated an effective approach to promote neuronal differentiation and maturation of human fetal tissue-derived neural stem cells (hNSCs) in a brain lesion site of a rat traumatic brain injury model using biodegradable nanoparticle-mediated transfection method to deliver key transcriptional factor neurogenin-2 to hNSCs when transplanted with a tailored hyaluronic acid (HA) hydrogel, generating larger number of more mature neurons engrafted to the host brain tissue than non-transfected cells. The nanoparticle-mediated transcription activation method together with an HA hydrogel delivery matrix provides a translatable approach for stem cell-based regenerative therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Leptin upregulates telomerase activity and transcription of human telomerase reverse transcriptase in MCF-7 breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Ren, He, E-mail: herenrh@yahoo.com.cn [Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University, Ministry of Education, Tianjin Medical University Cancer Hospital, Tianjin (China); Zhao, Tiansuo; Wang, Xiuchao; Gao, Chuntao; Wang, Jian; Yu, Ming [Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University, Ministry of Education, Tianjin Medical University Cancer Hospital, Tianjin (China); Hao, Jihui, E-mail: jihuihao@yahoo.com [Key Laboratory of Breast Cancer Prevention and Therapy, Tianjin Medical University, Ministry of Education, Tianjin Medical University Cancer Hospital, Tianjin (China)

    2010-03-26

    The aim was to analyze the mechanism of leptin-induced activity of telomerase in MCF-7 breast cancer cells. We found that leptin activated telomerase in a dose-dependent manner; leptin upregulated the expression of Human Telomerase Reverse Transcriptase (hTERT) at mRNA and protein levels; blockade of signal transducer and activator of transcription 3 (STAT3) phosphorylation significantly counteracted leptin-induced hTERT transcription and protein expression; chromatin immunoprecipitation analysis showed that leptin enhanced the binding of STAT3 to the hTERT promoter. This study uncovers a new mechanism of the proliferative effect of leptin on breast cancer cells and provides a new explanation of obesity-related breast cancer.

  14. Crystal structure of the C-terminal domain of the RAP74 subunit of human transcription factor IIF

    Energy Technology Data Exchange (ETDEWEB)

    Kamada, Katsuhiko; De Angelis, Jacqueline; Roeder, Robert G.; Burley, Stephen K. (Rockefeller)

    2012-12-13

    The x-ray structure of a C-terminal fragment of the RAP74 subunit of human transcription factor (TF) IIF has been determined at 1.02-{angstrom} resolution. The {alpha}/{beta} structure is strikingly similar to the globular domain of linker histone H5 and the DNA-binding domain of hepatocyte nuclear factor 3{gamma} (HNF-3{gamma}), making it a winged-helix protein. The surface electrostatic properties of this compact domain differ significantly from those of bona fide winged-helix transcription factors (HNF-3{gamma} and RFX1) and from the winged-helix domains found within the RAP30 subunit of TFIIF and the {beta} subunit of TFIIE. RAP74 has been shown to interact with the TFIIF-associated C-terminal domain phosphatase FCP1, and a putative phosphatase binding site has been identified within the RAP74 winged-helix domain.

  15. RUNX1 promotes cell growth in human T-cell acute lymphoblastic leukemia by transcriptional regulation of key target genes.

    Science.gov (United States)

    Jenkins, Catherine E; Gusscott, Samuel; Wong, Rachel J; Shevchuk, Olena O; Rana, Gurneet; Giambra, Vincenzo; Tyshchenko, Kateryna; Islam, Rashedul; Hirst, Martin; Weng, Andrew P

    2018-05-04

    RUNX1 is frequently mutated in T-cell acute lymphoblastic leukemia (T-ALL). The spectrum of RUNX1 mutations has led to the notion that it acts as a tumor suppressor in this context; however, other studies have placed RUNX1 along with transcription factors TAL1 and NOTCH1 as core drivers of an oncogenic transcriptional program. To reconcile these divergent roles, we knocked down RUNX1 in human T-ALL cell lines and deleted Runx1 or Cbfb in primary mouse T-cell leukemias. RUNX1 depletion consistently resulted in reduced cell proliferation and increased apoptosis. RUNX1 upregulated variable sets of target genes in each cell line, but consistently included a core set of oncogenic effectors including IGF1R and NRAS. Our results support the conclusion that RUNX1 has a net positive effect on cell growth in the context of established T-ALL. Copyright © 2018. Published by Elsevier Inc.

  16. The forkhead transcription factor Foxl2 is sumoylated in both human and mouse: sumoylation affects its stability, localization, and activity.

    Directory of Open Access Journals (Sweden)

    Mara Marongiu

    2010-03-01

    Full Text Available The FOXL2 forkhead transcription factor is expressed in ovarian granulosa cells, and mutated FOXL2 causes the blepharophimosis, ptosis and epicanthus inversus syndrome (BPES and predisposes to premature ovarian failure. Inactivation of Foxl2 in mice demonstrated its indispensability for female gonadal sex determination and ovary development and revealed its antagonism of Sox9, the effector of male testis development. To help to define the regulatory activities of FOXL2, we looked for interacting proteins. Based on yeast two-hybrid screening, we found that FOXL2 interacts with PIAS1 and UBC9, both parts of the sumoylation machinery. We showed that human FOXL2 is sumoylated in transfected cell lines, and that endogenous mouse Foxl2 is comparably sumoylated. This modification changes its cellular localization, stability and transcriptional activity. It is intriguing that similar sumoylation and regulatory consequences have also been reported for SOX9, the male counterpart of FOXL2 in somatic gonadal tissues.

  17. Measurements of vitamin B12 in human blood serum using resonance Raman spectroscopy

    Science.gov (United States)

    Tsiminis, G.; Schartner, E. P.; Brooks, J. L.; Hutchinson, M. R.

    2016-12-01

    Vitamin B12 (cobalamin and its derivatives) deficiency has been identified as a potential modifiable risk factor for dementia and Alzheimer's disease. Chronic deficiency of vitamin B12 has been significantly associated with an increased risk of cognitive decline. An effective and efficient method for measuring vitamin B12 concentration in human blood would enable ongoing tracking and assessment of this potential modifiable risk factor. In this work we present an optical sensor based on resonance Raman spectroscopy for rapid measurements of vitamin B12 in human blood serum. The measurement takes less than a minute and requires minimum preparation (centrifuging) of the collected blood samples.

  18. Study of terahertz-radiation-induced DNA damage in human blood leukocytes

    Energy Technology Data Exchange (ETDEWEB)

    Angeluts, A A; Esaulkov, M N; Kosareva, O G; Solyankin, P M; Shkurinov, A P [International Laser Center, M. V. Lomonosov Moscow State University, Moscow (Russian Federation); Gapeyev, A B; Pashovkin, T N [Institute of Cell Biophysics, Russian Academy of Sciences, Pushchino, Moscow Region (Russian Federation); Matyunin, S N [Section of Applied Problems at the Presidium of the Russian Academy of Sciences, Moscow (Russian Federation); Nazarov, M M [Institute on Laser and Information Technologies, Russian Academy of Sciences, Shatura, Moscow Region (Russian Federation); Cherkasova, O P [Institute of Laser Physics, Siberian Branch, Russian Academy of Sciences, Novosibirsk (Russian Federation)

    2014-03-28

    We have carried out the studies aimed at assessing the effect of terahertz radiation on DNA molecules in human blood leukocytes. Genotoxic testing of terahertz radiation was performed in three different oscillation regimes, the blood leukocytes from healthy donors being irradiated for 20 minutes with the mean intensity of 8 – 200 μW cm{sup -2} within the frequency range of 0.1 – 6.5 THz. Using the comet assay it is shown that in the selected regimes such radiation does not induce a direct DNA damage in viable human blood leukocytes. (biophotonics)

  19. Optical diagnosis of dengue virus infection in human blood serum using Raman spectroscopy

    International Nuclear Information System (INIS)

    Saleem, M; Bilal, M; Anwar, S; Rehman, A; Ahmed, M

    2013-01-01

    We present the optical diagnosis of dengue virus infection in human blood serum using Raman spectroscopy. Raman spectra were acquired from 18 blood serum samples using a laser at 532 nm as the excitation source. A multivariate regression model based on partial least-squares regression is developed that uses Raman spectra to predict dengue infection with leave-one-sample-out cross validation. The prediction of dengue infection by our model yields correlation coefficient r 2 values of 0.9998 between the predicted and reference clinical results. The model was tested for six unknown human blood sera and found to be 100% accurate in accordance with the clinical results. (letter)

  20. Transcription and splicing regulation in human umbilical vein endothelial cells under hypoxic stress conditions by exon array

    Directory of Open Access Journals (Sweden)

    Wu Yonghong

    2009-03-01

    Full Text Available Abstract Background The balance between endothelial cell survival and apoptosis during stress is an important cellular process for vessel integrity and vascular homeostasis, and it is also pivotal in angiogenesis during the development of many vascular diseases. However, the underlying molecular mechanisms remain largely unknown. Although both transcription and alternative splicing are important in regulating gene expression in endothelial cells under stress, the regulatory mechanisms underlying this state and their interactions have not yet been studied on a genome-wide basis. Results Human umbilical vein endothelial cells (HUVECs were treated with cobalt chloride (CoCl2 both to mimic hypoxia and to induce cell apoptosis and alternative splicing responses. Cell apoptosis rate analysis indicated that HUVECs exposed to 300 μM CoCl2 for 24 hrs were initially counterbalancing apoptosis with cell survival. We therefore used the Affymetrix exon array system to determine genome-wide transcript- and exon-level differential expression. Other than 1583 differentially expressed transcripts, 342 alternatively spliced exons were detected and classified by different splicing types. Sixteen alternatively spliced exons were validated by RT-PCR. Furthermore, direct evidence for the ongoing balance between HUVEC survival and apoptosis was provided by Gene Ontology (GO and protein function, as well as protein domain and pathway enrichment analyses of the differentially expressed transcripts. Importantly, a novel molecular module, in which the heat shock protein (HSP families play a significant role, was found to be activated under mimicked hypoxia conditions. In addition, 46% of the transcripts containing stress-modulated exons were differentially expressed, indicating the possibility of combinatorial regulation of transcription and splicing. Conclusion The exon array system effectively profiles gene expression and splicing on the genome-wide scale. Based on

  1. The human tartrate-resistant acid phosphatase (TRAP): involvement of the hemin responsive elements (HRE) in transcriptional regulation.

    Science.gov (United States)

    Fleckenstein, E C; Dirks, W G; Drexler, H G

    2000-02-01

    The biochemical properties and protein structure of the tartrate-resistant acid phosphatase (TRAP), an iron-containing lysosomal glycoprotein in cells of the mononuclear phagocyte system, are well known. In contrast, little is known about the physiology and genic structure of this unique enzyme. In some diseases, like hairy cell leukemia, Gaucher's disease and osteoclastoma, cytochemically detected TRAP expression is used as a disease-associated marker. In order to begin to elucidate the regulation of this gene we generated different deletion constructs of the TRAP 5'-flanking region, placed them upstream of the luciferase reporter gene and assayed them for their ability to direct luciferase expression in human 293 cells. Treatment of these cells with the iron-modulating reagents transferrin and hemin causes opposite effects on the TRAP promoter activity. Two regulatory GAGGC tandem repeat sequences (the hemin responsive elements, HRE) within the 5'-flanking region of the human TRAP gene were identified. Studies with specific HRE-deletion constructs of the human TRAP 5'-flanking region upstream of the luciferase reporter gene document the functionality of these HRE-sequences which are apparently responsible for mediating transcriptional inhibition upon exposure to hemin. In addition to the previously published functional characterization of the murine TRAP HRE motifs, these results provide the first description of a new iron/hemin-responsive transcriptional regulation in the human TRAP gene.

