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Sample records for expanded human primary

  1. Expanding Human Cognition and Communication

    Energy Technology Data Exchange (ETDEWEB)

    Spohrer, Jim [IBM, North Castle, NY (United States); Pierce, Brian M. [Raytheon Co., Waltham, MA (United States); Murray, Cherry A. [Lucent Technologies, Murray Hill, NJ (United States); Golledge, Reginald G. [Univ. of California, Santa Barbara, CA (United States); Horn, Robert E. [Stanford Univ., CA (United States); Turkle, Sherry [Massachusetts Inst. of Technology (MIT), Cambridge, MA (United States); Yonas, Gerold [Sandia National Lab. (SNL-NM), Albuquerque, NM (United States); Glicken Turnley, Jessica [Galisteo Consulting Group, Inc., Albuquerque, NM (United States); Pollack, Jordan [Brandeis Univ., Waltham, MA (United States); Burger, Rudy [Massachusetts Inst. of Technology (MIT), Cambridge, MA (United States); Robinett, Warren; Wilson, Larry Todd [Inst. of Electrical and Electronics Engineers (IEEE), Piscataway, NJ (United States); Bainbridge, W. S.; Canton, J.; Kuekes, P.; Loomis, J.; Penz, P.

    2013-01-01

    To be able to chart the most profitable future directions for societal transformation and corresponding scientific research, five multidisciplinary themes focused on major goals have been identified to fulfill the overall motivating vision of convergence described in the previous pages. The first, “Expanding Human Cognition and Communication,” is devoted to technological breakthroughs that have the potential to enhance individuals’ mental and interaction abilities. Throughout the twentieth century, a number of purely psychological techniques were offered for strengthening human character and personality, but evaluation research has generally failed to confirm the alleged benefits of these methods (Druckman and Bjork 1992; 1994). Today, there is good reason to believe that a combination of methods, drawing upon varied branches of converging science and technology, would be more effective than attempts that rely upon mental training alone.

  2. Primary hyperaldosteronism in cats: expanding the diagnostic net

    NARCIS (Netherlands)

    Djajadiningrat-Laanen, S.C.

    2014-01-01

    Primary hyperaldosteronism or low-renin hyperaldosteronism in cats is characterized by inappropriately high aldosterone secretion from one or both adrenal glands, with systemic arterial hypertension and hypokalemia as leading clinical manifestations. In this thesis, non-tumorous primary

  3. Functional evolution of new and expanded attention networks in humans

    Science.gov (United States)

    Patel, Gaurav H.; Yang, Danica; Jamerson, Emery C.; Snyder, Lawrence H.; Corbetta, Maurizio; Ferrera, Vincent P.

    2015-01-01

    Macaques are often used as a model system for invasive investigations of the neural substrates of cognition. However, 25 million years of evolution separate humans and macaques from their last common ancestor, and this has likely substantially impacted the function of the cortical networks underlying cognitive processes, such as attention. We examined the homology of frontoparietal networks underlying attention by comparing functional MRI data from macaques and humans performing the same visual search task. Although there are broad similarities, we found fundamental differences between the species. First, humans have more dorsal attention network areas than macaques, indicating that in the course of evolution the human attention system has expanded compared with macaques. Second, potentially homologous areas in the dorsal attention network have markedly different biases toward representing the contralateral hemifield, indicating that the underlying neural architecture of these areas may differ in the most basic of properties, such as receptive field distribution. Third, despite clear evidence of the temporoparietal junction node of the ventral attention network in humans as elicited by this visual search task, we did not find functional evidence of a temporoparietal junction in macaques. None of these differences were the result of differences in training, experimental power, or anatomical variability between the two species. The results of this study indicate that macaque data should be applied to human models of cognition cautiously, and demonstrate how evolution may shape cortical networks. PMID:26170314

  4. Water extraction on Mars for an expanding human colony.

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    Ralphs, M; Franz, B; Baker, T; Howe, S

    2015-11-01

    In-situ water extraction is necessary for an extended human presence on Mars. This study looks at the water requirements of an expanding human colony on Mars and the general systems needed to supply that water from the martian atmosphere and regolith. The proposed combination of systems in order to supply the necessary water includes a system similar to Honeybee Robotics' Mobile In-Situ Water Extractor (MISWE) that uses convection, a system similar to MISWE but that directs microwave energy down a borehole, a greenhouse or hothouse type system, and a system similar to the Mars Atmospheric Resource Recovery System (MARRS). It is demonstrated that a large water extraction system that can take advantage of large deposits of water ice at site specific locations is necessary to keep up with the demands of a growing colony. Copyright © 2015 The Committee on Space Research (COSPAR). Published by Elsevier Ltd. All rights reserved.

  5. An Expanded Conceptual Framework of Medical Students' Primary Care Career Choice.

    Science.gov (United States)

    Pfarrwaller, Eva; Audétat, Marie-Claude; Sommer, Johanna; Maisonneuve, Hubert; Bischoff, Thomas; Nendaz, Mathieu; Baroffio, Anne; Junod Perron, Noëlle; Haller, Dagmar M

    2017-11-01

    In many countries, the number of graduating medical students pursuing a primary care career does not meet demand. These countries face primary care physician shortages. Students' career choices have been widely studied, yet many aspects of this process remain unclear. Conceptual models are useful to plan research and educational interventions in such complex systems.The authors developed a framework of primary care career choice in undergraduate medical education, which expands on previously published models. They used a group-based, iterative approach to find the best way to represent the vast array of influences identified in previous studies, including in a recent systematic review of the literature on interventions to increase the proportion of students choosing a primary care career. In their framework, students enter medical school with their personal characteristics and initial interest in primary care. They complete a process of career decision making, which is subject to multiple interacting influences, both within and outside medical school, throughout their medical education. These influences are stratified into four systems-microsystem, mesosystem, exosystem, and macrosystem-which represent different levels of interaction with students' career choices.This expanded framework provides an updated model to help understand the multiple factors that influence medical students' career choices. It offers a guide for the development of new interventions to increase the proportion of students choosing primary care careers and for further research to better understand the variety of processes involved in this decision.

  6. TOURISM AS A WAY TO EXPAND THE HUMAN LIFE SPACE

    Directory of Open Access Journals (Sweden)

    L. V. Martseniuk

    2017-06-01

    Full Text Available Purpose of the work is to substantiate the development of railway tourism in the context of human needs in accordance with the theory of individual life space. Methodology. Theoretical and methodological basis of the study is the provisions of the economic theory, management theory, corporate governance. The rational activity of travel agencies is based on the principles of economic equilibrium; Ukrainian population demand for railway tourism was determined with the help of market research and anonymous survey; to explore the real balance between the demand for tourist rail transport and the potential of the required volume of services the paper suggests the balance method. Since any travel company is an open system and is completely dependent on environmental factors, we proposed a method for estimating the factors of internal and external environment. Originality. The element of originality is compilation of existing concepts to the definition of the individual as a subject of life property, for the understanding of human relationships and its external environment. The paper developed the issue of the ability to influence the value of human life space with the help of tourist services. Conclusions. Market research conducted by the author has shown that in Ukraine there is a certain demand for tourist transport by rail, because it is more reliable, safer and more comfortable than the road transport. It is proved that the development of a new innovative project is very timely, as it will allow: to develop tourist infrastructure of Ukraine and bring it to the domestic and foreign tourists; replenish the state and local budgets by tourists; create new jobs for the population and improve their living level; partially reduce the loss-making passenger sector by increasing the volume of rail transport; expand life space for the people of Ukraine that will allow raising the intellectual level of the individual.

  7. Expanded benefits for humanity from the International Space Station

    Science.gov (United States)

    Rai, Amelia; Robinson, Julie A.; Tate-Brown, Judy; Buckley, Nicole; Zell, Martin; Tasaki, Kazuyuki; Karabadzhak, Georgy; Sorokin, Igor V.; Pignataro, Salvatore

    2016-09-01

    examples of materials improvements that are being developed using data from ISS. The publication also expands the benefits of research results in human health, environmental change and disaster response and in education activities developed to capture student imaginations in support of science, technology, engineering and mathematics, or STEM, education internationally. Applications to human health of the knowledge gained on ISS continue to grow and improve healthcare technologies and our understanding of human physiology. Distinct benefits return to Earth from the only orbiting multi-disciplinary laboratory of its kind. The ISS is a stepping stone for future space exploration by providing findings that develop LEO and improve life on our planet.

  8. Expanded Benefits for Humanity from the International Space Station

    Science.gov (United States)

    Rai, Amelia; Robinson, Julie A.; Tate-Brown, Judy; Buckley, Nicole; Zell, Martin; Tasaki, Kazuyuki; Karabadzhak, Georgy; Sorokin, Igor V.; Pignataro, Salvatore

    2016-01-01

    examples of materials improvements that are being developed using data from ISS. The publication also expands the benefits of research results in human health, environmental change and disaster response and in education activities developed to capture student imaginations in support of science, technology, engineering and mathematics, or STEM, education internationally. Applications to human health of the knowledge gained on ISS continues to grow and improve healthcare technologies and our understanding of human physiology. Distinct benefits return to Earth from the only orbiting multi-disciplinary laboratory of its kind. The ISS is a stepping stone for future space exploration by providing findings that develop LEO and improve life on our planet.

  9. Ex Vivo Expanded Human NK Cells Survive and Proliferate in Humanized Mice with Autologous Human Immune Cells.

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    Vahedi, Fatemeh; Nham, Tina; Poznanski, Sophie M; Chew, Marianne V; Shenouda, Mira M; Lee, Dean; Ashkar, Ali A

    2017-09-21

    Adoptive immune cell therapy is emerging as a promising immunotherapy for cancer. Particularly, the adoptive transfer of NK cells has garnered attention due to their natural cytotoxicity against tumor cells and safety upon adoptive transfer to patients. Although strategies exist to efficiently generate large quantities of expanded NK cells ex vivo, it remains unknown whether these expanded NK cells can persist and/or proliferate in vivo in the absence of exogenous human cytokines. Here, we have examined the adoptive transfer of ex vivo expanded human cord blood-derived NK cells into humanized mice reconstituted with autologous human cord blood immune cells. We report that ex vivo expanded NK cells are able to survive and possibly proliferate in vivo in humanized mice without exogenous cytokine administration, but not in control mice that lack human immune cells. These findings demonstrate that the presence of autologous human immune cells supports the in vivo survival of ex vivo expanded human NK cells. These results support the application of ex vivo expanded NK cells in cancer immunotherapy and provide a translational humanized mouse model to test the lifespan, safety, and functionality of adoptively transferred cells in the presence of autologous human immune cells prior to clinical use.

  10. Human inmmunoglobulin in primary immunodeficiencies

    Directory of Open Access Journals (Sweden)

    Sheffler Mendoza Selma

    2014-07-01

    Full Text Available Human immunoglobulin replacement therapy has become a corner- stone in the treatment of patients with primary immunodeficiencies (PID. Currently indicated as first-line therapy for predominantly antibody deficiencies, severe combined immunodeficiencies, and some well-defined syndromes with immunodeficiencies, it is also indicated as adjunct therapy in many other PID. Although considered a high-cost medication, elegant studies had showed that patients correctly treated with human immunoglobulin replacement therapy result in lower costs derived from their health- attention. Major benefits of immunoglobulin replacement therapy include but are not restricted to: protection against infectious processes, organ damage progression-arrest, immune modulation and quality-of-life improvement. Two modalities of treatment are currently used, intravenous and subcutaneous, each has clear advantages and disadvantages when compared to the other, which are presented in this article. The correct use of human immunoglobulin for the treatment of patients with PID translates in better medical-practices improving survival and quality of life of affected patients.

  11. RNA FISH for detecting expanded repeats in human diseases.

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    Urbanek, Martyna O; Krzyzosiak, Wlodzimierz J

    2016-04-01

    RNA fluorescence in situ hybridization (FISH) is a widely used technique for detecting transcripts in fixed cells and tissues. Many variants of RNA FISH have been proposed to increase signal strength, resolution and target specificity. The current variants of this technique facilitate the detection of the subcellular localization of transcripts at a single molecule level. Among the applications of RNA FISH are studies on nuclear RNA foci in diseases resulting from the expansion of tri-, tetra-, penta- and hexanucleotide repeats present in different single genes. The partial or complete retention of mutant transcripts forming RNA aggregates within the nucleoplasm has been shown in multiple cellular disease models and in the tissues of patients affected with these atypical mutations. Relevant diseases include, among others, myotonic dystrophy type 1 (DM1) with CUG repeats, Huntington's disease (HD) and spinocerebellar ataxia type 3 (SCA3) with CAG repeats, fragile X-associated tremor/ataxia syndrome (FXTAS) with CGG repeats, myotonic dystrophy type 2 (DM2) with CCUG repeats, amyotrophic lateral sclerosis/frontotemporal dementia (ALS/FTD) with GGGGCC repeats and spinocerebellar ataxia type 32 (SCA32) with GGCCUG. In this article, we summarize the results obtained with FISH to examine RNA nuclear inclusions. We provide a detailed protocol for detecting RNAs containing expanded CAG and CUG repeats in different cellular models, including fibroblasts, lymphoblasts, induced pluripotent stem cells and murine and human neuronal progenitors. We also present the results of the first single-molecule FISH application in a cellular model of polyglutamine disease. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Plastic compressed collagen as a novel carrier for expanded human corneal endothelial cells for transplantation.

    Directory of Open Access Journals (Sweden)

    Hannah J Levis

    Full Text Available Current treatments for reversible blindness caused by corneal endothelial cell failure involve replacing the failed endothelium with donor tissue using a one donor-one recipient strategy. Due to the increasing pressure of a worldwide donor cornea shortage there has been considerable interest in developing alternative strategies to treat endothelial disorders using expanded cell replacement therapy. Protocols have been developed which allow successful expansion of endothelial cells in vitro but this approach requires a supporting material that would allow easy transfer of cells to the recipient. We describe the first use of plastic compressed collagen as a highly effective, novel carrier for human corneal endothelial cells. A human corneal endothelial cell line and primary human corneal endothelial cells retained their characteristic cobblestone morphology and expression of tight junction protein ZO-1 and pump protein Na+/K+ ATPase α1 after culture on collagen constructs for up to 14 days. Additionally, ultrastructural analysis suggested a well-integrated endothelial layer with tightly opposed cells and apical microvilli. Plastic compressed collagen is a superior biomaterial in terms of its speed and ease of production and its ability to be manipulated in a clinically relevant manner without breakage. This method provides expanded endothelial cells with a substrate that could be suitable for transplantation allowing one donor cornea to potentially treat multiple patients.

  13. Plastic compressed collagen as a novel carrier for expanded human corneal endothelial cells for transplantation.

    Science.gov (United States)

    Levis, Hannah J; Peh, Gary S L; Toh, Kah-Peng; Poh, Rebekah; Shortt, Alex J; Drake, Rosemary A L; Mehta, Jodhbir S; Daniels, Julie T

    2012-01-01

    Current treatments for reversible blindness caused by corneal endothelial cell failure involve replacing the failed endothelium with donor tissue using a one donor-one recipient strategy. Due to the increasing pressure of a worldwide donor cornea shortage there has been considerable interest in developing alternative strategies to treat endothelial disorders using expanded cell replacement therapy. Protocols have been developed which allow successful expansion of endothelial cells in vitro but this approach requires a supporting material that would allow easy transfer of cells to the recipient. We describe the first use of plastic compressed collagen as a highly effective, novel carrier for human corneal endothelial cells. A human corneal endothelial cell line and primary human corneal endothelial cells retained their characteristic cobblestone morphology and expression of tight junction protein ZO-1 and pump protein Na+/K+ ATPase α1 after culture on collagen constructs for up to 14 days. Additionally, ultrastructural analysis suggested a well-integrated endothelial layer with tightly opposed cells and apical microvilli. Plastic compressed collagen is a superior biomaterial in terms of its speed and ease of production and its ability to be manipulated in a clinically relevant manner without breakage. This method provides expanded endothelial cells with a substrate that could be suitable for transplantation allowing one donor cornea to potentially treat multiple patients.

  14. 75 FR 51273 - Expanded Human Immunodeficiency Virus (HIV) Testing for Disproportionately Affected Populations

    Science.gov (United States)

    2010-08-19

    ... Announcement CDC-RFA-PS10-10138, ``Expanded Human Immunodeficiency Virus (HIV) Testing for Disproportionately... Assistance Number: 93.523 The Affordable Care Act: Human Immunodeficiency Virus (HIV) Prevention and Public... Number 93.523 The Affordable Care Act: Human Immunodeficiency Virus (HIV) Prevention and Public Health...

  15. Roadmap for human resources for expanded Indian nuclear industry

    Energy Technology Data Exchange (ETDEWEB)

    Singh, R.K. [Bhabha Atomic Research Centre, Mumbai (India); Indian Nuclear Society (India); Srinivasan, G.R.; Goyal, O.P. [Bhabha Atomic Research Centre, Mumbai (India)

    2011-07-01

    This paper deals with detailed requirement of human resources for all phases of nuclear power plant, for the manufacturing sector and the probable roadmap for achieving India's target. The accident in Fukushima has brought out that only nuclear power that avoids being a threat to the health and safety of the population and the environmental will be acceptable to the society and for this to be achieved human resources could be a single major contributor. India has ambitious plan of achieving 20,000MW by 2020 and 63,000MW by 2050. It is felt out of the three resources men, material and money; the critical shortage would be human resources both in quality and quantity. As per IAEA report (Publication of 2008 edition of energy, electricity and nuclear power estimates for the period of 2030), nuclear capacity must grow to at least 1.8 times current capacity by 2030 if global temperature rises are to be kept at 2°C. Objective of recruiting and training human resources for Indian Industry can be as follows: a) For catering domestic market. b) For catering international market later on for nuclear industries outside India. As India will be an important future international player. The above would require a multiplication of human resources by nearly seven times. In addition it has to be wholesome covering all levels and all skills and all disciplines and stages covering the whole nuclear cycle including regulators. Human resources are required for design and engineering, construction, commissioning, operation, manufacturing and for support services. The manpower for these has to be trained to achieve high quality of nuclear standards. Presently Indian Department of Atomic Energy(DAE) runs several training schools giving one year Post Graduate, tailor made courses. This needs to be multiplied by Joint efforts. Training should be on 'SAT (Systematic Approach to Training)' methodology to ensure focussed, specific, needed to culminate in safe, reliable and

  16. Technology Investment Agendas to Expand Human Space Futures

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    Sherwood, Brent

    2012-01-01

    The paper develops four alternative core-technology advancement specifications, one for each of the four strategic goal options for government investment in human space flight. Already discussed in the literature, these are: Explore Mars; Settle the Moon; accelerate commercial development of Space Passenger Travel; and enable industrial scale-up of Space Solar Power for Earth. In the case of the Explore Mars goal, the paper starts with the contemporary NASA accounting of ?55 Mars-enabling technologies. The analysis decomposes that technology agenda into technologies applicable only to the Explore Mars goal, versus those applicable more broadly to the other three options. Salient technology needs of all four options are then elaborated to a comparable level of detail. The comparison differentiates how technologies or major developments that may seem the same at the level of budget lines or headlines (e.g., heavy-lift Earth launch) would in fact diverge widely if developed in the service of one or another of the HSF goals. The paper concludes that the explicit choice of human space flight goal matters greatly; an expensive portfolio of challenging technologies would not only enable a particular option, it would foreclose the others. Technologies essential to enable human exploration of Mars cannot prepare interchangeably for alternative futures; they would not allow us to choose later to Settle the Moon, unleash robust growth of Space Passenger Travel industries, or help the transition to a post-petroleum future with Space Solar Power for Earth. The paper concludes that a decades-long decision in the U.S.--whether made consciously or by default--to focus technology investment toward achieving human exploration of Mars someday would effectively preclude the alternative goals in our lifetime.

  17. First-in-Human Experience With the Gore Balloon-Expandable Covered Endoprosthesis in Iliac Artery Occlusive Disease.

    Science.gov (United States)

    Holden, Andrew; Merrilees, Stephen; Buckley, Brendan; Connor, Brigid; Colgan, Frances; Hill, Andrew

    2017-02-01

    To report the first-in-human iliac artery experience of a new balloon-expandable covered endoprosthesis. A prospective, single-center pilot study recruited 30 symptomatic patients (mean age 64 years; 18 men) to evaluate the safety and early efficacy of the new Gore balloon-expandable covered endoprosthesis for the treatment of de novo or restenotic common and/or external iliac artery lesions. According to protocol, up to 2 discrete lesions could be treated with a maximum total treated length ≤110 mm. Follow-up included clinical evaluation with duplex ultrasound at 1, 6, and 12 months. Data are presented through 12-month follow-up. The primary safety endpoint was a composite of device- or procedure-related death, myocardial infarction, or amputation in the treated leg within 30 days of the index procedure. Multiple performance outcomes were also evaluated. The primary 30-day safety endpoint was 0%. Per-subject estimates of primary patency, freedom from target lesion revascularization, and freedom from target vessel revascularization were 100% at 1 and 6 months and 96.6% at 12 months. Estimates of assisted primary and secondary patency were both 100% at 12 months. Freedom from major adverse events at 12 months was 100%. Most patients experienced improvements in Rutherford category, ankle-brachial index, and functional status that were sustained to 12 months. This positive first-in-human experience with the Gore balloon-expandable covered endoprosthesis suggests this device will have an important role in the management of aortoiliac occlusive disease.

  18. High-expanding cortical regions in human development and evolution are related to higher intellectual abilities.

    Science.gov (United States)

    Fjell, Anders M; Westlye, Lars T; Amlien, Inge; Tamnes, Christian K; Grydeland, Håkon; Engvig, Andreas; Espeseth, Thomas; Reinvang, Ivar; Lundervold, Astri J; Lundervold, Arvid; Walhovd, Kristine B

    2015-01-01

    Cortical surface area has tremendously expanded during human evolution, and similar patterns of cortical expansion have been observed during childhood development. An intriguing hypothesis is that the high-expanding cortical regions also show the strongest correlations with intellectual function in humans. However, we do not know how the regional distribution of correlations between intellectual function and cortical area maps onto expansion in development and evolution. Here, in a sample of 1048 participants, we show that regions in which cortical area correlates with visuospatial reasoning abilities are generally high expanding in both development and evolution. Several regions in the frontal cortex, especially the anterior cingulate, showed high expansion in both development and evolution. The area of these regions was related to intellectual functions in humans. Low-expanding areas were not related to cognitive scores. These findings suggest that cortical regions involved in higher intellectual functions have expanded the most during development and evolution. The radial unit hypothesis provides a common framework for interpretation of the findings in the context of evolution and prenatal development, while additional cellular mechanisms, such as synaptogenesis, gliogenesis, dendritic arborization, and intracortical myelination, likely impact area expansion in later childhood. © The Author 2013. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  19. Primary Humanities: A Perspective from Wales

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    Jones, Mark; Whitehouse, Sarah

    2017-01-01

    How the humanities subjects are represented in primary schools in Wales has been influenced by curriculum developments including Curriculum Cymraeg, the Skills Framework and the Foundation Phase. A central tenet of Welsh Government policy has been to actively encourage schools to promote a sense of "Welshness" through curriculum content,…

  20. Expanding the Range of Text Types Used in the Primary Grades

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    Strachan, Stephanie L.

    2015-01-01

    Primary-grade students' experiences with text should prepare them to critically read an extensive range of text types throughout their schooling and career, a primary goal of the Common Core State Standards (CCSS). However, research demonstrates that narrative text overshadows other text types in the primary grades. The purpose of this…

  1. HANPP Collection: Human Appropriation of Net Primary Productivity as a Percentage of Net Primary Productivity

    Data.gov (United States)

    National Aeronautics and Space Administration — The Human Appropriation of Net Primary Productivity (HANPP) as a Percentage of Net Primary Productivity (NPP) portion of the Human Appropriation of Net Primary...

  2. Potency testing of mesenchymal stromal cell growth expanded in human platelet lysate from different human tissues.

    Science.gov (United States)

    Fazzina, R; Iudicone, P; Fioravanti, D; Bonanno, G; Totta, P; Zizzari, I G; Pierelli, L

    2016-08-25

    Mesenchymal stromal cells (MSCs) have been largely investigated, in the past decade, as potential therapeutic strategies for various acute and chronic pathological conditions. MSCs isolated from different sources, such as bone marrow (BM), umbilical cord tissue (UCT) and adipose tissue (AT), share many biological features, although they may show some differences on cumulative yield, proliferative ability and differentiation potential. The standardization of MSCs growth and their functional amplification is a mandatory objective of cell therapies. The aim of this study was to evaluate the cumulative yield and the ex vivo amplification potential of MSCs obtained from various sources and different subjects, using defined culture conditions with a standardized platelet lysate (PL) as growth stimulus. MSCs isolated from BM, UCT and AT and expanded in human PL were compared in terms of cumulative yield and growth potential per gram of starting tissue. MSCs morphology, phenotype, differentiation potential, and immunomodulatory properties were also investigated to evaluate their biological characteristics. The use of standardized PL-based culture conditions resulted in a very low variability of MSC growth. Our data showed that AT has the greater capacity to generate MSC per gram of initial tissue, compared to BM and UCT. However, UCT-MSCs replicated faster than AT-MSCs and BM-MSCs, revealing a greater proliferation capacity of this source irrespective of its lower MSC yield. All MSCs exhibited the typical MSC phenotype and the ability to differentiate into all mesodermal lineages, while BM-MSCs showed the most prominent immunosuppressive effect in vitro. The adoption of standardized culture conditions may help researchers and clinicians to reveal particular characteristics and inter-individual variability of MSCs sourced from different tissues. These data will be beneficial to set the standards for tissue collection and MSCs clinical-scale expansion both for cell banking

  3. Human factors and ergonomics for primary care.

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    Bowie, Paul; Jeffcott, Shelly

    2016-03-01

    In the second paper of this series, we provide a brief overview of the scientific discipline of human factors and ergonomics (HFE). Traditionally the HFE focus in healthcare has been in acute hospital settings which are perceived to exhibit characteristics more similar to other high-risk industries already applying related principles and methods. This paper argues that primary care is an area which could benefit extensively from an HFE approach, specifically in improving the performance and well-being of people and organisations. To this end, we define the purpose of HFE, outline its three specialist sub-domains (physical, cognitive and organisational HFE) and provide examples of guiding HFE principles and practices. Additionally, we describe HFE issues of significance to primary care education, improvement and research and outline early plans for building capacity and capability in this setting.

  4. In vivo prevention of transplant arteriosclerosis by ex vivo-expanded human regulatory T cells.

    Science.gov (United States)

    Nadig, Satish N; Wieckiewicz, Joanna; Wu, Douglas C; Warnecke, Gregor; Zhang, Wei; Luo, Shiqiao; Schiopu, Alexandru; Taggart, David P; Wood, Kathryn J

    2010-07-01

    Transplant arteriosclerosis is the hallmark of chronic allograft dysfunction (CAD) affecting transplanted organs in the long term. These fibroproliferative lesions lead to neointimal thickening of arteries in all transplanted allografts. Luminal narrowing then leads to graft ischemia and organ demise. To date, there are no known tolerance induction strategies that prevent transplant arteriosclerosis. Therefore, we designed this study to test the hypothesis that human regulatory T cells (T(reg) cells) expanded ex vivo can prevent transplant arteriosclerosis. Here we show the comparative capacity of T(reg) cells, sorted via two separate strategies, to prevent transplant arteriosclerosis in a clinically relevant chimeric humanized mouse system. We found that the in vivo development of transplant arteriosclerosis in human arteries was prevented by treatment of ex vivo-expanded human T(reg) cells. Additionally, we show that T(reg) cells sorted on the basis of low expression of CD127 provide a more potent therapy to conventional T(reg) cells. Our results demonstrate that human T(reg) cells can inhibit transplant arteriosclerosis by impairing effector function and graft infiltration. We anticipate our findings to serve as a foundation for the clinical development of therapeutics targeting transplant arteriosclerosis in both allograft transplantation and other immune-mediated causes of vasculopathy.

  5. In vivo Prevention of Transplant Arteriosclerosis by ex vivo Expanded Human Regulatory T Cells

    Science.gov (United States)

    Nadig, Satish N.; Więckiewicz, Joanna; Wu, Douglas C.; Warnecke, Gregor; Zhang, Wei; Luo, Shiqiao; Schiopu, Alexandru; Taggart, David P.; Wood, Kathryn J.

    2010-01-01

    Transplant arteriosclerosis (TA) is the hallmark of chronic allograft dysfunction (CAD) affecting transplanted organs in the long term [1,2]. These fibroproliferative lesions lead to neointimal thickening of arteries in all transplanted allografts [2]. Luminal narrowing then leads to graft ischemia and organ demise. To date, there are no known tolerance induction strategies that prevent TA [3,4]. Therefore, this study was designed to test the hypothesis that human regulatory T cells (Treg cells) expanded ex vivo could prevent TA. Here we show the comparative capacity of Treg cells, sorted via two separate strategies, to prevent TA in a clinically relevant chimeric humanized mouse system. We found that the in vivo development of TA in human arteries was prevented with the treatment of ex vivo expanded human Treg cells. Additionally, we show that Treg cells sorted based on the low expression of CD127 (IL-7Rα) provide a more potent therapy to conventional Treg cells. Our results demonstrate, for the first time, that human Treg cells can inhibit TA by impairing effector function and graft infiltration. We anticipate our findings to serve as a foundation for the clinical development of therapeutics targeting TA in both allograft transplantation and other immune-mediated causes of vasculopathy [5]. PMID:20473306

  6. Expanding choice of primary care in Finland: much debate but little change so far.

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    Tynkkynen, Liina-Kaisa; Chydenius, Miisa; Saloranta, Anna; Keskimäki, Ilmo

    2016-03-01

    "Putting the patient in the driver's seat" is one of the top issues on the health policy agenda in Finland. One of the means believed to promote patient empowerment and patient centeredness is the introduction and further expansion of choice policies with accompanying competition between public and private service providers. However, the Finnish health care system has a highly decentralized administration with multiple funding sources and three different types of providers that people can seek primary care from (municipal health centers, occupational health care services, and private sector providers). This complicates the implementation of choice at the level of primary health care. In this paper, we describe the current policy debates and initiatives promoting the expansion of the choice of primary care provider in Finland. We examine the legislation and policies that have contributed to the current, complex service system in Finland. In light of this examination, we critically discuss the current debate on choice policies as well as the introduction of choice in the context of primary health care. Copyright © 2016 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  7. Expanding Global Language Education in Public Primary Schools: The National English Programme in Mexico

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    Sayer, Peter

    2015-01-01

    The paper examines the recent national programme of English language instruction in the Mexican public primary schools, called the "Programa Nacional de Inglés en Educación Básica" (PNIEB). The programme, initiated in 2009 by the Ministry of Education as part of the national curriculum, represents the largest expansion of English…

  8. Human-started wildfires expand the fire niche across the United States

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    Balch, Jennifer K.; Bradley, Bethany A.; Nagy, R. Chelsea; Fusco, Emily J.; Mahood, Adam L.

    2017-01-01

    The economic and ecological costs of wildfire in the United States have risen substantially in recent decades. Although climate change has likely enabled a portion of the increase in wildfire activity, the direct role of people in increasing wildfire activity has been largely overlooked. We evaluate over 1.5 million government records of wildfires that had to be extinguished or managed by state or federal agencies from 1992 to 2012, and examined geographic and seasonal extents of human-ignited wildfires relative to lightning-ignited wildfires. Humans have vastly expanded the spatial and seasonal “fire niche” in the coterminous United States, accounting for 84% of all wildfires and 44% of total area burned. During the 21-y time period, the human-caused fire season was three times longer than the lightning-caused fire season and added an average of 40,000 wildfires per year across the United States. Human-started wildfires disproportionally occurred where fuel moisture was higher than lightning-started fires, thereby helping expand the geographic and seasonal niche of wildfire. Human-started wildfires were dominant (>80% of ignitions) in over 5.1 million km2, the vast majority of the United States, whereas lightning-started fires were dominant in only 0.7 million km2, primarily in sparsely populated areas of the mountainous western United States. Ignitions caused by human activities are a substantial driver of overall fire risk to ecosystems and economies. Actions to raise awareness and increase management in regions prone to human-started wildfires should be a focus of United States policy to reduce fire risk and associated hazards. PMID:28242690

  9. Expanding the clinical and neuroradiologic phenotype of primary microcephaly due to ASPM mutations.

    Science.gov (United States)

    Passemard, S; Titomanlio, L; Elmaleh, M; Afenjar, A; Alessandri, J-L; Andria, G; de Villemeur, T Billette; Boespflug-Tanguy, O; Burglen, L; Del Giudice, E; Guimiot, F; Hyon, C; Isidor, B; Mégarbané, A; Moog, U; Odent, S; Hernandez, K; Pouvreau, N; Scala, I; Schaer, M; Gressens, P; Gerard, B; Verloes, Alain

    2009-09-22

    To determine the spectrum of clinical, neuropsychological, and neuroradiologic features in patients with autosomal recessive primary microcephaly (MCPH) due to ASPM gene mutations. ASPM was sequenced in 52 unrelated MCPH probands. In patients with ASPM mutations, we evaluated the clinical phenotype, cognition, behavior, brain MRI, and family. We found homozygous or compound heterozygous ASPM loss-of-function mutations in 11 (22%) probands and 5 siblings. The probands harbored 18 different mutations, of which 16 were new. Microcephaly was severe after 1 year of age in all 16 patients, although in 4 patients the occipital-frontal circumference (OFC) at birth was decreased by only 2 SD. The OFC Z score consistently decreased after birth. Late-onset seizures occurred in 3 patients and significant pyramidal tract involvement in 1 patient. Intellectual quotients ranged from borderline-normal to severe mental retardation. Mild motor delay was noted in 7/16 patients. Language development was delayed in all patients older than 3 years. Brain MRI (n = 12) showed a simplified gyral pattern in 9 patients and several malformations including ventricle enlargement (n = 7), partial corpus callosum agenesis (n = 3), mild cerebellar hypoplasia (n = 1), focal cortical dysplasia (n = 1), and unilateral polymicrogyria (n = 1). Non-neurologic abnormalities consisted of short stature (n = 1), idiopathic premature puberty (n = 1), and renal dysplasia (n = 1). We provide a detailed description of features associated with ASPM mutations. Borderline microcephaly at birth, borderline-normal intellectual efficiency, and brain malformations can occur in ASPM-related primary hereditary microcephaly.

  10. Expanding the Human Performance Technologist's Repertoire: Knowledge Management, Organizational Learning and Human Performance Technology Literature Review.

    Science.gov (United States)

    Doucette, Mary Ann

    Successful performance improvement efforts draw from such disciplines as psychology and systems theory, and from the fields of instructional design and human resource development. Both knowledge management and organizational learning are valuable additions to the human performance technologist's repertoire for performance analysis and intervention…

  11. Expanding the landscape of chromatin modification (CM-related functional domains and genes in human.

    Directory of Open Access Journals (Sweden)

    Shuye Pu

    2010-11-01

    Full Text Available Chromatin modification (CM plays a key role in regulating transcription, DNA replication, repair and recombination. However, our knowledge of these processes in humans remains very limited. Here we use computational approaches to study proteins and functional domains involved in CM in humans. We analyze the abundance and the pair-wise domain-domain co-occurrences of 25 well-documented CM domains in 5 model organisms: yeast, worm, fly, mouse and human. Results show that domains involved in histone methylation, DNA methylation, and histone variants are remarkably expanded in metazoan, reflecting the increased demand for cell type-specific gene regulation. We find that CM domains tend to co-occur with a limited number of partner domains and are hence not promiscuous. This property is exploited to identify 47 potentially novel CM domains, including 24 DNA-binding domains, whose role in CM has received little attention so far. Lastly, we use a consensus Machine Learning approach to predict 379 novel CM genes (coding for 329 proteins in humans based on domain compositions. Several of these predictions are supported by very recent experimental studies and others are slated for experimental verification. Identification of novel CM genes and domains in humans will aid our understanding of fundamental epigenetic processes that are important for stem cell differentiation and cancer biology. Information on all the candidate CM domains and genes reported here is publicly available.

  12. HANPP Collection: Human Appropriation of Net Primary Productivity as a Percentage of Net Primary Productivity

    Data.gov (United States)

    National Aeronautics and Space Administration — The Human Appropriation of Net Primary Productivity (HANPP) as a Percentage of Net Primary Product (NPP) portion of the HANPP Collection represents a map identifying...

  13. Human Papillomavirus Infections in Primary Care

    Science.gov (United States)

    Ogunmodede, Folashade; Yale, Steven H.; Krawisz, Bruce; Tyler, Gregory C.; Evans, Anthony C.

    2007-01-01

    Cervical cancer continues to be a leading cause of mortality worldwide. The incidence and mortality associated with invasive cervical cancer have declined significantly in developed countries due to widespread availability of screening with the Papanicolaou (Pap) test. However, the incidence and prevalence of non-invasive cervical intraepithelial neoplasms and genital warts related to oncogenic and nononcogenic strains of human papilloma viruses (HPV) have remained relatively stable. Recent advances in molecular diagnostics have resulted in improved characterization of various HPV types and have led to changes in terminology of Pap test findings. Changes in nomenclature may lead to confusion among primary care providers regarding how best to further evaluate abnormal cytological results. This article provides a concise overview of the approach to the treatment of genital warts and management of abnormal cervical cytology based on guidelines from the American Society of Colposcopy and Cervical Pathology. It also reviews advances in HPV vaccine development and the new recombinant vaccine recently approved for use in the United States. PMID:18086908

  14. Improving washing strategies of human mesenchymal stem cells using negative mode expanded bed chromatography.

    Science.gov (United States)

    Cunha, Bárbara; Silva, Ricardo J S; Aguiar, Tiago; Serra, Margarida; Daicic, John; Maloisel, Jean-Luc; Clachan, John; Åkerblom, Anna; Carrondo, Manuel J T; Peixoto, Cristina; Alves, Paula M

    2016-01-15

    The use of human mesenchymal stem cells (hMSC) in clinical applications has been increasing over the last decade. However, to be applied in a clinical setting hMSC need to comply with specific requirements in terms of identity, potency and purity. This study reports the improvement of established tangential flow filtration (TFF)-based washing strategies, further increasing hMSC purity, using negative mode expanded bed adsorption (EBA) chromatography with a new multimodal prototype matrix based on core-shell bead technology. The matrix was characterized and a stable, expanded bed could be obtained using standard equipment adapted from what is used for conventional packed bed chromatography processes. The effect of different expansion rates on cell recovery yield and protein removal capacity was assessed. The best trade-off between cell recovery (89%) and protein clearance (67%) was achieved using an intermediate expansion bed rate (1.4). Furthermore, we also showed that EBA chromatography can be efficiently integrated on the already established process for the downstream processing (DSP) of hMSC, where it improved the washing efficiency more than 10-fold, recovering approximately 70% of cells after global processing. This strategy showed not to impact cell viability (>95%), neither hMSC's characteristics in terms of morphology, immunophenotype, proliferation, adhesion capacity and multipotent differentiation potential. Copyright © 2015 Elsevier B.V. All rights reserved.

  15. Role of Geography and Nurse Practitioner Scope-of-Practice in Efforts to Expand Primary Care System Capacity: Health Reform and the Primary Care Workforce.

    Science.gov (United States)

    Graves, John A; Mishra, Pranita; Dittus, Robert S; Parikh, Ravi; Perloff, Jennifer; Buerhaus, Peter I

    2016-01-01

    Little is known about the geographic distribution of the overall primary care workforce that includes both physician and nonphysician clinicians--particularly in areas with restrictive nurse practitioner scope-of-practice laws and where there are relatively large numbers of uninsured. We investigated whether geographic accessibility to primary care clinicians (PCCs) differed across urban and rural areas and across states with more or less restrictive scope-of-practice laws. An observational study. 2013 Area Health Resource File (AHRF) and US Census Bureau county travel data. The measures included percentage of the population in low-accessibility, medium-accessibility, and high-accessibility areas; number of geographically accessible primary care physicians (PCMDs), nurse practitioners (PCNPs), and physician assistants (PCPAs) per 100,000 population; and number of uninsured per PCC. We found divergent patterns in the geographic accessibility of PCCs. PCMDs constituted the largest share of the workforce across all settings, but were relatively more concentrated within urban areas. Accessibility to nonphysicians was highest in rural areas: there were more accessible PCNPs per 100,000 population in rural areas of restricted scope-of-practice states (21.4) than in urban areas of full practice states (13.9). Despite having more accessible nonphysician clinicians, rural areas had the largest number of uninsured per PCC in 2012. While less restrictive scope-of-practice states had up to 40% more PCNPs in some areas, we found little evidence of differences in the share of the overall population in low-accessibility areas across scope-of-practice categorizations. Removing restrictive scope-of-practice laws may expand the overall capacity of the primary care workforce, but only modestly in the short run. Additional efforts are needed that recognize the locational tendencies of physicians and nonphysicains.

  16. Global Patterns in Human Consumption of Net Primary Production

    Science.gov (United States)

    Imhoff, Marc L.; Bounoua, Lahouari; Ricketts, Taylor; Loucks, Colby; Harriss, Robert; Lawrence William T.

    2004-01-01

    The human population and its consumption profoundly affect the Earth's ecosystems. A particularly compelling measure of humanity's cumulative impact is the fraction of the planet's net primary production that we appropriate for our Net primary production-the net amount of solar energy converted to plant organic matter through photosynthesis-can be measured in units of elemental carbon and represents the primary food energy source for the world's ecosystems. Human appropriation of net primary production, apart from leaving less for other species to use, alters the composition of the atmosphere, levels of biodiversity, flows within food webs and the provision of important primary production required by humans and compare it to the total amount generated on the landscape. We then derive a spatial ba!mce sheet of net primary production supply and demand for the world. We show that human appropriation of net primary production varies spatially from almost zero to many times the local primary production. These analyses reveal the uneven footprint of human consumption and related environmental impacts, indicate the degree to which human populations depend on net primary production "imports" and suggest policy options for slowing future growth of human appropriation of net primary production.

  17. SPATIAL DEVELOPMENT QUALITIES AND HUMAN CAPITAL EXPANDED REPRODUCTION IN THE REPUBLIC OF KARELIA AND THE RUSSIA EUROPEAN NORTH ARCTIC REGIONS

    Directory of Open Access Journals (Sweden)

    Kekkonen A. L.

    2017-06-01

    Full Text Available The article presents the results of studies of the spatial development qualities and the human capital expanded reproduction of the Republic of Karelia and the Arctic regions of Russia’s European North. The aim of the article is the identification of characteristics, opportunities and threats of human capital reproduction in terms of region spatial development. The application of the research results is to provide recommendations for improving reproduction of the human capital and development of the education and vocational guidance systems, human capital management in the region. The study of spatial development and reproduction of the human capital used methods of logical, statistical and comparative analysis and SWOT- analysis principles. The results of the study allowed revealing the peculiarities of spatial development and human capital expanded reproduction of the Republic of Karelia in comparison with the Russian Arctic regions of the European North (Arkhangelsk Oblast, Murmansk Oblast, Komi Republic and Nenets Autonomous District. The Republic of Karelia has a special geographical position, and based on many socio-economic indicators is considered to be a «depressive» region. The expanded reproduction of human capital will bring the Republic of Karelia to the next level of development, taking into consideration external and internal factors as well as opportunities and threats. The results obtained in the study will expand the existing system of knowledge about reproduction of the human capital in the framework of spatial development, and the review of the expanded reproduction of human capital on the regional level will allow the government authorities to develop objective measures of quality formation of the region's human resources. It is proved that the introduction of the vocational guidance system in the region could lead to minimization of the threats associated with depopulation and the gap between education system and the

  18. Human Rights Texts: Converting Human Rights Primary Source Documents into Data

    National Research Council Canada - National Science Library

    Fariss, Christopher J; Linder, Fridolin J; Jones, Zachary M; Crabtree, Charles D; Biek, Megan A; Ross, Ana-Sophia M; Kaur, Taranamol; Tsai, Michael

    2015-01-01

    We introduce and make publicly available a large corpus of digitized primary source human rights documents which are published annually by monitoring agencies that include Amnesty International, Human...

  19. Expanding the spectrum of human ganglionic eminence region anomalies on fetal magnetic resonance imaging

    Energy Technology Data Exchange (ETDEWEB)

    Righini, Andrea; Parazzini, Cecilia; Izzo, Giana [Children' s Hospital ' ' V. Buzzi' ' , Department of Radiology and Neuroradiology, Milan (Italy); Cesaretti, Claudia [Children' s Hospital ' ' V. Buzzi' ' , Department of Radiology and Neuroradiology, Milan (Italy); Ospedale Maggiore Policlinico, Medical Genetics Unit, Fondazione I.R.C.C.S. Ca' Granda, Milan (Italy); Conte, Giorgio [Children' s Hospital ' ' V. Buzzi' ' , Department of Radiology and Neuroradiology, Milan (Italy); University of Milan, Department of Health Sciences, Milan (Italy); Frassoni, Carolina; Inverardi, Francesca [Fondazione I.R.C.C.S. Istituto Neurologico ' ' C. Besta' ' , Clinical Epileptology and Experimental Neurophysiology Unit, Milan (Italy); Bulfamante, Gaetano; Avagliano, Laura [San Paolo Hospital, Division of Human Pathology, Milan (Italy); Rustico, Mariangela [Children' s Hospital ' ' V. Buzzi' ' , Department of Obstetrics and Gynaecology, Prenatal Diagnosis, Milan (Italy)

    2016-03-15

    Ganglionic eminence (GE) is a transient fetal brain structure that harvests a significant amount of precursors of cortical GABA-ergic interneurons. Prenatal magnetic resonance (MR) imaging features of GE anomalies (i.e., cavitations) have already been reported associated with severe micro-lissencephaly. The purpose of this report was to illustrate the MR imaging features of GE anomalies in conditions other than severe micro-lissencephalies. Among all the fetuses submitted to prenatal MR imaging at our center from 2005 to 2014, we collected eight cases with GE anomalies and only limited associated brain anomalies. The median gestational age at the time of MR imaging was 21 weeks ranging from 19 to 29 weeks. Two senior pediatric neuroradiologists categorized the anomalies of the GE region in two groups: group one showing cavitation in the GE region and group two showing enlarged GE region. For each fetal case, associated cranial anomalies were also reported. Five out of the eight cases were included in group one and three in group two. Besides the GE region abnormality, all eight cases had additional intracranial anomalies, such as mild partial callosal agenesis, vermian hypoplasia and rotation, cerebellar hypoplasia, ventriculomegaly, enlarged subarachnoid spaces, molar tooth malformation. Ultrasound generally detected most of the associated intracranial anomalies, prompting the MR investigation; on the contrary in none of the cases, GE anomalies had been detected by ultrasound. Our observation expands the spectrum of human GE anomalies, demonstrating that these may take place also without associated severe micro-lissencephalies. (orig.)

  20. Expanding access to pain care for frail, older people in primary care: a cross-sectional study.

    Science.gov (United States)

    Muntinga, M E; Jansen, A P D; Schellevis, F G; Nijpels, G

    2016-01-01

    Although untreated pain has a negative impact on quality of life and health outcomes, research has shown that older people do not always have access to adequate pain care. Practice nurse-led, comprehensive geriatric assessments (CGAs) may increase access to tailored pain care for frail, older people who live at home. To explore this, we investigated whether new pain cases were identified by practice nurses during CGAs administered as part of an intervention with the Geriatric Care Model, a comprehensive care model based on the Chronic Care Model, and whether the intervention led to tailored pain action plans in care plans of frail, older people. We used cross-sectional data from the older Adults: Care in Transition (ACT) study, a 2-year clinical trial carried out in two regions of the Netherlands. Practice nurses proactively visited older people at home and administered an in-home CGA that included an assessment of pain. Pain care-related agreements and actions (pain action plans) based on CGA results were described in a tailored care plan. We analyzed care plans of 781 older people who received a first-time CGA by a practice nurse for the presence of pain, pain location and cause, new pain cases, and pain action plans. We used descriptive statistics to analyze our data. We found that 315 (40.3 %) older people experienced any type of pain. Practice nurses identified 20 (10.6 %) new pain cases, and 188 (59.7 %) older people with pain formulated at least one therapeutic or non-therapeutic pain action plan together with a practice nurse. More than half of the older people whose pain had already been identified by a primary care physician wanted a pain action plan. Most pain action plans consisted of actions or agreements related to continuity of care. Practice nurses in primary care can contribute to expanding older people's access to tailored pain care. Future researchers should continue to direct their focus at ways to overcome the barriers that restrict older

  1. Functional Analysis of Serially Expanded Human iPS Cell-Derived RPE Cultures

    Science.gov (United States)

    Singh, Ruchira; Phillips, M. Joseph; Kuai, David; Meyer, Jackelyn; Martin, Jessica M.; Smith, Molly A.; Perez, Enio T.; Shen, Wei; Wallace, Kyle A.; Capowski, Elizabeth E.; Wright, Lynda S.; Gamm, David M.