  2. Renal cortical and medullary blood flow responses to altered NO availability in humans

    DEFF Research Database (Denmark)

    Damkjær, Mads; Vafaee, Manoucher; Møller, Michael L

    2010-01-01

    The objective of this study was to quantify regional renal blood flow in humans. In nine young volunteers on a controlled diet, the lower abdomen was CT-scanned, and regional renal blood flow was determined by positron emission tomography (PET) scanning using H(2)(15)O as tracer. Measurements were......-NMMA injection to 1.57 ± 0.17 ml·g tissue(-1)·min(-1) (P blood flow was 4.67 ± 0.31 ml·g tissue(-1)·min(-1) during control, unchanged by glyceryl nitrate, and decreased after L-NMMA [3.48 ± 0.23 ml·(g·min)(-1), P renal medullary region in which...... the measured blood flow is 1) low, 2) independent of reduction in the VOI, and 3) reactive to changes in systemic NO supply. The technique seems to provide indices of renal medullary blood flow in humans....

  3. Pattern of distribution of blood group antigens on human epidermal cells during maturation

    DEFF Research Database (Denmark)

    Dabelsteen, Erik; Buschard, Karsten; Hakomori, Sen-Itiroh

    1984-01-01

    The distribution in human epidermis of A, B, and H blood group antigens and of a precursor carbohydrate chain, N-acetyl-lactosamine, was examined using immunofluorescence staining techniques. The material included tissue from 10 blood group A, 4 blood group B, and 9 blood group O persons. Murine...... on the lower spinous cells whereas H antigen was seen predominantly on upper spinous cells or on the granular cells. Epithelia from blood group A or B persons demonstrated A or B antigens, respectively, but only if the tissue sections were trypsinized before staining. In such cases A or B antigens were found...... monoclonal antibodies were used to identify H antigen (type 2 chain) and N-acetyl-lactosamine. Human antisera were used to identify A and B antigens. In all groups N-acetyl-lactosamine and H antigen were found on the cell membranes of the spinous cell layer. N-acetyl-lactosamine was present mainly...

  4. Blood temperature and perfusion to exercising and non-exercising human limbs

    DEFF Research Database (Denmark)

    González-Alonso, José; Calbet, José Al; Boushel, Robert

    2015-01-01

    Temperature-sensitive mechanisms may contribute to blood flow regulation, but the influence of temperature on perfusion to exercising and non-exercising human limbs is not established. Blood temperature (TB ), blood flow and oxygen uptake (VO2 ) in the legs and arms were measured in 16 healthy...... humans during 90 min of leg and arm exercise and during exhaustive incremental leg or arm exercise. During prolonged exercise, leg blood flow (LBF) was 4-fold higher than arm blood flow (ABF) in association with higher TB and limb VO2 . Leg and arm vascular conductance during exercise compared to rest...... was related closely to TB (R(2) = 0.91; P exercise, LBF increased in association with elevations in TB and limb VO2 whereas ABF, arm TB and VO2 remained largely unchanged. During...

  5. “Data characterizing microfabricated human blood vessels created via hydrodynamic focusing”

    Directory of Open Access Journals (Sweden)

    Kyle A. DiVito

    2017-10-01

    Full Text Available This data article provides further detailed information related to our research article titled “Microfabricated Blood Vessels Undergo Neovascularization” (DiVito et al., 2017 [1], in which we report fabrication of human blood vessels using hydrodynamic focusing (HDF. Hydrodynamic focusing with advection inducing chevrons were used in concert to encase one fluid stream within another, shaping the inner core fluid into ‘bullseye-like” cross-sections that were preserved through click photochemistry producing streams of cellularized hollow 3-dimensional assemblies, such as human blood vessels (Daniele et al., 2015a, 2015b, 2014, 2016; Roberts et al., 2016 [2–6]. Applications for fabricated blood vessels span general tissue engineering to organ-on-chip technologies, with specific utility in in vitro drug delivery and pharmacodynamics studies. Here, we report data regarding the construction of blood vessels including cellular composition and cell positioning within the engineered vascular construct as well as functional aspects of the tissues.

  6. Global transcriptional analysis of psoriatic skin and blood confirms known disease-associated pathways and highlights novel genomic "hot spots" for differentially expressed genes.

    Science.gov (United States)

    Coda, Alvin B; Icen, Murat; Smith, Jason R; Sinha, Animesh A

    2012-07-01

    There are major gaps in our knowledge regarding the exact mechanisms and genetic basis of psoriasis. To investigate the pathogenesis of psoriasis, gene expression in 10 skin (5 lesional, 5 nonlesional) and 11 blood (6 psoriatic, 5 nonpsoriatic) samples were examined using Affymetrix HG-U95A microarrays. We detected 535 (425 upregulated, 110 downregulated) DEGs in lesional skin at 1% false discovery rate (FDR). Combining nine microarray studies comparing lesional and nonlesional psoriatic skin, 34.5% of dysregulated genes were overlapped in multiple studies. We further identified 20 skin and 2 blood associated transcriptional "hot spots" at specified genomic locations. At 5% FDR, 11.8% skin and 10.4% blood DEGs in our study mapped to one of the 12 PSORS loci. DEGs that overlap with PSORS loci may offer prioritized targets for downstream genetic fine mapping studies. Novel DEG "hot spots" may provide new targets for defining susceptibility loci in future studies. Copyright © 2012 Elsevier Inc. All rights reserved.

  7. In vitro differentiation of human umbilical cord blood mesenchymal ...

    African Journals Online (AJOL)

    May H. Hasan

    2016-08-05

    Aug 5, 2016 ... c Experimental Biology, Urology & Nephrology Center, Mansoura University, ... Liver is dedicated to perform an extensive range of tissue speci- ..... tors to repair hepatic injury.29 Adult human bone marrow .... regeneration.

  8. Study of Engraftment of human cord blood cells to rescue the sublethal radiation damage mice

    International Nuclear Information System (INIS)

    Cao Xiangshan; Zou Zhenghui; Yu Fei; Zhang Zhilin; Lin Baojue

    1997-01-01

    To investigate alternative source of hematopoiesis stem cells to rescue the sublethal radiation damage (SRD) casualties. Human-umbilical cord blood hematopoietic cells were transplanted into SRD mice, the survival rate and the hematopoiesis reconstitution of bone marrow were assessed. The survival rate, in the mice transplanted and the untransplanted, were 90% and 10% respectively. Bone marrow and spleen of survival mice showed human leukocytic antigen CD45 + cells. Presence of multilineage engraftment, including myeloid and erythroid lineages, were found indicating that immature human cells home to the mouse bone marrow. conclusion: engraftment of umbilical cord blood cells is very useful to reconstitute hematopoiesis of SRD casualties. As cord blood has many advantages over bone marrow and peripheral blood, it is important in rescuing radiation accidental casualties

  9. Transcriptional activity of telomerase complex in CD34- stem cells of cord blood in dependence of preparation time.

    Directory of Open Access Journals (Sweden)

    M Bojdys-Szyndlar

    2009-12-01

    Full Text Available The aim of the study was to determine whether the expression of telomerase subunits encoding genes changes during the process of cord blood preparation. It should establish if the commonly accepted 24 hours time interval in stem cells kriopreservation procedure significantly influences their immortalization and so decreases the "quality" of cord blood stem cells. Investigation includes 69 women. Spontaneous labour was the inclusion condition. The material was collected at birth after clamping of umbilical cord by direct vasopuncture. CD34- cells were extracted from cord blood (MACS, Miltenyi Biotec; Bisley, Surrey, UK. The expression profile of telomerase activators and inhibitors encoding genes was determined using HG_U133A oligonucleotide microarray (Affymetrix. We used a real-time quantitative RT-PCR assay to quantify the telomerase TERT, hTR and TP1 subunits mRNA copy numbers in CD34- cells in 0, 6, 12 and 24 hours after cord blood collection. We observed significant decrease of numbers of copies of TERTA+B mRNA within the successive hours of observation. Significant decrease of numbers of TERTA mRNA copies was confirmed after 24 hours. However, we observed significant increase of numbers of copies of TERTB mRNA after 6 hours of observation. Similar level was maintained during another 6h. The significantly lower number of copies of TERTB mRNA was observed after 24h. We also observed significant increase of number of copies of TERT mRNA after 6 hours. Number of copies of TERT mRNA significantly decreased after another 6h, remaining, however, on a higher then initial one. The significant lower number of copies of TERT mRNA was observed 24h after delivery. The possible explanation of those results is discussed in the paper.

  10. Biochemical analysis of CTLA-4 immunoreactive material from human blood

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    Dennert Kate

    2009-09-01

    Full Text Available Abstract Background CTLA-4 was initially described as a membrane-bound molecule that inhibited lymphocyte activation by interacting with B7.1 and B7.2 molecules on antigen presenting cells. Alternative splicing of mRNA encoding the CTLA-4 receptor leads to the production of a molecule (sCTLA-4 that lacks a membrane anchor and is therefore secreted into the extracellular space. Despite studies finding that people with autoimmune disease more frequently express high levels of sCTLA-4 in their blood than apparently healthy people, the significance of these findings is unclear. Methods Molecules isolated from blood using CTLA-4 specific antibodies were analyzed with ligand binding assays, mass spectroscopy, and biochemical fractionation in an effort to increase our understanding of CTLA-4 immunoreactive material. Results Mass spectroscopy analysis of the molecules recognized by multiple CTLA-4-specific antibodies failed to identify any CTLA-4 protein. Even though these molecules bind to the CTLA-4 receptors B7.1 and B7.2, they also exhibit properties common to immunoglobulins. Conclusion We have identified molecules in blood that are recognized by CTLA-4 specific antibodies but also exhibit properties of immunoglobulins. Our data indicates that what has been called sCTLA-4 is not a direct product of the CTLA-4 gene, and that the CTLA-4 protein is not part of this molecule. These results may explain why the relationship of sCTLA-4 to immune system activity has been difficult to elucidate.

  11. Turnover of adenosine in plasma of human and dog blood

    International Nuclear Information System (INIS)

    Moeser, G.H.S.; Schrader, J.; Deussen, A.

    1989-01-01

    To determine half-life and turnover of plasma adenosine, heparinized blood from healthy volunteers was incubated with radiolabeled adenosine in the physiological concentration range of 0.1-1 microM. Plasma levels of adenosine in vitro were 82 +/- 14 nM and were similar to those determined immediately after blood collection with a ''stopping solution.'' Dipyridamole (83 microM) and erythro-9(2-hydroxynon-3yl)-adenine (EHNA) (8 microM) did not measurably alter basal adenosine levels but completely blocked the uptake of added adenosine. Inhibition of ecto-5'-nucleotidase with 100 microM alpha, beta-methyleneadenosine 5'-diphosphate (AOPCP) reduced plasma adenosine to 22 +/- 6 nM. For the determination of adenosine turnover, the decrease in specific radioactivity of added [ 3 H]adenosine was measured using a dipyridamole-containing stopping solution. Without altering basal adenosine levels, the half-life was estimated to be 0.6 s. Similar experiments were carried out with washed erythrocytes or in the presence of AOPCP, yielding half-lives of 0.7 and 0.9 s, respectively. When the initial adenosine concentration was 1 microM, its specific activity decreased by only 11% within 5 s, whereas total plasma adenosine exponentially decreased with a half-life of 1.5 s. Venous plasma concentrations were measured after relief of a 3-min forearm ischemia. Changes in plasma adenosine did not correlate well with changes in blood flow but were augmented in the presence of dipyridamole

  12. Degradation and Stabilization of Peptide Hormones in Human Blood Specimens.

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    Jizu Yi

    Full Text Available Plasma hormone peptides, including GLP-1, GIP, Glucagon, and OXM, possess multiple physiological roles and potential therapeutic and diagnostic utility as biomarkers in the research of metabolic disorders. These peptides are subject to proteolytic degradation causing preanalytical variations. Stabilization for accurate quantitation of these active peptides in ex vivo blood specimens is essential for drug and biomarker development. We investigated the protease-driven instability of these peptides in conventional serum, plasma, anticoagulated whole blood, as well as whole blood and plasma stabilized with protease inhibitors. The peptide was monitored by both time-course Matrix-Assisted Laser Desorption Ionization Time-to-Flight Mass Spectrometry (MALDI -TOF MS and Ab-based assay (ELISA or RIA. MS enabled the identification of proteolytic fragments. In non-stabilized blood samples, the results clearly indicated that dipeptidyl peptidase-IV (DPP-IV removed the N-terminal two amino acid residues from GLP-1, GIP and OXM(1-37 and not-yet identified peptidase(s cleave(s the full-length OXM(1-37 and its fragments. DPP-IV also continued to remove two additional N-terminal residues of processed OXM(3-37 to yield OXM(5-37. Importantly, both DPP-IV and other peptidase(s activities were inhibited efficiently by the protease inhibitors included in the BD P800* tube. There was preservation of GLP-1, GIP, OXM and glucagon in the P800 plasma samples with half-lives > 96, 96, 72, and 45 hours at room temperature (RT, respectively. In the BD P700* plasma samples, the stabilization of GLP-1 was also achieved with half-life > 96 hours at RT. The stabilization of these variable peptides increased their utility in drug and/or biomarker development. While stability results of GLP-1 obtained with Ab-based assay were consistent with those obtained by MS analysis, the Ab-based results of GIP, Glucagon, and OXM did not reflect the time-dependent degradations revealed by MS

  13. Assessment of in vitro radiosensitivity of human peripheral blood lymphocytes

    International Nuclear Information System (INIS)

    Knox, S.J.; Shifrine, M.; Rosenblatt, L.S.

    1980-01-01

    The proliferative capacity of sensitive lymphocyte progenitor cells, from thirty-one clinically normal adults, was evaluated following in vitro x-irradiation (0-400R). Radiation effects were studied using both whole blood and lymphocyte-enriched mononuclear cell fractions in the lymphocyte stimulation test and colony formation assay with 6 different mitogens and antigens. Radiation dose-response survival curves were determined for the different test groups. The sensitivity of the different assay systems is compared and normative values are presented that may be used for comparison purposes to determine the relative radiosensitivity of atypical individuals and groups of individuals

  14. An in vitro study of the effects of low-level laser radiation on human blood

    Science.gov (United States)

    Siposan, Dan G.; Lukacs, Adalbert

    2003-12-01

    In the last time the study of the effects of LLLR on the blood is considered to be a subject of great importance in elucidating the mechanisms of action between LLLR and biologic tissues. Different methods of therapy by blood irradiation have been developed and used in clinical purposes with benefic effects. This study investigates some in vitro effects of LLLR on some selected rheologic indices of human blood. After establishing whether or not damaging effects could appear due to laser irradiation of the blood, we tried to find a new method for rejuvenating the blood preserved in MacoPharma-type bags. Blood samples were obtained from adult regular donors (volunteers). HeNe laser and laser diodes were used as radiation source, in a wide range of wavelengths, power densities, doses and other parameters of irradiation protocol. In the first series of experiments we established that LLLR does not alter the fresh blood from healthy donors, for doses between 0 and 10 J/cm3 and power densities between 30 and 180 mW/cm3. In the second series of experiments we established that LLLR does have, in some specific conditions, a revitalizing effect on the erythrocytes in preserved blood. We concluded that laser irradiation of the preserved blood, following a selected protocol of irradiation, could be used as a new method to improve the performances of preservation: prolonging the period of storage and blood rejuvenation before transfusion.

  15. The application of silicon sol-gel technology to forensic blood substitute development: Mimicking aspects of whole human blood rheology.

    Science.gov (United States)

    Stotesbury, Theresa; Illes, Mike; Wilson, Paul; Vreugdenhil, Andrew J

    2017-01-01

    Solution-gelation chemistry has promising applications in forensic synthetic blood substitute development. This research offers a silicon-based sol-gel approach to creating stable materials that share similar rheological properties to that of whole human blood samples. Room temperature, high water content, silicon sol-gels were created using the organosilane precursors 3-glycidoxypropyltrimethoxysilane and tetraethylorthosilicate along with various concentrations of filler and pigment. Shear-thinning non-Newtonian properties were observed within most formulations of the presented materials. The effects of colloidal concentration, temperature, age and filler addition on the viscosity of the sol-gels were investigated. SEM-EDS analysis was used to identify the behavior of the fillers within the film and support their inclusion for basic bloodstain pattern simulation. A final proposed candidate sol-gel was assessed using a previously reported passive drip simulation test on a hard, dry surface and passed. This works represents encouraging development in providing safe material alternatives to using whole human blood for forensic training and research. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  16. Noninvasive MRI measurement of the absolute cerebral blood volume-cerebral blood flow relationship during visual stimulation in healthy humans.

    Science.gov (United States)

    Ciris, Pelin Aksit; Qiu, Maolin; Constable, R Todd

    2014-09-01

    The relationship between cerebral blood volume (CBV) and cerebral blood flow (CBF) underlies blood oxygenation level-dependent functional MRI signal. This study investigates the potential for improved characterization of the CBV-CBF relationship in humans, and examines sex effects as well as spatial variations in the CBV-CBF relationship. Healthy subjects were imaged noninvasively at rest and during visual stimulation, constituting the first MRI measurement of the absolute CBV-CBF relationship in humans with complete coverage of the functional areas of interest. CBV and CBF estimates were consistent with the literature, and their relationship varied both spatially and with sex. In a region of interest with stimulus-induced activation in CBV and CBF at a significance level of the P < 0.05, a power function fit resulted in CBV = 2.1 CBF(0.32) across all subjects, CBV = 0.8 CBF(0.51) in females and CBV = 4.4 CBF(0.15) in males. Exponents decreased in both sexes as ROIs were expanded to include less significantly activated regions. Consideration for potential sex-related differences, as well as regional variations under a range of physiological states, may reconcile some of the variation across literature and advance our understanding of the underlying cerebrovascular physiology. Copyright © 2013 Wiley Periodicals, Inc.

  17. A Premature Termination of Human Epidermal Growth Factor Receptor Transcription in Escherichia coli

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    Jihene Elloumi-Mseddi

    2014-01-01

    Full Text Available Our success in producing an active epidermal growth factor receptor (EGFR tyrosine kinase in Escherichia coli encouraged us to express the full-length receptor in the same host. Despite its large size, we were successful at producing the full-length EGFR protein fused to glutathione S-transferase (GST that was detected by Western blot analysis. Moreover, we obtained a majoritarian truncated GST-EGFR form detectable by gel electrophoresis and Western blot. This truncated protein was purified and confirmed by MALDI-TOF/TOF analysis to belong to the N-terminal extracellular region of the EGFR fused to GST. Northern blot analysis showed two transcripts suggesting the occurrence of a transcriptional arrest.

  18. Genome-wide transcriptional reorganization associated with senescence-to-immortality switch during human hepatocellular carcinogenesis.

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    Gokhan Yildiz

    Full Text Available Senescence is a permanent proliferation arrest in response to cell stress such as DNA damage. It contributes strongly to tissue aging and serves as a major barrier against tumor development. Most tumor cells are believed to bypass the senescence barrier (become "immortal" by inactivating growth control genes such as TP53 and CDKN2A. They also reactivate telomerase reverse transcriptase. Senescence-to-immortality transition is accompanied by major phenotypic and biochemical changes mediated by genome-wide transcriptional modifications. This appears to happen during hepatocellular carcinoma (HCC development in patients with liver cirrhosis, however, the accompanying transcriptional changes are virtually unknown. We investigated genome-wide transcriptional changes related to the senescence-to-immortality switch during hepatocellular carcinogenesis. Initially, we performed transcriptome analysis of senescent and immortal clones of Huh7 HCC cell line, and identified genes with significant differential expression to establish a senescence-related gene list. Through the analysis of senescence-related gene expression in different liver tissues we showed that cirrhosis and HCC display expression patterns compatible with senescent and immortal phenotypes, respectively; dysplasia being a transitional state. Gene set enrichment analysis revealed that cirrhosis/senescence-associated genes were preferentially expressed in non-tumor tissues, less malignant tumors, and differentiated or senescent cells. In contrast, HCC/immortality genes were up-regulated in tumor tissues, or more malignant tumors and progenitor cells. In HCC tumors and immortal cells genes involved in DNA repair, cell cycle, telomere extension and branched chain amino acid metabolism were up-regulated, whereas genes involved in cell signaling, as well as in drug, lipid, retinoid and glycolytic metabolism were down-regulated. Based on these distinctive gene expression features we developed a 15

  19. Construction and analysis of the transcription factor-microRNA co-regulatory network response to Mycobacterium tuberculosis: a view from the blood.