    2013-01-01

    Purpose. To determine the effects of serial expansion on the cellular, molecular, and functional properties of human iPS cell (hiPSC)-derived RPE cultures. Methods. Fibroblasts obtained from four individuals were reprogrammed into hiPSCs and differentiated to RPE cells using previously described methods. Patches of deeply pigmented hiPSC-RPE were dissected, dissociated, and grown in culture until they re-formed pigmented monolayers. Subsequent passages were obtained by repeated dissociation, expansion, and maturation of RPE into pigmented monolayers. Gene and protein expression profiles and morphological and functional characteristics of hiPSC-RPE at different passages were compared with each other and to human fetal RPE (hfRPE). Results. RPE from all four hiPSC lines could be expanded more than 1000-fold when serially passaged as pigmented monolayer cultures. Importantly, expansion of hiPSC-RPE monolayers over the first three passages (P1–P3) resulted in decreased expression of pluripotency and neuroretinal markers and maintenance of characteristic morphological features and gene and protein expression profiles. Furthermore, P1 to P3 hiPSC-RPE monolayers reliably demonstrated functional tight junctions, G-protein–coupled receptor-mediated calcium transients, phagocytosis and degradation of photoreceptor outer segments, and polarized secretion of biomolecules. In contrast, P4 hiPSC-RPE cells failed to form monolayers and possessed altered morphological and functional characteristics and gene expression levels. Conclusions. Highly differentiated, pigmented hiPSC-RPE monolayers can undergo limited serial expansion while retaining key cytological and functional attributes. However, passaging hiPSC-RPE cultures beyond senescence leads to loss of such features. Our findings support limited, controlled passaging of patient-specific hiPSC-RPE to procure cells needed for in vitro disease modeling, drug screening, and cellular transplantation. PMID:24030465

  2. Integrated Modular Teaching of Human Biology for Primary Care Practitioners

    Science.gov (United States)

    Glasgow, Michael S.

    1977-01-01

    Describes the use of integrated modular teaching of the human biology component of the Health Associate Program at Johns Hopkins University, where the goal is to develop an understanding of the sciences as applied to primary care. Discussion covers the module sequence, the human biology faculty, goals of the human biology faculty, laboratory…

  3. Family and Consumer Sciences Focus on the Human Dimension: The Expanded Food and Nutrition Education Program Example

    Science.gov (United States)

    Cason, Katherine L.; Chipman, Helen; Forstadt, Leslie A.; Rasco, Mattie R.; Sellers, Debra M.; Stephenson, Laura; York, De'Shoin A.

    2017-01-01

    The history of family and consumer sciences (FCS) and the Expanded Food and Nutrition Education Program (EFNEP) is discussed with an emphasis on the critical importance of the human dimension. EFNEP's focus on people, education for change, accountability, strategic partnerships, and public value are highlighted as an example and model for…

  4. Galectin-9 activates and expands human T-helper 1 cells.

    Directory of Open Access Journals (Sweden)

    Marloes J M Gooden

    Full Text Available Galectin-9 (Gal-9 is known for induction of apoptosis in IFN-γ and IL-17 producing T-cells and amelioration of autoimmunity in murine models. On the other hand, Gal-9 induced IFN-γ positive T-cells in a sarcoma mouse model and in food allergy, suggesting that Gal-9 can have diametric effects on T-cell immunity. Here, we aimed to delineate the immunomodulatory effect of Gal-9 on human resting and ex vivo activated peripheral blood lymphocytes. Treatment of resting lymphocytes with low concentrations of Gal-9 (5-30 nM induced apoptosis in ∼60% of T-cells after 1 day, but activated the surviving T-cells. These viable T-cells started to expand after 4 days with up to 6 cell divisions by day 7 and an associated shift from naïve towards central memory and IFN-γ producing phenotype. In the presence of T-cell activation signals (anti-CD3/IL-2 Gal-9 did not induce T-cell expansion, but shifted the CD4/CD8 balance towards a CD4-dominated T-cell response. Thus, Gal-9 activates resting T-cells in the absence of typical T-cell activating signals and promotes their transition to a TH1/C1 phenotype. In the presence of T-cell activating signals T-cell immunity is directed towards a CD4-driven response by Gal-9. Thus, Gal-9 may specifically enhance reactive immunological memory.

  5. Human ovarian surface epithelium in primary culture.

    Science.gov (United States)

    Auersperg, N; Siemens, C H; Myrdal, S E

    1984-10-01

    The ovarian surface epithelium (OSE) represents a minute fraction of the cell mass of the ovary but gives rise to over 80% of human ovarian carcinomas. No experimental models for the study of human OSE exist. To characterize OSE cells in culture, explants of ovarian surface from normal ovary of premenopausal women were grown on plastic, glass, and collagen gel in 25% fetal bovine serum/Waymouth's medium 752/1. About 25% of explants produced epithelial outgrowths. Morphologically, these outgrowths resembled OSE in vivo and endothelial and mesothelial cells in culture, but they differed from cultured ovarian stromal, granulosa, and luteal cells. Only OSE among ovarian cell types were intensely keratin positive by immunofluorescence. Keratin also distinguished OSE cells from the keratin-negative endothelial cells. Most but not all OSE colonies tested showed 17 beta-hydroxysteroid dehydrogenase (HSD) activity, which was absent in peritoneal mesothelial cells. Colonies from most patients were limited to a few millimetres and became stationary within a few weeks. Changes that accompanied cessation of growth included senescence, increased keratin content, or the formation of multicellular papillary aggregates. With time, OSE cells tended to assume a fibroblast-like morphology but remained keratin positive and continued to resemble OSE by scanning electron microscopy (SEM). Subcultured OSE cells persisted in a stationary keratin-positive form for many weeks. Throughout this study, all pavementlike epithelial outgrowths that were contiguous with an explant stained for keratin; thus, such colonies can be assumed to be OSE. Conversely, fibroblast-shaped cells may represent OSE as indicated by keratin content and SEM appearance. The methods presented here permit culture of normal human OSE under conditions in which the cells exhibit morphologic plasticity, variable 17 beta-HSD activity, and presence of keratin.

  6. Paracrine interactions between primary human macrophages and human fibroblasts enhance murine mammary gland humanization in vivo.

    Science.gov (United States)

    Fleming, Jodie M; Miller, Tyler C; Kidacki, Michal; Ginsburg, Erika; Stuelten, Christina H; Stewart, Delisha A; Troester, Melissa A; Vonderhaar, Barbara K

    2012-06-25

    Macrophages comprise an essential component of the mammary microenvironment necessary for normal gland development. However, there is no viable in vivo model to study their role in normal human breast function. We hypothesized that adding primary human macrophages to the murine mammary gland would enhance and provide a novel approach to examine immune-stromal cell interactions during the humanization process. Primary human macrophages, in the presence or absence of ectopic estrogen stimulation, were used to humanize mouse mammary glands. Mechanisms of enhanced humanization were identified by cytokine/chemokine ELISAs, zymography, western analysis, invasion and proliferation assays; results were confirmed with immunohistological analysis. The combined treatment of macrophages and estrogen stimulation significantly enhanced the percentage of the total gland humanized and the engraftment/outgrowth success rate. Timecourse analysis revealed the disappearance of the human macrophages by two weeks post-injection, suggesting that the improved overall growth and invasiveness of the fibroblasts provided a larger stromal bed for epithelial cell proliferation and structure formation. Confirming their promotion of fibroblasts humanization, estrogen-stimulated macrophages significantly enhanced fibroblast proliferation and invasion in vitro, as well as significantly increased proliferating cell nuclear antigen (PCNA) positive cells in humanized glands. Cytokine/chemokine ELISAs, zymography and western analyses identified TNFα and MMP9 as potential mechanisms by which estrogen-stimulated macrophages enhanced humanization. Specific inhibitors to TNFα and MMP9 validated the effects of these molecules on fibroblast behavior in vitro, as well as by immunohistochemical analysis of humanized glands for human-specific MMP9 expression. Lastly, glands humanized with macrophages had enhanced engraftment and tumor growth compared to glands humanized with fibroblasts alone. Herein, we

  7. The Adolescent "Expanded Medical Home": School-Based Health Centers Partner with a Primary Care Clinic to Improve Population Health and Mitigate Social Determinants of Health.

    Science.gov (United States)

    Riley, Margaret; Laurie, Anna R; Plegue, Melissa A; Richarson, Caroline R

    2016-01-01

    Access to high-quality health care is a crucial social determinant of health. We describe the implementation of an "expanded medical home" partnering a primary care practice (the Ypsilanti Health Center [YHC]) with local school-based health centers (the Regional Alliance for Healthy Schools [RAHS]), and to assess whether this model improves access to and quality of care for shared patients. Using the Consolidated Framework for Implementation Research, we define the steps in, barriers to, and facilitating factors in implementing the expanded medical home model. Visits and quality measures were assessed for patients seen by YHC only versus YHC/RAHS at baseline and during the intervention. At baseline, patients seen at YHC/RAHS had higher compliance with most quality metrics compared with those seen at YHC only. The proportion of shared patients significantly increased because of the intervention (P < .001). Overall, patients seen in the expanded medical home had a higher likelihood of receiving quality metric services than patients in YHC only (odds ratio, 1.8; 95% confidence interval, 1.57-2.05) across all measures. Thoughtful and intentional implementation of an expanded medical home partnership between primary care physicians and school-based health centers increases the number of shared high-risk adolescent patients. Shared patients have improved compliance with quality measures, which may lead to long-term improved health equity. © Copyright 2016 by the American Board of Family Medicine.

  8. The Humanities in English Primary Schools: Struggling to Survive

    Science.gov (United States)

    Barnes, Jonathan; Scoffham, Stephen

    2017-01-01

    This article surveys the state of the humanities in English primary schools drawing on evidence from serving head teachers, current literature and policy documents. The findings suggest that whilst the humanities are highly valued in schools, there are serious challenges which threaten the "broad and balanced" curriculum. It is suggested…

  9. Eugenol Toxicity in Human Dental Pulp Fibroblasts of Primary Teeth.

    Science.gov (United States)

    Escobar-García, Maria; Rodríguez-Contreras, Karen; Ruiz-Rodríguez, Socorro; Pierdant-Pérez, Mauricio; Cerda-Cristerna, Bernardino; Pozos-Guillén, Amaury

    2016-01-01

    The aim of the study was to determine the eugenol concentrations at which toxicity occurs in human dental pulp fibroblasts of primary teeth. Samples of primary dental pulp tissue were taken. Tissue samples were seeded by means of explant technique and used in the 4(th)-5th pass. Single Cell Gel Electrophoresis (Comet), phenazine MeThoSulfate (MTS), LIVE/DEAD Cell Viability/Toxicity and trypan blue assays for evaluation of the cytotoxicity of increasing concentrations of eugenol (0.06 to 810 μM) were performed. The results of toxicity tests showed toxic effects on dental pulp fibroblasts, even at very low concentrations of eugenol (0.06 μM). Very low concentrations of eugenol produce high toxicity in human dental pulp fibroblasts. All of the concentrations of eugenol that we evaluated produced high toxicity in human dental pulp fibroblasts of primary teeth.

  10. Successful transplantation of in vitro expanded human corneal endothelial precursors to corneal endothelial surface using a nanocomposite sheet

    Directory of Open Access Journals (Sweden)

    Parikumar P

    2011-01-01

    Full Text Available Background: Though the transplantation of in vitro expanded human corneal endothelial precursors in animal models of endothelial damage by injecting into the anterior chamber has been reported, the practical difficulties of accomplishing such procedure in human patients have been a hurdle to clinical translation. Here we report the successful transplantation of in vitro expanded human corneal precursor cells to an animal eye using a transparent Nano-composite sheet and their engraftment.Materials and Methods: Human Corneal endothelial cells (HCEC were isolated from human cadaver eyes with informed consent and expanded in the lab using a sphere forming assay in a novel Thermoreversible Gelation Polymer (TGP for 26 days. HCEC obtained by sphere forming assay were seeded in a novel Nano-composite sheet, which was made of PNIPA-NC gels by in-situ, free-radical polymerization of NIPA monomer in the presence of exfoliated clay (synthetic hectorite “Laponite XLG” uniformly dispersed in aqueous media. After a further seven days in vitro culture of HCEC in the Nano-composite sheet, cells were harvested and transplanted on cadaver-bovine eyes (n=3. The cells were injected between the corneal endothelial layer and the Nano-composite sheet that had been placed prior to the injection in close proximity to the endothelial layer. After three hours, the transplanted Nano-composite sheets were removed from the bovine eyes and subjected to microscopic examination. The corneas were subjected to Histo-pathological studies along with controls. Results: HCEC formed sphere like colonies in TGP which expressed relevant markers as confirmed by RT-PCR. Microscopic studies of the Nanosheets and histopathological studies of the cornea of the Bull’s eye revealed that the HCEC got engrafted to the corneal endothelial layer of the bovine eyes with no remnant cells in the Nanosheet. Conclusion: Transplantation of in vitro expanded donor human corneal endothelial cells

  11. Ago HITS-CLIP expands understanding of Kaposi's sarcoma-associated herpesvirus miRNA function in primary effusion lymphomas.

    Directory of Open Access Journals (Sweden)

    Irina Haecker

    Full Text Available KSHV is the etiological agent of Kaposi's sarcoma (KS, primary effusion lymphoma (PEL, and a subset of multicentricCastleman's disease (MCD. The fact that KSHV-encoded miRNAs are readily detectable in all KSHV-associated tumors suggests a potential role in viral pathogenesis and tumorigenesis. MiRNA-mediated regulation of gene expression is a complex network with each miRNA having many potential targets, and to date only few KSHV miRNA targets have been experimentally determined. A detailed understanding of KSHV miRNA functions requires high-through putribonomics to globally analyze putative miRNA targets in a cell type-specific manner. We performed Ago HITS-CLIP to identify viral and cellular miRNAs and their cognate targets in two latently KSHV-infected PEL cell lines. Ago HITS-CLIP recovered 1170 and 950 cellular KSHV miRNA targets from BCBL-1 and BC-3, respectively. Importantly, enriched clusters contained KSHV miRNA seed matches in the 3'UTRs of numerous well characterized targets, among them THBS1, BACH1, and C/EBPβ. KSHV miRNA targets were strongly enriched for genes involved in multiple pathways central for KSHV biology, such as apoptosis, cell cycle regulation, lymphocyte proliferation, and immune evasion, thus further supporting a role in KSHV pathogenesis and potentially tumorigenesis. A limited number of viral transcripts were also enriched by HITS-CLIP including vIL-6 expressed only in a subset of PEL cells during latency. Interestingly, Ago HITS-CLIP revealed extremely high levels of Ago-associated KSHV miRNAs especially in BC-3 cells where more than 70% of all miRNAs are of viral origin. This suggests that in addition to seed match-specific targeting of cellular genes, KSHV miRNAs may also function by hijacking RISCs, thereby contributing to a global de-repression of cellular gene expression due to the loss of regulation by human miRNAs. In summary, we provide an extensive list of cellular and viral miRNA targets representing an

  12. An expanded One Health model: integrating social science and One Health to inform study of the human-animal interface.

    Science.gov (United States)

    Woldehanna, Sara; Zimicki, Susan

    2015-03-01

    Zoonotic disease emergence is not a purely biological process mediated only by ecologic factors; opportunities for transmission of zoonoses from animals to humans also depend on how people interact with animals. While exposure is conditioned by the type of animal and the location in which interactions occur, these in turn are influenced by human activity. The activities people engage in are determined by social as well as contextual factors including gender, age, socio-economic status, occupation, social norms, settlement patterns and livelihood systems, family and community dynamics, as well as national and global influences. This paper proposes an expanded "One Health" conceptual model for human-animal exposure that accounts for social as well as epidemiologic factors. The expanded model informed a new study approach to document the extent of human exposure to animals and explore the interplay of social and environmental factors that influence risk of transmission at the individual and community level. The approach includes a formative phase using qualitative and participatory methods, and a representative, random sample survey to quantify exposure to animals in a variety of settings. The paper discusses the different factors that were considered in developing the approach, including the range of animals asked about and the parameters of exposure that are included, as well as factors to be considered in local adaptation of the generic instruments. Illustrative results from research using this approach in Lao PDR are presented to demonstrate the effect of social factors on how people interact with animals. We believe that the expanded model can be similarly operationalized to explore the interactions of other social and policy-level determinants that may influence transmission of zoonoses. Copyright © 2014 Elsevier Ltd. All rights reserved.

  13. Physical benefits of expanded physical education in primary school: findings from a 3-year intervention study in Sweden.

    Science.gov (United States)

    Sollerhed, A-C; Ejlertsson, G

    2008-02-01

    The aim of this study was to assess whether a school-based program with expanded physical education lessons was effective in increasing children's physical capacity and in preventing excessive weight gain in children. The study performed in 2000-2003 comprised 132 children, 73 boys and 59 girls at baseline 6-9 years and in follow-up 9-12 years, attending two different schools with a similar size, appearance and structure in a rural area. The norm school (N-school) followed the stipulated curricular time, one to two physical education lessons a week, while the intervention school (I-school) increased it to four lessons. More positive changes in physical index (the sum of the age-standardized results in 11 physical tests) were found among children in the I-school than in the N-school. The number of children who increased body mass index (BMI) increased in both schools, but a lower increase in BMI could be seen in the I-school. Expanded physical education lessons could increase physical status among both overweight and normal-weight children, in particular aerobic fitness. The weekly dose of physical activity must be higher than 40 min a day and must start earlier in children's life to be more effective in combating BMI increase.

  14. Primary and secondary infection with human parvovirus B19 in ...

    African Journals Online (AJOL)

    Primary and secondary infection with human parvovirus. B19 in pregnant women in South Africa ... I Infection during pregnancy may lead to fetal loss, especially during the second trimester, because of severe ... IgG antibodies) with viruses such as rubella: cyto- megalovirus' and varicella." Subjects and methods. A total of 1 ...

  15. Functional sex differences in human primary auditory cortex

    NARCIS (Netherlands)

    Ruytjens, Liesbet; Georgiadis, Janniko R.; Holstege, Gert; Wit, Hero P.; Albers, Frans W. J.; Willemsen, Antoon T. M.

    2007-01-01

    Background We used PET to study cortical activation during auditory stimulation and found sex differences in the human primary auditory cortex (PAC). Regional cerebral blood flow (rCBF) was measured in 10 male and 10 female volunteers while listening to sounds (music or white noise) and during a

  16. Adaptation and cross-cultural validationof the United States Primary Care Assessment Tool (expanded version for use in South Africa

    Directory of Open Access Journals (Sweden)

    Graham Bresick

    2015-02-01

    Full Text Available Background: Measuring primary care is important for health sector reform. The Primary Care Assessment Tool (PCAT measures performance of elements essential for cost-effective care. Following minor adaptations prior to use in Cape Town in 2011, a few findings indicated a need to improve the contentand cross-cultural validity for wider use in South Africa (SA.Aim: This study aimed to validate the United States of America-developed PCAT before being used in a baseline measure of primary care performance prior to major reform.Setting: Public sector primary care clinics, users, practitioners and managers in urban and rural districtsin the Western Cape Province.Methods: Face value evaluation of item phrasing and a combination of Delphi and Nominal Group Technique (NGT methods with an expert panel and user focus group were used to obtain consensus on content relevant to SA. Original and new domains and items with > = 70% agreement were included in the South African version – ZA PCAT.Results: All original PCAT domains achieved consensus on inclusion. One new domain, the primary healthcare (PHC team, was added. Three of 95 original items achieved < 70% agreement, that is consensus to exclude as not relevant to SA; 19 new items were added. A few items needed minor rephrasing with local healthcare jargon. The demographic section was adapted to local socio-economic conditions. The adult PCAT was translated into isiXhosa and Afrikaans.Conclusion: The PCAT is a valid measure of primary care performance in SA. The PHC team domainis an important addition, given its emphasis in PHC re-engineering. A combination of Delphi and NGT methods succeeded in obtaining consensus on a multi-domain, multi-item instrument in a resource-constrained environment.

  17. Disease Ontology 2015 update: an expanded and updated database of human diseases for linking biomedical knowledge through disease data.

    Science.gov (United States)

    Kibbe, Warren A; Arze, Cesar; Felix, Victor; Mitraka, Elvira; Bolton, Evan; Fu, Gang; Mungall, Christopher J; Binder, Janos X; Malone, James; Vasant, Drashtti; Parkinson, Helen; Schriml, Lynn M

    2015-01-01

    The current version of the Human Disease Ontology (DO) (http://www.disease-ontology.org) database expands the utility of the ontology for the examination and comparison of genetic variation, phenotype, protein, drug and epitope data through the lens of human disease. DO is a biomedical resource of standardized common and rare disease concepts with stable identifiers organized by disease etiology. The content of DO has had 192 revisions since 2012, including the addition of 760 terms. Thirty-two percent of all terms now include definitions. DO has expanded the number and diversity of research communities and community members by 50+ during the past two years. These community members actively submit term requests, coordinate biomedical resource disease representation and provide expert curation guidance. Since the DO 2012 NAR paper, there have been hundreds of term requests and a steady increase in the number of DO listserv members, twitter followers and DO website usage. DO is moving to a multi-editor model utilizing Protégé to curate DO in web ontology language. This will enable closer collaboration with the Human Phenotype Ontology, EBI's Ontology Working Group, Mouse Genome Informatics and the Monarch Initiative among others, and enhance DO's current asserted view and multiple inferred views through reasoning. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

  18. Expand and Regularize Federal Funding for Human Pluripotent Stem Cell Research

    Science.gov (United States)

    Owen-Smith, Jason; Scott, Christopher Thomas; McCormick, Jennifer B.

    2012-01-01

    Human embryonic stem cell (hESC) research has sparked incredible scientific and public excitement, as well as significant controversy. hESCs are pluripotent, which means, in theory, that they can be differentiated into any type of cell found in the human body. Thus, they evoke great enthusiasm about potential clinical applications. They are…

  19. Humanization policy in primary health care: a systematic review

    Science.gov (United States)

    Nora, Carlise Rigon Dalla; Junges, José Roque

    2013-01-01

    OBJECTIVE To analyze humanization practices in primary health care in the Brazilian Unified Health System according to the principles of the National Humanization Policy. METHODS A systematic review of the literature was carried out, followed by a meta-synthesis, using the following databases: BDENF (nursing database), BDTD (Brazilian digital library of theses and dissertations), CINAHL (Cumulative Index to nursing and allied health literature), LILACS (Latin American and Caribbean health care sciences literature), MedLine (International health care sciences literature), PAHO (Pan-American Health Care Organization Library) and SciELO (Scientific Electronic Library Online). The following descriptors were used: Humanization; Humanizing Health Care; Reception: Humanized care: Humanization in health care; Bonding; Family Health Care Program; Primary Care; Public Health and Sistema Único de Saúde (the Brazilian public health care system). Research articles, case studies, reports of experiences, dissertations, theses and chapters of books written in Portuguese, English or Spanish, published between 2003 and 2011, were included in the analysis. RESULTS Among the 4,127 publications found on the topic, 40 studies were evaluated and included in the analysis, producing three main categories: the first referring to the infrastructure and organization of the primary care service, made clear the dissatisfaction with the physical structure and equipment of the services and with the flow of attendance, which can facilitate or make difficult the access. The second, referring to the health work process, showed issues about the insufficient number of professionals, fragmentation of the work processes, the professional profile and responsibility. The third category, referring to the relational technologies, indicated the reception, bonding, listening, respect and dialog with the service users. CONCLUSIONS Although many practices were cited as humanizing they do not produce changes

  20. [Humanization policy in primary health care: a systematic review].

    Science.gov (United States)

    Nora, Carlise Rigon Dalla; Junges, Jose Roque

    2013-12-01

    To analyze humanization practices in primary health care in the Brazilian Unified Health System according to the principles of the National Humanization Policy. A systematic review of the literature was carried out, followed by a meta-synthesis, using the following databases: BDENF (nursing database), BDTD (Brazilian digital library of theses and dissertations), CINAHL (Cumulative Index to nursing and allied health literature), LILACS (Latin American and Caribbean health care sciences literature), MedLine (International health care sciences literature), PAHO (Pan-American Health Care Organization Library) and SciELO (Scientific Electronic Library Online). The following descriptors were used: Humanization; Humanizing Health Care; Reception: Humanized care: Humanization in health care; Bonding; Family Health Care Program; Primary Care; Public Health and Sistema Único de Saúde (the Brazilian public health care system). Research articles, case studies, reports of experiences, dissertations, theses and chapters of books written in Portuguese, English or Spanish, published between 2003 and 2011, were included in the analysis. Among the 4,127 publications found on the topic, 40 studies were evaluated and included in the analysis, producing three main categories: the first referring to the infrastructure and organization of the primary care service, made clear the dissatisfaction with the physical structure and equipment of the services and with the flow of attendance, which can facilitate or make difficult the access. The second, referring to the health work process, showed issues about the insufficient number of professionals, fragmentation of the work processes, the professional profile and responsibility. The third category, referring to the relational technologies, indicated the reception, bonding, listening, respect and dialog with the service users. Although many practices were cited as humanizing they do not produce changes in the health services because of the

  1. Anti-leukemia activity of in vitro-expanded human gamma delta T cells in a xenogeneic Ph+ leukemia model.

    Directory of Open Access Journals (Sweden)

    Gabrielle M Siegers

    Full Text Available Gamma delta T cells (GDTc lyse a variety of hematological and solid tumour cells in vitro and in vivo, and are thus promising candidates for cellular immunotherapy. We have developed a protocol to expand human GDTc in vitro, yielding highly cytotoxic Vgamma9/Vdelta2 CD27/CD45RA double negative effector memory cells. These cells express CD16, CD45RO, CD56, CD95 and NKG2D. Flow cytometric, clonogenic, and chromium release assays confirmed their specific cytotoxicity against Ph(+ cell lines in vitro. We have generated a fluorescent and bioluminescent Ph(+ cell line, EM-2eGFPluc, and established a novel xenogeneic leukemia model. Intravenous injection of EM-2eGFPluc into NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG mice resulted in significant dose-dependent bone marrow engraftment; lower levels engrafted in blood, lung, liver and spleen. In vitro-expanded human GDTc injected intraperitoneally were found at higher levels in blood and organs compared to those injected intravenously; GDTc survived at least 33 days post-injection. In therapy experiments, we documented decreased bone marrow leukemia burden in mice treated with GDTc. Live GDTc were found in spleen and bone marrow at endpoint, suggesting the potential usefulness of this therapy.

  2. Hypoxia Upregulates Mitotic Cyclins Which Contribute to the Multipotency of Human Mesenchymal Stem Cells by Expanding Proliferation Lifespan.

    Science.gov (United States)

    Lee, Janet; Kim, Hyun-Soo; Kim, Su-Min; Kim, Dong-Ik; Lee, Chang-Woo

    2018-02-21

    Hypoxic culture is widely recognized as a method to efficiently expand human mesenchymal stem cells (MSCs) without loss of stem cell properties. However, the molecular basis of how hypoxia priming benefits MSC expansion remains unclear. In this report, our systemic quantitative proteomic and RT-PCR analyses revealed the involvement of hypoxic conditioning activated genes in the signaling process of the mitotic cell cycle. Introduction of screened two mitotic cyclins, CCNA2 and CCNB1, significantly extended the proliferation lifespan of MSCs in normoxic condition. Our results provide important molecular evidence that multipotency of human MSCs by hypoxic conditioning is determined by the mitotic cell cycle duration. Thus, the activation of mitotic cyclins could be a potential strategy to the application of stem cell therapy.

  3. Expanding Role of T Cells in Human Autoimmune Diseases of the Central Nervous System

    Directory of Open Access Journals (Sweden)

    Deepti Pilli

    2017-06-01

    Full Text Available It is being increasingly recognized that a dysregulation of the immune system plays a vital role in neurological disorders and shapes the treatment of the disease. Aberrant T cell responses, in particular, are key in driving autoimmunity and have been traditionally associated with multiple sclerosis. Yet, it is evident that there are other neurological diseases in which autoreactive T cells have an active role in pathogenesis. In this review, we report on the recent progress in profiling and assessing the functionality of autoreactive T cells in central nervous system (CNS autoimmune disorders that are currently postulated to be primarily T cell driven. We also explore the autoreactive T cell response in a recently emerging group of syndromes characterized by autoantibodies against neuronal cell-surface proteins. Common methodology implemented in T cell biology is further considered as it is an important determinant in their detection and characterization. An improved understanding of the contribution of autoreactive T cells expands our knowledge of the autoimmune response in CNS disorders and can offer novel methods of therapeutic intervention.

  4. Human primary liver cancer-derived organoid cultures for disease modeling and drug screening.

    Science.gov (United States)

    Broutier, Laura; Mastrogiovanni, Gianmarco; Verstegen, Monique Ma; Francies, Hayley E; Gavarró, Lena Morrill; Bradshaw, Charles R; Allen, George E; Arnes-Benito, Robert; Sidorova, Olga; Gaspersz, Marcia P; Georgakopoulos, Nikitas; Koo, Bon-Kyoung; Dietmann, Sabine; Davies, Susan E; Praseedom, Raaj K; Lieshout, Ruby; IJzermans, Jan N M; Wigmore, Stephen J; Saeb-Parsy, Kourosh; Garnett, Mathew J; van der Laan, Luc Jw; Huch, Meritxell

    2017-12-01

    Human liver cancer research currently lacks in vitro models that can faithfully recapitulate the pathophysiology of the original tumor. We recently described a novel, near-physiological organoid culture system, wherein primary human healthy liver cells form long-term expanding organoids that retain liver tissue function and genetic stability. Here we extend this culture system to the propagation of primary liver cancer (PLC) organoids from three of the most common PLC subtypes: hepatocellular carcinoma (HCC), cholangiocarcinoma (CC) and combined HCC/CC (CHC) tumors. PLC-derived organoid cultures preserve the histological architecture, gene expression and genomic landscape of the original tumor, allowing for discrimination between different tumor tissues and subtypes, even after long-term expansion in culture in the same medium conditions. Xenograft studies demonstrate that the tumorogenic potential, histological features and metastatic properties of PLC-derived organoids are preserved in vivo. PLC-derived organoids are amenable for biomarker identification and drug-screening testing and led to the identification of the ERK inhibitor SCH772984 as a potential therapeutic agent for primary liver cancer. We thus demonstrate the wide-ranging biomedical utilities of PLC-derived organoid models in furthering the understanding of liver cancer biology and in developing personalized-medicine approaches for the disease.

  5. BiodentineTM is cytocompatible with human primary osteoblasts

    Directory of Open Access Journals (Sweden)

    Miriam Zaccaro SCELZA

    2017-10-01

    Full Text Available Abstract Calcium silicate-based materials have been widely studied due to their resemblance to, and similar applicability of, mineral trioxide aggregate (MTA. Among these, Biodentine™ (BD was specifically designed as a “dentin replacement” material for applications such as root perforations, apexification, treatment of resorptive lesions, and as a retrograde filling material. The present study aimed to assess the in vitro response of human primary osteoblasts to BD using MTA AngelusTM as a reference material, by simultaneously analyzing three different cell viability parameters, namely mitochondrial activity, membrane integrity, and cell density. BD and MTA extracts were prepared by incubation on culture media for 24 h or 42 days after mixing. Primary human osteoblasts were exposed to extracts for 24 h, at 37oC with 5% CO2, and cell viability was evaluated by the XTT, NRU, and CVDE assays. Both materials induced cell viability levels higher than 70% when extracted for 24 h. However, when cells were exposed to extracts with increased conditioning times, MTA presented significant cytotoxic effects (p < 0.05 in comparison to the control and MTA at 24 h. After 42 days, the XTT assay identified a significant reduction in cell viability by BD when compared to the control (p<0.05, despite the fact that levels above the 70% viability cutoff were attained for biocompatible materials. It can be concluded that BD is cytocompatible with human primary osteoblasts, indicating its adequacy in direct contact with bone tissues.

  6. Expandable and Rapidly Differentiating Human Induced Neural Stem Cell Lines for Multiple Tissue Engineering Applications

    Directory of Open Access Journals (Sweden)

    Dana M. Cairns

    2016-09-01

    Full Text Available Limited availability of human neurons poses a significant barrier to progress in biological and preclinical studies of the human nervous system. Current stem cell-based approaches of neuron generation are still hindered by prolonged culture requirements, protocol complexity, and variability in neuronal differentiation. Here we establish stable human induced neural stem cell (hiNSC lines through the direct reprogramming of neonatal fibroblasts and adult adipose-derived stem cells. These hiNSCs can be passaged indefinitely and cryopreserved as colonies. Independently of media composition, hiNSCs robustly differentiate into TUJ1-positive neurons within 4 days, making them ideal for innervated co-cultures. In vivo, hiNSCs migrate, engraft, and contribute to both central and peripheral nervous systems. Lastly, we demonstrate utility of hiNSCs in a 3D human brain model. This method provides a valuable interdisciplinary tool that could be used to develop drug screening applications as well as patient-specific disease models related to disorders of innervation and the brain.

  7. The human health effects of Florida red tide (FRT) blooms: an expanded analysis.

    Science.gov (United States)

    Hoagland, Porter; Jin, Di; Beet, Andrew; Kirkpatrick, Barbara; Reich, Andrew; Ullmann, Steve; Fleming, Lora E; Kirkpatrick, Gary

    2014-07-01

    Human respiratory and digestive illnesses can be caused by exposures to brevetoxins from blooms of the marine alga Karenia brevis, also known as Florida red tide (FRT). K. brevis requires macro-nutrients to grow; although the sources of these nutrients have not been resolved completely, they are thought to originate both naturally and anthropogenically. The latter sources comprise atmospheric depositions, industrial effluents, land runoffs, or submerged groundwater discharges. To date, there has been only limited research on the extent of human health risks and economic impacts due to FRT. We hypothesized that FRT blooms were associated with increases in the numbers of emergency room visits and hospital inpatient admissions for both respiratory and digestive illnesses. We sought to estimate these relationships and to calculate the costs of associated adverse health impacts. We developed environmental exposure-response models to test the effects of FRT blooms on human health, using data from diverse sources. We estimated the FRT bloom-associated illness costs, using extant data and parameters from the literature. When controlling for resident population, a proxy for tourism, and seasonal and annual effects, we found that increases in respiratory and digestive illnesses can be explained by FRT blooms. Specifically, FRT blooms were associated with human health and economic effects in older cohorts (≥55 years of age) in six southwest Florida counties. Annual costs of illness ranged from $60,000 to $700,000 annually, but these costs could exceed $1.0 million per year for severe, long-lasting FRT blooms, such as the one that occurred during 2005. Assuming that the average annual illness costs of FRT blooms persist into the future, using a discount rate of 3%, the capitalized costs of future illnesses would range between $2 and 24 million. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Replacement of nickel sealing rings by expanded graphite sealing rings -upgrading of SG primary collector flange connection

    Energy Technology Data Exchange (ETDEWEB)

    Cikryt, F.; Bednarek, L.; Kusyn, L. [Vitkovice, Ostrava (Switzerland)

    1997-12-31

    One of the most loaded parts of a steam generator of VVER 440 MW type are the bolts and thread holes of the primary collector cover sealing set. The strength calculations and tensometric measurings performed during operation proved the high degree of a load on the bolts. The conditions of the stress limitation are not met in some cases according to the pertinent standards. The untightnesses at nickel rings occurred during putting the units of Jaslovske Bohunice and Dukovany nuclear power stations into operation. With regard to improve the reliability, the producer has taken measures to improve the quality of the rings and users have introduced more strict regulations for bolts tightening. Due to these measures the high reliability of the set has been obtained from point of view of the tightness, but substantial reduction of bolts and holes threads loading have not been obtained. Several years operation experience proved relatively low service of bolts, damage of thread holes and sealing grooves. As the degree of mechanical load is one of the vital parameters influencing the damage of sealing set, in 1996 we started with the works aimed at a possibility of nickel sealing rings replacement for a more modern type of sealing which assure the higher reliability and service life of the individual part of sealing set under the reduced load.

  9. Human primary auditory cortex follows the shape of Heschl's gyrus.

    Science.gov (United States)

    Da Costa, Sandra; van der Zwaag, Wietske; Marques, Jose P; Frackowiak, Richard S J; Clarke, Stephanie; Saenz, Melissa

    2011-10-05

    The primary auditory cortex (PAC) is central to human auditory abilities, yet its location in the brain remains unclear. We measured the two largest tonotopic subfields of PAC (hA1 and hR) using high-resolution functional MRI at 7 T relative to the underlying anatomy of Heschl's gyrus (HG) in 10 individual human subjects. The data reveals a clear anatomical-functional relationship that, for the first time, indicates the location of PAC across the range of common morphological variants of HG (single gyri, partial duplications, and complete duplications). In 20/20 individual hemispheres, two primary mirror-symmetric tonotopic maps were clearly observed with gradients perpendicular to HG. PAC spanned both divisions of HG in cases of partial and complete duplications (11/20 hemispheres), not only the anterior division as commonly assumed. Specifically, the central union of the two primary maps (the hA1-R border) was consistently centered on the full Heschl's structure: on the gyral crown of single HGs and within the sulcal divide of duplicated HGs. The anatomical-functional variants of PAC appear to be part of a continuum, rather than distinct subtypes. These findings significantly revise HG as a marker for human PAC and suggest that tonotopic maps may have shaped HG during human evolution. Tonotopic mappings were based on only 16 min of fMRI data acquisition, so these methods can be used as an initial mapping step in future experiments designed to probe the function of specific auditory fields.

  10. Expanding the limits of human blood metabolite quantitation using NMR spectroscopy.

    Science.gov (United States)

    Nagana Gowda, G A; Gowda, Yashas N; Raftery, Daniel

    2015-01-06

    A current challenge in metabolomics is the reliable quantitation of many metabolites. Limited resolution and sensitivity combined with the challenges associated with unknown metabolite identification have restricted both the number and the quantitative accuracy of blood metabolites. Focused on alleviating this bottleneck in NMR-based metabolomics, investigations of pooled human serum combining an array of 1D/2D NMR experiments at 800 MHz, database searches, and spiking with authentic compounds enabled the identification of 67 blood metabolites. Many of these (∼1/3) are new compared with those reported previously as a part of the Human Serum Metabolome Database. In addition, considering both the high reproducibility and quantitative nature of NMR as well as the sensitivity of NMR chemical shifts to altered sample conditions, experimental protocols and comprehensive peak annotations are provided here as a guide for identification and quantitation of the new pool of blood metabolites for routine applications. Further, investigations focused on the evaluation of quantitation using organic solvents revealed a surprisingly poor performance for protein precipitation using acetonitrile. One-third of the detected metabolites were attenuated by 10-67% compared with methanol precipitation at the same solvent-to-serum ratio of 2:1 (v/v). Nearly 2/3 of the metabolites were further attenuated by up to 65% upon increasing the acetonitrile-to-serum ratio to 4:1 (v/v). These results, combined with the newly established identity for many unknown metabolites in the NMR spectrum, offer new avenues for human serum/plasma-based metabolomics. Further, the ability to quantitatively evaluate nearly 70 blood metabolites that represent numerous classes, including amino acids, organic acids, carbohydrates, and heterocyclic compounds, using a simple and highly reproducible analytical method such as NMR may potentially guide the evaluation of samples for analysis using mass spectrometry.

  11. Cost and yield considerations when expanding recruitment for genetic studies: the primary open-angle African American glaucoma genetics study

    Directory of Open Access Journals (Sweden)

    Rebecca Salowe

    2017-07-01

    Full Text Available Abstract Background African Americans have been historically under-represented in genetic studies. More research is needed on effective recruitment strategies for this population, especially on approaches that supplement traditional clinic enrollment. This study evaluates the cost and efficacy of four supplemental recruitment methods employed by the Primary Open-Angle African American Glaucoma Genetics (POAAGG study. Methods After enrolling 2304 patients from University of Pennsylvania ophthalmology clinics, the POAAGG study implemented four new recruitment methods to supplement clinic enrollment. These methods included: 1 outreach in the local community, 2 in-house screening of community members (“in-reach”, 3 expansion to two external sites, and 4 sampling of the Penn Medicine Biobank. The cost per subject was calculated for each method and enrollment among cases, controls, and suspects was reported. Results The biobank offered the lowest cost ($5/subject and highest enrollment yield (n = 2073 of the four methods, but provided very few glaucoma cases (n = 31. External sites provided 88% of cases recruited from the four methods (n = 388; $85/subject, but case enrollment at these sites declined over the next 9 months as the pool of eligible subjects was depleted. Outreach and in-reach screenings of community members were very high cost for low return on enrollment ($569/subject for 102 subjects and $606/subject for 45 subjects, respectively. Conclusions The biobank offered the most cost-effective method for control enrollment, while expansion to external sites was necessary to recruit richly phenotyped cases. These recruitment methods helped the POAAGG study to exceed enrollment of the discovery cohort (n = 5500 6 months in advance of the predicated deadline and could be adopted by other large genetic studies seeking to supplement clinic enrollment.

  12. Diabetes Case Management in Primary Care: The New Brunswick Experience and Expanding the Practice of the Certified Diabetes Educator Nurse into Primary Care.

    Science.gov (United States)

    Jones, Shelley L

    2015-08-01

    The role of the outreach diabetes case manager in New Brunswick, Canada, was first developed in the Moncton Area of Horizon Health Network in response to a physician-identified gap between patients' diagnoses of diabetes and their attendance at the local diabetes education centre. This model of collaborative interprofessional practice increases support for primary care providers and people living with diabetes in that they are being provided the services of certified diabetes educators who can address knowledge gaps with respect to evidence-based guidelines and best practice, promote advancement of diabetes and chronic-disease management therapies and support adherence to treatment plans and self-management practices. This report chronicles a review of the implementation, expansion and evaluation of the outreach diabetes case manager model in the province of New Brunswick, Canada, along with the rationale for development of the role for registered nurses in other jurisdictions. Crown Copyright © 2015. Published by Elsevier Inc. All rights reserved.

  13. Microstructure of mineralized tissues in human primary teeth.

    Science.gov (United States)

    Ruschel, H C; Ligocki, G D; Flaminghi, D L; Fossati, A C M

    2011-01-01

    The aim of this study was to analyze the structural characteristics of the mineralized dental tissues--enamel, dentin and cementum--in primary teeth and to correlate the histological aspects observed in function of the dental type--single-rooted or multi-rooted. Eighteen human primary noncarious teeth were sectioned in facial-lingual (single-rooted) and mesio-distal direction (multi-rooted). One to three samples from each tooth were obtained. The samples were prepared by the ground technique and analyzed under light microscopy at different magnifications. A quantitative and descriptive analysis of the morphology of the mineralized tissues was performed. Spindles, tufts and lamellae were consistently observed mainly in the occlusal surface of the primary molars. The scalloped pattern of the dentinoenamel junction was not always present. The same was seen for zones of interglobular dentin. Dead tracts in dentin and tertiary dentin were observed mainly in single-rooted teeth below areas of dental attrition. Areas of cellular and acellular cementum were observed in the two dental types. Primary teeth have some structural peculiarities and these should be investigated concerning the clinical repercussion.

  14. Perceptions of registered nurses regarding factors influencing service delivery in expanding programmes in a primary healthcare setting

    Directory of Open Access Journals (Sweden)

    Nnoi. A. Xaba

    2012-11-01

    Full Text Available The aim of this study was to explore and describe the perceptions of registered nurses regarding factors influencing service delivery regarding expansion programmes in a primary healthcare setting, using a qualitative approach. The registered nurses, who have been working in the clinics for more than two years and have been exposed to the expansion programmes there, were purposively sampled. Two focus group interviews were conducted in a neutral place and the data collected by the researcher Nnoi A. Xaba (N.A.X.. Data were analysed by the researcher and an independent co-coder using the Tesch method. Categories, subcategories and themes were identified; those that formed the basis of discussion were disabling factors, enabling factors, client-related factors, service-related factors and solutions to problems. It is recommended that integration of programmes and coordination be done at a provincial level and planned together with the training centres in order to alleviate problems in service delivery. Training on expansion programmes in the form of in-service education should be carried out continually in the region.Die doel van die studie was om die persepsie van geregistreerde verpleegkundiges met betrekking tot die faktore wat dienslewering van die uitbreidingsprogramme in ‘n primêre gesondheid opset beinvloed; te eksploreer en te beskryf. ‘n Kwalitatiewe benadering is gevolg in die iutvoering van die studie. ‘n Doelgerigte steekproef is uitgevoer vanuit geregistreerde verpleegkundiges wat vir langer as twee jaar in die klinieke werksaam was en blootgestel is aan die uitbreiding programme. Twee fokus groep onderhoude is deur die navorser Nnoi A. Xaba (N.A.X. in ‘n neutrale opset uitgevoer. Data is deur die navorser en ʼn onafhanklike kodeerder ontleed volgens Tesch se metode van analise. Kategorieë, sub-kategorieë en temas was geidentifiseer. Die kategorieë fundamenteel tot die bespreking behels: remmende faktore, bydraende faktore

  15. Aquatic Insects and their Potential to Contribute to the Diet of the Globally Expanding Human Population

    Science.gov (United States)

    Williams, D. Dudley; Williams, Siân S.

    2017-01-01

    Of the 30 extant orders of true insect, 12 are considered to be aquatic, or semiaquatic, in either some or all of their life stages. Out of these, six orders contain species engaged in entomophagy, but very few are being harvested effectively, leading to over-exploitation and local extinction. Examples of existing practices are given, ranging from the extremes of including insects (e.g., dipterans) in the dietary cores of many indigenous peoples to consumption of selected insects, by a wealthy few, as novelty food (e.g., caddisflies). The comparative nutritional worth of aquatic insects to the human diet and to domestic animal feed is examined. Questions are raised as to whether natural populations of aquatic insects can yield sufficient biomass to be of practicable and sustained use, whether some species can be brought into high-yield cultivation, and what are the requirements and limitations involved in achieving this? PMID:28754025

  16. Aquatic Insects and their Potential to Contribute to the Diet of the Globally Expanding Human Population.

    Science.gov (United States)

    Williams, D Dudley; Williams, Siân S

    2017-07-21

    Of the 30 extant orders of true insect, 12 are considered to be aquatic, or semiaquatic, in either some or all of their life stages. Out of these, six orders contain species engaged in entomophagy, but very few are being harvested effectively, leading to over-exploitation and local extinction. Examples of existing practices are given, ranging from the extremes of including insects (e.g., dipterans) in the dietary cores of many indigenous peoples to consumption of selected insects, by a wealthy few, as novelty food (e.g., caddisflies). The comparative nutritional worth of aquatic insects to the human diet and to domestic animal feed is examined. Questions are raised as to whether natural populations of aquatic insects can yield sufficient biomass to be of practicable and sustained use, whether some species can be brought into high-yield cultivation, and what are the requirements and limitations involved in achieving this?

  17. Cellular Proteome Dynamics during Differentiation of Human Primary Myoblasts

    DEFF Research Database (Denmark)

    Le Bihan, Marie-Catherine; Barrio, Inigo; Mortensen, Tenna Pavia

    2015-01-01

    Muscle stem cells, or satellite cells, play an important role in the maintenance and repair of muscle tissue and have the capacity to proliferate and differentiate in response to physiological or environmental changes. Although they have been extensively studied, the key regulatory steps and the ......Muscle stem cells, or satellite cells, play an important role in the maintenance and repair of muscle tissue and have the capacity to proliferate and differentiate in response to physiological or environmental changes. Although they have been extensively studied, the key regulatory steps...... and the complex temporal protein dynamics accompanying the differentiation of primary human muscle cells remain poorly understood. Here, we demonstrate the advantages of applying a MS-based quantitative approach, stable isotope labeling by amino acids in cell culture (SILAC), for studying human myogenesis...... in vitro and characterize the fine-tuned changes in protein expression underlying the dramatic phenotypic conversion of primary mononucleated human muscle cells during in vitro differentiation to form multinucleated myotubes. Using an exclusively optimized triple encoding SILAC procedure, we generated...

  18. Pulpotomies with portland cement in human primary molars

    Directory of Open Access Journals (Sweden)

    Taísa Regina Conti

    2009-02-01

    Full Text Available Two clinical cases in which Portland cement (PC was applied as a medicament after pulpotomy of mandibular primary molars in children are presented. Pulpotomy using PC was carried out in two mandibular first molars and one mandibular second molar, which were further followed-up. At the 3, 6 and 12-month follow-up appointments, clinical and radiographic examinations of the pulpotomized teeth and their periradicular area revealed that the treatments were successful in maintaining the teeth asymptomatic and preserving pulpal vitality. Additionally, the formation of a dentin bridge immediately below the PC could be observed in the three molars treated. PC may be considered as an effective alternative for primary molar pulpotomies, at least in a short-term period. Randomized clinical trials with human teeth are required in order to determine the suitability of PC before unlimited clinical use can be recommended.

  19. ISOLATION AND PRIMARY CULTURES OF HUMAN INTRAHEPATIC BILE DUCTULAR EPITHELIUM

    Science.gov (United States)

    Demetris, A. J.; Markus, B. H.; Saidman, S.; Fung, J. J.; Makowka, L.; Graner, S.; Duquesnoy, R.; Starzl, T. E.