    Science.gov (United States)

    Lin, Yan; Duan, Zipeng; Xu, Feng; Zhang, Jiayuan; Shulgina, Marina V; Li, Fan

    2017-01-01

    Mycobacterium tuberculosis ( Mtb ) infection has been regional outbreak, recently. The traditional focus on the patterns of "reductionism" which was associated with single molecular changes has been unable to meet the demand of early diagnosis and clinical application when current tuberculosis infection happened. In this study, we employed a systems biology approach to collect large microarray data sets including mRNAs and microRNAs (miRNAs) to identify the differentially expressed mRNAs and miRNAs in the whole blood of TB patients. The aim was to identify key genes associated with the immune response in the pathogenic process of tuberculosis by analyzing the co-regulatory network that was consisted of transcription factors and miRNAs as well as their target genes. The network along with their co-regulatory genes was analyzed utilizing Transcriptional Regulatory Element Database (TRED) and Database for Annotation, Visualization and Integrated Discovery (DAVID). We got 21 (19 up-regulated and 2 down-regulated) differentially expressed genes that were co-regulated by transcription factors and miRNAs. KEGG pathway enrichment analysis showed that the 21 differentially expressed genes were predominantly involved in Tuberculosis signaling pathway, which may play a major role in tuberculosis biological process. Quantitative real-time PCR was performed to verify the over expression of co-regulatory genes ( FCGR1A and CEBPB ). The genetic expression was correlated with clinicopathological characteristics in TB patients and inferences drawn. Our results suggest the TF-miRNA gene co-regulatory network may help us further understand the molecular mechanism of immune response to tuberculosis and provide us a new angle of future biomarker and therapeutic targets.

  20. Collaborating functions of BLM and DNA topoisomerase I in regulating human rDNA transcription

    International Nuclear Information System (INIS)

    Grierson, Patrick M.; Acharya, Samir; Groden, Joanna

    2013-01-01

    Bloom's syndrome (BS) is an inherited disorder caused by loss of function of the recQ-like BLM helicase. It is characterized clinically by severe growth retardation and cancer predisposition. BLM localizes to PML nuclear bodies and to the nucleolus; its deficiency results in increased intra- and inter-chromosomal recombination, including hyper-recombination of rDNA repeats. Our previous work has shown that BLM facilitates RNA polymerase I-mediated rRNA transcription in the nucleolus (Grierson et al., 2012 [18]). This study uses protein co-immunoprecipitation and in vitro transcription/translation (IVTT) to identify a direct interaction of DNA topoisomerase I with the C-terminus of BLM in the nucleolus. In vitro helicase assays demonstrate that DNA topoisomerase I stimulates BLM helicase activity on a nucleolar-relevant RNA:DNA hybrid, but has an insignificant effect on BLM helicase activity on a control DNA:DNA duplex substrate. Reciprocally, BLM enhances the DNA relaxation activity of DNA topoisomerase I on supercoiled DNA substrates. Our study suggests that BLM and DNA topoisomerase I function coordinately to modulate RNA:DNA hybrid formation as well as relaxation of DNA supercoils in the context of nucleolar transcription

  1. Collaborating functions of BLM and DNA topoisomerase I in regulating human rDNA transcription

    Energy Technology Data Exchange (ETDEWEB)

    Grierson, Patrick M. [Department of Microbiology, Immunology and Medical Genetics, The Ohio State University College of Medicine, Columbus, OH 43210 (United States); Acharya, Samir, E-mail: samir.acharya@osumc.edu [Department of Microbiology, Immunology and Medical Genetics, The Ohio State University College of Medicine, Columbus, OH 43210 (United States); Groden, Joanna [Department of Microbiology, Immunology and Medical Genetics, The Ohio State University College of Medicine, Columbus, OH 43210 (United States)

    2013-03-15

    Bloom's syndrome (BS) is an inherited disorder caused by loss of function of the recQ-like BLM helicase. It is characterized clinically by severe growth retardation and cancer predisposition. BLM localizes to PML nuclear bodies and to the nucleolus; its deficiency results in increased intra- and inter-chromosomal recombination, including hyper-recombination of rDNA repeats. Our previous work has shown that BLM facilitates RNA polymerase I-mediated rRNA transcription in the nucleolus (Grierson et al., 2012 [18]). This study uses protein co-immunoprecipitation and in vitro transcription/translation (IVTT) to identify a direct interaction of DNA topoisomerase I with the C-terminus of BLM in the nucleolus. In vitro helicase assays demonstrate that DNA topoisomerase I stimulates BLM helicase activity on a nucleolar-relevant RNA:DNA hybrid, but has an insignificant effect on BLM helicase activity on a control DNA:DNA duplex substrate. Reciprocally, BLM enhances the DNA relaxation activity of DNA topoisomerase I on supercoiled DNA substrates. Our study suggests that BLM and DNA topoisomerase I function coordinately to modulate RNA:DNA hybrid formation as well as relaxation of DNA supercoils in the context of nucleolar transcription.

  2. Nucleotide sequence of the gene coding for human factor VII, a vitamin K-dependent protein participating in blood coagulation

    International Nuclear Information System (INIS)

    O'Hara, P.J.; Grant, F.J.; Haldeman, B.A.; Gray, C.L.; Insley, M.Y.; Hagen, F.S.; Murray, M.J.

    1987-01-01

    Activated factor VII (factor VIIa) is a vitamin K-dependent plasma serine protease that participates in a cascade of reactions leading to the coagulation of blood. Two overlapping genomic clones containing sequences encoding human factor VII were isolated and characterized. The complete sequence of the gene was determined and found to span about 12.8 kilobases. The mRNA for factor VII as demonstrated by cDNA cloning is polyadenylylated at multiple sites but contains only one AAUAAA poly(A) signal sequence. The mRNA can undergo alternative splicing, forming one transcript containing eight segments as exons and another with an additional exon that encodes a larger prepro leader sequence. The latter transcript has no known counterpart in the other vitamin K-dependent proteins. The positions of the introns with respect to the amino acid sequence encoded by the eight essential exons of factor VII are the same as those present in factor IX, factor X, protein C, and the first three exons of prothrombin. These exons code for domains generally conserved among members of this gene family. The comparable introns in these genes, however, are dissimilar with respect to size and sequence, with the exception of intron C in factor VII and protein C. The gene for factor VII also contains five regions made up of tandem repeats of oligonucleotide monomer elements. More than a quarter of the intron sequences and more than a third of the 3' untranslated portion of the mRNA transcript consist of these minisatellite tandem repeats

  3. Multiple correlation analyses revealed complex relationship between DNA methylation and mRNA expression in human peripheral blood mononuclear cells.

    Science.gov (United States)

    Xie, Fang-Fei; Deng, Fei-Yan; Wu, Long-Fei; Mo, Xing-Bo; Zhu, Hong; Wu, Jian; Guo, Yu-Fan; Zeng, Ke-Qin; Wang, Ming-Jun; Zhu, Xiao-Wei; Xia, Wei; Wang, Lan; He, Pei; Bing, Peng-Fei; Lu, Xin; Zhang, Yong-Hong; Lei, Shu-Feng

    2018-01-01

    DNA methylation is an important regulator on the mRNA expression. However, a genome-wide correlation pattern between DNA methylation and mRNA expression in human peripheral blood mononuclear cells (PBMCs) is largely unknown. The comprehensive relationship between mRNA and DNA methylation was explored by using four types of correlation analyses and a genome-wide methylation-mRNA expression quantitative trait locus (eQTL) analysis in PBMCs in 46 unrelated female subjects. An enrichment analysis was performed to detect biological function for the detected genes. Single pair correlation coefficient (r T1 ) between methylation level and mRNA is moderate (-0.63-0.62) in intensity, and the negative and positive correlations are nearly equal in quantity. Correlation analysis on each gene (T4) found 60.1% genes showed correlations between mRNA and gene-based methylation at P correlation (R T4  > 0.8). Methylation sites have regulation effects on mRNA expression in eQTL analysis, with more often observations in region of transcription start site (TSS). The genes under significant methylation regulation both in correlation analysis and eQTL analysis tend to cluster to the categories (e.g., transcription, translation, regulation of transcription) that are essential for maintaining the basic life activities of cells. Our findings indicated that DNA methylation has predictive regulation effect on mRNA with a very complex pattern in PBMCs. The results increased our understanding on correlation of methylation and mRNA and also provided useful clues for future epigenetic studies in exploring biological and disease-related regulatory mechanisms in PBMC.

  4. Phenobarbital induces cell cycle transcriptional responses in mouse liver humanized for constitutive androstane and pregnane x receptors.

    Science.gov (United States)

    Luisier, Raphaëlle; Lempiäinen, Harri; Scherbichler, Nina; Braeuning, Albert; Geissler, Miriam; Dubost, Valerie; Müller, Arne; Scheer, Nico; Chibout, Salah-Dine; Hara, Hisanori; Picard, Frank; Theil, Diethilde; Couttet, Philippe; Vitobello, Antonio; Grenet, Olivier; Grasl-Kraupp, Bettina; Ellinger-Ziegelbauer, Heidrun; Thomson, John P; Meehan, Richard R; Elcombe, Clifford R; Henderson, Colin J; Wolf, C Roland; Schwarz, Michael; Moulin, Pierre; Terranova, Rémi; Moggs, Jonathan G

    2014-06-01

    The constitutive androstane receptor (CAR) and the pregnane X receptor (PXR) are closely related nuclear receptors involved in drug metabolism and play important roles in the mechanism of phenobarbital (PB)-induced rodent nongenotoxic hepatocarcinogenesis. Here, we have used a humanized CAR/PXR mouse model to examine potential species differences in receptor-dependent mechanisms underlying liver tissue molecular responses to PB. Early and late transcriptomic responses to sustained PB exposure were investigated in liver tissue from double knock-out CAR and PXR (CAR(KO)-PXR(KO)), double humanized CAR and PXR (CAR(h)-PXR(h)), and wild-type C57BL/6 mice. Wild-type and CAR(h)-PXR(h) mouse livers exhibited temporally and quantitatively similar transcriptional responses during 91 days of PB exposure including the sustained induction of the xenobiotic response gene Cyp2b10, the Wnt signaling inhibitor Wisp1, and noncoding RNA biomarkers from the Dlk1-Dio3 locus. Transient induction of DNA replication (Hells, Mcm6, and Esco2) and mitotic genes (Ccnb2, Cdc20, and Cdk1) and the proliferation-related nuclear antigen Mki67 were observed with peak expression occurring between 1 and 7 days PB exposure. All these transcriptional responses were absent in CAR(KO)-PXR(KO) mouse livers and largely reversible in wild-type and CAR(h)-PXR(h) mouse livers following 91 days of PB exposure and a subsequent 4-week recovery period. Furthermore, PB-mediated upregulation of the noncoding RNA Meg3, which has recently been associated with cellular pluripotency, exhibited a similar dose response and perivenous hepatocyte-specific localization in both wild-type and CAR(h)-PXR(h) mice. Thus, mouse livers coexpressing human CAR and PXR support both the xenobiotic metabolizing and the proliferative transcriptional responses following exposure to PB.

  5. Proneural transcription factor Atoh1 drives highly efficient differentiation of human pluripotent stem cells into dopaminergic neurons.

    Science.gov (United States)

    Sagal, Jonathan; Zhan, Xiping; Xu, Jinchong; Tilghman, Jessica; Karuppagounder, Senthilkumar S; Chen, Li; Dawson, Valina L; Dawson, Ted M; Laterra, John; Ying, Mingyao

    2014-08-01

    Human pluripotent stem cells (PSCs) are a promising cell resource for various applications in regenerative medicine. Highly efficient approaches that differentiate human PSCs into functional lineage-specific neurons are critical for modeling neurological disorders and testing potential therapies. Proneural transcription factors are crucial drivers of neuron development and hold promise for driving highly efficient neuronal conversion in PSCs. Here, we study the functions of proneural transcription factor Atoh1 in the neuronal differentiation of PSCs. We show that Atoh1 is induced during the neuronal conversion of PSCs and that ectopic Atoh1 expression is sufficient to drive PSCs into neurons with high efficiency. Atoh1 induction, in combination with cell extrinsic factors, differentiates PSCs into functional dopaminergic (DA) neurons with >80% purity. Atoh1-induced DA neurons recapitulate key biochemical and electrophysiological features of midbrain DA neurons, the degeneration of which is responsible for clinical symptoms in Parkinson's disease (PD). Atoh1-induced DA neurons provide a reliable disease model for studying PD pathogenesis, such as neurotoxin-induced neurodegeneration in PD. Overall, our results determine the role of Atoh1 in regulating neuronal differentiation and neuron subtype specification of human PSCs. Our Atoh1-mediated differentiation approach will enable large-scale applications of PD patient-derived midbrain DA neurons in mechanistic studies and drug screening for both familial and sporadic PD. ©AlphaMed Press.

  6. Molecular cloning of the human gene for von Willebrand factor and identification of the transcription initiation site

    International Nuclear Information System (INIS)

    Collins, C.J.; Underdahl, J.P.; Levene, R.B.; Ravera, C.P.; Morin, M.J.; Dombalagian, M.J.; Ricca, G.; Livingston, D.M.; Lynch, D.C.

    1987-01-01

    A series of overlapping cosmid genomic clones have been isolated that contain the entire coding unit of the human gene for van Willebrand factor (vWf), a major component of the hemostatic system. The cloned segments span ≅ 175 kilobases of human DNA sequence, and hybridization analysis suggest that the vWf coding unit is ≅150 kilobases in length. Within one of these clones, the vWF transcription initiation site has been mapped and a portion of the vWf promoter region has been sequenced, revealing a typical TATA box, a downstream CCAAT box, and a perfect downstream repeat of the 8 base pairs containing the transcription start site. Sequencing of a segment of another genomic clone has revealed the vWF translation termination codon. Where tested, comparative restriction analysis of cloned and chromosomal DNA segments strongly suggests that no major alterations occurred during cloning and that there is only one complete copy of the vWf gene in the human haploid genome. Similar analyses of DNA from vWf-producing endothelial cells and nonexpressing leukocytes suggest that vWf gene expression is not accompanied by gross genomic rearrangements. In addition, there is significant homology of C-terminal coding sequences among the vWf genes of several vertebrate species

  7. TRIM45, a novel human RBCC/TRIM protein, inhibits transcriptional activities of ElK-1 and AP-1

    International Nuclear Information System (INIS)

    Wang Yuequn; Li Yongqing; Qi Xinzhu; Yuan Wuzhou; Ai Jianping; Zhu Chuanbing; Cao Lei; Yang Hong; Liu Fang; Wu Xiushan; Liu Mingyao

    2004-01-01

    The tripartite motif (TRIM) proteins play important roles in a variety of cellular functions including cell proliferation, differentiation, development, oncogenesis, and apoptosis. In this study, we report the identification and characterization of the human tripartite motif-containing protein 45 (TRIM45), a novel member of the TRIM family, from a human embryonic heart cDNA library. TRIM45 has a predicted 580 amino acid open reading frame, encoding a putative 64-kDa protein. The N-terminal region harbors a RING finger, two B-boxes, and a predicted α-helical coiled-coil domain, which together form the RBCC/TRIM motif found in a large family of proteins, whereas the C-terminal region contains a filamin-type immunoglobulin (IG-FLMN) domain. Northern blot analysis indicates that TRIM45 is expressed in a variety of human adult and embryonic tissues. In the cell, TRIM45 protein is expressed both in cytoplasm and in cell nucleus. Overexpression of TRIM45 in COS-7 cells inhibits the transcriptional activities of ElK-1 and AP-1. These results suggest that TRIM45 may act as a new transcriptional repressor in mitogen-activated protein kinase signaling pathway

  8. A synthetic interaction screen identifies factors selectively required for proliferation and TERT transcription in p53-deficient human cancer cells.

    Directory of Open Access Journals (Sweden)

    Li Xie

    Full Text Available Numerous genetic and epigenetic alterations render cancer cells selectively dependent on specific genes and regulatory pathways, and represent potential vulnerabilities that can be therapeutically exploited. Here we describe an RNA interference (RNAi-based synthetic interaction screen to identify genes preferentially required for proliferation of p53-deficient (p53- human cancer cells. We find that compared to p53-competent (p53+ human cancer cell lines, diverse p53- human cancer cell lines are preferentially sensitive to loss of the transcription factor ETV1 and the DNA damage kinase ATR. In p53- cells, RNAi-mediated knockdown of ETV1 or ATR results in decreased expression of the telomerase catalytic subunit TERT leading to growth arrest, which can be reversed by ectopic TERT expression. Chromatin immunoprecipitation analysis reveals that ETV1 binds to a region downstream of the TERT transcriptional start-site in p53- but not p53+ cells. We find that the role of ATR is to phosphorylate and thereby stabilize ETV1. Our collective results identify a regulatory pathway involving ETV1, ATR, and TERT that is preferentially important for proliferation of diverse p53- cancer cells.

  9. Transcriptional Response of Human Cells to Microbeam Irradiation with 2.1 MeV Alpha Particles

    Science.gov (United States)

    Hellweg, C. E.; Bogner, S.; Spitta, L.; Arenz, A.; Baumstark-Khan, C.; Greif, K. D.; Giesen, U.

    Within the next decades an increasing number of human beings in space will be simultaneously exposed to different stimuli especially microgravity and radiation To assess the risks for humans during long-duration space missions the complex interplay of these parameters at the cellular level must be understood Cellular stress protection responses lead to increased transcription of several genes via modulation of transcription factors Activation of the Nuclear Factor kappa B NF- kappa B pathway as a possible anti-apoptotic route represents such an important cellular stress response A screening assay for detection of NF- kappa B-dependent gene activation using the destabilized variant of Enhanced Green Fluorescent Protein d2EGFP as reporter protein had been developed It consists of Human Embryonic Kidney HEK 293 Cells stably transfected with a receptor-reporter-construct carrying d2EGFP under the control of a NF- kappa B response element Clones positive for Tumor Necrosis Factor alpha TNF- alpha inducible d2EGFP expression were selected as cellular reporters Irradiation was performed either with X-rays 150 kV 19 mA at DLR Cologne or with 2 1 MeV alpha particles LET sim 160 keV mu m at PTB Braunschweig After irradiation the following biological endpoints were determined i cell survival via the colony forming ability test ii time-dependent activation of NF- kappa B dependent d2EGFP gene expression using flow cytometry iii quantitative RT-PCR

  10. Analysis of cytotoxic effects of chlorhexidine gluconate as antiseptic agent on human blood lymphocytes.

    Science.gov (United States)

    Salimi, Ahmad; Alami, Bahare; Pourahmad, Jalal

    2017-08-01

    The aim of this study was to assess the cytotoxicity of chlorhexidine gluconate (CHG) on human blood lymphocytes as a useful ex vivo model for accelerated human toxicity studies. Using biochemical and flow cytometry assessments, we demonstrated that addition of CHG at 1 μM concentration to human blood lymphocytes induced cytotoxicity following 6 h. The CHG-induced cytotoxicity on human blood lymphocytes was associated with intracellular reactive oxygen species generation, mitochondrial membrane potential collapse, lysosomal membrane injury, lipid peroxidation, and depletion of glutathione. According to our results, CHG triggers oxidative stress and organelles damages in lymphocytes which are important cells in defense against foreign agents. Finally our findings suggest that using of antioxidants and mitochondrial/lysosomal protective agents could be of benefit for the people in the exposure with CHG. © 2017 Wiley Periodicals, Inc.

  11. [Inhibitory effect and mechanism of tofacitinib on the secretion of cytokines by T cells in human peripheral blood].

    Science.gov (United States)

    Wu, Kunlun; Zhao, Jun; Wu, Qiongli; Wu, Changyou

    2017-11-01

    Objective To study the inhibitory effect of tofacitinib on the production of cytokines by T cells in human peripheral blood and its mechanism. Methods Peripheral blood mononuclear cells (PBMCs) and purified T cells were cultured and stimulated with anti-CD3 plus anti-CD28 antibodies in the presence or absence of tofacitinib (0.5 μmol/L). The levels of interferon γ (IFN-γ), tumor necrosis factor α (TNF-α) and interleukin 2 (IL-2) in the culture supernatants were detected by ELISA, and the expressions of activated molecules CD69 and CD25 on the surface of CD4 + and CD8 + T cells, the production of cytokines and the phosphorylation of signal transducers and transcriptional activators STAT1, STAT3, STAT4 in T cells were examined by flow cytometry. At the same time, the proliferation and apoptosis of T cells were observed by 5- (and 6-) carboxyfluorescein diacetate succinimidyl ester (CFSE) staining and the flow cy tometry with annexin V-FITC/PI, respectively. Results Tofacitinib inhibited the production of IFN-γ, TNF-α and the expression of CD25 on T cells from the peripheral blood. In addition, the proliferation and the phosphorylation of STAT1, STAT3, STAT4 by T cells were also depressed. However, tofacitinib had no effect on the secretion of IL-2, the expression of CD69 and the apoptosis of T cells. Conclusion Tofacitinib can inhibit the secretion of IFN-γ and TNF-α by T cells in the peripheral blood, and its mechanism might be related to the inhibitory effect of tofacitinib on the activation, proliferation and signal transduction in T cells.

  12. Transcription Profiling Demonstrates Epigenetic Control of Non-retroviral RNA Virus-Derived Elements in the Human Genome

    Directory of Open Access Journals (Sweden)

    Kozue Sofuku

    2015-09-01

    Full Text Available Endogenous bornavirus-like nucleoprotein elements (EBLNs are DNA sequences in vertebrate genomes formed by the retrotransposon-mediated integration of ancient bornavirus sequence. Thus, EBLNs evidence a mechanism of retrotransposon-mediated RNA-to-DNA information flow from environment to animals. Although EBLNs are non-transposable, they share some features with retrotransposons. Here, to test whether hosts control the expression of EBLNs similarly to retrotransposons, we profiled the transcription of all Homo sapiens EBLNs (hsEBLN-1 to hsEBLN-7. We could detect transcription of all hsEBLNs in at least one tissue. Among them, hsEBLN-1 is transcribed almost exclusively in the testis. In most tissues, expression from the hsEBLN-1 locus is silenced epigenetically. Finally, we showed the possibility that hsEBLN-1 integration at this locus affects the expression of a neighboring gene. Our results suggest that hosts regulate the expression of endogenous non-retroviral virus elements similarly to how they regulate the expression of retrotransposons, possibly contributing to new transcripts and regulatory complexity to the human genome.