    2010-01-01

    SUMMARY A technique for the isolation of human intrahepatic bile ductular epithelium, and the establishment of primary cultures using a serum- and growth-factor-supplemented medium combined with a connective tissue substrata is described. Initial cell isolates and monolayer cultures display phenotypic characteristics of biliary epithelial cells (low molecular weight prekeratin positive; albumin, alphafetoprotein, and Factor VIII-related antigen negative). Ultrastructural features of the cultured cells show cell polarization with surface microvilli, numerous interepithelial junctional complexes and cytoplasmic intermediate prekeratin filaments. PMID:3131298

  20. Differential effect of platelet-rich plasma and fetal calf serum on bone marrow-derived human mesenchymal stromal cells expanded in vitro.

    Science.gov (United States)

    Goedecke, Anja; Wobus, Manja; Krech, Mathias; Münch, Nadine; Richter, Katja; Hölig, Kristina; Bornhauser, Martin

    2011-08-01

    Mesenchymal stromal cells (MSCs) derived from various sources have great potential for use in cell-based therapies. Since the proportion of primary MSCs contained in bone marrow or adipose tissue is low, plastic adherence and in vitro expansion are necessary to expand MSCs prior to clinical application. Human platelet-rich plasma has been introduced as an alternative serum source but functional differences have so far not been described. Here we cultured MSCs derived from human bone marrow in medium supplemented with either 10% fetal calf serum (FCS) or 5% and 10% platelet-rich plasma (PRP) until the first or second passage. Parameters under investigation were cell yield, clonogenicity, phenotype as well as migratory and differentiation potential. In addition, the secretion of SDF-1α and the induced migration of CD34(+) haematopoietic stem cells (HSCs) were investigated with regard to the different serum source. The use of PRP resulted in a significantly higher expansion rate and yield at passages 0 and 1. In addition, the level of secreted SDF-1α was significantly increased in the supernatant of MSCs cultured with FCS instead of human PRP. Consistent with this, the migration capacity of MSCs cultured with 10% FCS as well as their capability to induce the migration of CD34(+) haematopoietic progenitors in a transwell assay was higher. Our results demonstrate that human PRP can be seen as an alternative serum source to FCS for MSC cultivation. However, the requirements of the specific clinical application must be carefully considered before the respective serum source is selected. Copyright © 2010 John Wiley & Sons, Ltd.

  1. Secretome Analysis of Human Primary Fibroblasts Undergoing Senescence

    DEFF Research Database (Denmark)

    Rogowska-Wrzesinska, Adelina; Micutkova, Lucia; Diener, Thomas

    Introduction Cultures of diploid human fibroblasts can replicate only a finite number of times; rapid proliferation is followed by decline in replicative frequency and finally cells become senescent and are incapable of further proliferation. Senescencent cells display altered growth, morphology......-degrading. In this study we use proteomic tools to characterise the secretome of young and senescent fibroblasts.   Methods Three independent preparations of primary human foreskin fibroblasts were grown to senescence. Young, rapidly proliferating cells at passage 11 and cells from passage 28 displaying senescent...... spectrometry; peptide count was used to estimate protein abundance.   Results 2DGE based analysis of secretion profiles of young and senescent cells derived from the same cell lineage revealed a number of protein spots differentially expressed. We have observed an increased secretion of matrix...

  2. Reframing cervical cancer prevention. Expanding the field towards prevention of human papillomavirus infections and related diseases.

    Science.gov (United States)

    Bosch, F Xavier; Tsu, Vivien; Vorsters, Alex; Van Damme, Pierre; Kane, Mark A

    2012-11-20

    The reframed paradigm of cervical cancer prevention will include strategic combinations of at least four major components: 1) routine introduction of human papillomavirus (HPV) vaccines to women in all countries, 2) extension and simplification of existing screening programs using HPV-based technology, 3) extension of adapted screening programs to developing populations, and 4) consideration of the broader spectrum of cancers and other diseases preventable by HPV vaccination in women, as well as in men. On a global scale, vaccination of newborns and infants is well established and has developed a successful working infrastructure. The hepatitis B virus (HBV) vaccination programs offer a model for HPV introduction in which newborn and infant immunization achieves a rapid reduction in the prevalence of the HBV carrier rates in immunized cohorts of children, and of liver cirrhosis and liver cancer decades later. In contrast, screening for cervical pre-cancer is largely restricted to industrialized populations and upper social classes in developing countries. The expertise gained by vaccination programs worldwide needs to be coordinated with the traditional cervical cancer prevention community of gynecologists and pathologists. Significant political and advocacy efforts at the Global level (World Health Organization, other United Nations agencies and The GAVI Alliance) need to be organized and reinforced to achieve a meaningful reduction in HPV transmission and its related health conditions and cancers. This desirable goal is now scientifically and technologically attainable, and great progress is being made in obtaining financing for global HPV immunization. This article forms part of a special supplement entitled "Comprehensive Control of HPV Infections and Related Diseases" Vaccine Volume 30, Supplement 5, 2012. Copyright © 2012 Elsevier Ltd. All rights reserved.

  3. Expanded human blood-derived γδT cells display potent antigen-presentation functions

    Directory of Open Access Journals (Sweden)

    Mohd Wajid Ali Khan

    2014-07-01

    Full Text Available Cell-based immunotherapy strategies target tumors directly (via cytolytic effector cells or aim at mobilizing endogenous anti-tumor immunity. The latter approach includes dendritic cells (DC, most frequently in the form of in vitro cultured peripheral blood monocytes-derived DC. Human blood γδT cells are selective for a single class of non-peptide agonists (phosphoantigens and develop into potent antigen-presenting cells (APC, termed γδT-APC, within 1-3 days of in vitro culture. Availability of large numbers of γδT-APC would be advantageous for use as a novel cellular vaccine. We here report optimal γδT cell expansion (>107 cells/ml blood when peripheral blood mononuclear cells (PBMC from healthy individuals and melanoma patients were stimulated with zoledronate and then cultured for 14 days in the presence of IL-2 and IL-15, yielding γδT cell cultures of variable purity (77±21% and 56±26%, respectively. They resembled effector-memory αβT (TEM cells and retained full functionality as assessed by in vitro tumor cell killing as well as secretion of proinflammatory cytokines (IFNγ, TNFα and cell proliferation in response to stimulation with phosphoantigens. Importantly, day 14 γδT cells expressed numerous APC-related cell surface markers and, in agreement, displayed potent in vitro APC functions. Day 14 γδT cells from PBMC of patients with cancer were equally effective as their counterparts derived from blood of healthy individuals and triggered potent CD8+ αβT cell responses following processing and cross-presentation of simple (influenza M1 and complex (tuberculin purified protein derivative protein antigens. Of note, and in clear contrast to peripheral blood γδT cells, the ability of day 14 γδT cells to trigger antigen-specific αβT cell responses did not depend on re-stimulation. We conclude that day 14 γδT cell cultures provide a convenient source of autologous APC for use in immunotherapy of

  4. Telomerase contributes to fludarabine resistance in primary human leukemic lymphocytes.

    Directory of Open Access Journals (Sweden)

    May Shawi

    Full Text Available We report that Imetelstat, a telomerase inhibitor that binds to the RNA component of telomerase (hTR, can sensitize primary CLL lymphocytes to fludarabine in vitro. This effect was observed in lymphocytes from clinically resistant cases and with cytogenetic abnormalities associated with bad prognosis. Imetelstat mediated-sensitization to fludarabine was not associated with telomerase activity, but with the basal expression of Ku80. Since both Imetelstat and Ku80 bind hTR, we assessed 1 if Ku80 and Imetelstat alter each other's binding to hTR in vitro and 2 the effect of an oligonucleotide complementary to the Ku binding site in hTR (Ku oligo on the survival of primary CLL lymphocytes exposed to fludarabine. We show that Imetelstat interferes with the binding of Ku70/80 (Ku to hTR and that the Ku oligo can sensitize CLL lymphocytes to FLU. Our results suggest that Ku binding to hTR may contribute to fludarabine resistance in CLL lmphocytes. This is the first report highlighting the potentially broad effectiveness of Imetelstat in CLL, and the potential biological and clinical implications of a functional interaction between Ku and hTR in primary human cancer cells.

  5. Pulpal status of human primary teeth with physiological root resorption.

    Science.gov (United States)

    Monteiro, Joana; Day, Peter; Duggal, Monty; Morgan, Claire; Rodd, Helen

    2009-01-01

    The overall aim of this study was to determine whether any changes occur in the pulpal structure of human primary teeth in association with physiological root resorption. The experimental material comprised 64 sound primary molars, obtained from children requiring routine dental extractions under general anaesthesia. Pulp sections were processed for indirect immunofluorescence using combinations of: (i) protein gene product 9.5 (a general neuronal marker); (ii) leucocyte common antigen CD45 (a general immune cell marker); and (iii) Ulex europaeus I lectin (a marker of vascular endothelium). Image analysis was then used to determine the percentage area of staining for each label within both the pulp horn and mid-coronal region. Following measurement of the greatest degree of root resorption in each sample, teeth were subdivided into three groups: those with physiological resorption involving less than one-third, one-third to two-thirds, and more than two-thirds of their root length. Wide variation was evident between different tooth samples with some resorbed teeth showing marked changes in pulpal histology. Decreased innervation density, increased immune cell accumulation, and increased vascularity were evident in some teeth with advanced root resorption. Analysis of pooled data, however, did not reveal any significant differences in mean percentage area of staining for any of these variables according to the three root resorption subgroups (P > 0.05, analysis of variance on transformed data). This investigation has revealed some changes in pulpal status of human primary teeth with physiological root resorption. These were not, however, as profound as one may have anticipated. It is therefore speculated that teeth could retain the potential for sensation, healing, and repair until advanced stages of root resorption.

  6. [Comparison of the characteristics of expanded cells in human PBMCs activated by combination of MTB-Ag and IL-15 or IL-2 in vitro].

    Science.gov (United States)

    Chen, Yong

    2016-10-01

    Objective To investigate the biological characteristics of expanded cells in human peripheral blood mononuclear cells (PBMCs) activated by the combination of Mycobacterium tuberculosis antigen (MTB-Ag) and IL-15 or IL-2 in vitro. Methods Fresh PBMCs of normal subjects were stimulated with MTB-Ag and IL-15 or MTB-Ag and IL-2 for 12 days. Proportions of γ δ T cells, natural killer (NK) cells and CD4 + T cells in the expanded cells were analyzed by flow cytometry; the numbers of the three lymphocytes were counted by an automated cell counter; the killing activity of the expanded cells against tumor cells was tested by MTT assay. Results Compared with MTB-Ag plus IL-2 group, proportion of γδ T cells was higher significantly, and proportion of NK cells was lower significantly in MTB-Ag plus IL-15 group. There was no statistically significant difference in CD4 + T cell proportion between the two groups. And the same results could be obtained in counting the numbers of the three lymphocytes in both groups. And killing activity of expanded cells from both groups against tumor cells was very strong, and there was no statistically significant difference between both groups. Conclusion γ δ T cells can be activated and expanded in human PBMCs stimulated by MTB-Ag and IL-15 greatly, and the total expanded cells have high cytotoxicity against tumor cells in vitro.

  7. RANK and RANK ligand expression in primary human osteosarcoma

    Directory of Open Access Journals (Sweden)

    Daniel Branstetter

    2015-09-01

    Our results demonstrate RANKL expression was observed in the tumor element in 68% of human OS using IHC. However, the staining intensity was relatively low and only 37% (29/79 of samples exhibited≥10% RANKL positive tumor cells. RANK expression was not observed in OS tumor cells. In contrast, RANK expression was clearly observed in other cells within OS samples, including the myeloid osteoclast precursor compartment, osteoclasts and in giant osteoclast cells. The intensity and frequency of RANKL and RANK staining in OS samples were substantially less than that observed in GCTB samples. The observation that RANKL is expressed in OS cells themselves suggests that these tumors may mediate an osteoclastic response, and anti-RANKL therapy may potentially be protective against bone pathologies in OS. However, the absence of RANK expression in primary human OS cells suggests that any autocrine RANKL/RANK signaling in human OS tumor cells is not operative, and anti-RANKL therapy would not directly affect the tumor.

  8. An expanded self-antigen peptidome is carried by the human lymph as compared to the plasma.

    Directory of Open Access Journals (Sweden)

    Cristina C Clement

    2010-03-01

    Full Text Available The pre-nodal afferent lymph is the fluid which directly derives from the extracellular milieu from every parenchymal organ and, as it continues to circulate between the cells, it collects products deriving from the organ metabolism/catabolism. A comprehensive qualitative and quantitative investigation of the self-antigenic repertoire transported by the human lymph is still missing.A major difference between lymph and plasma could be visualized by FPLC and 2D gel in the amount of low molecular weight products corresponding to peptide fragments. Naturally processed peptides in normal pre-nodal human lymph were then fractionated by HPLC and characterized by multidimensional mass spectrometry. Analysis of more then 300 sequences identified self-peptides derived from both intracellular and extracellular proteins revealing the variety of catabolic products transported by human lymph. Quantitative analysis established that at least some of these peptides are present in the circulating lymph in nanomolar concentration.The peptidome, generated by physiological tissue catabolism and transported by the pre-nodal lymph, is in addition to the self-peptidome generated in endosomal compartment. Unlike self antigen processed by local or nodal APC, which mostly produce epitopes constrained by the endosomal processing activity, self antigens present in the lymph could derived from a wider variety of processing pathways; including caspases, involved in cellular apoptosis, and ADAM and other metalloproteinases involved in surface receptor editing, cytokines processing and matrix remodeling. Altogether, expanding the tissue-specific self-repertoire available for the maintenance of immunological tolerance.

  9. Uptake of gold nanoparticles in primary human endothelial cells

    DEFF Research Database (Denmark)

    Klingberg, Henrik; Oddershede, Lene B.; Löschner, Katrin

    2015-01-01

    Gold nanoparticles (AuNPs) are relevant in nanomedicine for drug delivery in the vascular system, where endothelial cells are the first point of contact. We investigated the uptake of 80 nm AuNPs in primary human umbilical vein endothelial cells (HUVECs) by flow cytometry, 3D confocal microscopy......, nano-scale 3D-imaging using focused ion beam/scanning electron microscopy (FIB/SEM), and single particle inductively coupled plasma-mass spectrometry (spICP-MS). HUVECs were cultured for 3 h or 24 h in medium with AuNPs in a concentration range of 1.25–10 μg ml−1. There was a concentration...

  10. Optimized DNA electroporation for primary human T cell engineering.

    Science.gov (United States)

    Zhang, Zhang; Qiu, Shunfang; Zhang, Xiaopeng; Chen, Wei

    2018-01-30

    Effective gene-delivery systems for primary human T cell engineering are useful tools for both basic research and clinical immunotherapy applications. Pseudovirus-based systems and electro-transfection are the most popular strategies for genetic material transduction. Compared with viral-particle-mediated approaches, electro-transfection is theoretically safer, because it does not promote transgene integration into the host genome. Additionally, the simplicity and speed of the procedure increases the attractiveness of electroporation. Here, we developed and optimized an electro-transfection method for the production of engineered chimeric antigen receptor (CAR)-T cells. Stimulation of T cells had the greatest effect on their transfection, with stimulation of cells for up to 3 days substantially improving transfection efficiency. Additionally, the strength of the external electric field, input cell number, and the initial amount of DNA significantly affected transfection performance. The voltage applied during electroporation affected plasmid permeation and was negatively correlated with the number of viable cells after electroporation. Moreover, higher plasmid concentration increased the proportion of positively transfected cells, but decreased cell viability, and for single-activated cells, higher cell density enhanced their viability. We evaluated the effects of two clinically relevant factors, serum supplementation in the culture medium and cryopreservation immediately after the isolation of peripheral blood lymphocytes. Our findings showed that our protocol performed well using xeno-free cultured, fresh T cells, with application resulting in a lower but acceptable transfection efficiency of cells cultured with fetal bovine serum or thawed cells. Furthermore, we described an optimized procedure to generate CAR-T cells within 6 days and that exhibited cytotoxicity toward targeted cells. Our investigation of DNA electro-transfection for the use in human primary

  11. Youth who sexual offended: primary human goods and offense pathways.

    Science.gov (United States)

    Chu, Chi Meng; Koh, Li Lian; Zeng, Gerald; Teoh, Jennifer

    2015-04-01

    There has been an increased focus on understanding youth sexual offending in recent years, but there has been limited empirical research on the causes, pathways, and treatment of youth who have sexually offended-especially within a non-Western context. The Good Lives and Self-Regulation Models have often been used to understand and rehabilitate adult sexual offenders, but (unfortunately) there is scant research on youth who sexually offended using these models. The present study aims to describe the different primary goods that are associated with youth sexual offending behaviors in an Asian context. In addition, the study sought to explore whether the age of victim (child vs. nonchild) and nature of sexual offense (penetrative vs. nonpenetrative) influenced the youth's engagement in offense pathways. The results suggest that pleasure, relatedness, and inner peace were the primary human goods that were most sought after by a sample of 168 youth who sexually offended in Singapore. In addition, offender classification (in relation to the age of victim and nature of sexual offense) influenced the pathways to sexual offending. Therefore, these findings have important clinical implications for assessment, management, and intervention planning for youth who sexually offended. © The Author(s) 2013.

  12. The cutaneous rabbit illusion affects human primary sensory cortex somatotopically.

    Directory of Open Access Journals (Sweden)

    Felix Blankenburg

    2006-03-01

    Full Text Available We used functional magnetic resonance imaging (fMRI to study neural correlates of a robust somatosensory illusion that can dissociate tactile perception from physical stimulation. Repeated rapid stimulation at the wrist, then near the elbow, can create the illusion of touches at intervening locations along the arm, as if a rabbit hopped along it. We examined brain activity in humans using fMRI, with improved spatial resolution, during this version of the classic cutaneous rabbit illusion. As compared with control stimulation at the same skin sites (but in a different order that did not induce the illusion, illusory sequences activated contralateral primary somatosensory cortex, at a somatotopic location corresponding to the filled-in illusory perception on the forearm. Moreover, the amplitude of this somatosensory activation was comparable to that for veridical stimulation including the intervening position on the arm. The illusion additionally activated areas of premotor and prefrontal cortex. These results provide direct evidence that illusory somatosensory percepts can affect primary somatosensory cortex in a manner that corresponds somatotopically to the illusory percept.

  13. Dopamine Receptor Activation Increases HIV Entry into Primary Human Macrophages

    Science.gov (United States)

    Gaskill, Peter J.; Yano, Hideaki H.; Kalpana, Ganjam V.; Javitch, Jonathan A.; Berman, Joan W.

    2014-01-01

    Macrophages are the primary cell type infected with HIV in the central nervous system, and infection of these cells is a major component in the development of neuropathogenesis and HIV-associated neurocognitive disorders. Within the brains of drug abusers, macrophages are exposed to increased levels of dopamine, a neurotransmitter that mediates the addictive and reinforcing effects of drugs of abuse such as cocaine and methamphetamine. In this study we examined the effects of dopamine on HIV entry into primary human macrophages. Exposure to dopamine during infection increased the entry of R5 tropic HIV into macrophages, irrespective of the concentration of the viral inoculum. The entry pathway affected was CCR5 dependent, as antagonizing CCR5 with the small molecule inhibitor TAK779 completely blocked entry. The effect was dose-dependent and had a steep threshold, only occurring above 108 M dopamine. The dopamine-mediated increase in entry required dopamine receptor activation, as it was abrogated by the pan-dopamine receptor antagonist flupenthixol, and could be mediated through both subtypes of dopamine receptors. These findings indicate that the effects of dopamine on macrophages may have a significant impact on HIV pathogenesis. They also suggest that drug-induced increases in CNS dopamine may be a common mechanism by which drugs of abuse with distinct modes of action exacerbate neuroinflammation and contribute to HIV-associated neurocognitive disorders in infected drug abusers. PMID:25268786

  14. Human Rights Texts: Converting Human Rights Primary Source Documents into Data

    Science.gov (United States)

    Fariss, Christopher J.; Linder, Fridolin J.; Jones, Zachary M.; Crabtree, Charles D.; Biek, Megan A.; Ross, Ana-Sophia M.; Kaur, Taranamol; Tsai, Michael

    2015-01-01

    We introduce and make publicly available a large corpus of digitized primary source human rights documents which are published annually by monitoring agencies that include Amnesty International, Human Rights Watch, the Lawyers Committee for Human Rights, and the United States Department of State. In addition to the digitized text, we also make available and describe document-term matrices, which are datasets that systematically organize the word counts from each unique document by each unique term within the corpus of human rights documents. To contextualize the importance of this corpus, we describe the development of coding procedures in the human rights community and several existing categorical indicators that have been created by human coding of the human rights documents contained in the corpus. We then discuss how the new human rights corpus and the existing human rights datasets can be used with a variety of statistical analyses and machine learning algorithms to help scholars understand how human rights practices and reporting have evolved over time. We close with a discussion of our plans for dataset maintenance, updating, and availability. PMID:26418817

  15. Human Rights Texts: Converting Human Rights Primary Source Documents into Data.

    Science.gov (United States)

    Fariss, Christopher J; Linder, Fridolin J; Jones, Zachary M; Crabtree, Charles D; Biek, Megan A; Ross, Ana-Sophia M; Kaur, Taranamol; Tsai, Michael

    2015-01-01

    We introduce and make publicly available a large corpus of digitized primary source human rights documents which are published annually by monitoring agencies that include Amnesty International, Human Rights Watch, the Lawyers Committee for Human Rights, and the United States Department of State. In addition to the digitized text, we also make available and describe document-term matrices, which are datasets that systematically organize the word counts from each unique document by each unique term within the corpus of human rights documents. To contextualize the importance of this corpus, we describe the development of coding procedures in the human rights community and several existing categorical indicators that have been created by human coding of the human rights documents contained in the corpus. We then discuss how the new human rights corpus and the existing human rights datasets can be used with a variety of statistical analyses and machine learning algorithms to help scholars understand how human rights practices and reporting have evolved over time. We close with a discussion of our plans for dataset maintenance, updating, and availability.

  16. Quantitative effects of a nickel-titanium palatal expander on skeletal and dental structures in the primary and mixed dentition: a preliminary study.

    Science.gov (United States)

    Ferrario, Virgilio F; Garattini, Giovanna; Colombo, Anna; Filippi, Vittorio; Pozzoli, Silvio; Sforza, Chiarella

    2003-08-01

    The present study analysed the six-month effects of a nickel-titanium (NiTi) palatal expander on the dental and palatal structures of four primary (mean age 5.8 years) and nine mixed dentition children (mean age 8.7 years), with a posterior unilateral crossbite. Standardized dental and palatal landmarks were digitized using a three-dimensional (3D) electromagnetic instrument. Collected data were analysed with geometric-mathematical models. During a six-month interval, the natural growth and development of the dental arches and hard tissue palate was negligible, as assessed in seven control children (two in the primary dentition, mean age 4.4 years; five in the mixed dentition, mean age 7.7 years). In all children the crossbite was completely corrected. Indeed, dental expansion was always more than or corresponded to the palatal expansion. A smoothing of the size-independent (shape) palatal curvature in the transverse plane was observed. No differences in maximum palatal height were noted. Symmetrical derotation of the anchorage teeth in a distal direction occurred in almost all children. The inclination of the facial axis of the clinical crown (FACC) in the anatomical transverse plane of those teeth with differences between dental and palatal expansion always showed significant modifications (vestibular inclination up to 16.7 degrees). The clinical crown height of anchorage teeth remained nearly the same in all patients. No significant modifications in mandibular arch size were observed. The increase in maxillary arch width, especially in younger children, was probably due to a combination of different effects: opening of the midpalatal suture, tipping of the alveolar process, and molar tipping.

  17. Steroid synthesis by primary human keratinocytes; implications for skin disease

    Energy Technology Data Exchange (ETDEWEB)

    Hannen, Rosalind F., E-mail: r.f.hannen@qmul.ac.uk [Centre for Cutaneous Research, Institute of Cell and Molecular Science, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London E1 2AT (United Kingdom); Michael, Anthony E. [Centre for Developmental and Endocrine Signalling, Academic Section of Obstetrics and Gynaecology, Division of Clinical Developmental Sciences, 3rd Floor, Lanesborough Wing, St. George' s, University of London, Cranmer Terrace, Tooting, London SW17 0RE (United Kingdom); Jaulim, Adil [Centre for Cutaneous Research, Institute of Cell and Molecular Science, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London E1 2AT (United Kingdom); Bhogal, Ranjit [Life Science, Unilever R and D Colworth House, Sharnbrook, Bedfordshire MK44 1LQ (United Kingdom); Burrin, Jacky M. [Centre for Endocrinology, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London EC1M 6BQ (United Kingdom); Philpott, Michael P. [Centre for Cutaneous Research, Institute of Cell and Molecular Science, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London E1 2AT (United Kingdom)

    2011-01-07

    Research highlights: {yields} Primary keratinocytes express the steroid enzymes required for cortisol synthesis. {yields} Normal primary human keratinocytes can synthesise cortisol. {yields} Steroidogenic regulators, StAR and MLN64, are expressed in normal epidermis. {yields} StAR expression is down regulated in eczema and psoriatic epidermis. -- Abstract: Cortisol-based therapy is one of the most potent anti-inflammatory treatments available for skin conditions including psoriasis and atopic dermatitis. Previous studies have investigated the steroidogenic capabilities of keratinocytes, though none have demonstrated that these skin cells, which form up to 90% of the epidermis are able to synthesise cortisol. Here we demonstrate that primary human keratinocytes (PHK) express all the elements required for cortisol steroidogenesis and metabolise pregnenolone through each intermediate steroid to cortisol. We show that normal epidermis and cultured PHK express each of the enzymes (CYP11A1, CYP17A1, 3{beta}HSD1, CYP21 and CYP11B1) that are required for cortisol synthesis. These enzymes were shown to be metabolically active for cortisol synthesis since radiometric conversion assays traced the metabolism of [7-{sup 3}H]-pregnenolone through each steroid intermediate to [7-{sup 3}H]-cortisol in cultured PHK. Trilostane (a 3{beta}HSD1 inhibitor) and ketoconazole (a CYP17A1 inhibitor) blocked the metabolism of both pregnenolone and progesterone. Finally, we show that normal skin expresses two cholesterol transporters, steroidogenic acute regulatory protein (StAR), regarded as the rate-determining protein for steroid synthesis, and metastatic lymph node 64 (MLN64) whose function has been linked to cholesterol transport in steroidogenesis. The expression of StAR and MLN64 was aberrant in two skin disorders, psoriasis and atopic dermatitis, that are commonly treated with cortisol, suggesting dysregulation of epidermal steroid synthesis in these patients. Collectively these data

  18. Interhemispheric interactions between the human primary somatosensory cortices.

    Directory of Open Access Journals (Sweden)

    Patrick Ragert

    Full Text Available In the somatosensory domain it is still unclear at which processing stage information reaches the opposite hemispheres. Due to dense transcallosal connections, the secondary somatosensory cortex (S2 has been proposed to be the key candidate for interhemispheric information transfer. However, recent animal studies showed that the primary somatosensory cortex (S1 might as well account for interhemispheric information transfer. Using paired median nerve somatosensory evoked potential recordings in humans we tested the hypothesis that interhemispheric inhibitory interactions in the somatosensory system occur already in an early cortical processing stage such as S1. Conditioning right S1 by electrical median nerve (MN stimulation of the left MN (CS resulted in a significant reduction of the N20 response in the target (left S1 relative to a test stimulus (TS to the right MN alone when the interstimulus interval between CS and TS was between 20 and 25 ms. No such changes were observed for later cortical components such as the N20/P25, N30, P40 and N60 amplitude. Additionally, the subcortically generated P14 response in left S1 was also not affected. These results document the existence of interhemispheric inhibitory interactions between S1 in human subjects in the critical time interval of 20-25 ms after median nerve stimulation.

  19. High expression of DNA methyltransferases in primary human medulloblastoma.

    Science.gov (United States)

    Pócza, T; Krenács, T; Turányi, E; Csáthy, J; Jakab, Z; Hauser, P

    2016-01-01

    Epigenetic alterations have been implicated in cancer development. DNA methylation modulates gene expression, which is catalyzed by DNA methyltransferases (DNMTs). The objective of our study was to evaluate expression of DNMTs in medulloblastoma and analyze its correlation with clinical features. Nuclear expression of DNMT1, DNMT3A and DNMT3B was analyzed in human primary medulloblastoma of 44 patients using immunohistochemistry. Correlation of expression of DNMT levels with classical histological subtypes, novel molecular subgroups and survival of patients was analyzed. Elevated expression of DNMT1, DNMT3A and DNMT3B was observed in 63.64%, 68.18% and 72.73% of all cases, respectively. None of them showed a correlation with classical histology or survival. Concerning molecular subtypes, significantly higher expression of DNMT1 was observed in the SHH group compared to non-SHH samples (p = 0.02), but without significant difference in DNMT3A or DNMT3B levels between any subtypes. In conclusion, DNMT1, DNMT3A and DNMT3B are highly expressed in human medulloblastoma samples, suggesting that promoter hypermethylation may play a role in medulloblastoma development. Demethylation of tumor suppressor gene promoters may be considered as a possible future target in therapy of medulloblastoma.

  20. Primary cilia-mediated mechanotransduction in human mesenchymal stem cells.

    Science.gov (United States)

    Hoey, David A; Tormey, Shane; Ramcharan, Stacy; O'Brien, Fergal J; Jacobs, Christopher R

    2012-11-01

    Physical loading is a potent stimulus required to maintain bone homeostasis, partly through the renewal and osteogenic differentiation of mesenchymal stem cells (MSCs). However, the mechanism by which MSCs sense a biophysical force and translate that into a biochemical bone forming response (mechanotransduction) remains poorly understood. The primary cilium is a single sensory cellular extension, which has recently been shown to demonstrate a role in cellular mechanotransduction and MSC lineage commitment. In this study, we present evidence that short periods of mechanical stimulation in the form of oscillatory fluid flow (OFF) is sufficient to enhance osteogenic gene expression and proliferation of human MSCs (hMSCs). Furthermore, we demonstrate that the cilium mediates fluid flow mechanotransduction in hMSCs by maintaining OFF-induced increases in osteogenic gene expression and, surprisingly, to limit OFF-induced increases in proliferation. These data therefore demonstrate a pro-osteogenic mechanosensory role for the primary cilium, establishing a novel mechanotransduction mechanism in hMSCs. Based on these findings, the application of OFF may be a beneficial component of bioreactor-based strategies to form bone-like tissues suitable for regenerative medicine and also highlights the cilium as a potential therapeutic target for efforts to mimic loading with the aim of preventing bone loss during diseases such as osteoporosis. Furthermore, this study demonstrates a role for the cilium in controlling mechanically mediated increases in the proliferation of hMSCs, which parallels proposed models of polycystic kidney disease. Unraveling the mechanisms leading to rapid proliferation of mechanically stimulated MSCs with defective cilia could provide significant insights regarding ciliopathies and cystic diseases. Copyright © 2012 AlphaMed Press.

  1. CD8αα expression marks terminally differentiated human CD8+ T cells expanded in chronic viral infection

    Directory of Open Access Journals (Sweden)

    Lucy Jane Walker

    2013-08-01

    Full Text Available The T cell co-receptor CD8αβ enhances T cell sensitivity to antigen, however studies indicate CD8αα has the converse effect and acts as a co-repressor. Using a combination of Thymic Leukaemia antigen (TL tetramer, which directly binds CD8αα, anti-CD161 and anti-Vα7.2 antibodies we have been able for the first time to clearly define CD8αα expression on human CD8 T cells subsets. In healthy controls CD8αα is most highly expressed by CD161 bright (CD161++ mucosal associated invariant T (MAIT cells, with CD8αα expression highly restricted to the TCR Vα7.2+ cells of this subset. We also identified CD8αα-expressing populations within the CD161 mid (CD161+ and negative (CD161- non-MAIT CD8 T cell subsets and show TL-tetramer binding to correlate with expression of CD8β at low levels in the context of maintained CD8α expression (CD8α+CD8βlow. In addition, we found CD161-CD8α+CD8βlow populations to be significantly expanded in the peripheral blood of HIV-1 and hepatitis B (mean of 47% and 40% of CD161- T cells respectively infected individuals. Such CD8αα expressing T cells are an effector-memory population (CD45RA-, CCR7-, CD62L- that express markers of activation and maturation (HLA-DR+, CD28-, CD27-, CD57+ and are functionally distinct, expressing greater levels of TNF-α and IFN-γ on stimulation and perforin at rest than their CD8α+CD8βhigh counterparts. Antigen-specific T cells in HLA-B*4201+HIV-1 infected patients are found within both the CD161-CD8α+CD8βhigh and CD161-CD8α+CD8βlow populations. Overall we have clearly defined CD8αα expressing human T cell subsets using the TL-tetramer, and have demonstrated CD161-CD8α+CD8βlow populations, highly expanded in disease settings, to co-express CD8αβ and CD8αα. Co-expression of CD8αα on CD8αβ T cells may impact on their overall function in-vivo and contribute to the distinctive phenotype of highly differentiated populations in HBV and HIV-1 infection.

  2. Expanding subjectivities

    DEFF Research Database (Denmark)

    Lundgaard Andersen, Linda; Soldz, Stephen

    2012-01-01

    A major theme in recent psychoanalytic thinking concerns the use of therapist subjectivity, especially “countertransference,” in understanding patients. This thinking converges with and expands developments in qualitative research regarding the use of researcher subjectivity as a tool to understa...

  3. Expander Codes

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 10; Issue 1. Expander Codes - The Sipser–Spielman Construction. Priti Shankar. General Article Volume 10 ... Author Affiliations. Priti Shankar1. Department of Computer Science and Automation, Indian Institute of Science Bangalore 560 012, India.

  4. Long-term in-vivo tumorigenic assessment of human culture-expanded adipose stromal/stem cells

    Energy Technology Data Exchange (ETDEWEB)

    MacIsaac, Zoe Marie, E-mail: zmm4a@virgina.edu [University of Virginia (United States); Shang, Hulan, E-mail: shanghulan@gmail.com [Department of Plastic Surgery, University of Virginia (United States); Agrawal, Hitesh, E-mail: hiteshdos@hotmail.com [Department of Plastic Surgery, University of Virginia (United States); Yang, Ning, E-mail: ny6u@virgina.edu [Department of Plastic Surgery, University of Virginia (United States); Parker, Anna, E-mail: amp4v@virginia.edu [Department of Surgery, University of Virginia (United States); Katz, Adam J., E-mail: ajk2f@virginia.edu [Department of Plastic Surgery, University of Virginia (United States)

    2012-02-15

    After more than a decade of extensive experimentation, the promise of stem cells to revolutionize the field of medicine has negotiated their entry into clinical trial. Adipose tissue specifically holds potential as an attainable and abundant source of stem cells. Currently undergoing investigation are adipose stem cell (ASC) therapies for diabetes and critical limb ischemia, among others. In the enthusiastic pursuit of regenerative therapies, however, questions remain regarding ASC persistence and migration, and, importantly, their safety and potential for neoplasia. To date, assays of in vivo ASC activity have been limited by early end points. We hypothesized that with time, ASCs injected subcutaneously undergo removal by normal tissue turnover and homeostasis, and by the host's immune system. In this study, a high dose of culture expanded ASCs was formulated and implanted as multicellular aggregates into immunocompromised mice, which were maintained for over one year. Animals were monitored for toxicity, and surviving cells quantified at study endpoint. No difference in growth/weight or lifespan was found between cell-treated and vehicle treated animals, and no malignancies were detected in treated animals. Moreover, real-time PCR for a human specific sequence, ERV-3, detected no persistent ASCs. With the advent of clinical application, clarification of currently enigmatic stem cell properties has become imperative. Our study represents the longest duration determination of stem cell activity in vivo, and contributes strong evidence in support of the safety of adipose derived stem cell applications. -- Highlights: Black-Right-Pointing-Pointer Adipose stem cells promise novel clinical therapies. Black-Right-Pointing-Pointer Before clinical translation, safety profiles must be further elucidated. Black-Right-Pointing-Pointer Subcutaneously injected non-autologous adipose stem cells do not form tumors. Black-Right-Pointing-Pointer Subcutaneously injected non

  5. Functional sex differences in human primary auditory cortex

    Energy Technology Data Exchange (ETDEWEB)

    Ruytjens, Liesbet [University Medical Center Groningen, Department of Otorhinolaryngology, Groningen (Netherlands); University Medical Center Utrecht, Department Otorhinolaryngology, P.O. Box 85500, Utrecht (Netherlands); Georgiadis, Janniko R. [University of Groningen, University Medical Center Groningen, Department of Anatomy and Embryology, Groningen (Netherlands); Holstege, Gert [University of Groningen, University Medical Center Groningen, Center for Uroneurology, Groningen (Netherlands); Wit, Hero P. [University Medical Center Groningen, Department of Otorhinolaryngology, Groningen (Netherlands); Albers, Frans W.J. [University Medical Center Utrecht, Department Otorhinolaryngology, P.O. Box 85500, Utrecht (Netherlands); Willemsen, Antoon T.M. [University Medical Center Groningen, Department of Nuclear Medicine and Molecular Imaging, Groningen (Netherlands)

    2007-12-15

    We used PET to study cortical activation during auditory stimulation and found sex differences in the human primary auditory cortex (PAC). Regional cerebral blood flow (rCBF) was measured in 10 male and 10 female volunteers while listening to sounds (music or white noise) and during a baseline (no auditory stimulation). We found a sex difference in activation of the left and right PAC when comparing music to noise. The PAC was more activated by music than by noise in both men and women. But this difference between the two stimuli was significantly higher in men than in women. To investigate whether this difference could be attributed to either music or noise, we compared both stimuli with the baseline and revealed that noise gave a significantly higher activation in the female PAC than in the male PAC. Moreover, the male group showed a deactivation in the right prefrontal cortex when comparing noise to the baseline, which was not present in the female group. Interestingly, the auditory and prefrontal regions are anatomically and functionally linked and the prefrontal cortex is known to be engaged in auditory tasks that involve sustained or selective auditory attention. Thus we hypothesize that differences in attention result in a different deactivation of the right prefrontal cortex, which in turn modulates the activation of the PAC and thus explains the sex differences found in the activation of the PAC. Our results suggest that sex is an important factor in auditory brain studies. (orig.)

  6. Irreversible electroporation of human primary uveal melanoma in enucleated eyes.

    Directory of Open Access Journals (Sweden)

    Yossi Mandel

    Full Text Available Uveal melanoma (UM is the most common primary intraocular tumor in adults and is characterized by high rates of metastatic disease. Although brachytherapy is the most common globe-sparing treatment option for small- and medium-sized tumors, the treatment is associated with severe adverse reactions and does not lead to increased survival rates as compared to enucleation. The use of irreversible electroporation (IRE for tumor ablation has potential advantages in the treatment of tumors in complex organs such as the eye. Following previous theoretical work, herein we evaluate the use of IRE for uveal tumor ablation in human ex vivo eye model. Enucleated eyes of patients with uveal melanoma were treated with short electric pulses (50-100 µs, 1000-2000 V/cm using a customized electrode design. Tumor bioimpedance was measured before and after treatment and was followed by histopathological evaluation. We found that IRE caused tumor ablation characterized by cell membrane disruption while sparing the non-cellular sclera. Membrane disruption and loss of cellular capacitance were also associated with significant reduction in total tumor impedance and loss of impedance frequency dependence. The effect was more pronounced near the pulsing electrodes and was dependent on time from treatment to fixation. Future studies should further evaluate the potential of IRE as an alternative method of uveal melanoma treatment.

  7. Punicalagin promotes autophagy to protect primary human syncytiotrophoblasts from apoptosis.

    Science.gov (United States)

    Wang, Ying; Chen, Baosheng; Longtine, Mark S; Nelson, D Michael

    2016-02-01

    Punicalagin is a prominent polyphenol in pomegranate juice that protects cultured syncytiotrophoblasts from stress-induced apoptosis. Here, we test the hypothesis that punicalagin has this effect by inhibiting the mTOR kinase pathway to enhance autophagic turnover and limit apoptosis in cultured primary human syncytiotrophoblasts. In syncytiotrophoblasts, starvation, rapamycin, or punicalagin all decreased the expression of phosphorylated ribosomal protein S6, a downstream target of the mTOR kinase, and of the autophagy markers, LC3-II and p62. In contrast, in the presence of bafilomycin, an inhibitor of late stages of autophagy and degradation in the autophagolysosome, syncytiotrophoblasts exposed to starvation, rapamycin, or punicalagin all showed increased levels of LC3-II and p62. The number of LC3-II punctae also increased in punicalagin-treated syncytiotrophoblasts exposed to chloroquine, another inhibitor of autophagic degradation, and punicalagin increased the number of lysosomes. The apoptosis-reducing effect of punicalagin was attenuated by inhibition of autophagy using bafilomycin or knockdown of the autophagy related gene, ATG16L1. Collectively, these data support the hypothesis that punicalagin modulates the crosstalk between autophagy and apoptosis to promote survival in cultured syncytiotrophoblasts. © 2016 Society for Reproduction and Fertility.

  8. Functional sex differences in human primary auditory cortex.

    Science.gov (United States)

    Ruytjens, Liesbet; Georgiadis, Janniko R; Holstege, Gert; Wit, Hero P; Albers, Frans W J; Willemsen, Antoon T M

    2007-12-01

    We used PET to study cortical activation during auditory stimulation and found sex differences in the human primary auditory cortex (PAC). Regional cerebral blood flow (rCBF) was measured in 10 male and 10 female volunteers while listening to sounds (music or white noise) and during a baseline (no auditory stimulation). We found a sex difference in activation of the left and right PAC when comparing music to noise. The PAC was more activated by music than by noise in both men and women. But this difference between the two stimuli was significantly higher in men than in women. To investigate whether this difference could be attributed to either music or noise, we compared both stimuli with the baseline and revealed that noise gave a significantly higher activation in the female PAC than in the male PAC. Moreover, the male group showed a deactivation in the right prefrontal cortex when comparing noise to the baseline, which was not present in the female group. Interestingly, the auditory and prefrontal regions are anatomically and functionally linked and the prefrontal cortex is known to be engaged in auditory tasks that involve sustained or selective auditory attention. Thus we hypothesize that differences in attention result in a different deactivation of the right prefrontal cortex, which in turn modulates the activation of the PAC and thus explains the sex differences found in the activation of the PAC. Our results suggest that sex is an important factor in auditory brain studies.

  9. 3D Cultivation Techniques for Primary Human Hepatocytes

    Directory of Open Access Journals (Sweden)

    Anastasia Bachmann

    2015-02-01

    Full Text Available One of the main challenges in drug development is the prediction of in vivo toxicity based on in vitro data. The standard cultivation system for primary human hepatocytes is based on monolayer cultures, even if it is known that these conditions result in a loss of hepatocyte morphology and of liver-specific functions, such as drug-metabolizing enzymes and transporters. As it has been demonstrated that hepatocytes embedded between two sheets of collagen maintain their function, various hydrogels and scaffolds for the 3D cultivation of hepatocytes have been developed. To further improve or maintain hepatic functions, 3D cultivation has been combined with perfusion. In this manuscript, we discuss the benefits and drawbacks of different 3D microfluidic devices. For most systems that are currently available, the main issues are the requirement of large cell numbers, the low throughput, and expensive equipment, which render these devices unattractive for research and the drug-developing industry. A higher acceptance of these devices could be achieved by their simplification and their compatibility with high-throughput, as both aspects are of major importance for a user-friendly device.

  10. Linking human beta retrovirus infection with primary biliary cirrhosis.

    Science.gov (United States)

    Mason, A L; Zhang, G

    2010-01-01

    Several environmental agents have been linked with primary biliary cirrhosis (PBC) that include bacteria, xenobiotics and viruses. A human beta retrovirus (HBRV) related to mouse mammary tumor virus has been cloned and characterized from patients with PBC. This agent can be detected in the majority of patients' perihepatic lymph nodes by immunochemistry and RT-PCR. The HBRV has recently been isolated in culture and integration sites have been identified in the genome of patients to provide convincing evidence of beta retrovirus infection in patients. Three lines of evidence support a role for the virus in PBC. First, the beta retrovirus is linked with aberrant expression of mitochondrial protein(s) on the biliary epithelium cell (BEC) surface, a disease specific phenotype. Second, the related agent, mouse mammary tumor virus has been linked with autoimmune biliary disease in the NOD.c3c4 mouse model for PBC. In this mouse model, the virus is localized to diseased biliary epithelium that also display aberrant expression of the mitochondrial autoantigens. In translational studies, both patients with PBC and NOD.c3c4 mice demonstrate significant improvement in biliary disease with combination antiviral therapy. An overview of the biological relevance of the beta retrovirus infection in PBC will be discussed in this review. Copyright 2010 Elsevier Masson SAS. All rights reserved.

  11. Partition expanders

    Czech Academy of Sciences Publication Activity Database

    Gavinsky, Dmitry; Pudlák, Pavel

    2017-01-01

    Roč. 60, č. 3 (2017), s. 378-395 ISSN 1432-4350 R&D Projects: GA ČR GBP202/12/G061 Institutional support: RVO:67985840 Keywords : expanders * pseudorandomness * communication complexity Subject RIV: BA - General Mathematics Impact factor: 0.645, year: 2016 http://link.springer.com/article/10.1007%2Fs00224-016-9738-5

  12. HANPP Collection: Human Appropriation of Net Primary Productivity (HANPP) by Country and Product

    Data.gov (United States)

    National Aeronautics and Space Administration — The Global Patterns in Human Appropriation of Net Primary Productivity (HANPP) portion of the HANPP Collection represents a digital map of human appropriation of net...

  13. HANPP Collection: Global Patterns in Human Appropriation of Net Primary Productivity (HANPP)

    Data.gov (United States)

    National Aeronautics and Space Administration — The Global Patterns in Human Appropriation of Net Primary Productivity (HANPP) portion of the HANPP Collection represents a digital map of human appropriation of net...

  14. Primary Adult Human Retinal Pigment Epithelial Cell Cultures on Human Amniotic Membranes

    Directory of Open Access Journals (Sweden)

    Singhal Shweta

    2005-01-01

    Full Text Available Purpose: Retinal pigment epithelial (RPE cells grow well on surfaces that provide an extracellular matrix. Our aim was to establish primary adult human RPE cell cultures that retain their epithelial morphology in vitro using human amniotic membrane (hAM as substrate. Materials and Methods: Human cadaver eyeballs (16 were obtained from the eye bank after corneal trephination. RPE cells were harvested by a mechanical dissection of the inner choroid surface (10, group 1 or by b enzymatic digestion using 0.25% Trypsin/0.02% EDTA (6, group 2. The cells were explanted onto de-epithelialized hAM, nourished using DMEM/HAMS F-12 media and monitored for growth under the phase contrast microscope. Cell cultures were characterised by whole mount studies and paraffin sections. Growth data in the two groups were compared using the students′ ′t′ test. Results: Eleven samples (68.75% showed positive cultures with small, hexagonal cells arising from around the explant which formed a confluent and progressively pigmented monolayer. Whole mounts showed closely placed polygonal cells with heavily pigmented cytoplasm and indistinct nuclei. The histologic sections showed monolayers of cuboidal epithelium with variable pigmentation within the cytoplasm. Growth was seen by day 6-23 (average 11.5 days in the mechanical group, significantly earlier ( P Conclusions: Primary adult human RPE cell cultures retain epithelial morphology in vitro when cultured on human amniotic membranes . Mechanical dissection of the inner choroid surface appears to be an effective method of isolating RPE cells and yields earlier growth in cultures as compared to isolation by enzymatic digestion

  15. Expanding versus non expanding universe

    CERN Document Server

    Alfonso-Faus, Antonio

    2012-01-01

    In cosmology the number of scientists using the framework of an expanding universe is very high. This model, the big-bang, is now overwhelmingly present in almost all aspects of society. It is the main stream cosmology of today. A small number of scientists are researching on the possibility of a non-expanding universe. The existence of these two groups, one very large and the other very small, is a good proof of the use of the scientific method: it does not drive to an absolute certainty. All models have to be permanently validated, falsified. Ockham's razor, a powerful philosophical tool, will probably change the amount of scientists working in each of these groups. We present here a model where a big-bang is unnecessary. It ends, in a finite time, in a second INFLATION, or a disaggregation to infinity. We also discuss the possibilities of a non-expanding universe model. Only a few references will be cited, mainly concerned with our own work in the past, thus purposely avoiding citing the many thousands of ...