  13. Rearranged EML4-ALK fusion transcripts sequester in circulating blood platelets and enable blood-based crizotinib response monitoring in non-small-cell lung cancer

    Science.gov (United States)

    Nilsson, R. Jonas A.; Karachaliou, Niki; Berenguer, Jordi; Gimenez-Capitan, Ana; Schellen, Pepijn; Teixido, Cristina; Tannous, Jihane; Kuiper, Justine L.; Drees, Esther; Grabowska, Magda; van Keulen, Marte; Heideman, Danielle A.M.; Thunnissen, Erik; Dingemans, Anne-Marie C.; Viteri, Santiago; Tannous, Bakhos A.; Drozdowskyj, Ana; Rosell, Rafael; Smit, Egbert F.; Wurdinger, Thomas

    2016-01-01

    Purpose: Non-small-cell lung cancers harboring EML4-ALK rearrangements are sensitive to crizotinib. However, despite initial response, most patients will eventually relapse, and monitoring EML4-ALK rearrangements over the course of treatment may help identify these patients. However, challenges associated with serial tumor biopsies have highlighted the need for blood-based assays for the monitoring of biomarkers. Platelets can sequester RNA released by tumor cells and are thus an attractive source for the non-invasive assessment of biomarkers. Methods: EML4-ALK rearrangements were analyzed by RT-PCR in platelets and plasma isolated from blood obtained from 77 patients with non-small-cell lung cancer, 38 of whom had EML4-ALK-rearranged tumors. In a subset of 29 patients with EML4-ALK-rearranged tumors who were treated with crizotinib, EML4-ALK rearrangements in platelets were correlated with progression-free and overall survival. Results: RT-PCR demonstrated 65% sensitivity and 100% specificity for the detection of EML4-ALK rearrangements in platelets. In the subset of 29 patients treated with crizotinib, progression-free survival was 3.7 months for patients with EML4-ALK+ platelets and 16 months for those with EML4-ALK− platelets (hazard ratio, 3.5; P = 0.02). Monitoring of EML4-ALK rearrangements in the platelets of one patient over a period of 30 months revealed crizotinib resistance two months prior to radiographic disease progression. Conclusions: Platelets are a valuable source for the non-invasive detection of EML4-ALK rearrangements and may prove useful for predicting and monitoring outcome to crizotinib, thereby improving clinical decisions based on radiographic imaging alone. PMID:26544515

  14. Identification of NR4A2 as a transcriptional activator of IL-8 expression in human inflammatory arthritis.

    LENUS (Irish Health Repository)

    Aherne, Carol M

    2009-10-01

    Expression of the orphan nuclear receptor NR4A2 is controlled by pro-inflammatory mediators, suggesting that NR4A2 may contribute to pathological processes in the inflammatory lesion. This study identifies the chemoattractant protein, interleukin 8 (IL-8\\/CXCL8), as a molecular target of NR4A2 in human inflammatory arthritis and examines the mechanism through which NR4A2 modulates IL-8 expression. In TNF-alpha-activated human synoviocyte cells, enhanced expression of IL-8 mRNA and protein correspond to temporal changes in NR4A2 transcription and nuclear distribution. Ectopic expression of NR4A2 leads to robust changes in endogenous IL-8 mRNA levels and co-treatment with TNF-alpha results in significant (p<0.001) secretion of IL-8 protein. Transcriptional effects of NR4A2 on the human IL-8 promoter are enhanced in the presence of TNF-alpha, suggesting molecular crosstalk between TNF-alpha signalling and NR4A2. A dominant negative IkappaB kinase antagonizes the combined effects of NR4A2 and TNF-alpha on IL-8 promoter activity. Co-expression of NR4A2 and the p65 subunit of NF-kappaB enhances IL-8 transcription and functional studies indicate that transactivation occurs independently of NR4A2 binding to DNA or heterodimerization with additional nuclear receptors. The IL-8 minimal promoter region is sufficient to support NR4A2 and NF-kappaB\\/p65 co-operative activity and NR4A2 can interact with NF-kappaB\\/p65 on a 39bp sequence within this region. In patients treated with methotrexate for active inflammatory arthritis, a reduction in NR4A2 synovial tissue levels correlate significantly (n=10, r=0.73, p=0.002) with changes in IL-8 expression. Collectively, these data delineate an important role for NR4A2 in modulating IL-8 expression and reveal novel transcriptional responses to TNF-alpha in human inflammatory joint disease.

  15. Electron microscopic in situ hybridization and autoradiography: Localization and transcription of rDNA in human lymphocyte nucleoli

    International Nuclear Information System (INIS)

    Wachtler, F.; Mosgoeller, W.S.; Schwarzacher, H.G.

    1990-01-01

    The distribution of ribosomal DNA (rDNA) in the nucleoli of human lymphocytes was revealed by in situ hybridization with a nonautoradiographic procedure at the electron microscopic level. rDNA is located in the dense fibrillar component of the nucleolus but not in the fibrillar centers. In the same cells the incorporation of tritiated uridine takes place in the dense fibrillar component of the nucleolus as seen by autoradiography followed by gold latensification. From these findings it can be concluded that the transcription of ribosomal DNA takes place in the dense fibrillar component of the nucleolus

  16. Computed aided system for separation and classification of the abnormal erythrocytes in human blood

    Science.gov (United States)

    Wąsowicz, Michał; Grochowski, Michał; Kulka, Marek; Mikołajczyk, Agnieszka; Ficek, Mateusz; Karpieńko, Katarzyna; Cićkiewicz, Maciej

    2017-12-01

    The human peripheral blood consists of cells (red cells, white cells, and platelets) suspended in plasma. In the following research the team assessed an influence of nanodiamond particles on blood elements over various periods of time. The material used in the study consisted of samples taken from ten healthy humans of various age, different blood types and both sexes. The markings were leaded by adding to the blood unmodified diamonds and oxidation modified. The blood was put under an impact of two diamond concentrations: 20μl and 100μl. The amount of abnormal cells increased with time. The percentage of echinocytes as a result of interaction with nanodiamonds in various time intervals for individual specimens was scarce. The impact of the two diamond types had no clinical importance on red blood cells. It is supposed that as a result of longlasting exposure a dehydratation of red cells takes place, because of the function of the cells. The analysis of an influence of nanodiamond particles on blood elements was supported by computer system designed for automatic counting and classification of the Red Blood Cells (RBC). The system utilizes advanced image processing methods for RBCs separation and counting and Eigenfaces method coupled with the neural networks for RBCs classification into normal and abnormal cells purposes.

  17. Blood flow and microdialysis in the human femoral head

    DEFF Research Database (Denmark)

    Bøgehøj, Morten; Emmeluth, Claus; Overgaard, Søren

    2007-01-01

    BACKGROUND: If it would be possible to detect lack of flow and/or the development of ischemia in bone, we might have a way of predicting whether a broken bone will heal. We established microdialysis (MD) and laser Doppler (LD) flow measurement in the human femoral head in order to be able to detect...

  18. Effect of cholesterol and triglycerides levels on the rheological behavior of human blood

    Science.gov (United States)

    Moreno, Leonardo; Calderas, Fausto; Sanchez-Olivares, Guadalupe; Medina-Torres, Luis; Sanchez-Solis, Antonio; Manero, Octavio

    2015-02-01

    Important public health problems worldwide such as obesity, diabetes, hyperlipidemia and coronary diseases are quite common. These problems arise from numerous factors, such as hyper-caloric diets, sedentary habits and other epigenetic factors. With respect to Mexico, the population reference values of total cholesterol in plasma are around 200 mg/dL. However, a large proportion has higher levels than this reference value. In this work, we analyze the rheological properties of human blood obtained from 20 donors, as a function of cholesterol and triglyceride levels, upon a protocol previously approved by the health authorities. Samples with high and low cholesterol and triglyceride levels were selected and analyzed by simple-continuous and linear-oscillatory shear flow. Rheometric properties were measured and related to the structure and composition of human blood. In addition, rheometric data were modeled by using several constitutive equations: Bautista-Manero-Puig (BMP) and the multimodal Maxwell equations to predict the flow behavior of human blood. Finally, a comparison was made among various models, namely, the BMP, Carreau and Quemada equations for simple shear rate flow. An important relationship was found between cholesterol, triglycerides and the structure of human blood. Results show that blood with high cholesterol levels (400 mg/dL) has flow properties fully different (higher viscosity and a more pseudo-plastic behavior) than blood with lower levels of cholesterol (tendency to Newtonian behavior or viscosity plateau at low shear rates).

  19. Systems biology of coagulation initiation: kinetics of thrombin generation in resting and activated human blood.

    Directory of Open Access Journals (Sweden)

    Manash S Chatterjee

    2010-09-01

    Full Text Available Blood function defines bleeding and clotting risks and dictates approaches for clinical intervention. Independent of adding exogenous tissue factor (TF, human blood treated in vitro with corn trypsin inhibitor (CTI, to block Factor XIIa will generate thrombin after an initiation time (T(i of 1 to 2 hours (depending on donor, while activation of platelets with the GPVI-activator convulxin reduces T(i to ∼20 minutes. Since current kinetic models fail to generate thrombin in the absence of added TF, we implemented a Platelet-Plasma ODE model accounting for: the Hockin-Mann protease reaction network, thrombin-dependent display of platelet phosphatidylserine, VIIa function on activated platelets, XIIa and XIa generation and function, competitive thrombin substrates (fluorogenic detector and fibrinogen, and thrombin consumption during fibrin polymerization. The kinetic model consisting of 76 ordinary differential equations (76 species, 57 reactions, 105 kinetic parameters predicted the clotting of resting and convulxin-activated human blood as well as predicted T(i of human blood under 50 different initial conditions that titrated increasing levels of TF, Xa, Va, XIa, IXa, and VIIa. Experiments with combined anti-XI and anti-XII antibodies prevented thrombin production, demonstrating that a leak of XIIa past saturating amounts of CTI (and not "blood-borne TF" alone was responsible for in vitro initiation without added TF. Clotting was not blocked by antibodies used individually against TF, VII/VIIa, P-selectin, GPIb, protein disulfide isomerase, cathepsin G, nor blocked by the ribosome inhibitor puromycin, the Clk1 kinase inhibitor Tg003, or inhibited VIIa (VIIai. This is the first model to predict the observed behavior of CTI-treated human blood, either resting or stimulated with platelet activators. CTI-treated human blood will clot in vitro due to the combined activity of XIIa and XIa, a process enhanced by platelet activators and which proceeds

  20. Fiber-based optic sensor for detecting human blood clot: present and future revival

    Science.gov (United States)

    Elshikeri, Nada; Bakhtiar, Hazri

    2018-05-01

    Sustaining human’s life-frame away from being impeded by the clot - ghost term, we attempt to approach a mobile fiber-based optical sensor (f-s) for detecting blood clot in a blood vessel (intra-arteries/veins). Blood vessels are the part of the circulatory system that transport blood throughout the human body, thus their significance of being protected arise to the monograph focus. MRI (magnetic resonance imaging), X-rays and other medical instruments are diagnostic immobility techniques with a slackest interval. The corer causation of fiber-based optical sensor is to detect a clump of blood in the bloodstream by providing a prompt mobile diagnostic intervals preserving last-minutes-breath of human’s life. The detector (f-s) has been etched by diluting sulphuric acid ~10% at certain zone to sensate its function. The in-vitro monograph peaks its maximal monitoring when the sensor is attached to Raman Spectroscopy (RS) setup. RS quantifies the relative intensities of fibrinogen bond, which is the first type of blood coagulation elements of blood plasma. Blood coagulation parameters are the major concern of the monograph investigation, such as total haemoglobin (tHb), clotting reaction time (t), clot progression time (t2), maximum clot amplitude (ma) and mean refractive index (r). A blood sample will be drawn from the patient and after centrifugation to separate blood plasma from its constituents, then an immediate sloshing of blood plasma in the (f-s) packet which has its plug-in to RS. Estimating the quantitative analysis of blood sample concentration, RS will determine the presence of coagulation in terms of intensity and medical procedures will dominate the treatment process. Thus, the suggestive monograph provides a definite instrument for investigating blood coagulation intra-arteries/veins promptly.

  1. Human Blood Feeding Activity of Female Hybrids between Culex pipiens pipiens and Culex pipiens quinquefasciatus(Diptera: Culicidae)

    OpenAIRE

    Yoshii, Manabu; Mine, Mariko; Kurokawa, Kenji; Oda, Tsutomu; Kato, Katsutomo; Ogawa, Yasunori; Eshita, Yuuki; Uchida, Keikichi

    2007-01-01

    Human blood feeding activity was examined in females of hybrids between Culex pipiens pipiens and Culex pipiens quinquefasciatus during long photoperiod at 25℃. Blood feeding rates of hybrids were lower than in Culex pipiens quinquefasciatus and Culex pipiens pallens, and higher than in Culex pipiens pipiens, because no females fed on human blood in Culex pipiens pipiens.

  2. Polonium 210Po activities in human blood of patients with ischaemic heart disease from Gda?sk in Poland

    OpenAIRE

    Bory?o, Alicja; Skwarzec, Bogdan; Roma?czyk, Grzegorz; Siebert, Janusz

    2013-01-01

    The determination of polonium 210Po in human blood samples is presented and discussed in this paper. The human blood samples were collected from patients of Medical University of Gda?sk with ischaemic heart disease (morbus ischaemicus cordis, MIC). The polonium concentrations in analyzed human blood samples are very differentiated. 210Po is of particular interest in public health and although is present in the environment in extremely low amounts, it is easily bioaccumulated to the human body...

  3. T lymphocytes derived from human cord blood provide effective antitumor immunotherapy against a human tumor

    Directory of Open Access Journals (Sweden)

    Kim Tae-Sik

    2011-06-01

    Full Text Available Abstract Background Although the graft-versus-tumor (GVT effect of donor-derived T cells after allogeneic hematopoietic stem cell transplantation has been used as an effective adoptive immunotherapy, the antitumor effects of cord blood (CB transplantation have not been well studied. Methods We established the animal model by transplantation of CB mononuclear cells and/or tumor cells into NOD/SCID mice. The presence of CB derived T cells in NOD/SCID mice or tumor tissues were determined by flow cytometric and immunohistochemical analysis. The anti-tumor effects of CB derived T cells against tumor was determined by tumor size and weight, and by the cytotoxicity assay and ELISPOT assay of T cells. Results We found dramatic tumor remission following transfer of CB mononuclear cells into NOD/SCID mice with human cervical tumors with a high infiltration of CD3+ T cells in tumors. NOD/SCID mice that receive neonatal CB transplants have reconstituted T cells with significant antitumor effects against human cervical and lung tumors, with a high infiltration of CD3+ T cells showing dramatic induction of apoptotic cell death. We also confirmed that T cells showed tumor specific antigen cytotoxicity in vitro. In adoptive transfer of CD3+ T cells into mice with pre-established tumors, we observed much higher antitumor effects of HPV-specific T cells by ELISPOT assays. Conclusions Our results show that CB derived T lymphocytes will be useful for novel immunotherapeutic candidate cells for therapy of several tumors in clinic.

  4. Identification and transcription profiling of trypsin in Aedes taeniorhynchus (Diptera: Culicidae): developmental regulation, blood feeding, and permethrin exposure.

    Science.gov (United States)

    Zhao, Liming; Chen, Jian; Becnel, James J; Kline, Daniel L; Clark, Gary G; Linthicum, Kenneth J

    2011-05-01

    The cDNA of a trypsin gene from Aedes (Ochlerotatus) taeniorhynchus (Weidemann) was cloned and sequenced. The full-length mRNA sequence (890 bp) for trypsin from Ae. taeniorhynchus (AetTryp1) was obtained, which encodes an open reading frame of 765 bp (i.e., 255 amino acids). To detect whether AetTryp is developmentally regulated, a quantitative real-time polymerase chain reaction was used to examine AetTrypl mRNA expression levels in different developmental stages of Ae. taeniorhynchus. AetTryp1 was expressed at low levels in egg, larval, and pupal stages, but was differentially expressed in adult Ae. taeniorhynchus, with highest levels found in 5-d-old female adults when compared with teneral adults. In addition, AetTryp1 mRNA expression differed between sexes, with expression levels much lower in males. However, in both males and females, there was a significant increase in AetTryp1 transcription levels as age increased and peaked in 5-d-old adults. AetTrypl expressed in 5-d-old female Ae. taeniorhynchus significantly increased after 30 min postblood feeding compared with the control. The AetTryp1 mRNA expression in 5-d-old female Ae. taeniorhynchus was affected by different concentrations of permethrin.

  5. Transcriptional profiling in human HaCaT keratinocytes in response to kaempferol and identification of potential transcription factors for regulating differential gene expression

    Science.gov (United States)

    Kang, Byung Young; Lee, Ki-Hwan; Lee, Yong Sung; Hong, Il; Lee, Mi-Ock; Min, Daejin; Chang, Ihseop; Hwang, Jae Sung; Park, Jun Seong; Kim, Duck Hee

    2008-01-01

    Kaempferol is the major flavonol in green tea and exhibits many biomedically useful properties such as antioxidative, cytoprotective and anti-apoptotic activities. To elucidate its effects on the skin, we investigated the transcriptional profiles of kaempferol-treated HaCaT cells using cDNA microarray analysis and identified 147 transcripts that exhibited significant changes in expression. Of these, 18 were up-regulated and 129 were down-regulated. These transcripts were then classified into 12 categories according to their functional roles: cell adhesion/cytoskeleton, cell cycle, redox homeostasis, immune/defense responses, metabolism, protein biosynthesis/modification, intracellular transport, RNA processing, DNA modification/ replication, regulation of transcription, signal transduction and transport. We then analyzed the promoter sequences of differentially-regulated genes and identified over-represented regulatory sites and candidate transcription factors (TFs) for gene regulation by kaempferol. These included c-REL, SAP-1, Ahr-ARNT, Nrf-2, Elk-1, SPI-B, NF-κB and p65. In addition, we validated the microarray results and promoter analyses using conventional methods such as real-time PCR and ELISA-based transcription factor assay. Our microarray analysis has provided useful information for determining the genetic regulatory network affected by kaempferol, and this approach will be useful for elucidating gene-phytochemical interactions. PMID:18446059

  6. c-Jun binds the N terminus of human TAF(II)250 to derepress RNA polymerase II transcription in vitro.

    Science.gov (United States)

    Lively, T N; Ferguson, H A; Galasinski, S K; Seto, A G; Goodrich, J A

    2001-07-06

    c-Jun is an oncoprotein that activates transcription of many genes involved in cell growth and proliferation. We studied the mechanism of transcriptional activation by human c-Jun in a human RNA polymerase II transcription system composed of highly purified recombinant and native transcription factors. Transcriptional activation by c-Jun depends on the TATA-binding protein (TBP)-associated factor (TAF) subunits of transcription factor IID (TFIID). Protein-protein interaction assays revealed that c-Jun binds with high specificity to the largest subunit of human TFIID, TAF(II)250. The region of TAF(II)250 bound by c-Jun lies in the N-terminal 163 amino acids. This same region of TAF(II)250 binds to TBP and represses its interaction with TATA boxes, thereby decreasing DNA binding by TFIID. We hypothesized that c-Jun is capable of derepressing the effect of the TAF(II)250 N terminus on TFIID-driven transcription. In support of this hypothesis, we found that c-Jun increased levels of TFIID-driven transcription in vitro when added at high concentrations to a DNA template lacking activator protein 1 (AP-1) sites. Moreover, c-Jun blocked the repression of TBP DNA binding caused by the N terminus of TAF(II)250. In addition to revealing a mechanism by which c-Jun activates transcription, our studies provide the first evidence that an activator can bind directly to the N terminus of TAF(II)250 to derepress RNA polymerase II transcription in vitro.

  7. Gene transcript analysis blood values correlate with {sup 68}Ga-DOTA-somatostatin analog (SSA) PET/CT imaging in neuroendocrine tumors and can define disease status

    Energy Technology Data Exchange (ETDEWEB)

    Bodei, L. [European Institute of Oncology, Division of Nuclear Medicine, Milan (Italy); Kidd, M.; Modlin, I.M.; Drozdov, I. [Wren Laboratories, Branford, CT (United States); Prasad, V. [Charite University Hospital, Department of Nuclear Medicine, Berlin (Germany); Severi, S.; Paganelli, G. [Istituto Scientifico Romagnolo per lo Studio e la Cura dei Tumori (IRST) IRCCS, Nuclear Medicine and Radiometabolic Units, Meldola (Italy); Ambrosini, V. [S. Orsola-Malpighi University Hospital, Nuclear Medicine, Bologna (Italy); Kwekkeboom, D.J.; Krenning, E.P. [Erasmus Medical Center Rotterdam, Nuclear Medicine Department, Rotterdam (Netherlands); Baum, R.P. [Zentralklinik Bad Berka, THERANOSTICS Center for Molecular Radiotherapy and Imaging, Bad Berka (Germany)

    2015-08-15

    Precise determination of neuroendocrine tumor (NET) disease status and response to therapy remains a rate-limiting concern for disease management. This reflects limitations in biomarker specificity and resolution capacity of imaging. In order to evaluate biomarker precision and identify if combinatorial blood molecular markers and imaging could provide added diagnostic value, we assessed the concordance between {sup 68}Ga-somatostatin analog (SSA) positron emission tomography (PET), circulating NET gene transcripts (NETest), chromogranin A (CgA), and Ki-67 in NETs. We utilized two independent patient groups with positive {sup 68}Ga-SSA PET: data set 1 ({sup 68}Ga-SSA PETs undertaken for peptide receptor radionuclide therapy (PRRT), as primary or salvage treatment, n = 27) and data set 2 ({sup 68}Ga-SSA PETs performed in patients referred for initial disease staging or restaging after various therapies, n = 22). We examined the maximum standardized uptake value (SUV{sub max}), circulating gene transcripts, CgA levels, and baseline Ki-67. Regression analyses, generalized linear modeling, and receiver-operating characteristic (ROC) analyses were undertaken to determine the strength of the relationships. SUV{sub max} measured in two centers were mathematically evaluated (regression modeling) and determined to be comparable. Of 49 patients, 47 (96 %) exhibited a positive NETest. Twenty-six (54 %) had elevated CgA (χ{sup 2} = 20.1, p < 2.5 x 10{sup -6}). The majority (78 %) had Ki-67 < 20 %. Gene transcript scores were predictive of imaging with >95 % concordance and significantly correlated with SUV{sub max} (R {sup 2} = 0.31, root-mean-square error = 9.4). The genes MORF4L2 and somatostatin receptors SSTR1, 3, and 5 exhibited the highest correlation with SUV{sub max}. Progressive disease was identified by elevated levels of a quotient of MORF4L2 expression and SUV{sub max} [ROC-derived AUC (R {sup 2} = 0.7, p < 0.05)]. No statistical relationship was identified

  8. Gene transcript analysis blood values correlate with 68Ga-DOTA-somatostatin analog (SSA) PET/CT imaging in neuroendocrine tumors and can define disease status

    International Nuclear Information System (INIS)

    Bodei, L.; Kidd, M.; Modlin, I.M.; Drozdov, I.; Prasad, V.; Severi, S.; Paganelli, G.; Ambrosini, V.; Kwekkeboom, D.J.; Krenning, E.P.; Baum, R.P.