  16. Genome-wide binding and transcriptome analysis of human farnesoid X receptor in primary human hepatocytes.

    Directory of Open Access Journals (Sweden)

    Le Zhan

    Full Text Available Farnesoid X receptor (FXR, NR1H4 is a ligand-activated transcription factor, belonging to the nuclear receptor superfamily. FXR is highly expressed in the liver and is essential in regulating bile acid homeostasis. FXR deficiency is implicated in numerous liver diseases and mice with modulation of FXR have been used as animal models to study liver physiology and pathology. We have reported genome-wide binding of FXR in mice by chromatin immunoprecipitation - deep sequencing (ChIP-seq, with results indicating that FXR may be involved in regulating diverse pathways in liver. However, limited information exists for the functions of human FXR and the suitability of using murine models to study human FXR functions.In the current study, we performed ChIP-seq in primary human hepatocytes (PHHs treated with a synthetic FXR agonist, GW4064 or DMSO control. In parallel, RNA deep sequencing (RNA-seq and RNA microarray were performed for GW4064 or control treated PHHs and wild type mouse livers, respectively.ChIP-seq showed similar profiles of genome-wide FXR binding in humans and mice in terms of motif analysis and pathway prediction. However, RNA-seq and microarray showed more different transcriptome profiles between PHHs and mouse livers upon GW4064 treatment.In summary, we have established genome-wide human FXR binding and transcriptome profiles. These results will aid in determining the human FXR functions, as well as judging to what level the mouse models could be used to study human FXR functions.

  17. Inquiry-Based Learning in Teacher Education: A Primary Humanities Example

    Science.gov (United States)

    Preston, Lou; Harvie, Kate; Wallace, Heather

    2015-01-01

    Inquiry-based learning features strongly in the new Australian Humanities and Social Sciences curriculum and increasingly in primary school practice. Yet, there is little research into, and few exemplars of, inquiry approaches in the primary humanities context. In this article, we outline and explain the implementation of a place-based simulation…

  18. Availability and type of human resource for health in public primary ...

    African Journals Online (AJOL)

    Background: Health human resource capacity is one of the major determinants of an effective health care delivery system. It is more importantly so in the Primary Health Care system, which is the key component of every health system. This study assessed the available and type of human resource for health in public primary ...

  19. Expanded Dengue.

    Science.gov (United States)

    Kadam, D B; Salvi, Sonali; Chandanwale, Ajay

    2016-07-01

    The World Health Organization (WHO) has coined the term expanded dengue to describe cases which do not fall into either dengue shock syndrome or dengue hemorrhagic fever. This has incorporated several atypical findings of dengue. Dengue virus has not been enlisted as a common etiological agent in several conditions like encephalitis, Guillain Barre syndrome. Moreover it is a great mimic of co-existing epidemics like Malaria, Chikungunya and Zika virus disease, which are also mosquito-borne diseases. The atypical manifestations noted in dengue can be mutisystemic and multifacetal. In clinical practice, the occurrence of atypical presentation should prompt us to investigate for dengue. Knowledge of expanded dengue helps to clinch the diagnosis of dengue early, especially during ongoing epidemics, avoiding further battery of investigations. Dengue has proved to be the epidemic with the ability to recur and has a diverse array of presentation as seen in large series from India, Srilanka, Indonesia and Taiwan. WHO has given the case definition of dengue fever in their comprehensive guidelines. Accordingly, a probable case is defined as acute febrile illness with two or more of any findings viz. headache, retro-orbital pain, myalgia, arthralgia, rash, hemorrhagic manifestations, leucopenia and supportive serology. There have been cases of patients admitted with fever, altered mentation with or without neck stiffness and pyramidal tract signs. Some had seizures or status epilepticus as presentation. When they were tested for serology, dengue was positive. After ruling out other causes, dengue remained the only culprit. We have come across varied presentations of dengue fever in clinical practice and the present article throws light on atypical manifestations of dengue. © Journal of the Association of Physicians of India 2011.

  20. Humanities in the Primary School--Philosophical Considerations

    Science.gov (United States)

    Eaude, Tony

    2017-01-01

    Drawing on a range of philosophical traditions, this article argues that the humanities are essential aspects of the development of the whole child. The humanities help children to understand themselves and other people in relation to place, time, belief, identity and culture and to become empathetic, thoughtful and critical citizens. Learning the…

  1. Study of root canal accessibility in human primary molars.

    Science.gov (United States)

    Aminabadi, Naser A; Farahani, Ramin M Z; Gajan, Esrafil B

    2008-03-01

    The aim of the present study was to provide a general scheme for pulpectomy of primary molars that may be useful for decision-making about negotiation of root canals and selection of appropriate instruments. A total of 160 vital primary molars in 85 patients (40 males, 45 females) aged 4-6 years were selected. After taking primary radiographs, local anesthesia was induced, and the teeth were isolated using a rubber dam. Canal accessibility index (CAI) and tooth accessibility index (TAI) were calculated according to initial file size. Mandibular first molars had either three canals (79.2%) or four canals (20.8%), and all second molars had four canals. Maxillary first molars had three canals and second molars had either three canals (70.9%) or four canals (29.1%). Lower accessibility of the mandibular first molar distobuccal root accounted for the lower accessibility of these teeth in comparison with mandibular second molars. While three-canal maxillary second molars were more accessible due to the lower accessibility of the distobuccal canal of the maxillary first molar, poor accessibility of the distal canal in four-canal second molars was responsible for the difficult accessibility of these teeth. In conclusion, it seems that the accessibility of a single canal in each tooth determines the difficulty of accessibility for any given tooth. Moreover, while primary second molars are more accessible than first molars, all of them are negotiable.

  2. CD90 Expression on human primary cells and elimination of contaminating fibroblasts from cell cultures.

    Science.gov (United States)

    Kisselbach, Lynn; Merges, Michael; Bossie, Alexis; Boyd, Ann

    2009-01-01

    Cluster Differentiation 90 (CD90) is a cell surface glycoprotein originally identified on mouse thymocytes. Although CD90 has been identified on a variety of stem cells and at varying levels in non-lymphoid tissues such as on fibroblasts, brain cells, and activated endothelial cells, the knowledge about the levels of CD90 expression on different cell types, including human primary cells, is limited. The goal of this study was to identify CD90 as a human primary cell biomarker and to develop an efficient and reliable method for eliminating unwanted or contaminating fibroblasts from human primary cell cultures suitable for research pursuant to cell based therapy technologies.

  3. Expandable gastroretentive dosage forms.

    Science.gov (United States)

    Klausner, Eytan A; Lavy, Eran; Friedman, Michael; Hoffman, Amnon

    2003-06-24

    Expandable gastroretentive dosage forms (GRDFs) have been designed for the past 3 decades. They were originally created for possible veterinary use, but later the design was modified for enhanced drug therapy in humans. These GRDFs are easily swallowed and reach a significantly larger size in the stomach due to swelling or unfolding processes that prolong their gastric retention time (GRT). After drug release, their dimensions are minimized with subsequent evacuation from the stomach. Gastroretentivity is enhanced by the combination of substantial dimensions with high rigidity of the dosage form to withstand the peristalsis and mechanical contractility of the stomach. Positive results were obtained in preclinical and clinical studies evaluating GRT of expandable GRDFs. Narrow absorption window drugs compounded in such systems have improved in vivo absorption properties. These findings are an important step towards the implementation of expandable GRDFs in the clinical setting. The current review deals with expandable GRDFs reported in articles and patents, and describes the physiological basis of their design. Using the dog as a preclinical screening model prior to human studies, relevant imaging techniques and pharmacokinetic-pharmacodynamic aspects of such delivery systems are also discussed.

  4. Expanded Cinema.

    Science.gov (United States)

    Youngblood, Gene

    Today, our environment is created more by television and cinema than by nature. Our view of what human nature is and might be is conditioned by what we see in those media. Films which make use of traditional genres only reinforce traditional views. They do not force us to new ones. But new filmmakers are creating "synaesthetic" cinema, which…

  5. Primary Humanities--A Perspective from Northern Ireland

    Science.gov (United States)

    Greenwood, Richard; Richardson, Norman; Gracie, Anita

    2017-01-01

    The word "humanities" does not appear in the current Northern Ireland Curriculum (NIC). Geography and history are taught within an Area of Learning called "The World Around Us" which also contains science and technology. The curriculum has a strong emphasis on an integrated, "connected learning" way of teaching and…

  6. Human Resource Architectures for New Teachers in Flemish Primary Education

    Science.gov (United States)

    Vekeman, Eva; Devos, Geert; Valcke, Martin

    2016-01-01

    Since research shows that the quality of a school's teaching force is related to its personnel practices, there is a growing interest in human resource management (HRM) in education. Existing research has generated insights into the differences, constraints and effects of single and isolated HR practices. Yet, little research is available…

  7. Expanding the Scope of Organizational Behavior Management: Relational Frame Theory and the Experimental Analysis of Complex Human Behavior

    Science.gov (United States)

    Hayes, Steven C.; Bunting, Kara; Herbst, Scott; Bond, Frank W.; Barnes-Holmes, Dermot

    2006-01-01

    Behavior analysis in general and applied behavior analysis in particular requires a well developed, empirically supported, and useful approach to human language and cognition in order to fulfill its mission of providing a relatively adequate comprehensive account of complex human behavior. This article introduces a series of articles in which the…

  8. HANPP Collection: Human Appropriation of Net Primary Productivity (HANPP) by Country and Product

    Data.gov (United States)

    National Aeronautics and Space Administration — The Human Appropriation of Net Primary Productivity (HANPP) by Country and Product portion of the HANPP Collection contains tabular data on carbon-equivalents of...

  9. Referral population studies underestimate differences between human papillomavirus assays in primary cervical screening

    DEFF Research Database (Denmark)

    Rebolj, M; Njor, Sisse Helle; Lynge, E.

    2017-01-01

    OBJECTIVE: We studied how representative cytologically abnormal women ("referral populations") are with respect to uncovering differences between human papillomavirus (HPV) assays in the primary screening where most women are cytologically normal. METHODS: A total of 4997 women were tested...

  10. Human papillomavirus-driven neck lymph node metastases from oropharyngeal or unknown primary squamous cell carcinoma

    OpenAIRE

    Schroeder, Lea

    2017-01-01

    Patients with neck squamous cell carcinoma from unknown primary tumor (NSCCUP) present with lymph node metastases without evidence for a primary tumor. Most patients undergo an aggressive multimodal treatment, which induces severe toxicity. Primary tumors of NSCCUP can be hidden in the oropharynx. Human papillomavirus (HPV) is causally involved in a subgroup of oropharyngeal squamous cell carcinomas (OPSCC) associated with early lymph node metastasis and good prognosis. Detection of markers f...

  11. Immune Modulation in Primary Vaccinia virus Zoonotic Human Infections

    Directory of Open Access Journals (Sweden)

    Juliana Assis Silva Gomes

    2012-01-01

    Full Text Available In 2010, the WHO celebrated the 30th anniversary of the smallpox eradication. Ironically, infections caused by viruses related to smallpox are being increasingly reported worldwide, including Monkeypox, Cowpox, and Vaccinia virus (VACV. Little is known about the human immunological responses elicited during acute infections caused by orthopoxviruses. We have followed VACV zoonotic outbreaks taking place in Brazil and analyzed cellular immune responses in patients acutely infected by VACV. Results indicated that these patients show a biased immune modulation when compared to noninfected controls. Amounts of B cells are low and less activated in infected patients. Although present, T CD4+ cells are also less activated when compared to noninfected individuals, and so are monocytes/macrophages. Similar results were obtained when Balb/C mice were experimentally infected with a VACV sample isolated during the zoonotic outbreaks. Taking together, the data suggest that zoonotic VACVs modulate specific immune cell compartments during an acute infection in humans.

  12. The CEPH consortium primary linkage map of human chromosome 10.

    Science.gov (United States)

    White, R L; Lalouel, J M; Nakamura, Y; Donis-Keller, H; Green, P; Bowden, D W; Mathew, C G; Easton, D F; Robson, E B; Morton, N E

    1990-03-01

    The first CEPH consortium map, that of chromosome 10, is presented. This primary linkage map contains 28 continuously linked loci defined by genotypes generated from CEPH family DNAs with 37 probe and enzyme combinations. Cytogenetic localization of some of the genetic markers indicates that the consortium map extends, at least, from 10p13 to 10q26. The order of loci on the consortium map agrees with the physical localization data. The female map spans 309 cM (206 cM if an approximation of interference is included in the mapping function used to construct the map), and the mean genetic distance of intervals is 11 cM (7 cM). Also presented are maps of chromosome 10 from each of five CEPH collaborating laboratories, based on genotypes for all relevant markers in the CEPH database. The CEPH consortium map of chromosome 10 should be useful for localization of any gene of interest falling within the span covered. The genotypes in the chromosome 10 consortium map database are now available to the scientific community.

  13. Expanding the human-machine interface model to address the effect of leadership and management on performance

    Energy Technology Data Exchange (ETDEWEB)

    Briant, V.S.; Childress, J.R.; Hannaman, G.W.

    1988-01-01

    The US nuclear industry now focuses on improving plant performance in measurable areas such as availability, safety, and operations. Risk assessment and pioneering work in human reliability analysis (HRA) have provided methods to identify and prioritize numerous design improvements. Improvements such as control room design, training, and procedures have contributed positively to plant performance. Human performance is increasingly recognized as a fundamental contributor to safe, economic, and reliable operation. Industry leaders suggest that improved leadership and management are keys to enhanced plant performance. This paper identifies several critical aspects of individual and group behavior that, if managed, could significantly contribute to improved performance. Some existing tools for measuring performance are cited.

  14. Expanding horizons

    Directory of Open Access Journals (Sweden)

    Editor

    2009-01-01

    Full Text Available Dear Friends, The debate of whether Stem Cell Therapy is a hype or hope has been raging for quite some time and has been rekindled in the current year. Stem Cell Scientists have been particularly enthused by the bold standard taken by Barack Obama in passing an Executive Order that lifted the ban on federal funding of Research on Embryonic Stem Cell (ESC lines created after August 9, 2001. With the lifting of the ban more money is expected to be poured into ESC and Stem Cell Research in general and this augurs well for this emerging science. This is hope. Amariglio et al have reported the occurrence of a multifocal brain tumor in a boy with ataxia telengectasia four years after he was treated with intracerebella and intrathecal injection of human fetal neural stem cells. Molecular and Cytogenetic studies showed that the tumor was of non host origin raising the possibility of it being derived from transplanted neural stem cell. This is the first report of a donor-derived brain tumor in neural stem cell therapy and opens a Pandora’s Box of questions about the safety of such therapies. This signifies the hype surrounding the therapy. However controversies are a part of any emerging science. Our goals should be to march forward conducting our research under strict ethical principles and rigorous oversight, ironing out even minor flaws, always being on the lookout for adverse events and identifying ways and means of preventing their occurrence in future. JSRM has been in receipt of six articles, which speaks well for the interest people have for stem cell science in general and our journal in particular. The articles we have received for this edition of JSRM cover all aspects necessary for a science. Rosen et al have described the percentage variation of adipose stromal cells isolated from two different inbred mouse strains and Bhonde et al have reported the existence of multipotent stem cells in human fallopian tube. If cells can be identified

  15. Adaptation and cross-cultural validation of the United States Primary Care Assessment Tool (expanded version) for use in South Africa.

    Science.gov (United States)

    Bresick, Graham; Sayed, Abdul-Rauf; le Grange, Cynthia; Bhagwan, Susheela; Manga, Nayna

    2015-06-19

    Measuring primary care is important for health sector reform. The Primary Care Assessment Tool (PCAT) measures performance of elements essential for cost-effective care. Following minor adaptations prior to use in Cape Town in 2011, a few findings indicated a need to improve the content and cross-cultural validity for wider use in South Africa (SA). This study aimed to validate the United States of America-developed PCAT before being used in a baseline measure of primary care performance prior to major reform. Public sector primary care clinics, users, practitioners and managers in urban and rural districts in the Western Cape Province. Face value evaluation of item phrasing and a combination of Delphi and Nominal Group Technique (NGT) methods with an expert panel and user focus group were used to obtain consensus on content relevant to SA. Original and new domains and items with > = 70% agreement were included in the South African version--ZA PCAT. All original PCAT domains achieved consensus on inclusion. One new domain, the primary healthcare (PHC) team, was added. Three of 95 original items achieved < 70% agreement, that is consensus to exclude as not relevant to SA; 19 new items were added. A few items needed minor rephrasing with local healthcare jargon. The demographic section was adapted to local socio-economic conditions. The adult PCAT was translated into isiXhosa and Afrikaans. The PCAT is a valid measure of primary care performance in SA. The PHC team domain is an important addition, given its emphasis in PHC re-engineering. A combination of Delphi and NGT methods succeeded in obtaining consensus on a multi-domain, multi-item instrument in a resource-constrained environment.

  16. Assessing the Humanities in the Primary School Using a Portfolio-Based Approach

    Science.gov (United States)

    Eaude, Tony

    2017-01-01

    This article suggests that a portfolio-based approach to assessing the humanities in the primary school is appropriate and outlines what this might involve. It argues for a broad interpretation of "the humanities" and for adopting principles associated with formative assessment, where assessment is not equated with testing and a wide…

  17. High Quality in Primary Humanities: Insights from the UK's School Inspectorates

    Science.gov (United States)

    Catling, Simon

    2017-01-01

    The school inspectorates of the four jurisdictions of the UK are sources of evidence about the quality of humanities teaching, learning and curriculum in primary schools. The term "humanities" usually refers to the subjects of geography, history and Religious Education, but here they are considered holistically, not separately. Discrete…

  18. The Future of the Humanities in Primary Schools--Reflections in Troubled Times

    Science.gov (United States)

    Eaude, Tony; Butt, Graham; Catling, Simon; Vass, Peter

    2017-01-01

    This article reflects on the implications for practitioners, researchers and policy-makers of the future of the humanities in primary schools in the light of the challenges facing future generations. There is wide divergence in the four jurisdictions of the UK. The humanities are perceived as important, in principle, though curriculum frameworks…

  19. Human red blood cells alterations in primary aldosteronism.

    Science.gov (United States)

    Bordin, Luciana; Donà, Gabriella; Sabbadin, Chiara; Ragazzi, Eugenio; Andrisani, Alessandra; Ambrosini, Guido; Brunati, Anna Maria; Clari, Giulio; Armanini, Decio

    2013-06-01

    Aldosterone (Aldo) effects include NADPH oxidase activation involved in Aldo-related oxidative stress. Red blood cells (RBCs) are particularly sensitive to oxidative assault, and both the formation of high molecular weight aggregates (HMWAs) and the diamide-induced Tyr phosphorylation (Tyr-P) level of membrane band 3 can be used to monitor their redox status. The Aldo-related alterations in erythrocytes were evaluated by comparing in vitro evidence. This was a multicenter comparative study. The study included 12 patients affected by primary aldosteronism (PA) and 6 healthy control subjects (HCs), whose RBCs were compared with those of patients with PA. For in vitro experiments, RBCs from HCs were incubated with increasing Aldo concentrations. The Tyr-P level, band 3 HMWA formation, and autologous IgG binding were evaluated. In patients with PA, both Tyr-P levels and band 3 HMWAs were higher than those in HCs. RBCs from HCs were treated with increasing Aldo concentrations in both platelet-poor plasma (PPP) and charcoal-stripped (CS)-PPP. Results showed that Aldo had dose- and time-dependent effects on band 3 Tyr-P and HMWA formation in CS-PPP more than in PPP. These effects were almost completely prevented by canrenone or cortisol. Aldo-related membrane alterations led to increased autologous IgG binding. Erythrocytes from patients with PA show oxidative-like stress evidenced by increased HMWA content and diamide-induced band 3 Tyr-P level. Aldo effects are mediated by the mineralocorticoid receptor, as suggested by the inhibitory effects of canrenone, an antagonist of Aldo. In CS-PPP, in which Aldo induces remarkable membrane alterations leading to IgG binding, Aldo may be responsible for premature RBC removal from circulation.

  20. Analysis of the karyotype of expanded human adipose-derived stem cells for bone reconstruction of the maxillo-facial region.

    Science.gov (United States)

    Bellotti, C; Stanco, D; Ragazzini, S; Romagnoli, L; Martella, E; Lazzati, S; Marchetti, C; Donati, D; Lucarelli, E

    2013-01-01

    Mesenchymal stem cells (MSC) and adipose-derived stem cells (ASC) were recently proposed for bone maxillofacial reconstruction in association with biomaterials. For this application MSC must be ex-vivo expanded in order to obtain, for a given volume of implanted biomaterial, a relevant number of bone forming cells. Previously conducted pre-clinical studies suggested that a concentration of 6 x 10(8) ASC associated with 900 mg of anorganic bovine bone (ABB) could be effective for human maxillary sinus floor elevation. A keystone issue to guarantee the quality and safety of Advanced Therapy Medicinal Products containing expanded MSC and ASC is their chromosome stability in culture: this topic has been widely investigated and conflicting results have been published. Abnormal karyotype of human ex-vivo expanded MSC and ASC was found by some authors, while, at the same time, several other studies showed the MSC and ASC karyotype to be normal. It is therefore important that all the results obtained on MSC and ASC karyotype analysis be published. Given this context, the aim of this manuscript, aim of this manuscript is to verify the karyotype stability of ASC in view of their applications in clinical trials. ASC obtained from the adipose tissue of 4 donors were expanded over extended culture time. Based on previous ASC expansions we hypothesized to be able to obtain 6 x 10(8) cells by passage 7. Karyotype analysis of 30 metaphases was planned to be investigated at passage 2, 7, and 15 in all the cultures. No abnormalities were found in the karyotype of two donors at all the passages tested, while a translocation was found in 2 metaphases of a donor at passage 7, but not at passage 15, and in the fourth donor in 5 metaphases a trisomy was found at passage 15. Chromosomal abnormalities were detected only after extended ASC expansion. Whether these anomalies can be related to risk for the patient's safety will have to be demonstrated by in-vivo studies.

  1. Discovery of α-L-arabinopyranosidases from human gut microbiome expands the diversity within glycoside hydrolase family 42

    DEFF Research Database (Denmark)

    Viborg, Alexander Holm; Katayama, Takane; Arakawa, Takatoshi

    2017-01-01

    Enzymes of the glycoside hydrolase family 42 (GH42) are widespread in bacteria of the human gut microbiome and play fundamental roles in the decomposition of both milk and plant oligosaccharides. All GH42 enzymes characterized so far have β-galactosidase activity. Here, we report the existence...

  2. Human and animal infections by vaccinia-like viruses in the state of Rio de Janeiro: a novel expanding zoonosis.

    Science.gov (United States)

    Schatzmayr, H G; Costa, R V C; Gonçalves, M C R; D'Andréa, P S; Barth, O M

    2011-12-30

    Since 1999, vesicular infections caused by Orthopoxvirus in humans and animals, mainly in dairy cattle, have been identified in 20 municipalities in the Rio de Janeiro state of Brazil. This paper describes studies conducted in counties of the northwestern, middle-Paraíba Valley and southern regions of the Rio de Janeiro state where 77 human, 346 bovine and 78 rodent samples were collected over the past ten years. Laboratory investigations using virus isolation, electron microscopy, molecular biology (PCR) and serological analysis confirmed Orthopoxvirus infections in 77.9% of human, 49.2% of dairy cattle and 17.9% of rodent samples. The characterisation of the Cantagalo/IOC strain reconfirmed that this virus was a vaccinia-like virus. In other regions of the Rio de Janeiro state, vesicular/pustular infections in animals and humans are suspected but these have not yet been confirmed. A continuous surveillance system has been established to monitor these regions in addition to several other states of the Brazilian Federation. Copyright © 2011 Elsevier Ltd. All rights reserved.

  3. Reversal of hyperglycemia in mice by using human expandable insulin-producing cells differentiated from fetal liver progenitor cells

    Science.gov (United States)

    Zalzman, Michal; Gupta, Sanjeev; Giri, Ranjit K.; Berkovich, Irina; Sappal, Baljit S.; Karnieli, Ohad; Zern, Mark A.; Fleischer, Norman; Efrat, Shimon

    2003-06-01

    Beta-cell replacement is considered to be the most promising approach for treatment of type 1 diabetes. Its application on a large scale is hindered by a shortage of cells for transplantation. Activation of insulin expression, storage, and regulated secretion in stem/progenitor cells offers novel ways to overcome this shortage. We explored whether fetal human progenitor liver cells (FH) could be induced to differentiate into insulin-producing cells after expression of the pancreatic duodenal homeobox 1 (Pdx1) gene, which is a key regulator of pancreatic development and insulin expression in beta cells. FH cells possess a considerable replication capacity, and this was further extended by introduction of the gene for the catalytic subunit of human telomerase. Immortalized FH cells expressing Pdx1 activated multiple beta-cell genes, produced and stored considerable amounts of insulin, and released insulin in a regulated manner in response to glucose. When transplanted into hyperglycemic immunodeficient mice, the cells restored and maintained euglycemia for prolonged periods. Quantitation of human C-peptide in the mouse serum confirmed that the glycemia was normalized by the transplanted human cells. This approach offers the potential of a novel source of cells for transplantation into patients with type 1 diabetes.

  4. Food Safety - The Primary Objective of Human Society Existence

    Directory of Open Access Journals (Sweden)

    Tabita Cornelia Adamov

    2010-10-01

    Full Text Available Food has played, plays and will play a decisive role in the existence and development of human society. The level of food quality, causes physical, social and moral society health. In this sense food security is an essential goal of economic and social development, being an essential component of security of life and national security. Evolution of the Common Agricultural Policy (CAP, in terms of food security, was due not only changes in agriculture, but also came in response to the demands of society in general. Amoung these is growing concerns about hygiene and food safety and animal welfare. European consumers want safe and wholesome food and the EU wants to ensure that all its citizens consume food with high quality standards. Food safety policy has undergone an extensive refurbishment. The objective of this reform was to ensure that EU legislation on food safety is as complete as possible, and consumers benefit as much information about potential risks and measures to be taken to minimize them. The goal of a modern economy, is the correlation of quantitative and qualitative food production with consumer demand. It thus requires knowledge of the physiological needs of consumers, leading to demand for agricultural products.

  5. An expanding toolkit for preclinical pre-erythrocytic malaria vaccine development: bridging traditional mouse malaria models and human trials.

    Science.gov (United States)

    Steel, Ryan Wj; Kappe, Stefan Hi; Sack, Brandon K

    2016-12-01

    Malaria remains a significant public health burden with 214 million new infections and over 400,000 deaths in 2015. Elucidating relevant Plasmodium parasite biology can lead to the identification of novel ways to control and ultimately eliminate the parasite within geographic areas. Particularly, the development of an effective vaccine that targets the clinically silent pre-erythrocytic stages of infection would significantly augment existing malaria elimination tools by preventing both the onset of blood-stage infection/disease as well as spread of the parasite through mosquito transmission. In this Perspective, we discuss the role of small animal models in pre-erythrocytic stage vaccine development, highlighting how human liver-chimeric and human immune system mice are emerging as valuable components of these efforts.

  6. Maturation and demise of human primary monocytes by carbon nanotubes

    Science.gov (United States)

    De Nicola, Milena; Mirabile Gattia, Daniele; Traversa, Enrico; Ghibelli, Lina

    2013-06-01

    The possibility of exploiting carbon nanotubes (CNT) in biomedical practices requires thorough analysis of the chemical or bulk effects they may exert on the immune system, the complex network that recognizes and eliminates foreign particles. In particular, the phagocytosing ability of cells belonging to the monocyte/macrophage lineage may render these immune cells an ideal toxicological target of pristine CNT, which may form aggregates of size exceeding monocyte/macrophage phagocytosing plasticity. To shed light on this issue, we analyzed the effects that pristine multi-walled CNT (MWCNT) without metal or biological impurities exert on survival and activation of freshly explanted human peripheral blood monocytes, analyzing in parallel the non-phagocytosing lymphocytes, and using graphite as control carbon material. MWCNT (diameter 10-50 nm, length up to 10 μm) exert two different toxic effects on mononuclear leukocytes: a minor apoptogenic effect (on lymphocytes > monocytes), and a major, apoptosis-independent effect that exclusively and deeply affect monocyte homeostasis. Analysis of monocyte number, adhesion, redox equilibrium, and the differentiation markers CD14 and CD11b reveals that MWCNT cause the selective disappearance of phagocytosis-competent monocytes by mechanisms related to the presence of large nanoparticle aggregates, suggesting phenomena of bulk toxicity possibly consisting of frustrated phagocytosis. At the same time, MWCNT stimulate adhesion of the phagocytosis-incompetent monocytes, and their differentiation toward a peculiar maturation asset. These observations point out novel mechanisms of CNT toxicity, renewing concerns that they may impair the innate immune system deranging the inflammatory responses.

  7. Maturation and demise of human primary monocytes by carbon nanotubes

    Energy Technology Data Exchange (ETDEWEB)

    De Nicola, Milena, E-mail: milena.de.nicola@uniroma2.it [University of Rome ' Tor Vergata' , Department of Biology (Italy); Mirabile Gattia, Daniele, E-mail: daniele.mirabile@enea.it [UTTMAT, ENEA-C.R. Casaccia (Italy); Traversa, Enrico, E-mail: Enrico.Traversa@kaust.edu.sa [King Abdullah University of Science and Technology (KAUST), Division of Physical Science and Engineering (Saudi Arabia); Ghibelli, Lina, E-mail: ghibelli@uniroma2.it [University of Rome ' Tor Vergata' , Department of Biology (Italy)

    2013-06-15

    The possibility of exploiting carbon nanotubes (CNT) in biomedical practices requires thorough analysis of the chemical or bulk effects they may exert on the immune system, the complex network that recognizes and eliminates foreign particles. In particular, the phagocytosing ability of cells belonging to the monocyte/macrophage lineage may render these immune cells an ideal toxicological target of pristine CNT, which may form aggregates of size exceeding monocyte/macrophage phagocytosing plasticity. To shed light on this issue, we analyzed the effects that pristine multi-walled CNT (MWCNT) without metal or biological impurities exert on survival and activation of freshly explanted human peripheral blood monocytes, analyzing in parallel the non-phagocytosing lymphocytes, and using graphite as control carbon material. MWCNT (diameter 10-50 nm, length up to 10 {mu}m) exert two different toxic effects on mononuclear leukocytes: a minor apoptogenic effect (on lymphocytes > monocytes), and a major, apoptosis-independent effect that exclusively and deeply affect monocyte homeostasis. Analysis of monocyte number, adhesion, redox equilibrium, and the differentiation markers CD14 and CD11b reveals that MWCNT cause the selective disappearance of phagocytosis-competent monocytes by mechanisms related to the presence of large nanoparticle aggregates, suggesting phenomena of bulk toxicity possibly consisting of frustrated phagocytosis. At the same time, MWCNT stimulate adhesion of the phagocytosis-incompetent monocytes, and their differentiation toward a peculiar maturation asset. These observations point out novel mechanisms of CNT toxicity, renewing concerns that they may impair the innate immune system deranging the inflammatory responses.

  8. Maturation and demise of human primary monocytes by carbon nanotubes

    KAUST Repository

    De Nicola, Milena D.

    2013-05-17

    The possibility of exploiting carbon nanotubes (CNT) in biomedical practices requires thorough analysis of the chemical or bulk effects they may exert on the immune system, the complex network that recognizes and eliminates foreign particles. In particular, the phagocytosing ability of cells belonging to the monocyte/macrophage lineage may render these immune cells an ideal toxicological target of pristine CNT, which may form aggregates of size exceeding monocyte/macrophage phagocytosing plasticity. To shed light on this issue, we analyzed the effects that pristine multi-walled CNT (MWCNT) without metal or biological impurities exert on survival and activation of freshly explanted human peripheral blood monocytes, analyzing in parallel the non-phagocytosing lymphocytes, and using graphite as control carbon material. MWCNT (diameter 10-50 nm, length up to 10 μm) exert two different toxic effects on mononuclear leukocytes: a minor apoptogenic effect (on lymphocytes > monocytes), and a major, apoptosis-independent effect that exclusively and deeply affect monocyte homeostasis. Analysis of monocyte number, adhesion, redox equilibrium, and the differentiation markers CD14 and CD11b reveals that MWCNT cause the selective disappearance of phagocytosis-competent monocytes by mechanisms related to the presence of large nanoparticle aggregates, suggesting phenomena of bulk toxicity possibly consisting of frustrated phagocytosis. At the same time, MWCNT stimulate adhesion of the phagocytosis-incompetent monocytes, and their differentiation toward a peculiar maturation asset. These observations point out novel mechanisms of CNT toxicity, renewing concerns that they may impair the innate immune system deranging the inflammatory responses. © 2013 Springer Science+Business Media Dordrecht.

  9. Primary human cervical carcinoma cells require human papillomavirus E6 and E7 expression for ongoing proliferation

    OpenAIRE

    Magaldi, Thomas G.; Almstead, Laura L.; Bellone, Stefania; Prevatt, Edward G.; Santin, Alessandro D.; DiMaio, Daniel

    2012-01-01

    Repression of human papillomavirus (HPV) E6 and E7 oncogenes in established cervical carcinoma cell lines causes senescence due to reactivation of cellular tumor suppressor pathways. Here, we determined whether ongoing expression of HPV16 or HPV18 oncogenes is required for the proliferation of primary human cervical carcinoma cells in serum-free conditions at low passage number after isolation from patients. We used an SV40 viral vector expressing the bovine papillomavirus E2 protein to repre...

  10. Development of organoids from mouse and human endometrium showing endometrial epithelium physiology and long-term expandability.

    Science.gov (United States)

    Boretto, Matteo; Cox, Benoit; Noben, Manuel; Hendriks, Nikolai; Fassbender, Amelie; Roose, Heleen; Amant, Frédéric; Timmerman, Dirk; Tomassetti, Carla; Vanhie, Arne; Meuleman, Christel; Ferrante, Marc; Vankelecom, Hugo

    2017-05-15

    The endometrium, which is of crucial importance for reproduction, undergoes dynamic cyclic tissue remodeling. Knowledge of its molecular and cellular regulation is poor, primarily owing to a lack of study models. Here, we have established a novel and promising organoid model from both mouse and human endometrium. Dissociated endometrial tissue, embedded in Matrigel under WNT-activating conditions, swiftly formed organoid structures that showed long-term expansion capacity, and reproduced the molecular and histological phenotype of the tissue's epithelium. The supplemented WNT level determined the type of mouse endometrial organoids obtained: high WNT yielded cystic organoids displaying a more differentiated phenotype than the dense organoids obtained in low WNT. The organoids phenocopied physiological responses of endometrial epithelium to hormones, including increased cell proliferation under estrogen and maturation upon progesterone. Moreover, the human endometrial organoids replicated the menstrual cycle under hormonal treatment at both the morpho-histological and molecular levels. Together, we established an organoid culture system for endometrium, reproducing tissue epithelium physiology and allowing long-term expansion. This novel model provides a powerful tool for studying mechanisms underlying the biology as well as the pathology of this key reproductive organ. © 2017. Published by The Company of Biologists Ltd.

  11. Primary ciliogenesis defects are associated with human astrocytoma/glioblastoma cells

    Directory of Open Access Journals (Sweden)

    Rattner Jerome B

    2009-12-01

    Full Text Available Abstract Background Primary cilia are non-motile sensory cytoplasmic organelles that have been implicated in signal transduction, cell to cell communication, left and right pattern embryonic development, sensation of fluid flow, regulation of calcium levels, mechanosensation, growth factor signaling and cell cycle progression. Defects in the formation and/or function of these structures underlie a variety of human diseases such as Alström, Bardet-Biedl, Joubert, Meckel-Gruber and oral-facial-digital type 1 syndromes. The expression and function of primary cilia in cancer cells has now become a focus of attention but has not been studied in astrocytomas/glioblastomas. To begin to address this issue, we compared the structure and expression of primary cilia in a normal human astrocyte cell line with five human astrocytoma/glioblastoma cell lines. Methods Cultured normal human astrocytes and five human astrocytoma/glioblastoma cell lines were examined for primary cilia expression and structure using indirect immunofluorescence and electron microscopy. Monospecific antibodies were used to detect primary cilia and map the relationship between the primary cilia region and sites of endocytosis. Results We show that expression of primary cilia in normal astrocytes is cell cycle related and the primary cilium extends through the cell within a unique structure which we show to be a site of endocytosis. Importantly, we document that in each of the five astrocytoma/glioblastoma cell lines fully formed primary cilia are either expressed at a very low level, are completely absent or have aberrant forms, due to incomplete ciliogenesis. Conclusions The recent discovery of the importance of primary cilia in a variety of cell functions raises the possibility that this structure may have a role in a variety of cancers. Our finding that the formation of the primary cilium is disrupted in cells derived from astrocytoma/glioblastoma tumors provides the first

  12. Parallel Measurement of Circadian Clock Gene Expression and Hormone Secretion in Human Primary Cell Cultures.

    Science.gov (United States)

    Petrenko, Volodymyr; Saini, Camille; Perrin, Laurent; Dibner, Charna

    2016-11-11

    Circadian clocks are functional in all light-sensitive organisms, allowing for an adaptation to the external world by anticipating daily environmental changes. Considerable progress in our understanding of the tight connection between the circadian clock and most aspects of physiology has been made in the field over the last decade. However, unraveling the molecular basis that underlies the function of the circadian oscillator in humans stays of highest technical challenge. Here, we provide a detailed description of an experimental approach for long-term (2-5 days) bioluminescence recording and outflow medium collection in cultured human primary cells. For this purpose, we have transduced primary cells with a lentiviral luciferase reporter that is under control of a core clock gene promoter, which allows for the parallel assessment of hormone secretion and circadian bioluminescence. Furthermore, we describe the conditions for disrupting the circadian clock in primary human cells by transfecting siRNA targeting CLOCK. Our results on the circadian regulation of insulin secretion by human pancreatic islets, and myokine secretion by human skeletal muscle cells, are presented here to illustrate the application of this methodology. These settings can be used to study the molecular makeup of human peripheral clocks and to analyze their functional impact on primary cells under physiological or pathophysiological conditions.

  13. [A method for the primary culture of fibroblasts isolated from human airway granulation tissues].

    Science.gov (United States)

    Chen, Nan; Zhang, Jie; Xu, Min; Wang, Yu-ling; Pei, Ying-hua

    2013-04-01

    To establish a feasible method to culture primary fibroblasts isolated from human airway granulation tissues, and therefore to provide experimental data for the investigation of the pathogenesis of benign airway stenosis. The granulation tissues were collected from 6 patients during routine bronchoscopy at our department of Beijing Tiantan Hospital from April to June 2011. Primary fibroblasts were obtained by culturing the explanted tissues. Cell growth was observed under inverted microscope. All of these 6 primary cultures were successful. Fibroblast-like cells were observed to migrate from the tissue pieces 3 d after inoculation. After 9-11 d of culture, cells reached to 90% confluence and could be sub-cultured. After passage, the cells were still in a typical elongated spindle-shape and grew well. The cells could be sub-cultured further when they formed a monolayer. Explant culture is a reliable method for culturing primary fibroblasts from human airway granulation tissues.

  14. Human Immunodeficiency Virus Reverse Transcriptase and Protease Sequence Database: an expanded data model integrating natural language text and sequence analysis programs.

    Science.gov (United States)

    Kantor, R; Machekano, R; Gonzales, M J; Dupnik, K; Schapiro, J M; Shafer, R W

    2001-01-01

    The HIV Reverse Transcriptase and Protease Sequence Database is an on-line relational database that catalogs evolutionary and drug-related sequence variation in the human immunodeficiency virus (HIV) reverse transcriptase (RT) and protease enzymes, the molecular targets of anti-HIV therapy (http://hivdb.stanford.edu). The database contains a compilation of nearly all published HIV RT and protease sequences, including submissions from International Collaboration databases and sequences published in journal articles. Sequences are linked to data about the source of the sequence sample and the antiretroviral drug treatment history of the individual from whom the isolate was obtained. During the past year 3500 sequences have been added and the data model has been expanded to include drug susceptibility data on sequenced isolates. Database content has also been integrated with didactic text and the output of two sequence analysis programs.

  15. Regulatory T cells are expanded by Teriparatide treatment in humans and mediate intermittent PTH-induced bone anabolism in mice.

    Science.gov (United States)

    Yu, Mingcan; D'Amelio, Patrizia; Tyagi, Abdul Malik; Vaccaro, Chiara; Li, Jau-Yi; Hsu, Emory; Buondonno, Ilaria; Sassi, Francesca; Adams, Jonathan; Weitzmann, M Neale; DiPaolo, Richard; Pacifici, Roberto

    2017-11-20

    Teriparatide is a bone anabolic treatment for osteoporosis, modeled in animals by intermittent PTH (iPTH) administration, but the cellular and molecular mechanisms of action of iPTH are largely unknown. Here, we show that Teriparatide and iPTH cause a ~two-threefold increase in the number of regulatory T cells (Tregs) in humans and mice. Attesting in vivo relevance, blockade of the Treg increase in mice prevents the increase in bone formation and trabecular bone volume and structure induced by iPTH Therefore, increasing the number of Tregs is a pivotal mechanism by which iPTH exerts its bone anabolic activity. Increasing Tregs pharmacologically may represent a novel bone anabolic therapy, while iPTH-induced Treg increase may find applications in inflammatory conditions and transplant medicine. © 2017 The Authors.

  16. Trichomonas vaginalis and Human Immunodeficiency Virus Coinfection Among Women Under Community Supervision: A Call for Expanded T. vaginalis Screening.

    Science.gov (United States)

    Davis, Alissa; Dasgupta, Anindita; Goddard-Eckrich, Dawn; El-Bassel, Nabila

    2016-10-01

    The United States has a large community supervision population, a growing number of whom are women. Trichomonas vaginalis infection is strongly associated with an increased risk of human immunodeficiency virus (HIV) acquisition and transmission, particularly among women, but there is a paucity of research on HIV and T. vaginalis co-infection among women under community supervision. This article examines the prevalence of T. vaginalis infection and T. vaginalis and HIV coinfection at baseline among women under community supervision in New York City. It also examines the 12-month outcomes of women treated for T. vaginalis. Women received biological tests for HIV and T. vaginalis at baseline and 12 months follow-up. Of the 333 women tested for sexually transmitted infections, 77 women (23.1%) tested positive for T. vaginalis at baseline and 44 (13.3%) were HIV positive. Human immunodeficiency virus-positive women had significantly higher rates of T. vaginalis infection than HIV-negative women (36.4% vs 21.3%, P ≤ 0.05). Sixteen women (4.8%) were coinfected with T. vaginalis and HIV. Of the 77 women who were positive for T. vaginalis infection at baseline, 58 (75.3%) received treatment by a health care provider. Of those who received treatment, 17 (29.3%) tested positive for T. vaginalis at the 12-month follow-up. Given the high prevalence of T. vaginalis among this sample of women, particularly among HIV-positive women, and high levels of reinfection or persistent infection, screening for T. vaginalis among women under community supervision may have a substantial impact on reducing HIV acquisition and transmission among this high-risk population.

  17. Primary infection by type 1 human immunodeficiency virus: diagnosis and prognosis.

    OpenAIRE

    Vanhems, P.; Beaulieu, R.

    1997-01-01

    Primary infection by type 1 human immunodeficiency virus (HIV) is symptomatic in about 70% of cases. The acute illness is a mononucleosis-like syndrome with characteristics such as mucosal ulcerations. The duration and severity of the symptoms appear to be related to the prognosis. After reviewing the most frequent signs and symptoms of primary HIV infection, we report different prognostic studies which examined the association between the acute illness and the progression of HIV disease.

  18. Human enamel veneer restoration: an alternative technique to restore anterior primary teeth.

    Science.gov (United States)

    Oliveira, Luciana Butini; Tamay, Tereza Keiko; Oliveira, Marta Dutra Machado; Rodrigues, Célia Martins Delgado; Wanderley, Marcia Turolla

    2006-01-01

    Restoration of severely decayed primary teeth is a clinical challenge in Pediatric Dentistry. Among the restorative treatment options, the use of prefabricated crowns and resin composite restorations, either by means of direct or indirect techniques is mentioned in the literature. The purpose of this article is to describe the rehabilitation of primary anterior teeth in a 5-year-old patient. Dental treatment consisted on an anterior space maintainer prosthesis made with natural primary teeth, plus human dental enamel veneer (facet) restorations. The advantages of this technique are better esthetics and the natural enamel has physiologic wear and offers superficial smoothness and cervical adaptation compatible with those of the surrounding teeth.

  19. Bile acid-induced necrosis in primary human hepatocytes and in patients with obstructive cholestasis

    Energy Technology Data Exchange (ETDEWEB)

    Woolbright, Benjamin L.; Dorko, Kenneth [Department of Pharmacology, Toxicology & Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States); Antoine, Daniel J.; Clarke, Joanna I. [MRC Centre for Drug Safety Science, Department of Molecular and Clinical Pharmacology, Institute of Translational Medicine, University of Liverpool, Liverpool (United Kingdom); Gholami, Parviz [Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS (United States); Li, Feng [Department of Pharmacology, Toxicology & Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States); Kumer, Sean C.; Schmitt, Timothy M.; Forster, Jameson [Department of Surgery, University of Kansas Medical Center, Kansas City, KS (United States); Fan, Fang [Department of Pathology, University of Kansas Medical Center, Kansas City, KS (United States); Jenkins, Rosalind E.; Park, B. Kevin [MRC Centre for Drug Safety Science, Department of Molecular and Clinical Pharmacology, Institute of Translational Medicine, University of Liverpool, Liverpool (United Kingdom); Hagenbuch, Bruno [Department of Pharmacology, Toxicology & Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States); Olyaee, Mojtaba [Department of Internal Medicine, University of Kansas Medical Center, Kansas City, KS (United States); Jaeschke, Hartmut, E-mail: hjaeschke@kumc.edu [Department of Pharmacology, Toxicology & Therapeutics, University of Kansas Medical Center, Kansas City, KS (United States)

    2015-03-15

    Accumulation of bile acids is a major mediator of cholestatic liver injury. Recent studies indicate bile acid composition between humans and rodents is dramatically different, as humans have a higher percent of glycine conjugated bile acids and increased chenodeoxycholate content, which increases the hydrophobicity index of bile acids. This increase may lead to direct toxicity that kills hepatocytes, and promotes inflammation. To address this issue, this study assessed how pathophysiological concentrations of bile acids measured in cholestatic patients affected primary human hepatocytes. Individual bile acid levels were determined in serum and bile by UPLC/QTOFMS in patients with extrahepatic cholestasis with, or without, concurrent increases in serum transaminases. Bile acid levels increased in serum of patients with liver injury, while biliary levels decreased, implicating infarction of the biliary tracts. To assess bile acid-induced toxicity in man, primary human hepatocytes were treated with relevant concentrations, derived from patient data, of the model bile acid glycochenodeoxycholic acid (GCDC). Treatment with GCDC resulted in necrosis with no increase in apoptotic parameters. This was recapitulated by treatment with biliary bile acid concentrations, but not serum concentrations. Marked elevations in serum full-length cytokeratin-18, high mobility group box 1 protein (HMGB1), and acetylated HMGB1 confirmed inflammatory necrosis in injured patients; only modest elevations in caspase-cleaved cytokeratin-18 were observed. These data suggest human hepatocytes are more resistant to human-relevant bile acids than rodent hepatocytes, and die through necrosis when exposed to bile acids. These mechanisms of cholestasis in humans are fundamentally different to mechanisms observed in rodent models. - Highlights: • Cholestatic liver injury is due to cytoplasmic bile acid accumulation in hepatocytes. • Primary human hepatocytes are resistant to BA-induced injury

  20. Primary cilia are present on human blood and bone marrow cells and mediate Hedgehog signaling.

    Science.gov (United States)

    Singh, Mohan; Chaudhry, Parvesh; Merchant, Akil A

    2016-12-01

    Primary cilia are nonmotile, microtubule-based organelles that are present on the cellular membrane of all eukaryotic cells. Functional cilia are required for the response to developmental signaling pathways such as Hedgehog (Hh) and Wnt/β-catenin. Although the Hh pathway has been shown to be active in leukemia and other blood cancers, there have been no reports describing the presence of primary cilia in human blood or leukemia cells. In the present study, we show that nearly all human blood and bone marrow cells have primary cilia (97-99%). In contrast, primary cilia on AML cell lines (KG1, KG1a, and K562) were less frequent (10-36% of cells) and were often shorter and dysmorphic, with less well-defined basal bodies. Finally, we show that treatment of blood cells with the Hh pathway ligand Sonic Hedgehog (SHh) causes translocation of Smoothened (SMO) to the primary cilia and activation of Hh target genes, demonstrating that primary cilia in blood cells are functional and participate in Hh signaling. Loss of primary cilia on leukemia cells may have important implications for aberrant pathway activation and response to SMO inhibitors currently in clinical development. Copyright © 2016 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

  1. Expanding care for perinatal women with depression (EXPONATE): study protocol for a randomized controlled trial of an intervention package for perinatal depression in primary care.

    Science.gov (United States)

    Gureje, Oye; Oladeji, Bibilola Damilola; Araya, Ricardo; Montgomery, Alan A; Kola, Lola; Kirmayer, Laurence; Zelkowitz, Phyllis; Groleau, Danielle

    2015-06-30

    Depression is common among women during perinatal period and is associated with long-term adverse consequences for the mother and infant. In Nigeria, as in many other low- and-middle-income countries (LMIC), perinatal depression usually goes unrecognized and untreated. The aim of EXPONATE is to test the effectiveness and cost-effectiveness of an intervention package for perinatal depression delivered by community midwives in primary maternal care in which physician support and enhanced patient compliance are implemented using mobile phones. A pragmatic two-arm parallel cluster randomized controlled trial was designed. The units of allocation are the primary maternal care clinics. Thirty eligible and consenting clinics were randomized but, due to problems with logistics, 29 eventually participated. Consenting pregnant women with a gestational age between 16 and 28 weeks who screened positive on the Edinburgh Postnatal Depression Scale (EPDS score ≥12), absent psychosis or bipolar disorder, and not actively suicidal were recruited into the trial (N = 686). Midwives in the intervention arm were trained to deliver psychoeducation, problem solving treatment, and parenting skills. Eight weekly sessions were delivered following entry into the study. Further sessions during pregnancy and 6 weeks following childbirth were determined by level of depressive symptoms. Clinical support and supervision, delivered mainly by mobile phone, were provided by general physicians and psychiatrists. Automated text and voice messages, also delivered by mobile phones, were used to facilitate patient compliance with clinic appointments and 'homework' tasks. Patients in the control arm received care as usual enhanced by further training of the providers in that arm in the recognition and standard treatment of depression. Assessments are undertaken at baseline, 2 months following recruitment into the study and 3, 6, 9 and 12 months after childbirth. The primary outcome is recovery from

  2. Expanded CD8+ T cells of murine and human CLL are driven into a senescent KLRG1+ effector memory phenotype.

    Science.gov (United States)

    Göthert, Joachim Rudolf; Eisele, Lewin; Klein-Hitpass, Ludger; Weber, Stefanie; Zesewitz, Marie-Louise; Sellmann, Ludger; Röth, Alexander; Pircher, Hanspeter; Dührsen, Ulrich; Dürig, Jan

    2013-11-01

    Altered numbers and functions of T cells have previously been demonstrated in chronic lymphocytic leukemia (CLL) patients. However, dynamics and specific T-cell subset alterations have not been studied in great detail. Therefore, we studied CLL blood lymphocyte subsets of individual patients in a longitudinal manner. Dynamic expansions of blood CD4 + and CD8 + T-cell numbers were consistently associated with a progressively increasing CLL leukemic compartment. Interestingly, the T-cell subset expansion over time was more pronounced in CD38 + CLL. Additionally, we performed gene expression profiling of CD3 + T cells of CLL patients and normal donors. Using gene set enrichment analysis, we found significant enrichment of genes with higher expression in CLL T cells within CD8+ effector memory and terminal effector T-cell gene signatures. In agreement with these data, we observed a marked expansion of phenotypic CD8 + effector memory T cells in CLL by flow cytometry. Moreover, we observed that increments of CD8 + effector memory T cells in human CLL and also mouse CLL (Eμ-TCL1 model) were due to an expansion of the inhibitory killer cell lectin-like receptor G1 (KLRG1) expressing cellular subset. Furthermore, higher plasma levels of the natural KLRG1 ligand E-cadherin were detected in CLL patients compared to normal donor controls. The predominance of KLRG1+ expression within CD8+ T cells in conjunction with increased systemic soluble E-cadherin might significantly contribute to CLL immune dysfunction and might additionally represent an important component of the CLL microenvironment.

  3. [Ecotoxicology, human ecology, laser biotechnology in primary prevention of environmental health hazards].

    Science.gov (United States)

    Dobrowolski, J W

    2001-01-01

    Interdisciplinary studies in ecotoxicology (including the influence of complex physical, chemical and biological factors on ecosystems and human food chain), human ecology (related to estimation of individuals exposition to different pollutants both in the natural environment and in-door environment, as well as by food) and environmental biotechnology (based on application of high sensitive biological, especially embryological criteria in biotests for water quality, including image computisation and biosensors)--are scientific base for primary prevention of disturbances of the ecological balance as well as environmental risk factors for human health. Large scale applications of laser biostimulation in environmental engineering is a new chance for system approach in primary prevention by more efficient nutritional prevention, protection of proper water quality (including biotests) and protection of the human environment in working and living places.

  4. Lentiviral vector delivery of human interleukin-7 (hIL-7 to human immune system (HIS mice expands T lymphocyte populations.

    Directory of Open Access Journals (Sweden)

    Ryan M O'Connell

    2010-08-01

    Full Text Available Genetically modified mice carrying engrafted human tissues provide useful models to study human cell biology in physiologically relevant contexts. However, there remain several obstacles limiting the compatibility of human cells within their mouse hosts. Among these is inadequate cross-reactvitiy between certain mouse cytokines and human cellular receptors, depriving the graft of important survival and growth signals. To circumvent this problem, we utilized a lentivirus-based delivery system to express physiologically relevant levels of human interleukin-7 (hIL-7 in Rag2-/-gammac-/- mice following a single intravenous injection. hIL-7 promoted homeostatic proliferation of both adoptively transferred and endogenously generated T-cells in Rag2-/-gammac-/- Human Immune System (HIS mice. Interestingly, we found that hIL-7 increased T lymphocyte numbers in the spleens of HIV infected HIS mice without affecting viral load. Taken together, our study unveils a versatile approach to deliver human cytokines to HIS mice, to both improve engraftment and determine the impact of cytokines on human diseases.

  5. Primary cilia are increased in number and demonstrate structural abnormalities in human cancer.

    Science.gov (United States)

    Yasar, Binnaz; Linton, Kim; Slater, Christian; Byers, Richard

    2017-07-01

    Primary cilia play an important role in the regulation of cell signalling pathways and are thought to have a role in cancer but have seldom been studied in human cancer samples. Primary cilia were visualised by dual immunofluorescence for anti-CROCC (ciliary rootlet coiled-coil) and anti-tubulin in a range of human cancers (including carcinomas of stomach, pancreas, prostate, lung and colon, lobular and ductal breast cancers and follicular lymphoma) and in matched normal tissue (stomach, pancreas, lung, large and small intestines, breast and reactive lymph nodes) samples using a tissue microarray; their frequency, association with proliferation, was measured by Ki-67 staining and their structure was analysed. Compared with normal tissues, primary cilia frequency was significantly elevated in adenocarcinoma of the lung (2.75% vs 1.85%, p=0.016), adenocarcinoma of the colon (3.80% vs 2.43%, respectively, p=0.017), follicular lymphoma (1.18% vs 0.83%, p=0.003) and pancreatic adenocarcinoma (7.00% vs 5.26%, p=0.002); there was no statistically significant difference compared with normal control tissue for gastric and prostatic adenocarcinomas or for lobular and ductal breast cancers. Additionally, structural abnormalities of primary cilia were identified in cancer tissues, including elongation of the axoneme, multiple basal bodies and branching of the axoneme. Ki-67 scores ranged from 0.7% to 78.4% and showed no statistically significant correlation with primary cilia frequency across all tissues (p=0.1501). The results show upregulation of primary cilia and the presence of structural defects in a wide range of human cancer tissue samples demonstrating association of dysregulation of primary cilia with human cancer. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  6. High human papillomavirus (HPV prevalence in South African adolescents and young women encourages expanded HPV vaccination campaigns.

    Directory of Open Access Journals (Sweden)

    Zizipho Z A Mbulawa

    Full Text Available The objectives of the study were to investigate prevalence of cervical human papillomavirus (HPV genotypes to inform HPV vaccination strategy in South Africa and to study factors associated with HPV prevalence. Sexually active, HIV-negative women, aged 16-22 years recruited from Soweto (n = 143 and Cape Town (n = 148 were tested for cervical HPV and other genital infections. Overall HPV prevalence was 66.7% (194/291 in young women. Cape Town women were more likely to have multiple HPV infections than the Soweto women (48.0%, 71/148 versus 35.0%, 50/143 respectively, p = 0.033 and probable HR-HPV types (34.5%, 51/148 versus 21.7%, 31/143 respectively, p = 0.022. The most frequently detected HPV types were HPV-16 (11.7%, HPV-58 (10.3%, HPV-51 (8.9%, HPV-66 (8.6%, HPV-18 and HPV-81 (7.6% each. HPV types targeted by the bivalent HPV vaccine (HPV-16/18 were detected in 18.6% (54/291 of women, while those in the quadrivalent vaccine (HPV-6/11/16/18 were detected in 24.7% (72/291 of women; and those in the nonavalent vaccine (HPV-6/11/16/18/31/33/45/52/58 were detected in 38.5% (112/291 of women. In a multivariable analysis, bacterial vaginosis remained significantly associated with HPV infection (OR: 4.0, 95% CI: 1.4-12.6. Women were more likely to be HPV positive if they had received treatment for STI during the past 6-months (OR: 3.4, 95% CI: 1.1-12.4 or if they had ever been pregnant (OR: 2.3, 95% CI: 1.1-5.5. Compared to women who reported only one sexual partner, those with increased number of lifetime sex partners were more likely to have HPV (4-10 partners: OR: 2.9, 95% CI: 1.1-8.0. The high prevalence of HPV types targeted by the nonavalent HPV vaccine encourages the introduction of this vaccine and catch-up HPV vaccination campaigns in South Africa. The high burden of BV and concurrent STIs also highlights the need to improve the prevention and appropriate management of sexually-acquired and other genital tract infections in South African

  7. The Protein Content of Extracellular Vesicles Derived from Expanded Human Umbilical Cord Blood-Derived CD133+and Human Bone Marrow-Derived Mesenchymal Stem Cells Partially Explains Why both Sources are Advantageous for Regenerative Medicine.

    Science.gov (United States)

    Angulski, Addeli B B; Capriglione, Luiz G; Batista, Michel; Marcon, Bruna H; Senegaglia, Alexandra C; Stimamiglio, Marco A; Correa, Alejandro

    2017-04-01

    Adult stem cells have beneficial effects when exposed to damaged tissue due, at least in part, to their paracrine activity, which includes soluble factors and extracellular vesicles (EVs). Given the multiplicity of signals carried by these vesicles through the horizontal transfer of functional molecules, human mesenchymal stem cell (hMSCs) and CD133 + cell-derived EVs have been tested in various disease models and shown to recover damaged tissues. In this study, we profiled the protein content of EVs derived from expanded human CD133 + cells and bone marrow-derived hMSCs with the intention of better understanding the functions performed by these vesicles/cells and delineating the most appropriate use of each EV in future therapeutic procedures. Using LC-MS/MS analysis, we identified 623 proteins for expanded CD133 + -EVs and 797 proteins for hMSCs-EVs. Although the EVs from both origins were qualitatively similar, when protein abundance was considered, hMSCs-EVs and CD133 + -EVs were different. Gene Ontology (GO) enrichment analysis in CD133 + -EVs revealed proteins involved in a variety of angiogenesis-related functions as well proteins related to the cytoskeleton and highly implicated in cell motility and cellular activation. In contrast, when overrepresented proteins in hMSCs-EVs were analyzed, a GO cluster of immune response-related genes involved with immune response-regulating factors acting on phagocytosis and innate immunity was identified. Together our data demonstrate that from the point of view of protein content, expanded CD133 + -EVs and hMSCs-EVs are in part similar but also sufficiently different to reflect the main beneficial paracrine effects widely reported in pre-clinical studies using expanded CD133 + cells and/or hBM-MSCs.

  8. Primary human cervical carcinoma cells require human papillomavirus E6 and E7 expression for ongoing proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Magaldi, Thomas G.; Almstead, Laura L. [Department of Genetics, Yale School of Medicine, P.O. Box 208005, New Haven, CT 06520-8005 (United States); Bellone, Stefania [Department of Obstetrics and Gynecology and Reproductive Sciences, Yale School of Medicine, P.O. Box 208063, New Haven, CT 06520-8063 (United States); Prevatt, Edward G. [Department of Genetics, Yale School of Medicine, P.O. Box 208005, New Haven, CT 06520-8005 (United States); Santin, Alessandro D. [Department of Obstetrics and Gynecology and Reproductive Sciences, Yale School of Medicine, P.O. Box 208063, New Haven, CT 06520-8063 (United States); Yale Comprehensive Cancer Center, P.O. Box 208028, New Haven, CT 06520-8028 (United States); DiMaio, Daniel, E-mail: daniel.dimaio@yale.edu [Department of Genetics, Yale School of Medicine, P.O. Box 208005, New Haven, CT 06520-8005 (United States); Department of Therapeutic Radiology, Yale School of Medicine, P.O. Box 208040, New Haven, CT 06520-8040 (United States); Department of Molecular Biophysics and Biochemistry, Yale School of Medicine, P.O. Box 208024 (United States); Yale Comprehensive Cancer Center, P.O. Box 208028, New Haven, CT 06520-8028 (United States)

    2012-01-05

    Repression of human papillomavirus (HPV) E6 and E7 oncogenes in established cervical carcinoma cell lines causes senescence due to reactivation of cellular tumor suppressor pathways. Here, we determined whether ongoing expression of HPV16 or HPV18 oncogenes is required for the proliferation of primary human cervical carcinoma cells in serum-free conditions at low passage number after isolation from patients. We used an SV40 viral vector expressing the bovine papillomavirus E2 protein to repress E6 and E7 in these cells. To enable efficient SV40 infection and E2 gene delivery, we first incubated the primary cervical cancer cells with the ganglioside GM1, a cell-surface receptor for SV40 that is limiting in these cells. Repression of HPV in primary cervical carcinoma cells caused them to undergo senescence, but the E2 protein had little effect on HPV-negative primary cells. These data suggest that E6 and E7 dependence is an inherent property of human cervical cancer cells.

  9. Inhibition of highly productive HIV-1 infection in T cells, primary human macrophages, microglia, and astrocytes by Sargassum fusiforme

    Directory of Open Access Journals (Sweden)

    Veille Jean-Claude

    2006-05-01

    Full Text Available Abstract Background The high rate of HIV-1 mutation and increasing resistance to currently available antiretroviral (ART therapies highlight the need for new antiviral agents. Products derived from natural sources have been shown to inhibit HIV-1 replication during various stages of the virus life cycle, and therefore represent a potential source of novel therapeutic agents. To expand our arsenal of therapeutics against HIV-1 infection, we investigated aqueous extract from Sargassum fusiforme (S. fusiforme for ability to inhibit HIV-1 infection in the periphery, in T cells and human macrophages, and for ability to inhibit in the central nervous system (CNS, in microglia and astrocytes. Results S. fusiforme extract blocked HIV-1 infection and replication by over 90% in T cells, human macrophages and microglia, and it also inhibited pseudotyped HIV-1 (VSV/NL4-3 infection in human astrocytes by over 70%. Inhibition was mediated against both CXCR4 (X4 and CCR5 (R5-tropic HIV-1, was dose dependant and long lasting, did not inhibit cell growth or viability, was not toxic to cells, and was comparable to inhibition by the nucleoside analogue 2', 3'-didoxycytidine (ddC. S. fusiforme treatment blocked direct cell-to-cell infection spread. To investigate at which point of the virus life cycle this inhibition occurs, we infected T cells and CD4-negative primary human astrocytes with HIV-1 pseudotyped with envelope glycoprotein of vesicular stomatitis virus (VSV, which bypasses the HIV receptor requirements. Infection by pseudotyped HIV-1 (VSV/NL4-3 was also inhibited in a dose dependant manner, although up to 57% less, as compared to inhibition of native NL4-3, indicating post-entry interferences. Conclusion This is the first report demonstrating S. fusiforme to be a potent inhibitor of highly productive HIV-1 infection and replication in T cells, in primary human macrophages, microglia, and astrocytes. Results with VSV/NL4-3 infection, suggest inhibition

  10. Restricted spontaneous in vitro differentiation and region-specific migration of long-term expanded fetal human neural precursor cells after transplantation into the adult rat brain.

    Science.gov (United States)

    Maciaczyk, Jaroslaw; Singec, Ilyas; Maciaczyk, Donata; Klein, Alexander; Nikkhah, Guido

    2009-09-01

    Human fetal neural stem/progenitor cells (hNSCs) are investigated for their potential as a cell source for cell-based therapies in neurodegenerative diseases. However, the limited availability of fetal tissue and insufficient understanding of the lineage-dependent pattern of survival, migration, and differentiation following engraftment are still unresolved issues. In the current study hNSCs derived from different brain regions were long-term expanded in vitro to yield proliferating neurospheres giving rise to neurons, astro-, and oligodendroglial cells and assessed for their potential for migration, differentiation, and anatomical integration following intracerebral grafting into rats. hNSCs isolated from neocortex, striatum, midbrain, and spinal cord (SC) proliferated following in vitro differentiation, and showed a significant decrease of newly formed neurons along the rostrocaudal axis of the developing central nervous system (CNS). Most of the mature neurons were positive for the neurotransmitter GABA. In vivo all cell types survived up to 9 weeks posttransplantation. Intrastriatally grafted hNSCs migrated extensively along white matter tracts reaching both rostral (forceps minor) and caudal (midbrain, cerebral peduncle) brain regions. The majority of migratory cells expressed the stem cell marker, nestin. A fraction of grafted cells acquired a neuronal phenotype expressing doublecortin, beta-III-tubulin, or GABA. These data demonstrate efficient in vitro propagation, region-specific long-term survival, long-distance migration, and neuronal differentiation of hNSCs after transplantation into the adult rat brain. The availability of a large pool of in vitro expanded nestin-positive cells offers the possibility for further ex vivo manipulations and the recruitment of different neuronal phenotypes for cell replacement strategies for CNS disorders.

  11. Characteristics of human infant primary fibroblast cultures from Achilles tendons removed post-mortem

    DEFF Research Database (Denmark)

    Rohde, Marianne Cathrine; Corydon, Thomas Juhl; Hansen, Jakob

    2014-01-01

    Primary cell cultures were investigated as a tool for molecular diagnostics in a forensic setting. Fibroblast cultures had been established from human Achilles tendon resected at autopsies, from cases of sudden infant death syndrome and control infants who died in traumatic events (n=41). After...

  12. Arctigenin from Arctium lappa inhibits interleukin-2 and interferon gene expression in primary human T lymphocytes

    Science.gov (United States)

    2011-01-01

    Background Arctium lappa (Niubang), a Chinese herbal medicine, is used to treat tissue inflammation. This study investigates the effects of arctigenin (AC), isolated from A. lappa, on anti-CD3/CD28 Ab-stimulated cell proliferation and cytokine gene expression in primary human T lymphocytes. Methods Cell proliferation was determined with enzyme immunoassays and the tritiated thymidine uptake method. Cytokine production and gene expression were analyzed with reverse transcription-polymerase chain reaction. Results AC inhibited primary human T lymphocytes proliferation activated by anti-CD3/CD28 Ab. Cell viability test indicated that the inhibitory effects of AC on primary human T lymphocyte proliferation were not due to direct cytotoxicity. AC suppressed interleukin-2 (IL-2) and interferon-γ (IFN-γ) production in a concentration-dependent manner. Furthermore, AC decreased the IL-2 and IFN-γ gene expression in primary human T lymphocytes induced by anti-CD3/CD28 Ab. Reporter gene analyses revealed that AC decreased NF-AT-mediated reporter gene expression. Conclusion AC inhibited T lymphocyte proliferation and decreased the gene expression of IL-2, IFN-γ and NF-AT. PMID:21435270

  13. Humanities-Oriented Accents in Teaching Mathematics to Prospective Primary School Teachers

    Science.gov (United States)

    Tabov, Jordan; Gortcheva, Iordanka

    2016-01-01

    Our research includes undergraduate students who major in primary school education. Their academic background is prevailingly in the humanities. This poses specific demands on their mathematics instruction at university. To attract them to their mathematics course and raise its effectiveness, we use a series of activities. Writing assignments…

  14. Human disturbance provides foraging opportunities for birds in primary subalpine forest

    DEFF Research Database (Denmark)

    DuBay, Shane G.; Hart Reeve, Andrew; Wu, Yongjie

    2017-01-01

    to species that naturally occur in edge, open, or disturbed habitats. With observations and experiments we provide evidence of insectivorous birds exploiting human disturbance in primary subalpine forest in the mountains of southern China, displaying behavioral flexibility to gain novel foraging...

  15. Arctigenin from Arctium lappa inhibits interleukin-2 and interferon gene expression in primary human T lymphocytes.

    Science.gov (United States)

    Tsai, Wei-Jern; Chang, Chu-Ting; Wang, Guei-Jane; Lee, Tzong-Huei; Chang, Shwu-Fen; Lu, Shao-Chun; Kuo, Yuh-Chi

    2011-03-25

    Arctium lappa (Niubang), a Chinese herbal medicine, is used to treat tissue inflammation. This study investigates the effects of arctigenin (AC), isolated from A. lappa, on anti-CD3/CD28 Ab-stimulated cell proliferation and cytokine gene expression in primary human T lymphocytes. Cell proliferation was determined with enzyme immunoassays and the tritiated thymidine uptake method. Cytokine production and gene expression were analyzed with reverse transcription-polymerase chain reaction. AC inhibited primary human T lymphocytes proliferation activated by anti-CD3/CD28 Ab. Cell viability test indicated that the inhibitory effects of AC on primary human T lymphocyte proliferation were not due to direct cytotoxicity. AC suppressed interleukin-2 (IL-2) and interferon-γ (IFN-γ) production in a concentration-dependent manner. Furthermore, AC decreased the IL-2 and IFN-γ gene expression in primary human T lymphocytes induced by anti-CD3/CD28 Ab. Reporter gene analyses revealed that AC decreased NF-AT-mediated reporter gene expression. AC inhibited T lymphocyte proliferation and decreased the gene expression of IL-2, IFN-γ and NF-AT.

  16. Arctigenin from Arctium lappa inhibits interleukin-2 and interferon gene expression in primary human T lymphocytes

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    Chang Shwu-Fen

    2011-03-01

    Full Text Available Abstract Background Arctium lappa (Niubang, a Chinese herbal medicine, is used to treat tissue inflammation. This study investigates the effects of arctigenin (AC, isolated from A. lappa, on anti-CD3/CD28 Ab-stimulated cell proliferation and cytokine gene expression in primary human T lymphocytes. Methods Cell proliferation was determined with enzyme immunoassays and the tritiated thymidine uptake method. Cytokine production and gene expression were analyzed with reverse transcription-polymerase chain reaction. Results AC inhibited primary human T lymphocytes proliferation activated by anti-CD3/CD28 Ab. Cell viability test indicated that the inhibitory effects of AC on primary human T lymphocyte proliferation were not due to direct cytotoxicity. AC suppressed interleukin-2 (IL-2 and interferon-γ (IFN-γ production in a concentration-dependent manner. Furthermore, AC decreased the IL-2 and IFN-γ gene expression in primary human T lymphocytes induced by anti-CD3/CD28 Ab. Reporter gene analyses revealed that AC decreased NF-AT-mediated reporter gene expression. Conclusion AC inhibited T lymphocyte proliferation and decreased the gene expression of IL-2, IFN-γ and NF-AT.

  17. Primary human ovarian epithelial cancer cells broadly express HER2 at immunologically-detectable levels.

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    Evripidis Lanitis

    Full Text Available The breadth of HER2 expression by primary human ovarian cancers remains controversial, which questions its suitability as a universal antigen in this malignancy. To address these issues, we performed extensive HER2 expression analysis on a wide panel of primary tumors as well as established and short-term human ovarian cancer cell lines. Conventional immunohistochemical (IHC analysis of multiple tumor sites in 50 cases of high-grade ovarian serous carcinomas revealed HER2 overexpression in 29% of evaluated sites. However, more sensitive detection methods including flow cytometry, western blot analysis and q-PCR revealed HER2 expression in all fresh tumor cells derived from primary ascites or solid tumors as well as all established and short-term cultured cancer cell lines. Cancer cells generally expressed HER2 at higher levels than that found in normal ovarian surface epithelial (OSE cells. Accordingly, genetically-engineered human T cells expressing an HER2-specific chimeric antigen receptor (CAR recognized and reacted against all established or primary ovarian cancer cells tested with minimal or no reactivity against normal OSE cells. In conclusion, all human ovarian cancers express immunologically-detectable levels of HER2, indicating that IHC measurement underestimates the true frequency of HER2-expressing ovarian cancers and may limit patient access to otherwise clinically meaningful HER2-targeted therapies.

  18. Characterization of a unique technique for culturing primary adult human epithelial progenitor/“stem cells”

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    Marcelo Cynthia

    2012-06-01

    Full Text Available Abstract Background Primary keratinocytes derived from epidermis, oral mucosa, and urothelium are used in construction of cell based wound healing devices and in regenerative medicine. This study presents in vitro technology that rapidly expands keratinocytes in culture by growing monolayers under large volumes of serum-free, essential fatty acid free, low calcium medium that is replaced every 24 hrs. Methods Primary cell cultures were produced from epidermal skin, oral mucosa and ureter by trypsinization of tissue. Cells were grown using Epilife medium with growth factors under high medium volumes. Once densely confluent, the keratinocyte monolayer produced cells in suspension in the overlying medium that can be harvested every 24 hrs. over a 7–10 day period. The cell suspension (approximately 8 X 105 cells/ml is poured into a new flask to form another confluent monolayer over 2–4 days. This new culture, in turn produced additional cell suspensions that when serially passed expand the cell strain over 2–3 months, without the use of enzymes to split the cultures. The cell suspension, called epithelial Pop Up Keratinocytes (ePUKs were analyzed for culture expansion, cell size and glucose utilization, attachment to carrier beads, micro-spheroid formation, induction of keratinocyte differentiation, and characterized by immunohistochemistry. Results The ePUKs expanded greatly in culture, attached to carrier beads, did not form micro-spheroids, used approximately 50% of medium glucose over 24 hrs., contained a greater portion of smaller diameter cells (8–10 microns, reverted to classical appearing cultures when returned to routine feeding schedules (48 hrs. and 15 ml/T-75 flask and can be differentiated by either adding 1.2 mM medium calcium, or essential fatty acids. The ePUK cells are identified as cycling (Ki67 expressing basal cells (p63, K14 expressing. Conclusions Using this primary culture technique, large quantities

  19. Epidermal Growth Factor Rescues Endothelial Dysfunction in Primary Human Tissues In Vitro.

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    Hastie, Roxanne; Tong, Stephen; Hannan, Natalie J; Brownfoot, Fiona; Cannon, Ping; Kaitu'u-Lino, Tu'uhevaha J

    2017-09-01

    Preeclampsia is a hypertensive disorder of pregnancy, responsible for over 60 000 maternal deaths annually. Endothelial dysfunction is a central aspect to its pathophysiology, and currently, no medical therapeutic is available for its treatment. In this study, we aim to investigate the effect of epidermal growth factor (EGF) on endothelial dysfunction using primary human tissues. We performed a number of in vitro assays that mimic the vascular endothelial dysfunction that occurs in preeclampsia. Epidermal growth factor reduced the expression of vascular cell adhesion molecule-1, a marker of endothelial dysfunction, after insult with tumor necrosis factor α (TNF-α) or serum from women with preeclampsia. Additionally, after TNF-α insult, EGF reduced tube disruption and the adhesion of monocytes to primary human umbilical vein endothelial cells (HUVECs). Our findings suggest that EGF reduces endothelial dysfunction in primary HUVECs. Epidermal growth factor may have potential as a novel peptide treatment for preeclampsia and other diseases where there is endothelial dysfunction.

  20. Invariant NKT cells are expanded in peripheral blood but are undetectable in salivary glands of patients with primary Sjögren's syndrome.

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    Guggino, Giuliana; Ciccia, Francesco; Raimondo, Stefania; Giardina, Giuseppina; Alessandro, Riccardo; Dieli, Francesco; Sireci, Guido; Triolo, Giovanni

    2016-01-01

    Invariant NKT (iNKT) cells play a role in regulating the function of autoreactive B cells before their entry into germinal centres. Absence and/or reduction of iNKT cells have been demonstrated in patients with systemic lupus erythematosus (SLE) together with an increase of autoreactive B cell activity. Primary Sjögren's syndrome (pSS) is a systemic autoimmune disease in which lymphocyte infiltration and organisation in lymphoid structures of inflamed salivary glands occurs. The aim of the study was to investigate the percentage and function of iNKT in the salivary glands and peripheral blood of patients with pSS. Minor salivary gland biopsies were obtained from patients with pSS and with non-specific chronic sialoadenitis (nSS). Flow cytometry analysis of CD1d/α-GalactosylCeramide (α-GalCer) tetramer positive cells, producing IFN-γ and IL-17, and quantitative gene expression analysis by TaqMan real-time PCR for Vα24 were performed on salivary glands biopsies and peripheral blood samples obtained from patients and controls. Flow cytometry and immunofluorescence analysis for autoreactive B lymphocytes and ELISA for anti-SSA antibodies (Ab) detection were also performed. An increase of iNKT was detected ex vivo in peripheral blood of pSS patients; after α-GalCer stimulation this subset produce IL-17 and IFN-iNKT were undetectable in the salivary glands of pSS patients and anti-SSA specific B cells were found in target tissue. Invariant NKT cells were able to inhibit autoantibody production by B cells obtained from salivary glands of pSS. Impaired iNKT migration to inflamed sites might induce the activation of autoreactive B cells specific for SSA-antigen in salivary glands of pSS patients.

  1. The role of arginine vasopressin in electroacupuncture treatment of primary sciatica in human.

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    Zhao, Xue-Yan; Zhang, Qi-Shun; Yang, Jun; Sun, Fang-Jie; Wang, Da-Xin; Wang, Chang-Hong; He, Wei-Ya

    2015-08-01

    It has been implicated that electroacupuncture can relieve the symptoms of sciatica with the increase of pain threshold in human, and arginine vasopressin (AVP) in the brain rather than the spinal cord and blood circulation participates in antinociception. Our previous study has proven that AVP in the brain played a role in the process of electroacupuncture analgesia in rat. The goal of the present study was to investigate the role of AVP in electroacupuncture in treating primary sciatica in human. The results showed that (1) AVP concentration of cerebrospinal fluid (CSF) (7.5 ± 2.5 pg/ml), not plasma (13.2 ± 4.2 pg/ml) in primary sciatica patients was lower than that in health volunteers (16.1 ± 3.8 pg/ml and 12.3 ± 3.4 pg/ml), although the osmotic pressure in CSF and plasma did not change; (2) electroacupuncture of the bilateral "Zusanli" points (St. 36) for 60 min relieved the pain sensation in primary sciatica patients; (3) electroacupuncture increased the AVP level of CSF, not plasma in primary sciatica patients; and (4) there was the positive correlation between the effect of electroacupuncture relieving the pain and the AVP level of CSF in the primary sciatica patients. The data suggested that central AVP, not peripheral AVP might improve the effect of electroacupuncture treatment of primary sciatica in human, i.e., central AVP might take part in the electroacupuncture relieving the pain sensation in primary sciatica patients. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. DIFFERENTIAL-EFFECTS OF EICOSAPENTAENOIC ACID ON GLYCEROLIPID AND APOLIPOPROTEIN-B METABOLISM IN PRIMARY HUMAN HEPATOCYTES COMPARED TO HEPG2 CELLS AND PRIMARY RAT HEPATOCYTES

    NARCIS (Netherlands)

    LIN, YG; SMIT, MJ; HAVINGA, R; VERKADE, HJ; VONK, RJ; KUIPERS, F

    1995-01-01

    We compared the effects of eicosapentaenoic acid (EPA) and oleic acid (OA) on glycerolipid and apolipoprotein B (apoB) metabolism in primary human hepatocytes, HepG2 cells and primary rat hepatocytes. Cells were incubated for 1 to 5 h with 0.25 mM bovine serum albumin in the absence (control) or

  3. Quantifying and mapping the human appropriation of net primary production in earth's terrestrial ecosystems.

    Science.gov (United States)

    Haberl, Helmut; Erb, K Heinz; Krausmann, Fridolin; Gaube, Veronika; Bondeau, Alberte; Plutzar, Christoph; Gingrich, Simone; Lucht, Wolfgang; Fischer-Kowalski, Marina

    2007-07-31

    Human appropriation of net primary production (HANPP), the aggregate impact of land use on biomass available each year in ecosystems, is a prominent measure of the human domination of the biosphere. We present a comprehensive assessment of global HANPP based on vegetation modeling, agricultural and forestry statistics, and geographical information systems data on land use, land cover, and soil degradation that localizes human impact on ecosystems. We found an aggregate global HANPP value of 15.6 Pg C/yr or 23.8% of potential net primary productivity, of which 53% was contributed by harvest, 40% by land-use-induced productivity changes, and 7% by human-induced fires. This is a remarkable impact on the biosphere caused by just one species. We present maps quantifying human-induced changes in trophic energy flows in ecosystems that illustrate spatial patterns in the human domination of ecosystems, thus emphasizing land use as a pervasive factor of global importance. Land use transforms earth's terrestrial surface, resulting in changes in biogeochemical cycles and in the ability of ecosystems to deliver services critical to human well being. The results suggest that large-scale schemes to substitute biomass for fossil fuels should be viewed cautiously because massive additional pressures on ecosystems might result from increased biomass harvest.

  4. Maintenance of Hepatic Functions in Primary Human Hepatocytes Cultured on Xeno-Free and Chemical Defined Human Recombinant Laminins.

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    Watanabe, Masaaki; Zemack, Helen; Johansson, Helene; Hagbard, Louise; Jorns, Carl; Li, Meng; Ellis, Ewa

    2016-01-01

    Refined methods for maintaining specific functions of isolated hepatocytes under xeno-free and chemical defined conditions is of great importance for the development of hepatocyte research and regenerative therapy. Laminins, a large family of heterotrimeric basement membrane adhesion proteins, are highly cell and tissue type specific components of the extracellular matrix and strongly influence the behavior and function of associated cells and/or tissues. However, detailed biological functions of many laminin isoforms are still to be evaluated. In this study, we determined the distribution of laminin isoforms in human liver tissue and isolated primary human hepatocytes by western blot analysis, and investigated the efficacy of different human recombinant laminin isoforms on hepatic functions during culture. Protein expressions of laminin-chain α2, α3, α4, β1, β3, γ1, and γ2 were detected in both isolated human hepatocytes and liver tissue. No α1 and α5 expression could be detected in liver tissue or hepatocytes. Hepatocytes were isolated from five different individual livers, and cultured on human recombinant laminin isoforms -111, -211, -221, -332, -411, -421, -511, and -521 (Biolamina AB), matrigel (extracted from Engelbreth-Holm-Swarm sarcoma), or collagen type IV (Collagen). Hepatocytes cultured on laminin showed characteristic hexagonal shape in a flat cell monolayer. Viability, double stranded DNA concentration, and Ki67 expression for hepatocytes cultured for six days on laminin were comparable to those cultured on EHS and Collagen. Hepatocytes cultured on laminin also displayed production of human albumin, alpha-1-antitrypsin, bile acids, and gene expression of liver-enriched factors, such as hepatocyte nuclear factor 4 alpha, glucose-6-phosphate, cytochrome P450 3A4, and multidrug resistance-associated protein 2. We conclude that all forms of human recombinant laminin tested maintain cell viability and liver-specific functions of primary human

  5. One-step immortalization of primary human airway epithelial cells capable of oncogenic transformation.

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    Smith, Jordan L; Lee, Liam C; Read, Abigail; Li, Qiuning; Yu, Bing; Lee, Chih-Shia; Luo, Ji

    2016-01-01

    The ability to transform normal human cells into cancer cells with the introduction of defined genetic alterations is a valuable method for understanding the mechanisms of oncogenesis. Easy establishment of immortalized but non-transformed human cells from various tissues would facilitate these genetic analyses. We report here a simple, one-step immortalization method that involves retroviral vector mediated co-expression of the human telomerase protein and a shRNA targeting the CDKN2A gene locus. We demonstrate that this method could successfully immortalize human small airway epithelial cells while maintaining their chromosomal stability. We further showed that these cells retain p53 activity and can be transformed by the KRAS oncogene. Our method simplifies the immortalization process and is broadly applicable for establishing immortalized epithelial cell lines from primary human tissues for cancer research.

  6. Catalyst for Expanding Human Spaceflight

    Science.gov (United States)

    Lueders, Kathryn L.

    2014-01-01

    History supplies us with many models of how and how not to commercialize an industry. This presentation draws parallels between industries with government roots, like the railroad, air transport, communications and the internet, and NASAs Commercial Crew Program. In these examples, government served as a catalyst for what became a booming industry. The building block approach the Commercial Crew Program is taking is very simple -- establish a need, laying the groundwork, enabling industry and legal framework.

  7. Hyaline Cartilage Tissue Is Formed through the Co-culture of Passaged Human Chondrocytes and Primary Bovine Chondrocytes

    Science.gov (United States)

    Taylor, Drew W.; Ahmed, Nazish; Hayes, Anthony J.; Ferguson, Peter; Gross, Allan E.; Caterson, Bruce

    2012-01-01

    To circumvent the problem of a sufficient number of cells for cartilage engineering, the authors previously developed a two-stage culture system to redifferentiate monolayer culture-expanded dedifferentiated human articular chondrocytes by co-culture with primary bovine chondrocytes (bP0). The aim of this study was to analyze the composition of the cartilage tissue formed in stage 1 and compare it with bP0 grown alone to determine the optimal length of the co-culture stage of the system. Biochemical data show that extracellular matrix accumulation was evident after 2 weeks of co-culture, which was 1 week behind the bP0 control culture. By 3 to 4 weeks, the amounts of accumulated proteoglycans and collagens were comparable. Expression of chondrogenic genes, Sox 9, aggrecan, and collagen type II, was also at similar levels by week 3 of culture. Immunohistochemical staining of both co-culture and control tissues showed accumulation of type II collagen, aggrecan, biglycan, decorin, and chondroitin sulfate in appropriate zonal distributions. These data indicate that co-cultured cells form cartilaginous tissue that starts to resemble that formed by bP0 after 3 weeks, suggesting that the optimal time to terminate the co-culture stage, isolate the now redifferentiated cells, and start stage 2 is just after 3 weeks. PMID:22610463

  8. Screening for fecal carriage of MCR-producing Enterobacteriaceae in healthy humans and primary care patients

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    Katrin Zurfluh

    2017-03-01

    Full Text Available Abstract Background The extent of the occurrence of the plasmid-encoded colistin resistance genes mcr-1 and mcr-2 among humans is currently sparsely studied in Western Europe. Objectives To determine the occurrence of MCR-producing Enterobacteriaceae in fecal samples of healthy humans with high occupational exposure to food and primary care patients in Switzerland. Methods Stool samples from 1091 healthy individuals and fecal swabs from 53 primary care patients were screened for polymyxin-resistant Enterobacteriaceae using LB agar containing 4 mg/L colistin. Minimal inhibitory concentrations (MICs of colistin were determined for non-intrinsic colistin-resistant isolates. Isolates were screened by PCR for the presence of mcr-1 and mcr-2 genes. Results The fecal carriage rate of colistin resistant (MIC value >2 mg/l Enterobacteriaceae was 1.5% for healthy people and 3.8% for primary care patients. Isolates included Hafnia alvei (n = 9, Escherichia coli (n = 3, Enterobacter cloacae (n = 4, Klebsiella pneumoniae (n = 1 and Raoultella ornithinolytica (n = 1. None of the isolates harbored the mcr-1 or mcr-2 genes. Conclusions There is no evidence for the presence of MCR-producers in the fecal flora of healthy people or primary care patients. Therefore, the risk of transfer of mcr genes from animals, food or the environment to humans is likely to be low in Switzerland.

  9. Hypogammaglobulinemia in BLT humanized mice--an animal model of primary antibody deficiency.

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    Francisco Martinez-Torres

    Full Text Available Primary antibody deficiencies present clinically as reduced or absent plasma antibodies without another identified disorder that could explain the low immunoglobulin levels. Bone marrow-liver-thymus (BLT humanized mice also exhibit primary antibody deficiency or hypogammaglobulinemia. Comprehensive characterization of B cell development and differentiation in BLT mice revealed other key parallels with primary immunodeficiency patients. We found that B cell ontogeny was normal in the bone marrow of BLT mice but observed an absence of switched memory B cells in the periphery. PC-KLH immunizations led to the presence of switched memory B cells in immunized BLT mice although plasma cells producing PC- or KLH- specific IgG were not detected in tissues. Overall, we have identified the following parallels between the humoral immune systems of primary antibody deficiency patients and those in BLT mice that make this in vivo model a robust and translational experimental platform for gaining a greater understanding of this heterogeneous array of humoral immunodeficiency disorders in humans: (i hypogammaglobulinemia; (ii normal B cell ontogeny in bone marrow; and (iii poor antigen-specific IgG response to immunization. Furthermore, the development of strategies to overcome these humoral immune aberrations in BLT mice may in turn provide insights into the pathogenesis of some primary antibody deficiency patients which could lead to novel clinical interventions for improved humoral immune function.

  10. Indicator cell lines for detection of primary strains of human and simian immunodeficiency viruses.

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    Vodicka, M A; Goh, W C; Wu, L I; Rogel, M E; Bartz, S R; Schweickart, V L; Raport, C J; Emerman, M

    1997-06-23

    CCR5 and CXCR4 are the two major coreceptors that have been identified for human immunodeficiency virus (HIV) entry. We have modified several beta-galactosidase-based HIV indicator cell lines to express CCR5 and/or CXCR4. Using these new reagents, we have been able to detect all primary isolates tested using one or both of these cell lines. However, there is large variation in the absolute viral infectivity among primary strains. Furthermore, all HIV strains are capable of causing syncytia in the indicator cells when the coreceptor is present regardless of whether they had previously been characterized as "syncytia-inducing" or "non-syncytium-inducing."

  11. [Establishment of nude mice liver metastatic model of human primary malignant melanoma of the small intestine].

    Science.gov (United States)

    Tuo, Shuai; Zhang, Ning; Liu, Qiu-Zhen

    2008-07-01

    To provide ideal animal models for exploring pathogenesis and experimental therapy of primary malignant melanoma of the small intestine. The histologically intact primary and liver metastatic fragments derived from surgical specimens of one patient with metastatic malignant melanoma of the small intestine were orthotopically implanted in the small intestinal mucous layer of nude mice. The take rate, invasion and liver metastasis were observed. Morphology (light microscopy, electron microscopy), immunophenotype analysis, flow cytometry and karyotype analysis were applied for the original human tumors and the transplanted tumors. The primary and liver metastatic fragments of malignant melanoma of the small intestine were successfully implanted in nude mice. After continuous passages in nude mice,an orthotopic model of human primary malignant melanoma of the small intestine(from the primary focus)in nude mice (termed HSIM-0501) and a liver metastatic model of human primary malignant melanoma of the small intestine (from the liver metastatic focus) in nude mice (termed HSIM-0502) were established. Histological examination of transplanted tumors revealed high-grade melanoma. S-100 protein and HMB45 were positive. Massive melanin granules and melanin complex were seen in cytoplasm of tumor cells.Chromosomal modal number was between 55 and 59. DNA index (DI) was 1.49-1.61, representing heteroploid. HSIM-0501 and HSIM-0502 were maintained for 25 and 27 passages in nude mice respectively. Three hundred and seventeen nude mice were used for transplantation. Both the take rate after transplantation and resuscitation rate of liquid nitrogen cryopreservation were 100%. HSIM-0501 exhibited 46.2% liver metastasis and 36.7% lymph node metastases. In HSIM-0502, both liver and lymph node metastases were 100%.The transplanted tumors autonomically and invasively grew in the small intestines of nude mice and hematogenous metastasis, lymph node metastasis and celiac planting metastasis

  12. Mesenchymal stem cells expanded in human platelet lysate display a decreased inhibitory capacity on T- and NK-cell proliferation and function.

    Science.gov (United States)

    Abdelrazik, Heba; Spaggiari, Grazia M; Chiossone, Laura; Moretta, Lorenzo

    2011-11-01

    The use of fetal bovine serum (FBS) for the culture and expansion of mesenchymal stromal cells (MSCs) limits their possible clinical applications. Although some recent studies recommended substituting FBS with human platelet lysate (HPL) for the expansion of MSCs for clinical use, the functional capacity of the expanded cells has only been partially explored. 10% FBS and two other commercial FBS-containing media (MesenCult and MesenPro) were compared with 10% HPL-containing medium for their ability to support MSCs expansion and immunomodulation. We demonstrate that HPL sustained MSC proliferation and expansion in vitro. However, the cumulative cell numbers recovered were comparable with those obtained in MesenPro medium. Moreover, we show that HPL alters the expression of some relevant MSC surface molecules, namely the DNAM-1 ligands PVR and Nectin-2, the NKG2D ligand ULBP3, the adhesion molecules CD49d and αvβ3 and the fibroblast-associated protein. In addition, MSCs cultured in HPL displayed impaired inhibitory capacity on T-cell proliferation to alloantigen and NK-cell proliferation and cytotoxicity. Finally, they showed decreased constitutive PGE2 production while IL-6, IL-8 and RANTES secretion were upregulated. These results imply some limitations in the use of HPL for the expansion of MSCs to be used as immunomodulators in clinical applications. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Cell-to-Cell Contact and Nectin-4 Govern Spread of Measles Virus from Primary Human Myeloid Cells to Primary Human Airway Epithelial Cells

    Science.gov (United States)

    Singh, Brajesh K.; Li, Ni; Mark, Anna C.; Mateo, Mathieu; Cattaneo, Roberto

    2016-01-01

    ABSTRACT Measles is a highly contagious, acute viral illness. Immune cells within the airways are likely first targets of infection, and these cells traffic measles virus (MeV) to lymph nodes for amplification and subsequent systemic dissemination. Infected immune cells are thought to return MeV to the airways; however, the mechanisms responsible for virus transfer to pulmonary epithelial cells are poorly understood. To investigate this process, we collected blood from human donors and generated primary myeloid cells, specifically, monocyte-derived macrophages (MDMs) and dendritic cells (DCs). MDMs and DCs were infected with MeV and then applied to primary cultures of well-differentiated airway epithelial cells from human donors (HAE). Consistent with previous results obtained with free virus, infected MDMs or DCs were incapable of transferring MeV to HAE when applied to the apical surface. Likewise, infected MDMs or DCs applied to the basolateral surface of HAE grown on small-pore (0.4-μm) support membranes did not transfer virus. In contrast, infected MDMs and DCs applied to the basolateral surface of HAE grown on large-pore (3.0-μm) membranes successfully transferred MeV. Confocal microscopy demonstrated that MDMs and DCs are capable of penetrating large-pore membranes but not small-pore membranes. Further, by using a nectin-4 blocking antibody or recombinant MeV unable to enter cells through nectin-4, we demonstrated formally that transfer from immune cells to HAE occurs in a nectin-4-dependent manner. Thus, both infected MDMs and DCs rely on cell-to-cell contacts and nectin-4 to efficiently deliver MeV to the basolateral surface of HAE. IMPORTANCE Measles virus spreads rapidly and efficiently in human airway epithelial cells. This rapid spread is based on cell-to-cell contact rather than on particle release and reentry. Here we posit that MeV transfer from infected immune cells to epithelial cells also occurs by cell-to-cell contact rather than through cell

  14. Effect of phenylhexyl isothiocyanate on aberrant histone H3 methylation in primary human acute leukemia

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    Zou Yong

    2012-07-01

    Full Text Available Abstract Background We have previously studied the histone acetylation in primary human leukemia cells. However, histone H3 methylation in these cells has not been characterized. Methods This study examined the methylation status at histone H3 lysine 4 (H3K4 and histone H3 lysine 9 (H3K9 in primary acute leukemia cells obtained from patients and compared with those in the non-leukemia and healthy cells. We further characterized the effect of phenylhexyl isothiocyanate (PHI, Trichostatin A (TSA, and 5-aza-2’-deoxycytidine (5-Aza on the cells. Results We found that methylation of histone H3K4 was virtually undetectable, while methylation at H3K9 was significantly higher in primary human leukemia cells. The histone H3K9 hypermethylation and histone H3K4 hypomethylation were observed in both myeloid and lymphoid leukemia cells. PHI was found to be able to normalize the methylation level in the primary leukemia cells. We further showed that PHI was able to enhance the methyltransferase activity of H3K4 and decrease the activity of H3K9 methyltransferase. 5-Aza had similar effect on H3K4, but minimal effect on H3K9, whereas TSA had no effect on H3K4 and H3K9 methyltransferases. Conclusions This study revealed opposite methylation level of H3K4 and H3K9 in primary human leukemia cells and demonstrated for the first time that PHI has different effects on the methyltransferases for H3K4 and H3K9.