    2015-01-01

    Precise determination of neuroendocrine tumor (NET) disease status and response to therapy remains a rate-limiting concern for disease management. This reflects limitations in biomarker specificity and resolution capacity of imaging. In order to evaluate biomarker precision and identify if combinatorial blood molecular markers and imaging could provide added diagnostic value, we assessed the concordance between 68 Ga-somatostatin analog (SSA) positron emission tomography (PET), circulating NET gene transcripts (NETest), chromogranin A (CgA), and Ki-67 in NETs. We utilized two independent patient groups with positive 68 Ga-SSA PET: data set 1 ( 68 Ga-SSA PETs undertaken for peptide receptor radionuclide therapy (PRRT), as primary or salvage treatment, n = 27) and data set 2 ( 68 Ga-SSA PETs performed in patients referred for initial disease staging or restaging after various therapies, n = 22). We examined the maximum standardized uptake value (SUV max ), circulating gene transcripts, CgA levels, and baseline Ki-67. Regression analyses, generalized linear modeling, and receiver-operating characteristic (ROC) analyses were undertaken to determine the strength of the relationships. SUV max measured in two centers were mathematically evaluated (regression modeling) and determined to be comparable. Of 49 patients, 47 (96 %) exhibited a positive NETest. Twenty-six (54 %) had elevated CgA (χ 2 = 20.1, p < 2.5 x 10 -6 ). The majority (78 %) had Ki-67 < 20 %. Gene transcript scores were predictive of imaging with >95 % concordance and significantly correlated with SUV max (R 2 = 0.31, root-mean-square error = 9.4). The genes MORF4L2 and somatostatin receptors SSTR1, 3, and 5 exhibited the highest correlation with SUV max . Progressive disease was identified by elevated levels of a quotient of MORF4L2 expression and SUV max [ROC-derived AUC (R 2 = 0.7, p < 0.05)]. No statistical relationship was identified between CgA and Ki-67 and no relationship with imaging parameters

  9. Delivery of the Sox9 gene promotes chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells in an in vitro model

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Z.H. [Department of Otolaryngology - Head and Neck Surgery, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); Li, X.L. [Department of Dermatology, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); He, X.J. [Department of Orthopedics, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); Wu, B.J.; Xu, M. [Department of Otolaryngology - Head and Neck Surgery, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); Chang, H.M. [Department of Otolaryngology - Head and Neck Surgery, Affiliated Hospital of Xi' an Medical University, Xi' an (China); Zhang, X.H. [Department of Otolaryngology - Head and Neck Surgery, The Second Hospital, Xi' an Jiaotong University, Xi' an (China); Xing, Z. [Department of Clinical Dentistry, Faculty of Dentistry, Center for Clinical Dental Research, University of Bergen, Bergen (Norway); Jing, X.H.; Kong, D.M.; Kou, X.H.; Yang, Y.Y. [Department of Otolaryngology - Head and Neck Surgery, The Second Hospital, Xi' an Jiaotong University, Xi' an (China)

    2014-03-18

    SRY-related high-mobility-group box 9 (Sox9) gene is a cartilage-specific transcription factor that plays essential roles in chondrocyte differentiation and cartilage formation. The aim of this study was to investigate the feasibility of genetic delivery of Sox9 to enhance chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells (hUC-MSCs). After they were isolated from human umbilical cord blood within 24 h after delivery of neonates, hUC-MSCs were untreated or transfected with a human Sox9-expressing plasmid or an empty vector. The cells were assessed for morphology and chondrogenic differentiation. The isolated cells with a fibroblast-like morphology in monolayer culture were positive for the MSC markers CD44, CD105, CD73, and CD90, but negative for the differentiation markers CD34, CD45, CD19, CD14, or major histocompatibility complex class II. Sox9 overexpression induced accumulation of sulfated proteoglycans, without altering the cellular morphology. Immunocytochemistry demonstrated that genetic delivery of Sox9 markedly enhanced the expression of aggrecan and type II collagen in hUC-MSCs compared with empty vector-transfected counterparts. Reverse transcription-polymerase chain reaction analysis further confirmed the elevation of aggrecan and type II collagen at the mRNA level in Sox9-transfected cells. Taken together, short-term Sox9 overexpression facilitates chondrogenesis of hUC-MSCs and may thus have potential implications in cartilage tissue engineering.

  10. Delivery of the Sox9 gene promotes chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells in an in vitro model

    Directory of Open Access Journals (Sweden)

    Z.H. Wang

    2014-04-01

    Full Text Available SRY-related high-mobility-group box 9 (Sox9 gene is a cartilage-specific transcription factor that plays essential roles in chondrocyte differentiation and cartilage formation. The aim of this study was to investigate the feasibility of genetic delivery of Sox9 to enhance chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells (hUC-MSCs. After they were isolated from human umbilical cord blood within 24 h after delivery of neonates, hUC-MSCs were untreated or transfected with a human Sox9-expressing plasmid or an empty vector. The cells were assessed for morphology and chondrogenic differentiation. The isolated cells with a fibroblast-like morphology in monolayer culture were positive for the MSC markers CD44, CD105, CD73, and CD90, but negative for the differentiation markers CD34, CD45, CD19, CD14, or major histocompatibility complex class II. Sox9 overexpression induced accumulation of sulfated proteoglycans, without altering the cellular morphology. Immunocytochemistry demonstrated that genetic delivery of Sox9 markedly enhanced the expression of aggrecan and type II collagen in hUC-MSCs compared with empty vector-transfected counterparts. Reverse transcription-polymerase chain reaction analysis further confirmed the elevation of aggrecan and type II collagen at the mRNA level in Sox9-transfected cells. Taken together, short-term Sox9 overexpression facilitates chondrogenesis of hUC-MSCs and may thus have potential implications in cartilage tissue engineering.

  11. Delivery of the Sox9 gene promotes chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells in an in vitro model

    International Nuclear Information System (INIS)

    Wang, Z.H.; Li, X.L.; He, X.J.; Wu, B.J.; Xu, M.; Chang, H.M.; Zhang, X.H.; Xing, Z.; Jing, X.H.; Kong, D.M.; Kou, X.H.; Yang, Y.Y.

    2014-01-01

    SRY-related high-mobility-group box 9 (Sox9) gene is a cartilage-specific transcription factor that plays essential roles in chondrocyte differentiation and cartilage formation. The aim of this study was to investigate the feasibility of genetic delivery of Sox9 to enhance chondrogenic differentiation of human umbilical cord blood-derived mesenchymal stem cells (hUC-MSCs). After they were isolated from human umbilical cord blood within 24 h after delivery of neonates, hUC-MSCs were untreated or transfected with a human Sox9-expressing plasmid or an empty vector. The cells were assessed for morphology and chondrogenic differentiation. The isolated cells with a fibroblast-like morphology in monolayer culture were positive for the MSC markers CD44, CD105, CD73, and CD90, but negative for the differentiation markers CD34, CD45, CD19, CD14, or major histocompatibility complex class II. Sox9 overexpression induced accumulation of sulfated proteoglycans, without altering the cellular morphology. Immunocytochemistry demonstrated that genetic delivery of Sox9 markedly enhanced the expression of aggrecan and type II collagen in hUC-MSCs compared with empty vector-transfected counterparts. Reverse transcription-polymerase chain reaction analysis further confirmed the elevation of aggrecan and type II collagen at the mRNA level in Sox9-transfected cells. Taken together, short-term Sox9 overexpression facilitates chondrogenesis of hUC-MSCs and may thus have potential implications in cartilage tissue engineering

  12. Determination of mercury in human serum and packed blood cells by neutron activation analysis

    International Nuclear Information System (INIS)

    Versieck, J.; Vanballenberghe, L.; Wittoek, A.; Vermeir, G.; Vandecasteele, C.

    1990-01-01

    A method is described for the determination of mercury in human blood serum and packed blood cells employing neutron activation analysis. Great attention was devoted to the collection and manipulation of the samples. The accuracy and precision of the method were tested by analyzing biological reference materials and by comparing the concentrations measured in a number of serum samples to those obtained by another, independent technique (cold vapor atomic absorption spectrometry) in the same samples. The article reports the levels measured in blood serum and packed blood cells samples from 15 adult volunteers, as well as the figures determined in a open-quotes second-generationclose quotes biological reference material (freeze-dried human serum), prepared and conditioned at the University of Ghent

  13. Growth of human T lymphocyte colonies from whole blood: culture requirements and applications

    International Nuclear Information System (INIS)

    Knox, S.J.; Wilson, F.D.; Greenberg, B.R.; Shifrine, M.

    1982-01-01

    Growth of human lymphocyte colonies from whole blood following stimulation with PHA, Con A, or PPD is described. Individual colony cells were identified as T lymphocytes on the basis of surface marker and enzyme cytochemical characterizations. Colony formation increased as a power function over a wide range of cell concentrations above a critical minimal concentration. The whole blood culture system eliminates possible selective effects of lymphocyte colony techniques utilizing gradient-enriched lymphocyte fractions and more closely approximates the in vivo milieu. The whole blood colony method is more sensitive for the detection of low-level radiation effects on lymphocytes than widely used tests that measure 3 H-thymidine incorporation. In preliminary studies, researchers used the whole blood method to determine the relative radiosensitivity of lymphocytes from humans with various hematopoietic disorders, and observed abnormalities in mitogen responsiveness and colony formation in some of the patient groups. This method has wide application for studies in cellular and clinical immunology

  14. Cerebral blood flow reduction in Alzheimer's disease: impact of capillary occlusions on mice and humans

    Science.gov (United States)

    Berg, Maxime; Merlo, Adlan; Peyrounette, Myriam; Doyeux, Vincent; Smith, Amy; Cruz-Hernandez, Jean; Bracko, Oliver; Haft-Javaherian, Mohammad; Nishimura, Nozomi; Schaffer, Chris B.; Davit, Yohan; Quintard, Michel; Lorthois, Sylvie

    2017-11-01

    Alzheimer's disease may be the most common form of dementia, yet a satis