  15. Momordica charantia (bitter melon inhibits primary human adipocyte differentiation by modulating adipogenic genes

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    Nerurkar Vivek R

    2010-06-01

    Full Text Available Abstract Background Escalating trends of obesity and associated type 2 diabetes (T2D has prompted an increase in the use of alternative and complementary functional foods. Momordica charantia or bitter melon (BM that is traditionally used to treat diabetes and complications has been demonstrated to alleviate hyperglycemia as well as reduce adiposity in rodents. However, its effects on human adipocytes remain unknown. The objective of our study was to investigate the effects of BM juice (BMJ on lipid accumulation and adipocyte differentiation transcription factors in primary human differentiating preadipocytes and adipocytes. Methods Commercially available cryopreserved primary human preadipocytes were treated with and without BMJ during and after differentiation. Cytotoxicity, lipid accumulation, and adipogenic genes mRNA expression was measured by commercial enzymatic assay kits and semi-quantitative RT-PCR (RT-PCR. Results Preadipocytes treated with varying concentrations of BMJ during differentiation demonstrated significant reduction in lipid content with a concomitant reduction in mRNA expression of adipocyte transcription factors such as, peroxisome proliferator-associated receptor γ (PPARγ and sterol regulatory element-binding protein 1c (SREBP-1c and adipocytokine, resistin. Similarly, adipocytes treated with BMJ for 48 h demonstrated reduced lipid content, perilipin mRNA expression, and increased lipolysis as measured by the release of glycerol. Conclusion Our data suggests that BMJ is a potent inhibitor of lipogenesis and stimulator of lipolysis activity in human adipocytes. BMJ may therefore prove to be an effective complementary or alternative therapy to reduce adipogenesis in humans.

  16. A Primary Human Critical Success Factors Model for the ERP System Implementation

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    Jenko Aleksander

    2016-08-01

    Full Text Available Background and Purpose: Many researchers have investigated various Critical success factors (CSFs and the different causes of ERP implementation project failures. Despite a detailed literature preview, we were unable to find an appropriate research with a comprehensive overview of the true causes behind CSFs, observed from a human factors perspective. The objective of this research was therefore to develop and evaluate the Primary human factors (PHFs model and to confirm the significant impact of PHFs on traditional CSFs and on the project success.

  17. Phosphatase inhibitor 2 promotes acetylation of tubulin in the primary cilium of human retinal epithelial cells

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    Wang Weiping

    2008-11-01

    Full Text Available Abstract Background Primary cilia are flagella-like projections from the centriole of mammalian cells that have a key role in cell signaling. Human diseases are linked to defects in primary cilia. Microtubules make up the axoneme of cilia and are selectively acetylated and this is thought to contribute to the stability of the structure. However, mechanisms to regulate tubulin acetylation in cilia are poorly understood. Results Endogenous phosphatase inhibitor-2 (I-2 was found concentrated in cilia of human epithelial cells, and was localized to cilia early in the process of formation, prior to the full acetylation of microtubules. Knockdown of I-2 by siRNA significantly reduced the acetylation of microtubules in cilia, without a net decrease in whole cell tubulin acetylation. There was a reduction in the percentage of I-2 knockdown cells with a primary cilium, but no apparent alteration in the cilium length, suggesting no change in microtubule-based transport processes. Inhibition of either histone deacetylases with trichostatin A, or protein phosphatase-1 with calyculin A in I-2 knockdown cells partially rescued the acetylation of microtubules in cilia and the percentage of cells with a primary cilium. Conclusion The regulatory protein I-2 localizes to the primary cilium where it affects both Ser/Thr phosphorylation and is required for full tubulin acetylation. Rescue of tubulin acetylation in I-2 knockdown cells by different chemical inhibitors shows that deacetylases and phosphatases are functionally interconnected to regulate microtubules. As a multifunctional protein, I-2 may link cell cycle progression to structure and stability of the primary cilium.

  18. Sensitivity of endometrial cancer cells from primary human tumor samples to new potential anticancer peptide lactaptin.

    Science.gov (United States)

    Koval, Olga A; Sakaeva, Galiya R; Fomin, Alexander S; Nushtaeva, Anna A; Semenov, Dmitry V; Kuligina, Elena V; Gulyaeva, Ludmila F; Gerasimov, Alexey V; Richter, Vladimir A

    2015-01-01

    Endometrial carcinoma is the most common gynecologic malignancy which is associated with a poor prognosis when diagnosed at an advanced stage; therefore, the discovery of efficacious new drugs is required to reinforce conventional chemotherapy. Short-term cultures of primary cells from endometrial tumors could be used for testing new anticancer therapeutics as well as for the development of personalized cancer therapy strategy. Here, the antitumor effect of a recombinant analogue of lactaptin (RL2), a new potential anticancer molecule, was examined against primary human endometrial cancer cells. Primary cell cultures of malignant and normal human endometrium were performed by enzymatic digestion of endometrial tissue from biopsy material. Real-time quantitative reverse transcription polymerase chain reaction (RT-PCR) was performed to determine the messenger ribonucleic acid (mRNA) state of estrogen (ERs) and progesterone (PRs) hormone receptors and aromatase (Cyp 19) in cell cultures. Dynamic monitoring of cell adhesion and proliferation was made using the iCELLigence system (ASEA Biosciences). The sensitivity of cell cultures to conventional anticancer drugs and the lactaptin analog was estimated by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, flow cytometry, and the iCELLligence system. Established short-term primary cultures of endometrial cancer cells were ERα/ERβ/PR-positive and sensitive for RL2. The IC 50 values of doxorubicin and cisplatin were determined for all of the primary cultures designed. KE normal cells displaying low Cyp19 mRNA levels and high ERβ and PR mRNA levels were more resistant to RL2 treatment as well as to cisplatin and doxorubicin. Our results indicate that the recombinant analog of lactaptin, RL2, exerts cytotoxic effects against primary hormone-dependent endometrial tumor cells in vitro with features of apoptosis.

  19. Dissection of the transformation of primary human hematopoietic cells by the oncogene NUP98-HOXA9.

    Directory of Open Access Journals (Sweden)

    Enas R Yassin

    2009-08-01

    Full Text Available NUP98-HOXA9 is the prototype of a group of oncoproteins associated with acute myeloid leukemia. It consists of an N-terminal portion of NUP98 fused to the homeodomain of HOXA9 and is believed to act as an aberrant transcription factor that binds DNA through the homeodomain. Here we show that NUP98-HOXA9 can regulate transcription without binding to DNA. In order to determine the relative contributions of the NUP98 and HOXA9 portions to the transforming ability of NUP98-HOXA9, the effects of NUP98-HOXA9 on primary human CD34+ cells were dissected and compared to those of wild-type HOXA9. In contrast to previous findings in mouse cells, HOXA9 had only mild effects on the differentiation and proliferation of primary human hematopoietic cells. The ability of NUP98-HOXA9 to disrupt the differentiation of primary human CD34+ cells was found to depend primarily on the NUP98 portion, whereas induction of long-term proliferation required both the NUP98 moiety and an intact homeodomain. Using oligonucleotide microarrays in primary human CD34+ cells, a group of genes was identified whose dysregulation by NUP98-HOXA9 is attributable primarily to the NUP98 portion. These include RAP1A, HEY1, and PTGS2 (COX-2. Their functions may reflect the contribution of the NUP98 moiety of NUP98-HOXA9 to leukemic transformation. Taken together, these results suggest that the effects of NUP98-HOXA9 on gene transcription and cell transformation are mediated by at least two distinct mechanisms: one that involves promoter binding through the homeodomain with direct transcriptional activation, and another that depends predominantly on the NUP98 moiety and does not involve direct DNA binding.

  20. Human umbilical mesenchymal stem cells conditioned medium promote primary wound healing regeneration

    OpenAIRE

    Dwi Liliek Kusindarta3; Hevi Wihadmadyatami; Yuda Heru Fibrianto; Widagdo Sri Nugroho; Heru Susetya; Dewi Kania Musana; Hery Wijayanto; Surya Agus Prihatna; Wahyuni, A. E. T. H.

    2016-01-01

    Aim: This research was conducted to clarify the capability of human umbilical mesenchymal stem cells conditioned medium (HU-MSCM) to promote regenerations of primary wound healing on the incision skin injury. Materials and Methods: In this study, two approaches in vitro and in vivo already done. On in vitro analysis, tube formation was performed using HU vein endothelial cells in the presence of HU-MSCM, in some experiments cells line was incubated prior the presence of lipopolysaccharide ...

  1. Isolation and primary culture of various cell types from whole human endometrial biopsies

    OpenAIRE

    Barros, Flavio; Brosens, Jan J.; Brighton, Paul (Paul J.)

    2016-01-01

    The isolation and primary culture of cells from human endometrial biopsies provides valuable experimental material for reproductive and gynaecological research. Whole endometrial biopsies are collected from consenting women and digested with collagenase and DNase I to dissociate cells from the extracellular matrix. Cell populations are then isolated through culturing, filtering and magnetic separation using cell-surface antigen markers. Here we provide a comprehensive protocol on how to isola...

  2. A phenotypic comparison of osteoblast cell lines versus human primary osteoblasts for biomaterials testing.

    Science.gov (United States)

    Czekanska, E M; Stoddart, M J; Ralphs, J R; Richards, R G; Hayes, J S

    2014-08-01

    Immortalized cell lines are used more frequently in basic and applied biology research than primary bone-derived cells because of their ease of access and repeatability of results in experiments. It is clear that these cell models do not fully resemble the behavior of primary osteoblast cells. Although the differences will affect the results of biomaterials testing, they are not clearly defined. Here, we focused on comparing proliferation and maturation potential of three osteoblast cell lines, SaOs2, MG-63, and MC3T3-E1 with primary human osteoblast (HOb) cells to assess their suitability as in vitro models for biomaterials testing. We report similarities in cell proliferation and mineralization between primary cells and MC3T3-E1. Both, SaOs2 and MG-63 cells demonstrated a higher proliferation rate than HOb cells. In addition, SaOs2, but not MG-63, cells demonstrated similar ALP activity, mineralization potential and gene regulation to HOb's. Our results demonstrate that despite SaOs-2, MG63, and MC3T3 cells being popular choices for emulating osteoblast behavior, none can be considered appropriate replacements for HOb's. Nevertheless, these cell lines all demonstrated some distinct similarities with HOb's, thus when applied in the correct context are a valuable in vitro pilot model of osteoblast functionality, but should not be used to replace primary cell studies. © 2013 Wiley Periodicals, Inc.

  3. Highly efficient gene inactivation by adenoviral CRISPR/Cas9 in human primary cells.

    Science.gov (United States)

    Voets, Olaf; Tielen, Frans; Elstak, Edo; Benschop, Julian; Grimbergen, Max; Stallen, Jan; Janssen, Richard; van Marle, Andre; Essrich, Christian

    2017-01-01

    Phenotypic assays using human primary cells are highly valuable tools for target discovery and validation in drug discovery. Expression knockdown (KD) of such targets in these assays allows the investigation of their role in models of disease processes. Therefore, efficient and fast modes of protein KD in phenotypic assays are required. The CRISPR/Cas9 system has been shown to be a versatile and efficient means of gene inactivation in immortalized cell lines. Here we describe the use of adenoviral (AdV) CRISPR/Cas9 vectors for efficient gene inactivation in two human primary cell types, normal human lung fibroblasts and human bronchial epithelial cells. The effects of gene inactivation were studied in the TGF-β-induced fibroblast to myofibroblast transition assay (FMT) and the epithelial to mesenchymal transition assay (EMT), which are SMAD3 dependent and reflect pathogenic mechanisms observed in fibrosis. Co-transduction (co-TD) of AdV Cas9 with SMAD3-targeting guide RNAs (gRNAs) resulted in fast and efficient genome editing judged by insertion/deletion (indel) formation, as well as significant reduction of SMAD3 protein expression and nuclear translocation. This led to phenotypic changes downstream of SMAD3 inhibition, including substantially decreased alpha smooth muscle actin and fibronectin 1 expression, which are markers for FMT and EMT, respectively. A direct comparison between co-TD of separate Cas9 and gRNA AdV, versus TD with a single "all-in-one" Cas9/gRNA AdV, revealed that both methods achieve similar levels of indel formation. These data demonstrate that AdV CRISPR/Cas9 is a useful and efficient tool for protein KD in human primary cell phenotypic assays. The use of AdV CRISPR/Cas9 may offer significant advantages over the current existing tools and should enhance target discovery and validation opportunities.

  4. Highly efficient gene inactivation by adenoviral CRISPR/Cas9 in human primary cells

    Science.gov (United States)

    Tielen, Frans; Elstak, Edo; Benschop, Julian; Grimbergen, Max; Stallen, Jan; Janssen, Richard; van Marle, Andre; Essrich, Christian

    2017-01-01

    Phenotypic assays using human primary cells are highly valuable tools for target discovery and validation in drug discovery. Expression knockdown (KD) of such targets in these assays allows the investigation of their role in models of disease processes. Therefore, efficient and fast modes of protein KD in phenotypic assays are required. The CRISPR/Cas9 system has been shown to be a versatile and efficient means of gene inactivation in immortalized cell lines. Here we describe the use of adenoviral (AdV) CRISPR/Cas9 vectors for efficient gene inactivation in two human primary cell types, normal human lung fibroblasts and human bronchial epithelial cells. The effects of gene inactivation were studied in the TGF-β-induced fibroblast to myofibroblast transition assay (FMT) and the epithelial to mesenchymal transition assay (EMT), which are SMAD3 dependent and reflect pathogenic mechanisms observed in fibrosis. Co-transduction (co-TD) of AdV Cas9 with SMAD3-targeting guide RNAs (gRNAs) resulted in fast and efficient genome editing judged by insertion/deletion (indel) formation, as well as significant reduction of SMAD3 protein expression and nuclear translocation. This led to phenotypic changes downstream of SMAD3 inhibition, including substantially decreased alpha smooth muscle actin and fibronectin 1 expression, which are markers for FMT and EMT, respectively. A direct comparison between co-TD of separate Cas9 and gRNA AdV, versus TD with a single “all-in-one” Cas9/gRNA AdV, revealed that both methods achieve similar levels of indel formation. These data demonstrate that AdV CRISPR/Cas9 is a useful and efficient tool for protein KD in human primary cell phenotypic assays. The use of AdV CRISPR/Cas9 may offer significant advantages over the current existing tools and should enhance target discovery and validation opportunities. PMID:28800587

  5. Partial primary structure of human pregnancy zone protein: extensive sequence homology with human alpha 2-macroglobulin

    DEFF Research Database (Denmark)

    Sottrup-Jensen, Lars; Folkersen, J; Kristensen, Torsten

    1984-01-01

    the results of complete or partial sequence determination of a random selection of 38 tryptic peptides covering 685 residues of the subunit of PZP, that PZP and alpha 2M indeed are extensively homologous. In the stretches of PZP sequenced so far, the degree of identically placed residues in the two proteins......-mediated endocytosis. In this regard our studies indicate a bait region in PZP significantly different from that present in alpha 2M. PZP could be the human equivalent of the acute-phase alpha-macroglobulins (e.g., rat alpha 2M and rabbit alpha 1M) described earlier...

  6. Fibulin-5 is upregulated in decidualized human endometrial stromal cells and promotes primary human extravillous trophoblast outgrowth.

    Science.gov (United States)

    Winship, Amy; Cuman, Carly; Rainczuk, Katarzyna; Dimitriadis, Evdokia

    2015-12-01

    Interactions between the highly invasive trophoblasts and the maternal uterine decidual extracellular matrix (ECM) are crucial in the determination of a successful pregnancy. Fibulin-5 (FBLN5) is a member of the fibulin family that alters cell adhesive and invasive properties and is expressed in human villous cytotrophoblasts. We aimed to determine the expression and immunolocalization of FBLN5 in human first trimester decidua and examine the effect of FBLN5 in trophoblast invasion in vitro using a first trimester placental villous outgrowth assay. We demonstrated that FBLN5 mRNA expression is upregulated in response to cAMP-mediated decidualization of primary human endometrial stromal cells, although FBLN5 itself does not enhance decidualization. We reported for the first time, FBLN5 protein production in first trimester decidual cells and also co-localization to HLAG-positive EVTs in first trimester decidua. Consequently, we investigated the effects of exogenous FBLN5 on placental villous outgrowth in vitro and demonstrated that FBLN5 promotes EVT migration/invasion. This is the first study to identify FBLN5 in decidualized human endometrial stromal cells, first trimester decidua and EVT and determine a functional role for FBLN5 in human EVTs, suggesting that decidual and or EVT-derived FBLN5 regulates EVT invasion and placentation in women. Copyright © 2015 Elsevier Ltd. All rights reserved.

  7. Trehalose-mediated autophagy impairs the anti-viral function of human primary airway epithelial cells.

    Directory of Open Access Journals (Sweden)

    Qun Wu

    Full Text Available Human rhinovirus (HRV is the most common cause of acute exacerbations of chronic lung diseases including asthma. Impaired anti-viral IFN-λ1 production and increased HRV replication in human asthmatic airway epithelial cells may be one of the underlying mechanisms leading to asthma exacerbations. Increased autophagy has been shown in asthmatic airway epithelium, but the role of autophagy in anti-HRV response remains uncertain. Trehalose, a natural glucose disaccharide, has been recognized as an effective autophagy inducer in mammalian cells. In the current study, we used trehalose to induce autophagy in normal human primary airway epithelial cells in order to determine if autophagy directly regulates the anti-viral response against HRV. We found that trehalose-induced autophagy significantly impaired IFN-λ1 expression and increased HRV-16 load. Inhibition of autophagy via knockdown of autophagy-related gene 5 (ATG5 effectively rescued the impaired IFN-λ1 expression by trehalose and subsequently reduced HRV-16 load. Mechanistically, ATG5 protein interacted with retinoic acid-inducible gene I (RIG-I and IFN-β promoter stimulator 1 (IPS-1, two critical molecules involved in the expression of anti-viral interferons. Our results suggest that induction of autophagy in human primary airway epithelial cells inhibits the anti-viral IFN-λ1 expression and facilitates HRV infection. Intervention of excessive autophagy in chronic lung diseases may provide a novel approach to attenuate viral infections and associated disease exacerbations.

  8. Cytoskeletal stability and metabolic alterations in primary human macrophages in long-term microgravity.

    Directory of Open Access Journals (Sweden)

    Svantje Tauber

    Full Text Available The immune system is one of the most affected systems of the human body during space flight. The cells of the immune system are exceptionally sensitive to microgravity. Thus, serious concerns arise, whether space flight associated weakening of the immune system ultimately precludes the expansion of human presence beyond the Earth's orbit. For human space flight, it is an urgent need to understand the cellular and molecular mechanisms by which altered gravity influences and changes the functions of immune cells. The CELLBOX-PRIME (= CellBox-Primary Human Macrophages in Microgravity Environment experiment investigated for the first time microgravity-associated long-term alterations in primary human macrophages, one of the most important effector cells of the immune system. The experiment was conducted in the U.S. National Laboratory on board of the International Space Station ISS using the NanoRacks laboratory and Biorack type I standard CELLBOX EUE type IV containers. Upload and download were performed with the SpaceX CRS-3 and the Dragon spaceship on April 18th, 2014 / May 18th, 2014. Surprisingly, primary human macrophages exhibited neither quantitative nor structural changes of the actin and vimentin cytoskeleton after 11 days in microgravity when compared to 1g controls. Neither CD18 or CD14 surface expression were altered in microgravity, however ICAM-1 expression was reduced. The analysis of 74 metabolites in the cell culture supernatant by GC-TOF-MS, revealed eight metabolites with significantly different quantities when compared to 1g controls. In particular, the significant increase of free fucose in the cell culture supernatant was associated with a significant decrease of cell surface-bound fucose. The reduced ICAM-1 expression and the loss of cell surface-bound fucose may contribute to functional impairments, e.g. the activation of T cells, migration and activation of the innate immune response. We assume that the surprisingly small

  9. Development and validation of primary human myometrial cell culture models to study pregnancy and labour

    Directory of Open Access Journals (Sweden)

    Mosher Andrea A

    2013-01-01

    Full Text Available Abstract Background The development of the in vitro cell culture model has greatly facilitated the ability to study gene expression and regulation within human tissues. Within the human uterus, the upper (fundal segment and the lower segment may provide distinct functions throughout pregnancy and during labour. We have established primary cultured human myometrial cells, isolated from both upper and lower segment regions of the pregnant human uterus, and validated them for the purpose of studying human pregnancy and labour. The specific objectives of this study were to monitor the viability and characterize the expression profile using selected cellular, contractile and pregnancy associated markers in the primary cultured human myometrial cells. Labour has been described as an inflammatory process; therefore, the ability of these cells to respond to an inflammatory stimulus was also investigated. Methods Myometrial cells isolated from paired upper segment (US and lower segment (LS biopsies, obtained from women undergoing Caesarean section deliveries at term prior to the onset of labour, were used to identify expression of; α smooth muscle actin, calponin, caldesmon, connexin 43, cyclo-oxygenase-2 (COX-2, oxytocin receptor, tropomyosin and vimentin, by RT-PCR and/or immunocytochemistry. Interleukin (IL-1β was used to treat cells, subsequently expression of COX-2 mRNA and release of interleukin-8 (CXCL8, were measured. ANOVA followed by Bonferroni’s multiple comparisons test was performed. Results We demonstrate that US and LS human myometrial cells stably express all markers examined to at least passage ten (p10. Connexin 43, COX-2 and vimentin mRNA expression were significantly higher in LS cells compared to US cells. Both cell populations respond to IL-1β, demonstrated by a robust release of CXCL8 and increased expression of COX-2 mRNA from passage one (p1 through to p10. Conclusions Isolated primary myometrial cells maintain expression of

  10. Cytoskeletal stability and metabolic alterations in primary human macrophages in long-term microgravity.

    Science.gov (United States)

    Tauber, Svantje; Lauber, Beatrice A; Paulsen, Katrin; Layer, Liliana E; Lehmann, Martin; Hauschild, Swantje; Shepherd, Naomi R; Polzer, Jennifer; Segerer, Jürgen; Thiel, Cora S; Ullrich, Oliver

    2017-01-01

    The immune system is one of the most affected systems of the human body during space flight. The cells of the immune system are exceptionally sensitive to microgravity. Thus, serious concerns arise, whether space flight associated weakening of the immune system ultimately precludes the expansion of human presence beyond the Earth's orbit. For human space flight, it is an urgent need to understand the cellular and molecular mechanisms by which altered gravity influences and changes the functions of immune cells. The CELLBOX-PRIME (= CellBox-Primary Human Macrophages in Microgravity Environment) experiment investigated for the first time microgravity-associated long-term alterations in primary human macrophages, one of the most important effector cells of the immune system. The experiment was conducted in the U.S. National Laboratory on board of the International Space Station ISS using the NanoRacks laboratory and Biorack type I standard CELLBOX EUE type IV containers. Upload and download were performed with the SpaceX CRS-3 and the Dragon spaceship on April 18th, 2014 / May 18th, 2014. Surprisingly, primary human macrophages exhibited neither quantitative nor structural changes of the actin and vimentin cytoskeleton after 11 days in microgravity when compared to 1g controls. Neither CD18 or CD14 surface expression were altered in microgravity, however ICAM-1 expression was reduced. The analysis of 74 metabolites in the cell culture supernatant by GC-TOF-MS, revealed eight metabolites with significantly different quantities when compared to 1g controls. In particular, the significant increase of free fucose in the cell culture supernatant was associated with a significant decrease of cell surface-bound fucose. The reduced ICAM-1 expression and the loss of cell surface-bound fucose may contribute to functional impairments, e.g. the activation of T cells, migration and activation of the innate immune response. We assume that the surprisingly small and non

  11. Electronic cigarette liquid increases inflammation and virus infection in primary human airway epithelial cells.

    Directory of Open Access Journals (Sweden)

    Qun Wu

    Full Text Available The use of electronic cigarettes (e-cigarettes is rapidly increasing in the United States, especially among young people since e-cigarettes have been perceived as a safer alternative to conventional tobacco cigarettes. However, the scientific evidence regarding the human health effects of e-cigarettes on the lung is extremely limited. The major goal of our current study is to determine if e-cigarette use alters human young subject airway epithelial functions such as inflammatory response and innate immune defense against respiratory viral (i.e., human rhinovirus, HRV infection.We examined the effects of e-cigarette liquid (e-liquid on pro-inflammatory cytokine (e.g., IL-6 production, HRV infection and host defense molecules (e.g., short palate, lung, and nasal epithelium clone 1, SPLUNC1 in primary human airway epithelial cells from young healthy non-smokers. Additionally, we examined the role of SPLUNC1 in lung defense against HRV infection using a SPLUNC1 knockout mouse model. We found that nicotine-free e-liquid promoted IL-6 production and HRV infection. Addition of nicotine into e-liquid further amplified the effects of nicotine-free e-liquid. Moreover, SPLUNC1 deficiency in mice significantly increased lung HRV loads. E-liquid inhibited SPLUNC1 expression in primary human airway epithelial cells. These findings strongly suggest the deleterious health effects of e-cigarettes in the airways of young people. Our data will guide future studies to evaluate the impact of e-cigarettes on lung health in human populations, and help inform the public about potential health risks of e-cigarettes.

  12. Nek8, a NIMA family kinase member, is overexpressed in primary human breast tumors.

    Science.gov (United States)

    Bowers, Alex J; Boylan, John F

    2004-03-17

    The family of human Nek (NIMA Related Kinase) kinases currently contains 11 members. We have identified Nek8 as a new member of the Nek kinase family. For many of the Nek family members, primary tumor expression data and function have been limited. However, all of the Nek family proteins share considerable homology with the Never In Mitosis, gene A (NIMA) kinase from the filamentous fungus Aspergillus nidulans. NIMA, as well as its most closely related human ortholog, Nek2, are required for G(2)/M progression and promote centrosome maturation during mitosis. We isolated Nek8 from a primary human colon cDNA library, and found it to be highly homologous to murine Nek8. Recently, a previously named Nek8 sequence was renamed Nek9/Nercc1 in Genbank due to its lack of homology to murine Nek8 and its high homology to murine Nek9. Interestingly, in our study, phylogenetic analysis suggests that human Nek8 and Nek9 form a subfamily within the Nek family. Nek8 has high homology to the Nek family kinase domain as well as to a regulator of chromosome condensation domain (RCC1), which is also present in Nek9. The open reading frame of human Nek8 encodes a 692 amino-acid protein with a calculated molecular weight of 75 kDa. Nek8 is differently expressed between normal human breast tissue and breast tumors. Overexpression of a mutated kinase domain Nek8 in U2-0S cells led to a decrease in actin protein, and a small increase in the level of cdk1/cyclinB1. Our data demonstrate for the first time that Nek8 is a novel tumor associated gene, and shares considerable sequence homology with the Nek family of protein kinases and may be involved in G(2)/M progression.

  13. Resveratrol differentially regulates NAMPT and SIRT1 in Hepatocarcinoma cells and primary human hepatocytes.

    Directory of Open Access Journals (Sweden)

    Susanne Schuster

    Full Text Available Resveratrol is reported to possess chemotherapeutic properties in several cancers. In this study, we wanted to investigate the molecular mechanisms of resveratrol-induced cell cycle arrest and apoptosis as well as the impact of resveratrol on NAMPT and SIRT1 protein function and asked whether there are differences in hepatocarcinoma cells (HepG2, Hep3B cells and non-cancerous primary human hepatocytes. We found a lower basal NAMPT mRNA and protein expression in hepatocarcinoma cells compared to primary hepatocytes. In contrast, SIRT1 was significantly higher expressed in hepatocarcinoma cells than in primary hepatocytes. Resveratrol induced cell cycle arrest in the S- and G2/M- phase and apoptosis was mediated by activation of p53 and caspase-3 in HepG2 cells. In contrast to primary hepatocytes, resveratrol treated HepG2 cells showed a reduction of NAMPT enzymatic activity and increased p53 acetylation (K382. Resveratrol induced NAMPT release from HepG2 cells which was associated with increased NAMPT mRNA expression. This effect was absent in primary hepatocytes where resveratrol was shown to function as NAMPT and SIRT1 activator. SIRT1 inhibition by EX527 resembled resveratrol effects on HepG2 cells. Furthermore, a SIRT1 overexpression significantly decreased both p53 hyperacetylation and resveratrol-induced NAMPT release as well as S-phase arrest in HepG2 cells. We could show that NAMPT and SIRT1 are differentially regulated by resveratrol in hepatocarcinoma cells and primary hepatocytes and that resveratrol did not act as a SIRT1 activator in hepatocarcinoma cells.

  14. Resveratrol Differentially Regulates NAMPT and SIRT1 in Hepatocarcinoma Cells and Primary Human Hepatocytes

    Science.gov (United States)

    Schuster, Susanne; Penke, Melanie; Gorski, Theresa; Petzold-Quinque, Stefanie; Damm, Georg; Gebhardt, Rolf; Kiess, Wieland; Garten, Antje

    2014-01-01

    Resveratrol is reported to possess chemotherapeutic properties in several cancers. In this study, we wanted to investigate the molecular mechanisms of resveratrol-induced cell cycle arrest and apoptosis as well as the impact of resveratrol on NAMPT and SIRT1 protein function and asked whether there are differences in hepatocarcinoma cells (HepG2, Hep3B cells) and non-cancerous primary human hepatocytes. We found a lower basal NAMPT mRNA and protein expression in hepatocarcinoma cells compared to primary hepatocytes. In contrast, SIRT1 was significantly higher expressed in hepatocarcinoma cells than in primary hepatocytes. Resveratrol induced cell cycle arrest in the S- and G2/M- phase and apoptosis was mediated by activation of p53 and caspase-3 in HepG2 cells. In contrast to primary hepatocytes, resveratrol treated HepG2 cells showed a reduction of NAMPT enzymatic activity and increased p53 acetylation (K382). Resveratrol induced NAMPT release from HepG2 cells which was associated with increased NAMPT mRNA expression. This effect was absent in primary hepatocytes where resveratrol was shown to function as NAMPT and SIRT1 activator. SIRT1 inhibition by EX527 resembled resveratrol effects on HepG2 cells. Furthermore, a SIRT1 overexpression significantly decreased both p53 hyperacetylation and resveratrol-induced NAMPT release as well as S-phase arrest in HepG2 cells. We could show that NAMPT and SIRT1 are differentially regulated by resveratrol in hepatocarcinoma cells and primary hepatocytes and that resveratrol did not act as a SIRT1 activator in hepatocarcinoma cells. PMID:24603648

  15. Transcriptionally-active human papillomavirus is consistently retained in the distant metastases of primary oropharyngeal carcinomas.

    Science.gov (United States)

    Mehrad, Mitra; Zhao, Hongwei; Gao, Ge; Wang, Xiaowei; Lewis, James S

    2014-06-01

    High-risk human papillomavirus (HPV) is both causative and prognostic in the majority of oropharyngeal squamous cell carcinomas (OPSCCs). The aim of this study was to evaluate for transcriptionally-active HPV in matched primary OPSCCs and their distant metastases given the implications of HPV status for diagnosis and treatment. Twenty matched pairs of primary OPSCC and their distant metastases were retrieved from departmental files. Two study pathologists reviewed all cases to confirm the diagnoses and to evaluate histologic features. Real-time PCR (RT-PCR) for detection of E6/E7 mRNA for all major high-risk HPV types and p16 immunohistochemistry were performed. Distant metastases were to lung (70 %), bone (20 %), non-regional lymph nodes (5 %) and pericardium (5 %). Histologically, 15 primary tumors were nonkeratinizing, 3 nonkeratinizing with maturation, one basaloid, and one keratinizing. Seventeen (85 %) of the metastases had the same histologic type as the primary tumor. All 20 matched pairs were concordant for HPV status by RT-PCR and for p16 expression with 19 of 20 cases positive for high risk HPV and one negative. HPV types were concordant in all cases. These findings show that the distant metastases from HPV-related primary OPSCCs uniformly retain transcriptionally-active HPV and p16 overexpression. They also retain similar morphology. This argues that HPV status can be utilized to differentiate metastatic OPSCC from separate, new, primary squamous cell carcinomas in other organs, and that therapies specifically targeting HPV or virus-related proteins in patients with distant metastases can be utilized.

  16. Bortezomib sensitizes primary human esthesioneuroblastoma cells to TRAIL-induced apoptosis.

    Science.gov (United States)

    Koschny, Ronald; Holland, Heidrun; Sykora, Jaromir; Erdal, Hande; Krupp, Wolfgang; Bauer, Manfred; Bockmuehl, Ulrike; Ahnert, Peter; Meixensberger, Jürgen; Stremmel, Wolfgang; Walczak, Henning; Ganten, Tom M

    2010-04-01

    TNF-related apoptosis-inducing ligand (TRAIL), a promising novel anti-cancer cytokine of the TNF superfamily, and Bortezomib, the first-in-class clinically used proteasome inhibitor, alone or in combination have been shown to efficiently kill numerous tumor cell lines. However, data concerning primary human tumor cells are very rare. Using primary esthesioneuroblastoma cells we analyzed the anti-tumor potential and the mechanism employed by Bortezomib in combination with TRAIL for the treatment of this rare but aggressive tumor. Expression of components of the TRAIL pathway was analyzed in tumor specimens and isolated primary tumor cells at the protein level. Cells were treated with TRAIL, Bortezomib, and a combination thereof, and apoptosis induction was quantified. Clonogenicity assays were performed to elucidate the long-term effect of this treatment. Despite expressing all components of the TRAIL pathway, freshly isolated primary esthesioneuroblastoma cells were completely resistant to TRAIL-induced apoptosis. They could, however, be very efficiently sensitized by subtoxic doses of Bortezomib. The influence of Bortezomib on the TRAIL pathway was analyzed and showed upregulation of TRAIL death receptor expression, enhancement of the TRAIL death-inducing signaling complex (DISC), and downregulation of anti-apoptotic proteins of the TRAIL pathway. Of clinical relevance, TRAIL-resistant primary tumor cells could be repeatedly sensitized by Bortezomib, providing the basis for repeated clinical application schedules. This is the first report on the highly synergistic induction of apoptosis in primary esthesioneuroblastoma cells by Bortezomib and TRAIL. This combination, therefore, represents a promising novel therapeutic option for esthesioneuroblastoma.

  17. What Expands in an Expanding Universe?

    Directory of Open Access Journals (Sweden)

    JOSÉ A. DE FREITAS PACHECO

    2015-12-01

    Full Text Available ABSTRACT In the present investigation, the possible effects of the expansion of the Universe on systems bonded either by gravitational or electromagnetic forces, are reconsidered. It will be shown that the acceleration (positive or negative of the expanding background, is the determinant factor affecting planetary orbits and atomic sizes. In the presently accepted cosmology (ΛCDM all bonded systems are expanding at a decreasing rate that tends to be zero as the universe enters in a de Sitter phase. It is worth mentioning that the estimated expansion rates are rather small and they can be neglected for all practical purposes.

  18. What Expands in an Expanding Universe?

    Science.gov (United States)

    Pacheco, José A De Freitas

    2015-01-01

    In the present investigation, the possible effects of the expansion of the Universe on systems bonded either by gravitational or electromagnetic forces, are reconsidered. It will be shown that the acceleration (positive or negative) of the expanding background, is the determinant factor affecting planetary orbits and atomic sizes. In the presently accepted cosmology (ΛCDM) all bonded systems are expanding at a decreasing rate that tends to be zero as the universe enters in a de Sitter phase. It is worth mentioning that the estimated expansion rates are rather small and they can be neglected for all practical purposes.

  19. Knockdown of CDK2AP1 in primary human fibroblasts induces p53 dependent senescence.

    Directory of Open Access Journals (Sweden)

    Khaled N Alsayegh

    Full Text Available Cyclin Dependent Kinase-2 Associated Protein-1 (CDK2AP1 is known to be a tumor suppressor that plays a role in cell cycle regulation by sequestering monomeric CDK2, and targeting it for proteolysis. A reduction of CDK2AP1 expression is considered to be a negative prognostic indicator in patients with oral squamous cell carcinoma and also associated with increased invasion in human gastric cancer tissue. CDK2AP1 overexpression was shown to inhibit growth, reduce invasion and increase apoptosis in prostate cancer cell lines. In this study, we investigated the effect of CDK2AP1 downregulation in primary human dermal fibroblasts. Using a short-hairpin RNA to reduce its expression, we found that knockdown of CDK2AP1 in primary human fibroblasts resulted in reduced proliferation and in the induction of senescence associated beta-galactosidase activity. CDK2AP1 knockdown also resulted in a significant reduction in the percentage of cells in the S phase and an accumulation of cells in the G1 phase of the cell cycle. Immunocytochemical analysis also revealed that the CDK2AP1 knockdown significantly increased the percentage of cells that exhibited γ-H2AX foci, which could indicate presence of DNA damage. CDK2AP1 knockdown also resulted in increased mRNA levels of p53, p21, BAX and PUMA and p53 protein levels. In primary human fibroblasts in which p53 and CDK2AP1 were simultaneously downregulated, there was: (a no increase in senescence associated beta-galactosidase activity, (b decrease in the number of cells in the G1-phase and increase in number of cells in the S-phase of the cell cycle, and (c decrease in the mRNA levels of p21, BAX and PUMA when compared with CDK2AP1 knockdown only fibroblasts. Taken together, this suggests that the observed phenotype is p53 dependent. We also observed a prominent increase in the levels of ARF protein in the CDK2AP1 knockdown cells, which suggests a possible role of ARF in p53 stabilization following CDK2AP1

  20. Direct Phenotypical and Functional Dysregulation of Primary Human B Cells by Human Immunodeficiency Virus (HIV) Type 1 In Vitro

    Science.gov (United States)

    Perisé-Barrios, Ana Judith; Muñoz-Fernandez, María Ángeles; Pion, Marjorie

    2012-01-01

    Background Human immunodeficiency virus type 1 (HIV-1) induces a general dysregulation of immune system. Dysregulation of B cell compartment is generally thought to be induced by HIV-related immune activation and lymphopenia. However, a direct influence of HIV-1 particles on B cells was recently proposed as the third pathway of B cells dysregulation. Methods/Principal Findings We evaluated the direct and specific consequences of HIV-1 contact on activation, survival, proliferation and phenotype of primary B cells in vitro. Moreover, we examined expression of activation-induced cytidine deaminase (AID) mRNA that is responsible for class switch recombination (CSR) and somatic hypermutation (SHM). Here, we report that changes observed in cellular proliferation, phenotypes and activation of B cells could be caused by direct contact between HIV-1 particles and primary B cells in vitro. Finally, direct HIV-1-derived B cells activation led to the increase of AID mRNA expression and its subsequent CSR function was detected in vitro. Conclusion/Significance We showed that HIV-1 could directly induce primary B cells dysregulation triggering phenotypical and functional abilities of B cells in vitro that could explain in some extent early B-cell abnormalities in HIV disease. PMID:22768302

  1. Direct phenotypical and functional dysregulation of primary human B cells by human immunodeficiency virus (HIV) type 1 in vitro.

    Science.gov (United States)

    Perisé-Barrios, Ana Judith; Muñoz-Fernandez, María Ángeles; Pion, Marjorie

    2012-01-01

    Human immunodeficiency virus type 1 (HIV-1) induces a general dysregulation of immune system. Dysregulation of B cell compartment is generally thought to be induced by HIV-related immune activation and lymphopenia. However, a direct influence of HIV-1 particles on B cells was recently proposed as the third pathway of B cells dysregulation. We evaluated the direct and specific consequences of HIV-1 contact on activation, survival, proliferation and phenotype of primary B cells in vitro. Moreover, we examined expression of activation-induced cytidine deaminase (AID) mRNA that is responsible for class switch recombination (CSR) and somatic hypermutation (SHM). Here, we report that changes observed in cellular proliferation, phenotypes and activation of B cells could be caused by direct contact between HIV-1 particles and primary B cells in vitro. Finally, direct HIV-1-derived B cells activation led to the increase of AID mRNA expression and its subsequent CSR function was detected in vitro. We showed that HIV-1 could directly induce primary B cells dysregulation triggering phenotypical and functional abilities of B cells in vitro that could explain in some extent early B-cell abnormalities in HIV disease.

  2. Direct phenotypical and functional dysregulation of primary human B cells by human immunodeficiency virus (HIV type 1 in vitro.

    Directory of Open Access Journals (Sweden)

    Ana Judith Perisé-Barrios

    Full Text Available BACKGROUND: Human immunodeficiency virus type 1 (HIV-1 induces a general dysregulation of immune system. Dysregulation of B cell compartment is generally thought to be induced by HIV-related immune activation and lymphopenia. However, a direct influence of HIV-1 particles on B cells was recently proposed as the third pathway of B cells dysregulation. METHODS/PRINCIPAL FINDINGS: We evaluated the direct and specific consequences of HIV-1 contact on activation, survival, proliferation and phenotype of primary B cells in vitro. Moreover, we examined expression of activation-induced cytidine deaminase (AID mRNA that is responsible for class switch recombination (CSR and somatic hypermutation (SHM. Here, we report that changes observed in cellular proliferation, phenotypes and activation of B cells could be caused by direct contact between HIV-1 particles and primary B cells in vitro. Finally, direct HIV-1-derived B cells activation led to the increase of AID mRNA expression and its subsequent CSR function was detected in vitro. CONCLUSION/SIGNIFICANCE: We showed that HIV-1 could directly induce primary B cells dysregulation triggering phenotypical and functional abilities of B cells in vitro that could explain in some extent early B-cell abnormalities in HIV disease.

  3. Primary Cilia Negatively Regulate Melanogenesis in Melanocytes and Pigmentation in a Human Skin Model.

    Directory of Open Access Journals (Sweden)

    Hyunjung Choi

    Full Text Available The primary cilium is an organelle protruding from the cell body that senses external stimuli including chemical, mechanical, light, osmotic, fluid flow, and gravitational signals. Skin is always exposed to the external environment and responds to external stimuli. Therefore, it is possible that primary cilia have an important role in skin. Ciliogenesis was reported to be involved in developmental processes in skin, such as keratinocyte differentiation and hair formation. However, the relation between skin pigmentation and primary cilia is largely unknown. Here, we observed that increased melanogenesis in melanocytes treated with a melanogenic inducer was inhibited by a ciliogenesis inducer, cytochalasin D, and serum-free culture. However, these inhibitory effects disappeared in GLI2 knockdown cells. In addition, activation of sonic hedgehog (SHH-smoothened (Smo signaling pathway by a Smo agonist, SAG inhibited melanin synthesis in melanocytes and pigmentation in a human skin model. On the contrary, an inhibitor of primary cilium formation, ciliobrevin A1, activated melanogenesis in melanocytes. These results suggest that skin pigmentation may be regulated partly by the induction of ciliogenesis through Smo-GLI2 signaling.

  4. Adherence and viability of primary human keratinocytes and primary human dermal fibroblasts on acrylonitrile-based copolymers with different concentrations of positively charged functional groups.

    Science.gov (United States)

    Trescher, Karoline; Scharnagl, Nico; Kratz, Karl; Roch, Toralf; Lendlein, Andreas; Jung, Friedrich

    2012-01-01

    As shown in several studies, various properties of biomaterials such as stiffness, surface roughness, chemical composition or the amount of functional groups at the surface can influence adhesion, viability, proliferation and functionalities of cells. The aim of this work was to explore whether a cell-selective effect could be achieved for acrylonitrile-based copolymers containing different contents of positively charged functional groups, which were introduced by incorporation of methacrylic acid-2-aminoethylester hydrochloride (AEMA) units. The p(AN-co-AEMA) copolymers were synthesized by suspension polymerization in water and processed into disk shaped test specimen via a sintering process to ensure the absence of organic solvents in the copolymers. Copolymers with an AEMA content of 1.4, 1.6, and 4.4 mol-% were investigated according to their cell-selective capacity, which should support the adhesion, viability and proliferation of keratinocytes, while the adherence of fibroblasts should rather be disabled. The test samples were seeded with primary human keratinocytes and primary human dermal fibroblasts in mono- as well as in co-cultures. Tissue culture plate polystyrene (TCP) was used to control the physiologic growth of the cells. Density and viability of attached and non-adherent cells were analyzed by live/dead staining, lactate dehydrogenase (LDH) assay and flow cytometry with DAPI staining. For the assured discrimination of adherent cell types in coculture a keratin/vimentin-staining was performed. On copolymers with 4.4 mol-% AEMA adherent keratinocytes in monoculture and cocultured keratinocytes and fibroblasts showed a higher viability, a lower impairment of cell membranes and higher densities of viable cells compared to both other copolymers. For adherent fibroblasts these parameters did not differ between the copolymers and an increasing ratio of keratinocytes to fibroblasts in cocultures were found with increasing AEMA content. The results showed

  5. Cloning the human betaretrovirus proviral genome from patients with primary biliary cirrhosis.

    Science.gov (United States)

    Xu, Lizhe; Sakalian, Michael; Shen, Zhiwei; Loss, George; Neuberger, James; Mason, Andrew

    2004-01-01

    Patients with primary biliary cirrhosis (PBC) have both serologic and tissue evidence of infection. A recently identified human betaretrovirus was originally cloned from the biliary epithelium cDNA library of a patient with PBC. By conducting a BLASTN search, the initial partial pol gene fragment was found to have 95% to 97% nucleotide homology with mouse mammary tumor virus (MMTV) and with retrovirus sequences derived from human breast cancer samples. Using an anti-p27(CA) MMTV antibody, viral proteins were detected in the perihepatic lymph nodes but not in liver tissue samples from patients with PBC, suggesting a higher viral burden in lymphoid tissue. Therefore, in the current study, we used lymph node DNA to clone the proviral genome of the human betaretrovirus from two patients with PBC using a polymerase chain reaction (PCR) walking methodology with conserved primers complementary to MMTV. The human betaretrovirus genome contains five potential open reading frames (ORF) for Gag, protease (Pro), polymerase (Pol), envelope (Env), and superantigen (Sag) proteins that are collinear with their counterparts in MMTV. Alignment studies performed with characterized MMTV and human breast cancer betaretrovirus amino acid sequences revealed a 93% to 99% identity with the p27 capsid proteins, a 93% to 97% identity with the betaretrovirus envelope proteins, and a 76% to 85% identity with the more variable superantigen proteins. Phylogenetic analysis of known betaretrovirus superantigen proteins showed that the human and murine sequences did not cluster as two distinct species. In conclusion, human betaretrovirus nucleic acid sequences have been cloned from patients with PBC. They share marked homology with MMTV and human breast cancer-derived retrovirus sequences.

  6. High-throughput gene expression profiling of memory differentiation in primary human T cells

    Directory of Open Access Journals (Sweden)

    Russell Kate

    2008-08-01

    Full Text Available Abstract Background The differentiation of naive T and B cells into memory lymphocytes is essential for immunity to pathogens. Therapeutic manipulation of this cellular differentiation program could improve vaccine efficacy and the in vitro expansion of memory cells. However, chemical screens to identify compounds that induce memory differentiation have been limited by 1 the lack of reporter-gene or functional assays that can distinguish naive and memory-phenotype T cells at high throughput and 2 a suitable cell-line representative of naive T cells. Results Here, we describe a method for gene-expression based screening that allows primary naive and memory-phenotype lymphocytes to be discriminated based on complex genes signatures corresponding to these differentiation states. We used ligation-mediated amplification and a fluorescent, bead-based detection system to quantify simultaneously 55 transcripts representing naive and memory-phenotype signatures in purified populations of human T cells. The use of a multi-gene panel allowed better resolution than any constituent single gene. The method was precise, correlated well with Affymetrix microarray data, and could be easily scaled up for high-throughput. Conclusion This method provides a generic solution for high-throughput differentiation screens in primary human T cells where no single-gene or functional assay is available. This screening platform will allow the identification of small molecules, genes or soluble factors that direct memory differentiation in naive human lymphocytes.

  7. Electrotransfection and lipofection show comparable efficiency for in vitro gene delivery of primary human myoblasts.

    Science.gov (United States)

    Mars, Tomaz; Strazisar, Marusa; Mis, Katarina; Kotnik, Nejc; Pegan, Katarina; Lojk, Jasna; Grubic, Zoran; Pavlin, Mojca

    2015-04-01

    Transfection of primary human myoblasts offers the possibility to study mechanisms that are important for muscle regeneration and gene therapy of muscle disease. Cultured human myoblasts were selected here because muscle cells still proliferate at this developmental stage, which might have several advantages in gene therapy. Gene therapy is one of the most sought-after tools in modern medicine. Its progress is, however, limited due to the lack of suitable gene transfer techniques. To obtain better insight into the transfection potential of the presently used techniques, two non-viral transfection methods--lipofection and electroporation--were compared. The parameters that can influence transfection efficiency and cell viability were systematically approached and compared. Cultured myoblasts were transfected with the pEGFP-N1 plasmid either using Lipofectamine 2000 or with electroporation. Various combinations for the preparation of the lipoplexes and the electroporation media, and for the pulsing protocols, were tested and compared. Transfection efficiency and cell viability were inversely proportional for both approaches. The appropriate ratio of Lipofectamine and plasmid DNA provides optimal conditions for lipofection, while for electroporation, RPMI medium and a pulsing protocol using eight pulses of 2 ms at E = 0.8 kV/cm proved to be the optimal combination. The transfection efficiencies for the optimal lipofection and optimal electrotransfection protocols were similar (32 vs. 32.5%, respectively). Both of these methods are effective for transfection of primary human myoblasts; however, electroporation might be advantageous for in vivo application to skeletal muscle.

  8. The organisational and human resource challenges facing primary care trusts: protocol of a multiple case study

    Directory of Open Access Journals (Sweden)

    Tim Scott J

    2001-11-01

    Full Text Available Abstract Background The study is designed to assess the organisational and human resource challenges faced by Primary Care Trusts (PCTs. Its objectives are to: specify the organisational and human resources challenges faced by PCTs in fulfilling the roles envisaged in government and local policy; examine how PCTs are addressing these challenges, in particular, to describe the organisational forms they have adopted, and the OD/HR strategies and initiatives they have planned or in place; assess how effective these structures, strategies and initiatives have been in enabling the PCTs to meet the organisational and human resources challenges they face; identify the factors, both internal to the PCT and in the wider health community, which have contributed to the success or failure of different structures, strategies and initiatives. Methods The study will be undertaken in three stages. In Stage 1 the key literature on public sector and NHS organisational development and human resources management will be reviewed, and discussions will be held with key researchers and policy makers working in this area. Stage 2 will focus on detailed case studies in six PCTs designed to examine the organisational and human resources challenges they face. Data will be collected using semi-structured interviews, group discussion, site visits, observation of key meetings and examination of local documentation. The findings from the case study PCTs will be cross checked with a Reference Group of up to 20 other PCG/Ts, and key officers working in organisational development or primary care at local, regional and national level. In Stage 3 analysis of findings from the preparatory work, the case studies and the feedback from the Reference Group will be used to identify practical lessons for PCTs, key messages for policy makers, and contributions to further theoretical development.

  9. Characterization of primary cilia and Hedgehog signaling during development of the human pancreas and in human pancreatic duct cancer cell lines

    DEFF Research Database (Denmark)

    Nielsen, Sonja K; Møllgård, Kjeld; Clement, Christian A

    2008-01-01

    Hedgehog (Hh) signaling controls pancreatic development and homeostasis; aberrant Hh signaling is associated with several pancreatic diseases. Here we investigated the link between Hh signaling and primary cilia in the human developing pancreatic ducts and in cultures of human pancreatic duct...... adenocarcinoma cell lines, PANC-1 and CFPAC-1. We show that the onset of Hh signaling from human embryogenesis to fetal development is associated with accumulation of Hh signaling components Smo and Gli2 in duct primary cilia and a reduction of Gli3 in the duct epithelium. Smo, Ptc, and Gli2 localized to primary...

  10. Cancer-associated loss of TARSH gene expression in human primary lung cancer.

    Science.gov (United States)

    Terauchi, Kunihiko; Shimada, Junichi; Uekawa, Natsuko; Yaoi, Takeshi; Maruyama, Mitsuo; Fushiki, Shinji

    2006-01-01

    We have previously identified mouse Tarsh as one of the cellular senescence-related genes and showed the loss of expression of TARSH mRNA in four human lung cancer cell lines. TARSH is a presumptive signal transduction molecule interacting with NESH, which is implicated to have some roles in lung cancer metastasis. The amplification of complete ORF-encoding TARSH cDNA was done with reverse transcription-PCR. Northern blotting was carried out using TARSH cDNA probes. To clarify the relationship between TARSH and lung cancer, we quantified TARSH mRNA expression in 15 human lung cancer cell lines and 32 primary non-small cell lung cancers. We first determined the complete ORF-encoding cDNA sequence which is expressed in the human lung. On the Northern hybridization analysis, TARSH was strongly expressed in the human lung. The expression of TARSH mRNA is remarkably downregulated in all the lung cancer cell lines examined. Furthermore, TARSH expression was significantly low in all of the tumor specimens when compared to the expression in corresponding non-neoplastic lung tissue specimens. The cancer-associated transcriptional inactivation of TARSH suggests that TARSH could be used as a biomarker for lung cancer development as well as a molecular adjunct for lung carcinogenesis in human.

  11. Computational study of human head response to primary blast waves of five levels from three directions.

    Directory of Open Access Journals (Sweden)

    Chenzhi Wang

    Full Text Available Human exposure to blast waves without any fragment impacts can still result in primary blast-induced traumatic brain injury (bTBI. To investigate the mechanical response of human brain to primary blast waves and to identify the injury mechanisms of bTBI, a three-dimensional finite element head model consisting of the scalp, skull, cerebrospinal fluid, nasal cavity, and brain was developed from the imaging data set of a human female. The finite element head model was partially validated and was subjected to the blast waves of five blast intensities from the anterior, right lateral, and posterior directions at a stand-off distance of one meter from the detonation center. Simulation results show that the blast wave directly transmits into the head and causes a pressure wave propagating through the brain tissue. Intracranial pressure (ICP is predicted to have the highest magnitude from a posterior blast wave in comparison with a blast wave from any of the other two directions with same blast intensity. The brain model predicts higher positive pressure at the site proximal to blast wave than that at the distal site. The intracranial pressure wave invariably travels into the posterior fossa and vertebral column, causing high pressures in these regions. The severities of cerebral contusions at different cerebral locations are estimated using an ICP based injury criterion. Von Mises stress prevails in the cortex with a much higher magnitude than in the internal parenchyma. According to an axonal injury criterion based on von Mises stress, axonal injury is not predicted to be a cause of primary brain injury from blasts.

  12. Prognostic Implication of Human Papillomavirus Types and Species in Cervical Cancer Patients Undergoing Primary Treatment

    OpenAIRE

    Yat Ming Lau; Tak Hong Cheung; Winnie Yeo; Frankie Mo; Mei Yung Yu; Kun Min Lee; Ho, Wendy C. S.; Yeung, Apple C. M.; Priscilla T. Y. Law; Chan, Paul K. S.

    2015-01-01

    High-risk human papillomavirus (HPV) types are associated with cervical cancer. It is well established that individual HPV types vary in oncogenicity, but current data on their prognostic implication remain controversial. We examined the association between HPV types/species and the survival of 236 Chinese women aged 26-87 (mean 54.4) years after receiving primary treatment for cervical cancer. Overall, 45.8% were of FIGO stage I, 41.9% stage II, and 12.3% stage III. The four most prevalent t...

  13. Transcriptome analysis reveals a classical interferon signature induced by IFNλ4 in human primary cells

    DEFF Research Database (Denmark)

    Lauber, Chris; Vieyres, Gabrielle; Terczynska-Dyla, Ewa

    2015-01-01

    4 could be a tissue-specific regulation of an unknown subset of genes. To address both tissue and subtype specificity in the interferon response, we treated primary human hepatocytes and airway epithelial cells with IFNα, IFNλ3 or IFNλ4 and assessed interferon mediated gene regulation using...... genes being regulated in hepatocytes as well as airway epithelial cells. Thus we provide an in-depth analysis of the liver interferon response seen over an array of interferon subtypes and compare it to the response in the lung epithelium....

  14. Dormancy and activation of human oocytes from primordial and primary follicles: molecular clues to oocyte regulation

    DEFF Research Database (Denmark)

    Ernst, Emil Hagen; Grøndahl, Marie Louise; Grund, Simon

    2017-01-01

    as putative mediators of oocyte dormancy and activation. WHAT IS KNOWN ALREADY: Cellular signaling pathways including PI3K/AKT and AKT/mTOR as well as TGF-β and IGF signaling are known to regulate the primordial-to-primary transition in mammalian follicle development. STUDY DESIGN, SIZE, DURATION: We...... isolated by Laser Capture Microdissection, and submitted to the HiSeq Illumina platform. Data mapping, quality control, filtering and expression analysis were performed using Tophat (2.0.4), Cufflinks (2.0.2), BWA (0.6.2) and software R. Modeling of complex biological systems was performed using the IPA......® software. Finally, qPCR and immunohistochemistry were employed to explore expression and localization of selected genes and products in human ovarian tissue. MAIN RESULTS AND THE ROLE OF CHANCE: We found 223 and 268 genes down-regulated and up-regulated, respectively, in the oocytes during the human...

  15. Rabbit muscle creatine phosphokinase. CDNA cloning, primary structure and detection of human homologues.

    Science.gov (United States)

    Putney, S; Herlihy, W; Royal, N; Pang, H; Aposhian, H V; Pickering, L; Belagaje, R; Biemann, K; Page, D; Kuby, S

    1984-12-10

    A cDNA library was constructed from rabbit muscle poly(A) RNA. Limited amino acid sequence information was obtained on rabbit muscle creatine phosphokinase and this was the basis for design and synthesis of two oligonucleotide probes complementary to a creatine kinase cDNA sequence which encodes a pentapeptide. Colony hybridizations with the probes and subsequent steps led to isolation of two clones, whose cDNA segments partially overlap and which together encode the entire protein. The primary structure was established from the sequence of two cDNA clones and from independently determined sequences of scattered portions of the polypeptide. The reactive cysteine has been located to position 282 within the 380 amino acid polypeptide. The rabbit cDNA hybridizes to digests of human chromosomal DNA. This reveals a restriction fragment length polymorphism associated with the human homologue(s) which hybridizes to the rabbit cDNA.

  16. Injectable scaffold materials differ in their cell instructive effects on primary human myoblasts

    DEFF Research Database (Denmark)

    Hejbøl, Eva Kildall; Sellathurai, Jeeva; Nair, Prabha Damodaran

    2017-01-01

    a minimally invasive technique. In this study, we examined in vitro the cell instructive effects of three types of injectable scaffolds, fibrin, alginate, and poly(lactic-co-glycolic acid)-based microparticles on primary human myoblasts. The myoblast morphology and progression in the myogenic program differed......, depending on the type of scaffold material. In alginate gel, the cells obtained a round morphology, they ceased to proliferate, and entered quiescence. In the fibrin gels, differentiation was promoted, and myotubes were observed within a few days in culture, while poly(lactic-co-glycolic acid......)-based microparticles supported prolonged proliferation. Myoblasts released from the alginate and fibrin gels were studied, and cells released from these scaffolds had retained the ability to proliferate and differentiate. Thus, the study shows that human myogenic cells combined with injectable scaffold materials...

  17. Human primary adipocytes exhibit immune cell function: adipocytes prime inflammation independent of macrophages.

    Directory of Open Access Journals (Sweden)

    Kees Meijer

    Full Text Available BACKGROUND: Obesity promotes inflammation in adipose tissue (AT and this is implicated in pathophysiological complications such as insulin resistance, type 2 diabetes and cardiovascular disease. Although based on the classical hypothesis, necrotic AT adipocytes (ATA in obese state activate AT macrophages (ATM that then lead to a sustained chronic inflammation in AT, the link between human adipocytes and the source of inflammation in AT has not been in-depth and systematically studied. So we decided as a new hypothesis to investigate human primary adipocytes alone to see whether they are able to prime inflammation in AT. METHODS AND RESULTS: Using mRNA expression, human preadipocytes and adipocytes express the cytokines/chemokines and their receptors, MHC II molecule genes and 14 acute phase reactants including C-reactive protein. Using multiplex ELISA revealed the expression of 50 cytokine/chemokine proteins by human adipocytes. Upon lipopolysaccharide stimulation, most of these adipocyte-associated cytokines/chemokines and immune cell modulating receptors were up-regulated and a few down-regulated such as (ICAM-1, VCAM-1, MCP-1, IP-10, IL-6, IL-8, TNF-α and TNF-β highly up-regulated and IL-2, IL-7, IL-10, IL-13 and VEGF down-regulated. In migration assay, human adipocyte-derived chemokines attracted significantly more CD4+ T cells than controls and the number of migrated CD4+ cells was doubled after treating the adipocytes with LPS. Neutralizing MCP-1 effect produced by adipocytes reduced CD4+ migration by approximately 30%. CONCLUSION: Human adipocytes express many cytokines/chemokines that are biologically functional. They are able to induce inflammation and activate CD4+ cells independent of macrophages. This suggests that the primary event in the sequence leading to chronic inflammation in AT is metabolic dysfunction in adipocytes, followed by production of immunological mediators by these adipocytes, which is then exacerbated by

  18. Inhibition of aromatase activity by methyl sulfonyl PCB metabolites in primary culture of human mammary fibroblasts

    Energy Technology Data Exchange (ETDEWEB)

    Berg, M. van den; Heneweer, M.; Geest, M. de; Sanderson, T. [Inst. for Risk Assessment Sciences and Utrecht Univ. (Netherlands); Jong, P. de [St. Antonius Hospital, Nieuwegein (Netherlands); Bergman, A. [Stockholm Univ., Stockholm (Sweden)

    2004-09-15

    Methyl sulfonyl PCB metabolites (MeSO2-PCBs) are persistent contaminants and are ubiquitously present in humans and the environment. Lipophilicity of MeSO2- PCB metabolites is similar to the parent compounds and they have been detected in human milk, adipose, liver and lung tissue. 4- MeSO2-PCB-149 is the most abundant PCB metabolite in human adipose tissue and milk at a level of 1.5 ng/g lipids. Human blood concentration of 4-MeSO2-PCB-149 is approximately 0.03 nM. 3- MeSO2-PCB-101 is the predominant PCB metabolite in muscle and blubber in wildlife, such as otter, mink and grey seal. In the environment, they have been linked to chronic and reproductive toxicity in exposed mink. Additionaly, some MeSO{sub 2}-PCBs have been shown to be glucocorticoid receptor (GR) antagonists. Since approximately 60% of all breast tumors are estrogen responsive, exposure to compounds that are able to alter estrogen synthesis through interference with the aromatase enzyme, can lead to changes in estrogen levels and possibly to accelerated or inhibit breast tumor growth. Therefore, it is important to identify exogenous compounds that can alter aromatase activity in addition to those compounds which have direct interaction with the estrogen receptor (ER). Aromatase (CYP19) comprises the ubiquitous flavoprotein, NADPH-cytochrome P450 reductase, and a unique cytochrome P450 that is exclusively expressed in estrogen producing cells. Previous studies have revealed that expression of the aromatase gene is regulated in a species- and tissue specific manner. In healthy breast tissue, the predominantly active aromatase promoter region I.4 is regulated by glucocorticoids and class I cytokines. Therefore, it is important to investigate possible aromatase inhibiting properties of MeSO{sub 2}-PCBs (as anti glucocorticoids?) in relevant human tissues. We used primary human mammary fibroblasts because of their role in breast cancer development. We compared the results in primary fibroblasts with

  19. The Artful Universe Expanded

    Science.gov (United States)

    Barrow, John D.

    2005-07-01

    Our love of art, writes John Barrow, is the end product of millions of years of evolution. How we react to a beautiful painting or symphony draws upon instincts laid down long before humans existed. Now, in this enhanced edition of the highly popular The Artful Universe , Barrow further explores the close ties between our aesthetic appreciation and the basic nature of the Universe. Barrow argues that the laws of the Universe have imprinted themselves upon our thoughts and actions in subtle and unexpected ways. Why do we like certain types of art or music? What games and puzzles do we find challenging? Why do so many myths and legends have common elements? In this eclectic and entertaining survey, Barrow answers these questions and more as he explains how the landscape of the Universe has influenced the development of philosophy and mythology, and how millions of years of evolutionary history have fashioned our attraction to certain patterns of sound and color. Barrow casts the story of human creativity and thought in a fascinating light, considering such diverse topics as our instinct for language, the origins and uses of color in nature, why we divide time into intervals as we do, the sources of our appreciation of landscape painting, and whether computer-generated fractal art is really art. Drawing on a wide variety of examples, from the theological questions raised by St. Augustine and C.S. Lewis to the relationship between the pure math of Pythagoras and the music of the Beatles, The Artful Universe Expanded covers new ground and enters a wide-ranging debate about the meaning and significance of the links between art and science.

  20. Advances in the quantification of mitochondrial function in primary human immune cells through extracellular flux analysis.

    Science.gov (United States)

    Nicholas, Dequina; Proctor, Elizabeth A; Raval, Forum M; Ip, Blanche C; Habib, Chloe; Ritou, Eleni; Grammatopoulos, Tom N; Steenkamp, Devin; Dooms, Hans; Apovian, Caroline M; Lauffenburger, Douglas A; Nikolajczyk, Barbara S

    2017-01-01

    Numerous studies show that mitochondrial energy generation determines the effectiveness of immune responses. Furthermore, changes in mitochondrial function may regulate lymphocyte function in inflammatory diseases like type 2 diabetes. Analysis of lymphocyte mitochondrial function has been facilitated by introduction of 96-well format extracellular flux (XF96) analyzers, but the technology remains imperfect for analysis of human lymphocytes. Limitations in XF technology include the lack of practical protocols for analysis of archived human cells, and inadequate data analysis tools that require manual quality checks. Current analysis tools for XF outcomes are also unable to automatically assess data quality and delete untenable data from the relatively high number of biological replicates needed to power complex human cell studies. The objectives of work presented herein are to test the impact of common cellular manipulations on XF outcomes, and to develop and validate a new automated tool that objectively analyzes a virtually unlimited number of samples to quantitate mitochondrial function in immune cells. We present significant improvements on previous XF analyses of primary human cells that will be absolutely essential to test the prediction that changes in immune cell mitochondrial function and fuel sources support immune dysfunction in chronic inflammatory diseases like type 2 diabetes.

  1. Teaching about Human Relationships Education for Primary School Student-Teachers Using an Interactive CD-ROM

    Science.gov (United States)

    Goldman, Juliette D. G.; Torrisi-Steele, Geraldine

    2009-01-01

    Human Relationships Education is a very important part of primary school student-teacher education. All primary school children need sound guidance and enhanced knowledge about puberty, growing up successfully, and feeling competent and confident in themselves. An interactive multimedia CD-ROM was designed and developed for some Australian…

  2. Layouts of Expander Graphs

    OpenAIRE

    Dujmović, Vida; Sidiropoulos, Anastasios; Wood, David R.

    2015-01-01

    Bourgain and Yehudayoff recently constructed $O(1)$-monotone bipartite expanders. By combining this result with a generalisation of the unraveling method of Kannan, we construct 3-monotone bipartite expanders, which is best possible. We then show that the same graphs admit 3-page book embeddings, 2-queue layouts, 4-track layouts, and have simple thickness 2. All these results are best possible.

  3. Insulin promotes glycogen storage and cell proliferation in primary human astrocytes.

    Directory of Open Access Journals (Sweden)

    Martin Heni

    Full Text Available INTRODUCTION: In the human brain, there are at least as many astrocytes as neurons. Astrocytes are known to modulate neuronal function in several ways. Thus, they may also contribute to cerebral insulin actions. Therefore, we examined whether primary human astrocytes are insulin-responsive and whether their metabolic functions are affected by the hormone. METHODS: Commercially available Normal Human Astrocytes were grown in the recommended medium. Major players in the insulin signaling pathway were detected by real-time RT-PCR and Western blotting. Phosphorylation events were detected by phospho-specific antibodies. Glucose uptake and glycogen synthesis were assessed using radio-labeled glucose. Glycogen content was assessed by histochemistry. Lactate levels were measured enzymatically. Cell proliferation was assessed by WST-1 assay. RESULTS: We detected expression of key proteins for insulin signaling, such as insulin receptor β-subunit, insulin receptor substrat-1, Akt/protein kinase B and glycogen synthase kinase 3, in human astrocytes. Akt was phosphorylated and PI-3 kinase activity increased following insulin stimulation in a dose-dependent manner. Neither increased glucose uptake nor lactate secretion after insulin stimulation could be evidenced in this cell type. However, we found increased insulin-dependent glucose incorporation into glycogen. Furthermore, cell numbers increased dose-dependently upon insulin treatment. DISCUSSION: This study demonstrated that human astrocytes are insulin-responsive at the molecular level. We identified glycogen synthesis and cell proliferation as biological responses of insulin signaling in these brain cells. Hence, this cell type may contribute to the effects of insulin in the human brain.

  4. Heavy metals in human primary teeth: some factors influencing the metal concentrations.

    Science.gov (United States)

    Tvinnereim, H M; Eide, R; Riise, T

    2000-06-08

    Human primary teeth have been used as indicators of heavy metal exposure for several decades, but the knowledge about the influence of factors such as tooth type and the presence of caries and roots on metal concentrations is limited. Samples of tooth powder from more than 1200 Norwegian primary teeth without fillings have been analyzed for lead, zinc and cadmium content, and 554 of them for mercury. The material represents all groups of tooth types (incisors, canines and molars), carious and non-carious teeth, and teeth with and without roots. Here we investigate how tooth group and the presence of caries and roots are related to metal concentrations in the teeth. We find that carious teeth have higher metal concentrations than non-carious teeth; the difference was statistically significant for lead, mercury and zinc. Teeth with roots have higher lead and zinc concentrations than teeth without roots. We find differences in metal concentrations between the tooth groups for lead, mercury and zinc. Significant, positive correlations are found between lead and the three other metals and between mercury and zinc. We conclude that metal concentrations in primary teeth are affected by the presence of caries and roots and by tooth group.

  5. Effect of cell phone-like electromagnetic radiation on primary human thyroid cells.

    Science.gov (United States)

    Silva, Veronica; Hilly, Ohad; Strenov, Yulia; Tzabari, Cochava; Hauptman, Yirmi; Feinmesser, Raphael

    2016-01-01

    To evaluate the potential carcinogenic effects of radiofrequency energy (RFE) emitted by cell phones on human thyroid primary cells. Primary thyroid cell culture was prepared from normal thyroid tissue obtained from patients who underwent surgery at our department. Subconfluent thyroid cells were irradiated under different conditions inside a cell incubator using a device that simulates cell phone-RFE. Proliferation of control and irradiated cells was assessed by the immunohistochemical staining of antigen Kiel clone-67 (Ki-67) and tumor suppressor p53 (p53) expression. DNA ploidy and the stress biomarkers heat shock protein 70 (HSP70) and reactive oxygen species (ROS) was evaluated by fluorescence-activated cell sorting (FACS). Our cells highly expressed thyroglobulin (Tg) and sodium-iodide symporter (NIS) confirming the origin of the tissue. None of the irradiation conditions evaluated here had an effect neither on the proliferation marker Ki-67 nor on p53 expression. DNA ploidy was also not affected by RFE, as well as the expression of the biomarkers HSP70 and ROS. Our conditions of RFE exposure seem to have no potential carcinogenic effect on human thyroid cells. Moreover, common biomarkers usually associated to environmental stress also remained unchanged. We failed to find an association between cell phone-RFE and thyroid cancer. Additional studies are recommended.

  6. Primary pulmonary choriocarcinoma after human chorionic gonadotropin normalization following hydatidiform mole

    DEFF Research Database (Denmark)

    Maestá, Izildinha; Leite, Fábio Vicente; Michelin, Odair Carlito

    2010-01-01

    BACKGROUND: Primary pulmonary choriocarcinoma (PPC) is rare and frequently leads to death. CASES: Two young patients presented with previous molar pregnancy and spontaneous serum human chorionic gonadotropin (hCG) normalization. Patient 1 was referred to our center after partial response to chemo......BACKGROUND: Primary pulmonary choriocarcinoma (PPC) is rare and frequently leads to death. CASES: Two young patients presented with previous molar pregnancy and spontaneous serum human chorionic gonadotropin (hCG) normalization. Patient 1 was referred to our center after partial response...... to chemotherapy. Pulmonary lobectomy was performed, and hCG rapidly declined. During further chemotherapy, liver metastasis was detected by positron emission tomography. Right hepatectomy was performed, and hCG declined for 28 days, but increased again despite chemotherapy. This patient died from hepatic failure...... 3 years after diagnosis. Patient 2 presented with persistently high hCG, though the affected organ was not identified. Chemotherapy was unsuccessful. Patient reevaluation showed an isolated pulmonary mass. Pulmonary lobectomy was performed; 2 weeks later, hCG was normal and consolidation with 2...

  7. Knowledge about Human Papilloma Virus (HPV and HPV Vaccine at Reproductive Age in Primary Care

    Directory of Open Access Journals (Sweden)

    Ozde Onder

    2015-09-01

    Full Text Available Purpose: The aim of this study was to identify the knowledge and awareness about human papilloma virus (HPV and human papilloma virus (HPV vaccine of women in reproductive age. Material and Methods: The study covered 294 women aged between 15 and 49. A questionnaire was prepared by the researchers based on the literature review. Results: The mean age of the participants was 30.5+/-8.9 years. Only 24.5% had heard about HPV infection and 28.2% had heard HPV vaccine. Of the participants, 188 (63.9% got zero point from the knowledge questions. Conclusion: This study indicates that the women who apply primary care units have low knowledge levels; it is apperent that personal and social education is needed. Paying importance to patient education on HPV and cervical cancer in primary care health politics will increase knowledge and awareness for HPV infection and HPV vaccination. [Cukurova Med J 2015; 40(3.000: 517-524

  8. A method for establishing human primary gastric epithelial cell culture from fresh surgical gastric tissues.

    Science.gov (United States)

    Aziz, Faisal; Yang, Xuesong; Wen, Qingping; Yan, Qiu

    2015-08-01

    At present, biopsy specimens, cancer cell lines and tissues obtained by gastric surgery are used in the study and analysis of gastric cancer, including the molecular mechanisms and proteomics. However, fibroblasts and other tissue components may interfere with these techniques. Therefore, the present study aimed to develop a procedure for the isolation of viable human gastric epithelial cells from gastric surgical tissues. A method was developed to culture human gastric epithelial cells using fresh, surgically excised tissues and was evaluated using immunocytochemistry, periodic acid-Schiff (PAS) staining and cell viability assays. Low cell growth was observed surrounding the gastric tissue on the seventh day of tissue explant culture. Cell growth subsequently increased, and at 12 days post-explant a high number of pure epithelial cells were detected. The gastric cancer cells exhibited rapid growth with a doubling time of 13-52 h, as compared to normal cells, which had a doubling time of 20-53 h. Immunocytochemical analyses of primary gastric cells revealed positive staining for cytokeratin 18 and 19, which indicated that the culture was comprised of pure epithelial cells and contained no fibroblasts. Furthermore, PAS staining demonstrated that the cultured gastric cells produced neutral mucin. Granulin and carbohydrate antigen 724 staining confirmed the purity of gastric cancer and normal cells in culture. This method of cell culture indicated that the gastric cells in primary culture consisted of mucin-secreting gastric epithelial cells, which may be useful for the study of gastric infection with Helicobacter pylori and gastric cancer.

  9. Effects of Two Different Rhodiola rosea Extracts on Primary Human Visceral Adipocytes.

    Science.gov (United States)

    Pomari, Elena; Stefanon, Bruno; Colitti, Monica

    2015-05-11

    Rhodiola rosea (Rro) has been reported to have various pharmacological properties, including anti-fatigue, anti-stress and anti-inflammatory activity. It is also known to improve glucose and lipid metabolism, but the effects of Rhodiola rosea on adipocyte differentiation and metabolism are not still elucidated. In this study the anti-adipogenic and lipolytic activity of two extracts of Rhodiola rosea, containing 3% salidroside (RS) or 1% salidroside and 3% rosavines (RR) on primary human visceral adipocytes was investigated. Pre-adipocytes were analyzed after 10 and 20 days of treatment during differentiation and after 7 days of treatment when they reached mature shape. The RS extract significantly induced higher apoptosis and lipolysis in comparison to control cells and to RR extract. In contrast, RR extract significantly reduced triglyceride incorporation during maturation. Differentiation of pre-adipocytes in the presence of RS and RR extracts showed a significant decrease in expression of genes involved in adipocyte function such as SLC2A4 and the adipogenic factor FGF2 and significant increase in expression of genes involved in inhibition of adipogenesis, such as GATA3, WNT3A, WNT10B. Furthermore RR extract, in contrast to RS, significantly down-regulates PPARG, the master regulator of adipogenesis and FABP4. These data support the lipolytic and anti-adipogenetic activity of two different commercial extracts of Rhodiola rosea in primary human visceral pre-adipocytes during differentiation.

  10. Effects of Two Different Rhodiola rosea Extracts on Primary Human Visceral Adipocytes

    Directory of Open Access Journals (Sweden)

    Elena Pomari

    2015-05-01

    Full Text Available Rhodiola rosea (Rro has been reported to have various pharmacological properties, including anti-fatigue, anti-stress and anti-inflammatory activity. It is also known to improve glucose and lipid metabolism, but the effects of Rhodiola rosea on adipocyte differentiation and metabolism are not still elucidated. In this study the anti-adipogenic and lipolytic activity of two extracts of Rhodiola rosea, containing 3% salidroside (RS or 1% salidroside and 3% rosavines (RR on primary human visceral adipocytes was investigated. Pre-adipocytes were analyzed after 10 and 20 days of treatment during differentiation and after 7 days of treatment when they reached mature shape. The RS extract significantly induced higher apoptosis and lipolysis in comparison to control cells and to RR extract. In contrast, RR extract significantly reduced triglyceride incorporation during maturation. Differentiation of pre-adipocytes in the presence of RS and RR extracts showed a significant decrease in expression of genes involved in adipocyte function such as SLC2A4 and the adipogenic factor FGF2 and significant increase in expression of genes involved in inhibition of adipogenesis, such as GATA3, WNT3A, WNT10B. Furthermore RR extract, in contrast to RS, significantly down-regulates PPARG, the master regulator of adipogenesis and FABP4. These data support the lipolytic and anti-adipogenetic activity of two different commercial extracts of Rhodiola rosea in primary human visceral pre-adipocytes during differentiation.

  11. Progesterone receptor structure and protease activity in primary human endometrial carcinoma.

    Science.gov (United States)

    Feil, P D; Clarke, C L; Satyaswaroop, P G

    1988-03-01

    Monoclonal antibodies were used to investigate progesterone receptor structure (isoforms) in 33 primary human endometrial tumors. The monoclonal antibodies recognized on protein blots two progesterone receptor proteins with molecular weights of 116,000 and 81,000. The Mr 116,000 protein appeared as a triplet, while a single band was found for the Mr 81,000 protein. The triplet/singlet structure was found in all progesterone receptor-positive tumors, regardless of the degree of tumor differentiation. Protease activity, which gave rise to a false-negative pattern on protein blots, was found in approximately one-half of the tumors in which it was investigated. Inclusion of a cocktail of protease inhibitors during sample preparation resulted in the maintenance of the triplet/singlet progesterone receptor structure. Mixing experiments using a progesterone receptor-rich human endometrial carcinoma (EnCa 101), which lacks protease activity, and protease-containing primary tumor homogenates indicated that the protease was leupeptin sensitive. Interestingly, while the proteolytic activity reduced or eliminated the triplet/singlet progesterone receptor structure seen on protein blot analysis, it did not affect progesterone receptor concentration measured by Scatchard analysis. Sample preparation in the presence of protease inhibitors is therefore a requisite for structural analysis of the progesterone receptor in endometrial tumors.

  12. Efficient nanoparticle mediated sustained RNA interference in human primary endothelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Mukerjee, Anindita; Shankardas, Jwalitha; Ranjan, Amalendu P; Vishwanatha, Jamboor K, E-mail: Jamboor.vishwanatha@unthsc.edu [Department of Molecular Biology and Immunology and Institute for Cancer Research, Graduate School of Biomedical Sciences, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States)

    2011-11-04

    Endothelium forms an important target for drug and/or gene therapy since endothelial cells play critical roles in angiogenesis and vascular functions and are associated with various pathophysiological conditions. RNA mediated gene silencing presents a new therapeutic approach to overcome many such diseases, but the major challenge of such an approach is to ensure minimal toxicity and effective transfection efficiency of short hairpin RNA (shRNA) to primary endothelial cells. In the present study, we formulated shAnnexin A2 loaded poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles which produced intracellular small interfering RNA (siRNA) against Annexin A2 and brought about the downregulation of Annexin A2. The per cent encapsulation of the plasmid within the nanoparticle was found to be 57.65%. We compared our nanoparticle based transfections with Lipofectamine mediated transfection, and our studies show that nanoparticle based transfection efficiency is very high ({approx}97%) and is more sustained compared to conventional Lipofectamine mediated transfections in primary retinal microvascular endothelial cells and human cancer cell lines. Our findings also show that the shAnnexin A2 loaded PLGA nanoparticles had minimal toxicity with almost 95% of cells being viable 24 h post-transfection while Lipofectamine based transfections resulted in only 30% viable cells. Therefore, PLGA nanoparticle based transfection may be used for efficient siRNA transfection to human primary endothelial and cancer cells. This may serve as a potential adjuvant treatment option for diseases such as diabetic retinopathy, retinopathy of prematurity and age related macular degeneration besides various cancers.

  13. Efficient nanoparticle mediated sustained RNA interference in human primary endothelial cells

    Science.gov (United States)

    Mukerjee, Anindita; Shankardas, Jwalitha; Ranjan, Amalendu P.; Vishwanatha, Jamboor K.

    2011-11-01

    Endothelium forms an important target for drug and/or gene therapy since endothelial cells play critical roles in angiogenesis and vascular functions and are associated with various pathophysiological conditions. RNA mediated gene silencing presents a new therapeutic approach to overcome many such diseases, but the major challenge of such an approach is to ensure minimal toxicity and effective transfection efficiency of short hairpin RNA (shRNA) to primary endothelial cells. In the present study, we formulated shAnnexin A2 loaded poly(D,L-lactide-co-glycolide) (PLGA) nanoparticles which produced intracellular small interfering RNA (siRNA) against Annexin A2 and brought about the downregulation of Annexin A2. The per cent encapsulation of the plasmid within the nanoparticle was found to be 57.65%. We compared our nanoparticle based transfections with Lipofectamine mediated transfection, and our studies show that nanoparticle based transfection efficiency is very high (~97%) and is more sustained compared to conventional Lipofectamine mediated transfections in primary retinal microvascular endothelial cells and human cancer cell lines. Our findings also show that the shAnnexin A2 loaded PLGA nanoparticles had minimal toxicity with almost 95% of cells being viable 24 h post-transfection while Lipofectamine based transfections resulted in only 30% viable cells. Therefore, PLGA nanoparticle based transfection may be used for efficient siRNA transfection to human primary endothelial and cancer cells. This may serve as a potential adjuvant treatment option for diseases such as diabetic retinopathy, retinopathy of prematurity and age related macular degeneration besides various cancers.

  14. Differential effects of antidepressants on glucocorticoid receptors in human primary blood cells and human monocytic U-937 cells.

    Science.gov (United States)

    Heiske, Andreas; Jesberg, Jutta; Krieg, Jürgen-Christian; Vedder, Helmut

    2003-04-01

    A number of data support the assumption that antidepressants (ADs) normalize the altered function of the hypothalamic-pituitary-adrenocortical (HPA) system involved in the pathophysiology of depressive disorder via direct effects on glucocorticoid receptors (GRs). In the present study, we examined the tricyclic ADs desipramine (DESI) and imipramine (IMI), the noradrenaline reuptake inhibitor maprotiline (MAPRO), and the noradrenergic and specific serotonergic AD (NaSSA) mirtazapine (MIR) for their effects on GR expression in primary human leukocytes and in monocytic U-937 cells. Semiquantitative RT-PCR indicated that the ADs exert differential effects on GR-mRNA levels in both primary human leukocytes and U-937 cells: whereas MAPRO and IMI did not induce pronounced changes in GR-mRNA levels, DESI and MIR significantly decreased the amounts of GR-mRNA in both cell systems. Further characterization of the effects of MIR revealed a time dependency of the regulation with an initial increase of GR-mRNA levels above control levels after 2.5 h of treatment and a decrease after 4, 24, and 48 h of incubation. A dose-response analysis demonstrated maximal effects of MIR at a concentration of 10(-7) M. Immunohistochemical studies showed that MIR increased the GR protein levels in a time-dependent manner and that this upregulation appeared earlier by additional treatment with dexamethasone (DEX). A translocation of the GR protein from the cytoplasm to the nucleus was induced between 24 and 48 h of treatment with MIR and MIR/DEX, respectively. Taken together, our data further support the assumption that ADs influence the neuroendocrine and immune system via effects on cellular GRs.

  15. Propofol Treatment Inhibits Constitutive Apoptosis in Human Primary Neutrophils and Granulocyte-Differentiated Human HL60 Cells

    Science.gov (United States)

    Hsing, Chung-Hsi; Chen, Chia-Ling; Lin, Wei-Chieh; Lin, Chiou-Feng

    2015-01-01

    Apoptosis regulation is essential for neutrophil homeostasis. We previously demonstrated that a process involving glycogen synthase kinase (GSK)-3β determines neutrophil apoptosis. As for this apoptotic process, an overdose of propofol (2,6-Diisopropylphenol; 25 μg/ml or 140 μM) also causes GSK-3β-mediated macrophage apoptosis; however, the early deactivation of GSK-3β with low-dose propofol has been shown. Therefore, we hypothesize that low-dose propofol may induce neutrophil survival via GSK-3β inactivation. Following in vitro culture, the therapeutic concentration of propofol (10 μg/ml or 56 μM) treatment decreased constitutive apoptosis in isolated human primary neutrophils and in granulocyte-differentiated HL60 cells after all-trans retinoic acid (1 μM) treatment. The inactivation of phosphatidylinositol 3-kinase (PI3-kinase)/AKT and the activation of GSK-3β results in myeloid cell leukemia 1 (Mcl-1) down-regulation, the loss of the mitochondrial transmembrane potential, and caspase-3 activation in these cells, which is accompanied by apoptosis. Notably, propofol treatment attenuates these effects in a PI3-kinase-regulated manner. We found that propofol initiates PI3-kinase/AKT-mediated GSK-3β inactivation and Mcl-1 stabilization, rescuing the constitutive apoptosis in primary neutrophils and granulocyte-differentiated acute promyelocytic leukemia HL60 cells. PMID:26061531

  16. Evidence of Distinct Tumour-Propagating Cell Populations with Different Properties in Primary Human Hepatocellular Carcinoma

    Science.gov (United States)

    Colombo, Federico; Baldan, Francesca; Mazzucchelli, Silvia; Martin-Padura, Ines; Marighetti, Paola; Cattaneo, Alessandra; Foglieni, Barbara; Spreafico, Marta; Guerneri, Silvana; Baccarin, Marco; Bertolini, Francesco; Rossi, Giorgio; Mazzaferro, Vincenzo; Cadamuro, Massimiliano; Maggioni, Marco; Agnelli, Luca; Rebulla, Paolo

    2011-01-01

    Background and Aims Increasing evidence that a number of malignancies are characterised by tumour cell heterogeneity has recently been published, but there is still a lack of data concerning liver cancers. The aim of this study was to investigate and characterise tumour-propagating cell (TPC) compartments within human hepatocellular carcinoma (HCC). Methods After long-term culture, we identified three morphologically different tumour cell populations in a single HCC specimen, and extensively characterised them by means of flow cytometry, fluorescence microscopy, karyotyping and microarray analyses, single cell cloning, and xenotransplantation in NOD/SCID/IL2Rγ−/− mice. Results The primary cell populations (hcc-1, -2 and -3) and two clones generated by means of limiting dilutions from hcc-1 (clone-1/7 and -1/8) differently expressed a number of tumour-associated stem cell markers, including EpCAM, CD49f, CD44, CD133, CD56, Thy-1, ALDH and CK19, and also showed different doubling times, drug resistance and tumorigenic potential. Moreover, we found that ALDH expression, in combination with CD44 or Thy-1 negativity or CD56 positivity identified subpopulations with a higher clonogenic potential within hcc-1, hcc-2 and hcc-3 primary cell populations, respectively. Karyotyping revealed the clonal evolution of the cell populations and clones within the primary tumour. Importantly, the primary tumour cell population with the greatest tumorigenic potential and drug resistance showed more chromosomal alterations than the others and contained clones with epithelial and mesenchymal features. Conclusions Individual HCCs can harbor different self-renewing tumorigenic cell types expressing a variety of morphological and phenotypical markers, karyotypic evolution and different gene expression profiles. This suggests that the models of hepatic carcinogenesis should take into account TPC heterogeneity due to intratumour clonal evolution. PMID:21731718

  17. Evidence of distinct tumour-propagating cell populations with different properties in primary human hepatocellular carcinoma.

    Directory of Open Access Journals (Sweden)

    Federico Colombo

    Full Text Available Increasing evidence that a number of malignancies are characterised by tumour cell heterogeneity has recently been published, but there is still a lack of data concerning liver cancers. The aim of this study was to investigate and characterise tumour-propagating cell (TPC compartments within human hepatocellular carcinoma (HCC.After long-term culture, we identified three morphologically different tumour cell populations in a single HCC specimen, and extensively characterised them by means of flow cytometry, fluorescence microscopy, karyotyping and microarray analyses, single cell cloning, and xenotransplantation in NOD/SCID/IL2Rγ/⁻ mice.The primary cell populations (hcc-1, -2 and -3 and two clones generated by means of limiting dilutions from hcc-1 (clone-1/7 and -1/8 differently expressed a number of tumour-associated stem cell markers, including EpCAM, CD49f, CD44, CD133, CD56, Thy-1, ALDH and CK19, and also showed different doubling times, drug resistance and tumorigenic potential. Moreover, we found that ALDH expression, in combination with CD44 or Thy-1 negativity or CD56 positivity identified subpopulations with a higher clonogenic potential within hcc-1, hcc-2 and hcc-3 primary cell populations, respectively. Karyotyping revealed the clonal evolution of the cell populations and clones within the primary tumour. Importantly, the primary tumour cell population with the greatest tumorigenic potential and drug resistance showed more chromosomal alterations than the others and contained clones with epithelial and mesenchymal features.Individual HCCs can harbor different self-renewing tumorigenic cell types expressing a variety of morphological and phenotypical markers, karyotypic evolution and different gene expression profiles. This suggests that the models of hepatic carcinogenesis should take into account TPC heterogeneity due to intratumour clonal evolution.

  18. Human resources for primary health care in sub-Saharan Africa: progress or stagnation?

    Science.gov (United States)

    Willcox, Merlin L; Peersman, Wim; Daou, Pierre; Diakité, Chiaka; Bajunirwe, Francis; Mubangizi, Vincent; Mahmoud, Eman Hassan; Moosa, Shabir; Phaladze, Nthabiseng; Nkomazana, Oathokwa; Khogali, Mustafa; Diallo, Drissa; De Maeseneer, Jan; Mant, David

    2015-09-10

    The World Health Organization defines a "critical shortage" of health workers as being fewer than 2.28 health workers per 1000 population and failing to attain 80% coverage for deliveries by skilled birth attendants. We aimed to quantify the number of health workers in five African countries and the proportion of these currently working in primary health care facilities, to compare this to estimates of numbers needed and to assess how the situation has changed in recent years. This study is a review of published and unpublished "grey" literature on human resources for health in five disparate countries: Mali, Sudan, Uganda, Botswana and South Africa. Health worker density has increased steadily since 2000 in South Africa and Botswana which already meet WHO targets but has not significantly increased since 2004 in Sudan, Mali and Uganda which have a critical shortage of health workers. In all five countries, a minority of doctors, nurses and midwives are working in primary health care, and shortages of qualified staff are greatest in rural areas. In Uganda, shortages are greater in primary health care settings than at higher levels. In Mali, few community health centres have a midwife or a doctor. Even South Africa has a shortage of doctors in primary health care in poorer districts. Although most countries recognize village health workers, traditional healers and traditional birth attendants, there are insufficient data on their numbers. There is an "inverse primary health care law" in the countries studied: staffing is inversely related to poverty and level of need, and health worker density is not increasing in the lowest income countries. Unless there is money to recruit and retain staff in these areas, training programmes will not improve health worker density because the trained staff will simply leave to work elsewhere. Information systems need to be improved in a way that informs policy on the health workforce. It may be possible to use existing resources

  19. Primary cell culture from human oral tissue: gingival keratinocytes,gingival fibroblasts and periodontal ligament fibroblasts

    Directory of Open Access Journals (Sweden)

    Supreya Wanichpakorn

    2010-08-01

    Full Text Available Primary cell culture of human oral tissue has many applications for oral biology research. There are two techniques in primary culture, which includes the enzymatic and direct explant technique. The objectives of this study were (1 to isolate and investigate the difference in percentage the success in culturing three cell types from human oral tissue: gingival keratinocytes, gingival fibroblasts and periodontal ligament fibroblasts by using the direct explant technique; (2 to compare the effect of sex and age on the success of tissue culturing. Twenty seven tissue samples were obtained from healthy human gingival tissue, 19 female and 8 male patients aged 14-67 years (37.7±17.5. The tissue was cut into 1x1 mm pieces and placed on plastic culture plates containing Dulbecco’s Modified Eagle’s Medium supplemented with 10% fetal calf serum, 100 U/ml penicillin, 100 µg/ml streptomycin and 1% amphotericin B. For the keratinocytes culture, after the epithelial cells started to multiply around the gingival origin and the diameter was 2-5 mm., the fibroblasts were liminated by mechanical removal under inverted microscope to prevent fibroblast overgrowth and the medium was changed to keratinocyte-SFM (Gibco, BRL supplemented with 5 µg/ml gentamycin. The results revealed that gingival fibroblast gave the highest success rate in culture (96.3%, followed by gingival keratinocytes (88.9% and periodontal ligament fibroblasts (81.5%. There was no significant difference in the success rate of cultivation between younger and older individuals, as between sex of the subjects (p>0.05. The risk of failure in culture techniques is mainly caused by microbiological contamination from the tissue samples.

  20. UVA and UVB irradiation differentially regulate microRNA expression in human primary keratinocytes.

    Directory of Open Access Journals (Sweden)

    Anne Kraemer

    Full Text Available MicroRNA (miRNA-mediated regulation of the cellular transcriptome is an important epigenetic mechanism for fine-tuning regulatory pathways. These include processes related to skin cancer development, progression and metastasis. However, little is known about the role of microRNA as an intermediary in the carcinogenic processes following exposure to UV-radiation. We now show that UV irradiation of human primary keratinocytes modulates the expression of several cellular miRNAs. A common set of miRNAs was influenced by exposure to both UVA and UVB. However, each wavelength band also activated a distinct subset of miRNAs. Common sets of UVA- and UVB-regulated miRNAs harbor the regulatory elements GLYCA-nTRE, GATA-1-undefined-site-13 or Hox-2.3-undefined-site-2 in their promoters. In silico analysis indicates that the differentially expressed miRNAs responding to UV have potential functions in the cellular pathways of cell growth and proliferation. Interestingly, the expression of miR-23b, which is a differentiation marker of human keratinocytes, is remarkably up-regulated after UVA irradiation. Studying the interaction between miR-23b and its putative skin-relevant targets using a Luciferase reporter assay revealed that RRAS2 (related RAS viral oncogene homolog 2, which is strongly expressed in highly aggressive malignant skin cancer, to be a direct target of miR-23b. This study demonstrates for the first time a differential miRNA response to UVA and UVB in human primary keratinocytes. This suggests that selective regulation of signaling pathways occurs in response to different UV energies. This may shed new light on miRNA-regulated carcinogenic processes involved in UV-induced skin carcinogenesis.

  1. [Human ecology and interdisciplinary cooperation for primary prevention of environmental risk factors for public health].

    Science.gov (United States)

    Dobrowolski, Jan W

    2007-01-01

    training activity in ecologically-based primary prevention. Training in this important field is not adequate in medical, technological, and also natural subjects of studies. There is not enough opportunity for education of the students and graduates toward the application of integrated system approach of new achievements in different sciences and technologies. Interesting are experiences connected with long-term case studies in highly polluted regions in Poland, Japan, India, as well as exchange of methodological experiences during the series of International Summer Schools on the Human Environment from 1972, as well as during series of 11 International Conferences on Sustainable Development organized at AGH-UST from 1989 to 2006 and Polish Conferences in 2004 and 2007. It seems necessary not only to develop a training of experts that would be adequate to present needs, but also education of the whole society (including formal activities at all levels of education) as well as informal education (e.g. at Open Universities and Distance Education, based on the Internet) to achieve the integration of activity of scientists, practitioners and the whole society. It would be useful to focus this activity on crucial problems and selected regions. Let me propose as the top priority for inhabitants of Tarnow region as well as pilot projects for Poland; utilization of all possible achievements of science and technology for primary prevention of health hazard for inhabitants of Gmina Szczucin that is very polluted by asbestos, and also model management reducing risk factors for the natural environment and health of inhabitants in the regions of new motor-ways, as well as better primary prevention against flood accidents and connected with their effects (higher humidity of housing environment and its contamination by toxinogenic moulds) risk factors for health of communities living in rivers regions. For the purpose of optimisation of preventive action, it is necessary not only to

  2. Mechanisms of acetaminophen-induced cell death in primary human hepatocytes

    Energy Technology Data Exchange (ETDEWEB)

    Xie, Yuchao; McGill, Mitchell R.; Dorko, Kenneth [Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS 66160 (United States); Kumer, Sean C.; Schmitt, Timothy M.; Forster, Jameson [Department of Surgery, University of Kansas Medical Center, Kansas City, KS 66160 (United States); Jaeschke, Hartmut, E-mail: hjaeschke@kumc.edu [Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, KS 66160 (United States)

    2014-09-15

    Acetaminophen (APAP) overdose is the most prevalent cause of drug-induced liver injury in western countries. Numerous studies have been conducted to investigate the mechanisms of injury after APAP overdose in various animal models; however, the importance of these mechanisms for humans remains unclear. Here we investigated APAP hepatotoxicity using freshly isolated primary human hepatocytes (PHH) from either donor livers or liver resections. PHH were exposed to 5 mM, 10 mM or 20 mM APAP over a period of 48 h and multiple parameters were assessed. APAP dose-dependently induced significant hepatocyte necrosis starting from 24 h, which correlated with the clinical onset of human liver injury after APAP overdose. Interestingly, cellular glutathione was depleted rapidly during the first 3 h. APAP also resulted in early formation of APAP-protein adducts (measured in whole cell lysate and in mitochondria) and mitochondrial dysfunction, indicated by the loss of mitochondrial membrane potential after 12 h. Furthermore, APAP time-dependently triggered c-Jun N-terminal kinase (JNK) activation in the cytosol and translocation of phospho-JNK to the mitochondria. Both co-treatment and post-treatment (3 h) with the JNK inhibitor SP600125 reduced JNK activation and significantly attenuated cell death at 24 h and 48 h after APAP. The clinical antidote N-acetylcysteine offered almost complete protection even if administered 6 h after APAP and a partial protection when given at 15 h. Conclusion: These data highlight important mechanistic events in APAP toxicity in PHH and indicate a critical role of JNK in the progression of injury after APAP in humans. The JNK pathway may represent a therapeutic target in the clinic. - Highlights: • APAP reproducibly causes cell death in freshly isolated primary human hepatocytes. • APAP induces adduct formation, JNK activation and mitochondrial dysfunction in PHH. • Mitochondrial adducts and JNK translocation are delayed in PHH compared to

  3. Development of Functional Microfold (M Cells from Intestinal Stem Cells in Primary Human Enteroids.

    Directory of Open Access Journals (Sweden)

    Joshua D Rouch

    Full Text Available Intestinal microfold (M cells are specialized epithelial cells that act as gatekeepers of luminal antigens in the intestinal tract. They play a critical role in the intestinal mucosal immune response through transport of viruses, bacteria and other particles and antigens across the epithelium to immune cells within Peyer's patch regions and other mucosal sites. Recent studies in mice have demonstrated that M cells are generated from Lgr5+ intestinal stem cells (ISCs, and that infection with Salmonella enterica serovar Typhimurium increases M cell formation. However, it is not known whether and how these findings apply to primary human small intestinal epithelium propagated in an in vitro setting.Human intestinal crypts were grown as monolayers with growth factors and treated with recombinant RANKL, and assessed for mRNA transcripts, immunofluorescence and uptake of microparticles and S. Typhimurium.Functional M cells were generated by short-term culture of freshly isolated human intestinal crypts in a dose- and time-dependent fashion. RANKL stimulation of the monolayer cultures caused dramatic induction of the M cell-specific markers, SPIB, and Glycoprotein-2 (GP2 in a process primed by canonical WNT signaling. Confocal microscopy demonstrated a pseudopod phenotype of GP2-positive M cells that preferentially take up microparticles. Furthermore, infection of the M cell-enriched cultures with the M cell-tropic enteric pathogen, S. Typhimurium, led to preferential association of the bacteria with M cells, particularly at lower inoculum sizes. Larger inocula caused rapid induction of M cells.Human intestinal crypts containing ISCs can be cultured and differentiate into an epithelial layer with functional M cells with characteristic morphological and functional properties. This study is the first to demonstrate that M cells can be induced to form from primary human intestinal epithelium, and that S. Typhimurium preferentially infect these cells in an

  4. Susceptibility and response of human blood monocyte subsets to primary dengue virus infection.

    Directory of Open Access Journals (Sweden)

    Kok Loon Wong

    Full Text Available Human blood monocytes play a central role in dengue infections and form the majority of virus infected cells in the blood. Human blood monocytes are heterogeneous and divided into CD16(- and CD16(+ subsets. Monocyte subsets play distinct roles during disease, but it is not currently known if monocyte subsets differentially contribute to dengue protection and pathogenesis. Here, we compared the susceptibility and response of the human CD16(- and CD16(+ blood monocyte subsets to primary dengue virus in vitro. We found that both monocyte subsets were equally susceptible to dengue virus (DENV2 NGC, and capable of supporting the initial production of new infective virus particles. Both monocyte subsets produced anti-viral factors, including IFN-α, CXCL10 and TRAIL. However, CD16(+ monocytes were the major producers of inflammatory cytokines and chemokines in response to dengue virus, including IL-1β, TNF-α, IL-6, CCL2, 3 and 4. The susceptibility of both monocyte subsets to infection was increased after IL-4 treatment, but this increase was more profound for the CD16(+ monocyte subset, particularly at early time points after virus exposure. These findings reveal the differential role that monocyte subsets might play during dengue disease.

  5. Primary and secondary restraints of human and ovine knees for simulated in vivo gait kinematics.

    Science.gov (United States)

    Nesbitt, Rebecca J; Herfat, Safa T; Boguszewski, Daniel V; Engel, Andrew J; Galloway, Marc T; Shearn, Jason T

    2014-06-27

    Knee soft tissue structures are frequently injured, leading to the development of osteoarthritis even with treatment. Understanding how these structures contribute to knee function during activities of daily living (ADLs) is crucial in creating more effective treatments. This study was designed to determine the role of different knee structures during a simulated ADL in both human knees and ovine stifle joints. A six degree-of-freedom robot was used to reproduce each species' in vivo gait while measuring three-dimensional joint forces and torques. Using a semi-randomized selective cutting method, we determined the primary and secondary structures contributing to the forces and torques along and about each anatomical axis. In both species, the bony interaction, ACL, and medial meniscus provided most of the force contributions during stance, whereas the ovine MCL, human bone, and ACLs of both species were the key contributors during swing. This study contributes to our overarching goal of establishing functional tissue engineering parameters for knee structures by further validating biomechanical similarities between the ovine model and the human to provide a platform for measuring biomechanics during an in vivo ADL. These parameters will be used to develop more effective treatments for knee injuries to reduce or eliminate the incidence of osteoarthritis. © 2013 Published by Elsevier Ltd.

  6. Humans rather than climate the primary cause of Pleistocene megafaunal extinction in Australia

    Science.gov (United States)

    van der Kaars, Sander; Miller, Gifford H.; Turney, Chris S. M.; Cook, Ellyn J.; Nürnberg, Dirk; Schönfeld, Joachim; Kershaw, A. Peter; Lehman, Scott J.

    2017-01-01

    Environmental histories that span the last full glacial cycle and are representative of regional change in Australia are scarce, hampering assessment of environmental change preceding and concurrent with human dispersal on the continent ca. 47,000 years ago. Here we present a continuous 150,000-year record offshore south-western Australia and identify the timing of two critical late Pleistocene events: wide-scale ecosystem change and regional megafaunal population collapse. We establish that substantial changes in vegetation and fire regime occurred ~70,000 years ago under a climate much drier than today. We record high levels of the dung fungus Sporormiella, a proxy for herbivore biomass, from 150,000 to 45,000 years ago, then a marked decline indicating megafaunal population collapse, from 45,000 to 43,100 years ago, placing the extinctions within 4,000 years of human dispersal across Australia. These findings rule out climate change, and implicate humans, as the primary extinction cause.

  7. International validation of pyrogen tests based on cryopreserved human primary blood cells.

    Science.gov (United States)

    Schindler, Stefanie; Spreitzer, Ingo; Löschner, Bettina; Hoffmann, Sebastian; Hennes, Kilian; Halder, Marlies; Brügger, Peter; Frey, Esther; Hartung, Thomas; Montag, Thomas

    2006-10-20

    Pyrogens as fever-inducing agents can be a major health hazard in parenterally applied drugs. For the control of these contaminants, pyrogen testing for batch release is required by pharmacopoeias. This has been done either by the in vivo rabbit pyrogen test (since 1942) or the limulus amoebocyte lysate test (LAL), since 1976. New approaches include cell-based assays employing in vitro culture of human immune cells which respond e.g. by cytokine production (IL-1beta; IL-6) upon contact with pyrogens. Six variants of these assays have been validated in a collaborative international study. The recent successful development of cryopreservation methods promises to make standardized immunoreactive primary human blood cells available for widespread use. Furthermore, the pretesting of donors for infectious agents such as HIV or hepatitis has made it possible to develop a safe and standardised reagent for pyrogen testing. Using a total of 13 drugs, we have validated the pyrogen test based on fresh and cryopreserved human whole blood in four laboratories. The test reached >90% sensitivity and specificity. In contrast to the LAL, the test was capable of detecting non-endotoxin pyrogens derived from Gram-positive bacteria or fungi.

  8. Establishment and characterization of a human primary prostate carcinoma cell line, HH870.

    Science.gov (United States)

    Selvan, Senthamil R; Cornforth, Andrew N; Rao, Nagesh P; Reid, Yvonne A; Schiltz, Patric M; Liao, Ray P; Price, David T; Heinemann, F Scott; Dillman, Robert O

    2005-04-01

    Development of new therapeutic modalities for human prostate carcinoma has been impeded by a lack of adequate in vitro and in vivo models. Most in vitro studies have been carried out using a limited number of human prostate cancer cell lines that are mostly derived from metastatic tumors sites or are immortalized. Characterization of the prostate cancer cell line, HH870, included description of morphology, determination of doubling time, response to androgens, immunocytochemistry, and immunoblotting of proteins known to be associated with prostate carcinoma, karyotyping, fluorescence in situ hybridization (FISH), DNA profiling, and growth as xenograft in athymic rodents. HH870 expresses various epithelial marker antigens that correlate with known basic immunostaining profiles of prostate adenocarcinoma, although the cell line does not express PSA, PSMA, or PAP. HH870 exhibits complex chromosomal abnormalities and harbors no immortalizing HPV, BKV, JCV, and SV40 DNA. We report the successful establishment and characterization of a new long-term primary human prostate tumor cell line HH870. Copyright (c) 2004 Wiley-Liss, Inc.

  9. Gene expression in response to cyclic mechanical stretch in primary human dermal fibroblasts

    Directory of Open Access Journals (Sweden)

    Maria Reichenbach

    2014-12-01

    Full Text Available The human dermal skin is permanently exposed to mechanical stress, for instance during facial expression, which might cause wrinkles with age. Cyclic mechanical stretching of cells results in cellular and cytoskeleton alignment perpendicular to the stretch direction regulating cellular response. With gene expression profiling it was aimed to identify the differentially expressed genes associated with the regulation of the cytoskeleton to investigate the stretch-induced cell alignment mechanism. Here, the transcription activity of the genome in response to cyclic mechanical stress was measured using DNA microarray technology with Agilent SurePrint G3 Human GE 8x60k Microarrays, based on the overall measurement of the mRNA. Gene expression was measured at the beginning of the alignment process showing first reoriented cells after 5 h stretching and at the end after 24 h, where nearly all cells are aligned. Gene expression data of control vs. stretched primary human dermal fibroblasts after 5 h and 24 h demonstrated the regulation of differentially expressed genes associated with metabolism, differentiation and morphology and were deposited at http://www.ncbi.nlm.nih.gov/geo with the accession number GSE58389.

  10. Morphometric study of the two fused primary ossification centers of the clavicle in the human fetus.

    Science.gov (United States)

    Baumgart, Mariusz; Wiśniewski, Marcin; Grzonkowska, Magdalena; Badura, Mateusz; Dombek, Małgorzata; Małkowski, Bogdan; Szpinda, Michał

    2016-10-01

    A satisfactory understanding of the clavicle development may be contributing to both the diagnosis of its congenital defects and prevention of perinatal damage to the shoulder girdle. This study was carried out to examine the transverse and sagittal diameters, cross-sectional area and volume of the two fused primary ossification centers of the clavicle. Using the methods of CT, digital-image analysis and statistics, the size for two fused primary ossification centers of the clavicle in 42 spontaneously aborted human fetuses at ages of 18-30 weeks was studied. Without any male-female and right-left significant differences, the best fit growth models for two fused primary ossification centers of the clavicle were as follows: y = -31.373 + 15.243 × ln(age) ± 1.424 (R (2) = 0.74) for transverse diameter, y = -7.945 + 3.225 × ln(age) ± 0.262 (R (2) = 0.78), y = -4.503 + 2.007 × ln(age) ± 0.218 (R (2) = 0.68), and y = -4.860 + 2.117 × ln(age) ± 0.200 (R (2) = 0.73) for sagittal diameters of the lateral, middle and medial ends respectively, y = -31.390 + 2.432 × age ± 4.599 (R (2) = 0.78) for cross-sectional area, and y = 28.161 + 0.00017 × (age)(4) ± 15.357 (R (2) = 0.83) for volume. With no sex and laterality differences, the fused primary ossification centers of the clavicle grow logarithmically in both transverse and sagittal diameters, linearly in cross-sectional area, and fourth-degree polynomially in volume. Our normative quantitative findings may be conducive in monitoring normal fetal growth and screening for inherited faults and anomalies of the clavicle in European human fetuses.

  11. Xenobiotic-Metabolizing Enzyme and Transporter Gene Expression in Primary Cultures of Human Hepatocytes Modulated by Toxcast Chemicals

    Science.gov (United States)

    Primary human hepatocyte cultures are useful in vitro model systems of human liver because when cultured under appropriate conditions the hepatocytes retain liver-like functionality such as metabolism, transport, and cell signaling. This model system was used to characterize the ...

  12. Dexamethasone-Mediated Activation of Fibronectin Matrix Assembly Reduces Dispersal of Primary Human Glioblastoma Cells.

    Directory of Open Access Journals (Sweden)

    Stephen Shannon

    Full Text Available Despite resection and adjuvant therapy, the 5-year survival for patients with Glioblastoma multiforme (GBM is less than 10%. This poor outcome is largely attributed to rapid tumor growth and early dispersal of cells, factors that contribute to a high recurrence rate and poor prognosis. An understanding of the cellular and molecular machinery that drive growth and dispersal is essential if we are to impact long-term survival. Our previous studies utilizing a series of immortalized GBM cell lines established a functional causation between activation of fibronectin matrix assembly (FNMA, increased tumor cohesion, and decreased dispersal. Activation of FNMA was accomplished by treatment with Dexamethasone (Dex, a drug routinely used to treat brain tumor related edema. Here, we utilize a broad range of qualitative and quantitative assays and the use of a human GBM tissue microarray and freshly-isolated primary human GBM cells grown both as conventional 2D cultures and as 3D spheroids to explore the role of Dex and FNMA in modulating various parameters that can significantly influence tumor cell dispersal. We show that the expression and processing of fibronectin in a human GBM tissue-microarray is variable, with 90% of tumors displaying some abnormality or lack in capacity to secrete fibronectin or assemble it into a matrix. We also show that low-passage primary GBM cells vary in their capacity for FNMA and that Dex treatment reactivates this process. Activation of FNMA effectively "glues" cells together and prevents cells from detaching from the primary mass. Dex treatment also significantly increases the strength of cell-ECM adhesion and decreases motility. The combination of increased cohesion and decreased motility discourages in vitro and ex vivo dispersal. By increasing cell-cell cohesion, Dex also decreases growth rate of 3D spheroids. These effects could all be reversed by an inhibitor of FNMA and by the glucocorticoid receptor antagonist, RU

  13. Primary cilia: the chemical antenna regulating human adipose-derived stem cell osteogenesis.

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    Josephine C Bodle

    Full Text Available Adipose-derived stem cells (ASC are multipotent stem cells that show great potential as a cell source for osteogenic tissue replacements and it is critical to understand the underlying mechanisms of lineage specification. Here we explore the role of primary cilia in human ASC (hASC differentiation. This study focuses on the chemosensitivity of the primary cilium and the action of its associated proteins: polycystin-1 (PC1, polycystin-2 (PC2 and intraflagellar transport protein-88 (IFT88, in hASC osteogenesis. To elucidate cilia-mediated mechanisms of hASC differentiation, siRNA knockdown of PC1, PC2 and IFT88 was performed to disrupt cilia-associated protein function. Immunostaining of the primary cilium structure indicated phenotypic-dependent changes in cilia morphology. hASC cultured in osteogenic differentiation media yielded cilia of a more elongated conformation than those cultured in expansion media, indicating cilia-sensitivity to the chemical environment and a relationship between the cilium structure and phenotypic determination. Abrogation of PC1, PC2 and IFT88 effected changes in both hASC proliferation and differentiation activity, as measured through proliferative activity, expression of osteogenic gene markers, calcium accretion and endogenous alkaline phosphatase activity. Results indicated that IFT88 may be an early mediator of the hASC differentiation process with its knockdown increasing hASC proliferation and decreasing Runx2, alkaline phosphatase and BMP-2 mRNA expression. PC1 and PC2 knockdown affected later osteogenic gene and end-product expression. PC1 knockdown resulted in downregulation of alkaline phosphatase and osteocalcin gene expression, diminished calcium accretion and reduced alkaline phosphatase enzymatic activity. Taken together our results indicate that the structure of the primary cilium is intimately associated with the process of hASC osteogenic differentiation and that its associated proteins are critical

  14. Expanding Thurston maps

    CERN Document Server

    Bonk, Mario

    2017-01-01

    This monograph is devoted to the study of the dynamics of expanding Thurston maps under iteration. A Thurston map is a branched covering map on a two-dimensional topological sphere such that each critical point of the map has a finite orbit under iteration. It is called expanding if, roughly speaking, preimages of a fine open cover of the underlying sphere under iterates of the map become finer and finer as the order of the iterate increases. Every expanding Thurston map gives rise to a fractal space, called its visual sphere. Many dynamical properties of the map are encoded in the geometry of this visual sphere. For example, an expanding Thurston map is topologically conjugate to a rational map if and only if its visual sphere is quasisymmetrically equivalent to the Riemann sphere. This relation between dynamics and fractal geometry is the main focus for the investigations in this work.

  15. In Vitro experimental model of trained innate immunity in human primary monocytes

    DEFF Research Database (Denmark)

    Bekkering, S.; Blok, B. A.; Joosten, Leo A B

    2016-01-01

    training period with β-glucan, BCG, or oxLDL, followed by washing and resting of the cells and, thereafter, restimulation with secondary bacterial stimuli. The training and resting time intervals were varied to identify the optimal setting for the long-term induction of trained immunity. Trained immunity...... was assessed in terms of the secondary cytokine response, the production of reactive oxygen species, cell morphology, and induction of glycolysis. Monocytes primed with β-glucan, BCG, and oxLDL showed increased pro-and antiinflammatory cytokine responses upon restimulation with nonrelated stimuli. Also, all......-glucan led to the decreased production of reactive oxygen species. We describe the optimal conditions for an in vitro experimental model with human primary monocytes for study of the induction of trained innate immunity by microbial and metabolic stimuli. Copyright © 2016, American Society for Microbiology...

  16. Association of physical properties and maintenance of sterility of primary teeth in human tooth bank

    Directory of Open Access Journals (Sweden)

    Nitika Bajaj

    2014-01-01

    Full Text Available Background: This study was carried out to evaluate the sterility and structural integrity of stored primary teeth in artificial saliva over a storage period of 12 weeks (3 months in human tooth bank (HTB. Materials and Methods: A total of 80 freshly extracted teeth were taken and were categorized into four groups of 20 each. The samples were stored at 4°C for 12 weeks (3 months in the refrigerator. During their storage time of 3 months, they were repetitively evaluated for their sterility, enamel hardness, calcium and phosphate solubility and color stability at regular intervals of 3 weeks. Observations and Results: That teeth stored in artificial saliva maintained their sterility throughout the storage time of 12 weeks, but changes in their physical and chemical properties occurred with an increase in storage time. Conclusion: Storage time not more than 9 weeks is recommended for deciduous teeth to be stored in artificial saliva in a HTB.

  17. Human Papillomavirus Assays and Cytology in Primary Cervical Screening of Women Aged 30 Years and Above

    DEFF Research Database (Denmark)

    Rebolj, Matejka; Bonde, Jesper; Preisler, Sarah

    2016-01-01

    In women aged ≥30 years, Human Papillomavirus testing will replace cytology for primary cervical screening. We compared Hybrid Capture 2 (HC2), cobas, CLART, and APTIMA HPV assays with cytology on 2869 SurePath samples from women undergoing routine screening at 30-65 years in Copenhagen, Denmark....... Women with cytological abnormalities were managed according to routine recommendations, with 92% completeness. Those with cytology-normal/HPV-positive samples (on any of the four assays) were invited for repeated cytology and HPV testing in 1.5 year, and 58% had additional testing. HPV testing detected...... more ≥CIN3 than cytology (HC2: 35, cobas, CLART: 37, APTIMA: 34, cytology: 31), although statistically the differences were not significant. Cobas and CLART detected significantly more ≥CIN2 than cytology (cobas, CLART: 49, cytology: 39). The proportion of women with false-positive test results...

  18. Success Rate of Formocresol Pulpotomy versus Mineral Trioxide Aggregate in Human Primary Molar Tooth

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    S E Jabbarifar

    2004-12-01

    Full Text Available Background: In spite of long time and broad use of formaldehyde derivates (Fixation agent in primary tooth pulp treatment, There is some concerns about these derivates such as variability, inconsistency success rate, mutagenicity, cytotoxicity, alergenicity, and some other potential health hazards of them. Therefore other alternative pulpotomy procedures like Bioactive glass (BAG, Glutaraldehyde (2%, Hydroxyappetite (HA, Bone dried freezed (BDF, ferric sulfate (15%, laser, Electrosurgery (ES, Bone Morphogenic proteins (BMP, recombinant protein-1 (RP1, and Mineral Trioxide Aggregate (MTA have been compared. The purpose of this clinical trial is to assess radiographic and clinical success rate of Formocresol (FC pulpotomy in compare with MTA in human primary molar teeth. Methods: 64 molars were pulpotomized equally and randomly with mineral trioxide Aggregate and Formocresol. Prior to trial, we defined a case as failure, when one or more of the events such as external root resorption, internal root resorption, periapical and furca lucency, pain, swelling, mobility, dental abscess, or early extraction appeared. Every treated tooth was defined as successful, if any noted evident was not shown. Results: Totally, 60 teeth treatment (92.2 percent were successful and 7.8 percent were failed. Failure and success rates for MTA group were 6.3 and 93.7 percent, respectively. Failure and success rates in FC group were 8.4 and 90.2 percent respectively. The difference between MTA and FC treatment methods was not significant (Fisher Exact test. Conclusion: Findings of this study show that mineral trioxide aggregate can be an alternative procedure for FC pulpotomy of primary tooth. Keywords: Mineral trioxide aggregate, formocresol, pulpotomy, success and failure rate.

  19. Differential BCCIP gene expression in primary human ovarian cancer, renal cell carcinoma and colorectal cancer tissues.

    Science.gov (United States)

    Liu, Xiaoxia; Cao, Lingling; Ni, Jinsong; Liu, Ning; Zhao, Xiaoming; Wang, Yanfang; Zhu, Lin; Wang, Lingyao; Wang, Jin; Yue, Ying; Cai, Yong; Jin, Jingji

    2013-12-01

    Human BCCIP, a protein which interacts with BRCA2 and CDKN1A (Cip1, p21), has been implicated in many cellular processes including cell cycle regulation, DNA recombination and damage repair, telomere maintenance, embryonic development and genomic stability. BCCIP gene expression, which is an important BRCA2 cofactor in tumor suppression, has been identified in some primary cancers. Thus, we investigated the role of BCCIP expression in a large sample of clinically diagnosed primary ovarian cancer, renal cell carcinoma (RCC) and colorectal cancer (CRC) tissues. Using clinically diagnosed frozen primary cancer tissues, quantitative PCR (qPCR), western blot analysis (WB) and immunohistochemical staining (IHC) approaches were used to detect and measure gene expression. Reduced BCCIP gene expression in ovarian cancer, RCC and CRC tissues occurred in 74, 89 and 75% of tissue samples, respectively. qPCR analysis of mRNA expression in 54 ovarian cancer, 50 RCC and 44 CRC samples revealed significant (>2-fold decreased) BCCIP downregulation in 56, 70 and 46% of tissue samples, respectively. Although BCCIP expression in three different tumor tissues decreased, the relationship between BCCIP expression and clinicopathological features of each cancer was distinct. Compared to normal tissues, BCCIP expression in ovarian cancers was significantly downregulated in serous, endometrioid and mucinous carcinomas. Downregulation of BCCIP expression was strongly associated with clear cell RCC (ccRCC) and Fuhrman tumor grading, but significant differences in BCCIP expression between CRC and matched normal tissues occurred only in male CRC tissues (povarian cancer and RCC tissue samples (povarian cancer, RCC and CRC tissues, suggesting a role for the gene in the pathogenesis of these cancers.

  20. A novel three-dimensional cell culture method enhances antiviral drug screening in primary human cells.

    Science.gov (United States)

    Koban, Robert; Neumann, Markus; Daugs, Aila; Bloch, Oliver; Nitsche, Andreas; Langhammer, Stefan; Ellerbrok, Heinz

    2017-12-07

    Gefitinib is a specific inhibitor of the epidermal growth factor receptor (EGFR) and FDA approved for treatment of non-small cell lung cancer. In a previous study we could show the in vitro efficacy of gefitinib for treatment of poxvirus infections in monolayer (2D) cultivated cell lines. Permanent cell lines and 2D cultures, however, are known to be rather unphysiological; therefore it is difficult to predict whether determined effective concentrations or the drug efficacy per se are transferable to the in vivo situation. 3D cell cultures, which meanwhile are widely distributed across all fields of research, are a promising tool for more predictive in vitro investigations of antiviral compounds. In this study the spreading of cowpox virus and the antiviral efficacy of gefitinib were analyzed in primary human keratinocytes (NHEK) grown in a novel 3D extracellular matrix-based cell culture model and compared to the respective monolayer culture. 3D-cultivated NHEK grew in a polarized and thus a more physiological manner with altered morphology and close cell-cell contact. Infected cultures showed a strongly elevated sensitivity towards gefitinib. EGFR phosphorylation, cell proliferation, and virus replication were significantly reduced in 3D cultures at gefitinib concentrations which were at least 100-fold lower than those in monolayer cultures and well below the level of cytotoxicity. Our newly established 3D cell culture model with primary human cells is an easy-to-handle alternative to conventional monolayer cell cultures and previously described more complex 3D cell culture systems. It can easily be adapted to other cell types and a broad spectrum of viruses for antiviral drug screening and many other aspects of virus research under more in vivo-like conditions. In consequence, it may contribute to a more targeted realization of necessary in vivo experiments. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. In vitro culture of primary human myoblasts by using the dextran microcarriers Cytodex3®.

    Science.gov (United States)

    Rozwadowska, Natalia; Malcher, Agnieszka; Baumann, Ewa; Kolanowski, Tomasz Jan; Rucinski, Marek; Mietkiewski, Tomasz; Fiedorowicz, Katarzyna; Kurpisz, Maciej

    2016-01-01

    Primary cells in vitro culture scale-up is a crucial issue in cell-based tissue and organ regeneration therapy. Reducing costs and space occupied by the cells cultured in vitro has been an important target. Cells cultured in vitro with the use of bioreactor with dextran microcarriers (Cytodex®) have potentially a chance to meet many of the cell therapy requirements. We used collagen-coated carriers (Cytodex3®) and a spinner flask bioreactor to develop environment suitable for human myoblast proliferation. In parallel, standard adherent in vitro culture conditions for myoblasts propagation (T-flask) were conducted. Cell cycle characterization, senescence, myogenic gene ex-pression and cell apoptosis were evaluated in order to find differences between two culture systems under study. The number of cells obtained in bioreactor per 106 of starting cells population was approximately ten times lower in comparison with T-flask culture system. The microcarriers cultured adult myoblasts in compari-son with the regular T-flask culture showed faster and more advanced replicative aging and lower proliferative potential. Moreover, the percentage of the cells that entailed an irreversible cell arrest (G0 phase) was also significantly (p < 0.0001) increased. Our results suggest that population of primary human myoblasts obtained from adult individuals and propagated on dextran microcarriers did not meet the requirements of the regenerative medicine regarding quantity and quality of the cells obtained. Nonetheless, further optimization of the cell scaling up process including both microcarriers and/or bioreactor program is still an important option.

  2. Metaplasticity in human primary somatosensory cortex: effects on physiology and tactile perception

    Science.gov (United States)

    Jones, Christina B.; Lulic, Tea; Bailey, Aaron Z.; Mackenzie, Tanner N.; Mi, Yi Qun; Tommerdahl, Mark

    2016-01-01

    Theta-burst stimulation (TBS) over human primary motor cortex evokes plasticity and metaplasticity, the latter contributing to the homeostatic balance of excitation and inhibition. Our knowledge of TBS-induced effects on primary somatosensory cortex (SI) is limited, and it is unknown whether TBS induces metaplasticity within human SI. Sixteen right-handed participants (6 females, mean age 23 yr) received two TBS protocols [continuous TBS (cTBS) and intermittent TBS (iTBS)] delivered in six different combinations over SI in separate sessions. TBS protocols were delivered at 30 Hz and were as follows: a single cTBS protocol, a single iTBS protocol, cTBS followed by cTBS, iTBS followed by iTBS, cTBS followed by iTBS, and iTBS followed by cTBS. Measures included the amplitudes of the first and second somatosensory evoked potentials (SEPs) via median nerve stimulation, their paired-pulse ratio (PPR), and temporal order judgment (TOJ). Dependent measures were obtained before TBS and at 5, 25, 50, and 90 min following stimulation. Results indicate similar effects following cTBS and iTBS; increased amplitudes of the second SEP and PPR without amplitude changes to SEP 1, and impairments in TOJ. Metaplasticity was observed such that TOJ impairments following a single cTBS protocol were abolished following consecutive cTBS protocols. Additionally, consecutive iTBS protocols altered the time course of effects when compared with a single iTBS protocol. In conclusion, 30-Hz cTBS and iTBS protocols delivered in isolation induce effects consistent with a TBS-induced reduction in intracortical inhibition within SI. Furthermore, cTBS- and iTBS-induced metaplasticity appear to follow homeostatic and nonhomeostatic rules, respectively. PMID:26984422

  3. Glucosamine exposure reduces proteoglycan synthesis in primary human endothelial cells in vitro

    Directory of Open Access Journals (Sweden)

    Trine M. Reine

    2016-09-01

    Full Text Available Purpose: Glucosamine (GlcN supplements are promoted for medical reasons, for example, for patients with arthritis and other joint-related diseases. Oral intake of GlcN is followed by uptake in the intestine, transport in the circulation and thereafter delivery to chondrocytes. Here, it is postulated to have an effect on synthesis and turnover of extracellular matrix constituents expressed by these cells. Following uptake in the intestine, serum levels are transiently increased, and the endothelium is exposed to increased levels of GlcN. We investigated the possible effects of GlcN on synthesis of proteoglycans (PGs, an important matrix component, in primary human endothelial cells. Methods: Primary human endothelial cells were cultured in vitro in medium with 5 mM glucose and 0–10 mM GlcN. PGs were recovered and analysed by western blotting, or by SDS-PAGE, gel chromatography or ion-exchange chromatography of 35S-PGs after 35S-sulphate labelling of the cells. Results: The synthesis and secretion of 35S-PGs from cultured endothelial cells were reduced in a dose- and time-dependent manner after exposure to GlcN. PGs are substituted with sulphated glycosaminoglycan (GAG chains, vital for PG function. The reduction in 35S-PGs was not related to an effect on GAG chain length, number or sulphation, but rather to the total expression of PGs. Conclusion: Exposure of endothelial cells to GlcN leads to a general decrease in 35S-PG synthesis. These results suggest that exposure to high levels of GlcN can lead to decreased matrix synthesis, contrary to what has been claimed by supporters of such supplements.

  4. Helium generated cold plasma finely regulates activation of human fibroblast-like primary cells.

    Science.gov (United States)

    Brun, Paola; Pathak, Surajit; Castagliuolo, Ignazio; Palù, Giorgio; Brun, Paola; Zuin, Matteo; Cavazzana, Roberto; Martines, Emilio

    2014-01-01

    Non-thermal atmospheric pressure plasmas are being developed for a wide range of health care applications, including wound healing. However in order to exploit the potential of plasma for clinical applications, the understanding of the mechanisms involved in plasma-induced activation of fibroblasts, the cells active in the healing process, is mandatory. In this study, the role of helium generated plasma in the tissue repairing process was investigated in cultured human fibroblast-like primary cells, and specifically in hepatic stellate cells and intestinal subepithelial myofibroblasts. Five minutes after treatment, plasma induced formation of reactive oxygen species (ROS) in cultured cells, as assessed by flow cytometric analysis of fluorescence-activated 2',7'-dichlorofluorescein diacetate probe. Plasma-induced intracellular ROS were characterized by lower concentrations and shorter half-lives with respect to hydrogen peroxide-induced ROS. Moreover ROS generated by plasma treatment increased the expression of peroxisome proliferator activated receptor (PPAR)-γ, nuclear receptor that modulates the inflammatory responses. Plasma exposure promoted wound healing in an in vitro model and induced fibroblast migration and proliferation, as demonstrated, respectively, by trans-well assay and partitioning between daughter cells of carboxyfluorescein diacetate succinimidyl ester fluorescent dye. Plasma-induced fibroblast migration and proliferation were found to be ROS-dependent as cellular incubation with antioxidant agents (e.g. N-acetyl L-cysteine) cancelled the biological effects. This study provides evidence that helium generated plasma promotes proliferation and migration in liver and intestinal fibroblast-like primary cells mainly by increasing intracellular ROS levels. Since plasma-evoked ROS are time-restricted and elicit the PPAR-γ anti-inflammatory molecular pathway, this strategy ensures precise regulation of human fibroblast activation and can be considered a

  5. Targeting Nuclear Receptors with Lentivirus-Delivered Small RNAs in Primary Human Hepatocytes

    Directory of Open Access Journals (Sweden)

    Maria Thomas

    2014-07-01

    Full Text Available Background: RNA interference (RNAi has tremendous potential for investigating gene function and for developing new therapies. Primary human hepatocytes (PHH are the “gold standard” for studying the regulation of hepatic metabolism in vitro. However, application of RNAi in PHH has some technical hurdles. The objective of this study was to develop effective and robust protocol for transduction of PHH with lentiviral vectors. Methods: We used lentiviral vectors to transduce PHH for introduction of short hairpin RNAs (shRNAs targeting constitutive androstane receptor (CAR, peroxisome proliferator activated receptor alpha (PPARα, and microRNA, miR-143. Infection efficiency was quantitatively analyzed by flow cytometry and microscopy. Target gene expression was assessed using quantitative real-time (qRT-PCR method. Results: Lentiviral vector transduction resulted in ≥95% of infected cells at low multiplicity of infection (MOI of 3, which did not impair cellular viability. We demonstrated the feasibility of this technique in studies on targeting nuclear receptors, PPARα and CAR, with shRNAs as well as in lentivirus-mediated overexpression and knock-down of miRNA-143 experiments. Conclusions: We developed an efficient and robust protocol with standardized procedures for virus production, method of titer determination, and infection procedure for RNAi in primary human hepatocytes based on delivery of shRNAs, microRNAs or anti-microRNAs in different laboratory settings. This approach should be useful to study not only the regulation via nuclear receptors but also other biological, pharmacological, and toxicological aspects of drug metabolism.

  6. Biocompatibility of Polypyrrole with Human Primary Osteoblasts and the Effect of Dopants

    Science.gov (United States)

    Fahlgren, Anna; Bratengeier, Cornelia; Gelmi, Amy; Semeins, Cornelis M.; Klein-Nulend, Jenneke; Jager, Edwin W. H.; Bakker, Astrid D.

    2015-01-01

    Polypyrrole (PPy) is a conducting polymer that enables controlled drug release upon electrical stimulation. We characterized the biocompatibility of PPy with human primary osteoblasts, and the effect of dopants. We investigated the biocompatibility of PPy comprising various dopants, i.e. p-toluene sulfonate (PPy-pTS), chondroitin sulfate (PPy-CS), or dodecylbenzenesulfonate (PPy-DBS), with human primary osteoblasts. PPy-DBS showed the roughest appearance of all surfaces tested, and its wettability was similar to the gold-coated control. The average number of attached cells was 45% higher on PPy-DBS than on PPy-CS or PPy-pTS, although gene expression of the proliferation marker Ki-67 was similar in osteoblasts on all surfaces tested. Osteoblasts seeded on PPy-DBS or gold showed similar vinculin attachment points, vinculin area per cell area, actin filament structure, and Feret’s diameter, while cells seeded on PPY-CS or PPY-pTS showed disturbed focal adhesions and were enlarged with disorganized actin filaments. Osteoblasts grown on PPy-DBS or gold showed enhanced alkaline phosphatase activity and osteocalcin gene expression, but reduced osteopontin gene expression compared to cells grown on PPy-pTS and PPy-CS. In conclusion, PPy doped with DBS showed excellent biocompatibility, which resulted in maintaining focal adhesions, cell morphology, cell number, alkaline phosphatase activity, and osteocalcin gene expression. Taken together, conducting polymers doped with DBS are well tolerated by osteoblasts. Our results could provide a basis for the development of novel orthopedic or dental implants with controlled release of antibiotics and pharmaceutics that fight infections or focally enhance bone formation in a tightly controlled manner. PMID:26225862

  7. Human umbilical mesenchymal stem cells conditioned medium promote primary wound healing regeneration

    Directory of Open Access Journals (Sweden)

    Dwi Liliek Kusindarta

    2016-06-01

    Full Text Available Aim: This research was conducted to clarify the capability of human umbilical mesenchymal stem cells conditioned medium (HU-MSCM to promote regenerations of primary wound healing on the incision skin injury. Materials and Methods: In this study, two approaches in vitro and in vivo already done. On in vitro analysis, tube formation was performed using HU vein endothelial cells in the presence of HU-MSCM, in some experiments cells line was incubated prior the presence of lipopolysaccharide and HU-MSCM then apoptosis assay was performed. Furthermore, in vivo experiments 12 female rats (Rattus norvegicus were used after rats anesthetized, 7 mm wound was made by incision on the left side of the body. The wound was treated with HU-MSCM containing cream, povidone iodine was run as a control. Wound healing regenerations on the skin samples were visualized by hematoxylin-eosin staining. Results: In vitro models elucidate HU-MSCM may decreasing inflammation at the beginning of wound healing, promote cell migration and angiogenesis. In addition in vivo models show that the incision length on the skin is decreasing and more smaller, HE staining describe decreasing of inflammation phase, increasing of angiogenesis, accelerate fibroplasia, and maturation phase. Conclusions: Taken together our observation indicates that HU-MSCM could promote the acceleration of skin tissue regenerations in primary wound healing process.

  8. Human umbilical mesenchymal stem cells conditioned medium promote primary wound healing regeneration

    Science.gov (United States)

    Kusindarta, Dwi Liliek; Wihadmadyatami, Hevi; Fibrianto, Yuda Heru; Nugroho, Widagdo Sri; Susetya, Heru; Musana, Dewi Kania; Wijayanto, Hery; Prihatna, Surya Agus; Wahyuni, A. E. T. H.

    2016-01-01

    Aim: This research was conducted to clarify the capability of human umbilical mesenchymal stem cells conditioned medium (HU-MSCM) to promote regenerations of primary wound healing on the incision skin injury. Materials and Methods: In this study, two approaches in vitro and in vivo already done. On in vitro analysis, tube formation was performed using HU vein endothelial cells in the presence of HU-MSCM, in some experiments cells line was incubated prior the presence of lipopolysaccharide and HU-MSCM then apoptosis assay was performed. Furthermore, in vivo experiments 12 female rats (Rattus norvegicus) were used after rats anesthetized, 7 mm wound was made by incision on the left side of the body. The wound was treated with HU-MSCM containing cream, povidone iodine was run as a control. Wound healing regenerations on the skin samples were visualized by hematoxylin-eosin staining. Results: In vitro models elucidate HU-MSCM may decreasing inflammation at the beginning of wound healing, promote cell migration and angiogenesis. In addition in vivo models show that the incision length on the skin is decreasing and more smaller, HE staining describe decreasing of inflammation phase, increasing of angiogenesis, accelerate fibroplasia, and maturation phase. Conclusions: Taken together our observation indicates that HU-MSCM could promote the acceleration of skin tissue regenerations in primary wound healing process. PMID:27397984

  9. Interaction of Tissue Engineering Substrates with Serum Proteins and Its Influence on Human Primary Endothelial Cells.

    Science.gov (United States)

    Mohan, Tamilselvan; Niegelhell, Katrin; Nagaraj, Chandran; Reishofer, David; Spirk, Stefan; Olschewski, Andrea; Stana Kleinschek, Karin; Kargl, Rupert

    2017-02-13

    Polymer-based biomaterials particularly polycaprolactone (PCL) are one of the most promising substrates for tissue engineering. The surface chemistry of these materials plays a major role since it governs protein adsorption, cell adhesion, viability, degradation, and biocompatibility in the first place. This study correlates the interaction of the most abundant serum proteins (albumin, immunoglobulins, fibrinogen) with the surface properties of PCL and its influence on the morphology and metabolic activity of primary human arterial endothelial cells that are seeded on the materials. Prior to that, thin films of PCL are manufactured by spin-coating and characterized in detail. A quartz crystal microbalance with dissipation (QCM-D), a multiparameter surface plasmon resonance spectroscopy instrument (MP-SPR), wettability data, and atomic force microscopy are combined to elucidate the pH-dependent protein adsorption on the PCL substrates. Primary endothelial cells are cultured on the protein modified polymer, and conclusions are drawn on the significant impact of type and form of proteins coatings on cell morphology and metabolic activity.

  10. The Mucosal Innate Immune Response in Primary Human Papillomavirus Infection: A Pilot Study.

    Science.gov (United States)

    Gardella, Barbara; Iacobone, Anna Daniela; Musacchi, Valentina; Calvino, Isabel Giacoma; De Amici, Mara; Torre, Cristina; Bogliolo, Stefano; Spinillo, Arsenio

    2016-10-01

    The purpose of the study was to evaluate the mucosal immune response in women affected by primary human papillomavirus (HPV) infection, in comparison with HPV-negative women with no previous history of HPV. A case-control study comparing the activity of myeloperoxidase (MPO) and lactoferrin (LF) between 19 HPV-positive and 19 HPV-negative women matched for age. Plasmatic and cervicovaginal levels of polymorphonuclear neutrophils (PMN) exhibiting MPO and LF receptors were measured using cytofluorimetric analysis and expressed as mean of percentages. Cervicovaginal levels of MPO-/LF- PMN were lower among HPV-negative women, with a mean rate of 18.81% (SD, 21.38), as opposed to a mean rate of 35.56% (SD, 21.02) (P = 0.020) in HPV-positive women. A similar significant difference was not proven in plasma. The mean rates of plasmatic levels of MPO-/LF- PMN were 36.21% (SD, 16.87) and 36.93% (SD, 10.54) (P = 0.875) in cases and controls, respectively. All patients were evaluated 1 year later, and only 6 cases became negative. The presence of MPO-/LF- PMN has been considered as a marker of lower rate of apoptosis of HPV-infected cells. This could explain why HPV-positive women are less capable to deal with a primary infection.

  11. Drug screening of cancer cell lines and human primary tumors using droplet microfluidics.

    Science.gov (United States)

    Wong, Ada Hang-Heng; Li, Haoran; Jia, Yanwei; Mak, Pui-In; Martins, Rui Paulo da Silva; Liu, Yan; Vong, Chi Man; Wong, Hang Cheong; Wong, Pak Kin; Wang, Haitao; Sun, Heng; Deng, Chu-Xia

    2017-08-22

    Precision Medicine in Oncology requires tailoring of therapeutic strategies to individual cancer patients. Due to the limited quantity of tumor samples, this proves to be difficult, especially for early stage cancer patients whose tumors are small. In this study, we exploited a 2.4 × 2.4 centimeters polydimethylsiloxane (PDMS) based microfluidic chip which employed droplet microfluidics to conduct drug screens against suspended and adherent cancer cell lines, as well as cells dissociated from primary tumor of human patients. Single cells were dispersed in aqueous droplets and imaged within 24 hours of drug treatment to assess cell viability by ethidium homodimer 1 staining. Our results showed that 5 conditions could be screened for every 80,000 cells in one channel on our chip under current circumstances. Additionally, screening conditions have been adapted to both suspended and adherent cancer cells, giving versatility to potentially all types of cancers. Hence, this study provides a powerful tool for rapid, low-input drug screening of primary cancers within 24 hours after tumor resection from cancer patients. This paves the way for further technological advancement to cutting down sample size and increasing drug screening throughput in advent to personalized cancer therapy.

  12. MYOCILIN LEVELS IN PRIMARY OPEN-ANGLE GLAUCOMA AND PSEUDOEXFOLIATION GLAUCOMA HUMAN AQUEOUS HUMOR

    Science.gov (United States)

    Howell, Kyle G.; Vrabel, Anne M.; Chowdhury, Uttio Roy; Stamer, W. Daniel; Fautsch, Michael P.

    2010-01-01

    Purpose To determine the concentration of myocilin in primary open-angle and pseudoexfoliation glaucoma aqueous humor. Methods Aqueous humor was collected during surgery from patients with primary open-angle glaucoma (POAG), pseudoexfoliation glaucoma (PEXG), and elective cataract removal (control). Volume-equivalent aqueous samples were separated on SDS-PAGE gradient gels. Quantification of myocilin levels was performed using Western blots probed with two independent N-terminal polyclonal anti-myocilin antibodies (AB1 and AB2) followed by densitometry. Myocilin levels in aqueous humor were quantified by plotting the densitometry readings of the aqueous samples against a recombinant myocilin standard curve. Total protein concentration was determined by Bradford protein assay. Transforming growth factor beta 2 (TGFβ2) levels were assessed by ELISA. Results Myocilin levels are significantly elevated in human POAG aqueous humor when compared to control aqueous humor (AB1: 0.66 ± 0.53 ng/μl vs. 0.23 ± 0.20 ng/μl, phumor compared to control aqueous (AB1: 0.26 ± 0.20% vs. 0.10 ± 0.20%, phumor when compared to control aqueous humor. No correlation between myocilin and TGFβ2 levels was observed. Conclusions Myocilin is elevated in POAG, but not PEXG aqueous humor. PMID:20179615

  13. Comparison of anti-CD3 and anti-CD28-coated beads with soluble anti-CD3 for expanding human T cells: Differing impact on CD8 T cell phenotype and responsiveness to restimulation

    Science.gov (United States)

    2010-01-01

    Background The ability to expand virus- or tumor-specific T cells without damaging their functional capabilities is critical for success adoptive transfer immunotherapy of patients with opportunistic infection or tumor. Careful comparisons can help identify expansion methods better suited for particular clinical settings and identify recurrent deficiencies requiring new innovation. Methods We compared the efficacy of magnetic beads coated with anti-CD3 and anti-CD28 (anti-CD3/CD28 beads), and soluble anti-CD3 plus mixed mononuclear cells (designated a rapid expansion protocol or REP) in expanding normal human T cells. Results Both anti-CD3/CD28 beads and soluble anti-CD3 promoted extensive expansion. Beads stimulated greater CD4 cell growth (geometric mean of 56- versus 27-fold (p CD3-treated CD4 cells typically assumed an effector/effector memory phenotype by day 14. By comparison, a subset of anti-CD3-treated CD8 cells, derived from naïve cells, retained much greater expression of CD45RA, CD27 and CCR7, than matched bead-treated cells despite comparable expansion. These cells were clearly distinguishable from CD45RA+ terminally differentiated effector cells by the presence of CD27, the absence of CD57 and their inability to produce cytokines after stimulation. When used to expand previously stimulated cells, anti-CD3 plus autologous MNCs produced much less antigen-induced cell death of CD8 cells and significantly more CD8 expansion than beads. Conclusions Anti-CD3/CD28 beads are highly effective for expanding CD4 cells, but soluble anti-CD3 has significant potential advantages for expanding CD8 T cells, particularly where preservation of phenotypically "young" CD8 cells would be desirable, or where the T cells of interest have been antigen-stimulated in vitro or in vivo in the recent past. PMID:20977748

  14. II Brazilian Consensus on the use of human immunoglobulin in patients with primary immunodeficiencies.

    Science.gov (United States)

    Goudouris, Ekaterini Simões; Rego Silva, Almerinda Maria do; Ouricuri, Aluce Loureiro; Grumach, Anete Sevciovic; Condino-Neto, Antonio; Costa-Carvalho, Beatriz Tavares; Prando, Carolina Cardoso; Kokron, Cristina Maria; Vasconcelos, Dewton de Moraes; Tavares, Fabíola Scancetti; Silva Segundo, Gesmar Rodrigues; Barreto, Irma Cecília; Dorna, Mayra de Barros; Barros, Myrthes Anna; Forte, Wilma Carvalho Neves

    2017-01-01

    In the last few years, new primary immunodeficiencies and genetic defects have been described. Recently, immunoglobulin products with improved compositions and for subcutaneous use have become available in Brazil. In order to guide physicians on the use of human immunoglobulin to treat primary immunodeficiencies, based on a narrative literature review and their professional experience, the members of the Primary Immunodeficiency Group of the Brazilian Society of Allergy and Immunology prepared an updated document of the 1st Brazilian Consensus, published in 2010. The document presents new knowledge about the indications and efficacy of immunoglobulin therapy in primary immunodeficiencies, relevant production-related aspects, mode of use (routes of administration, pharmacokinetics, doses and intervals), adverse events (major, prevention, treatment and reporting), patient monitoring, presentations available and how to have access to this therapeutic resource in Brazil. RESUMO Nos últimos anos, novas imunodeficiências primárias e defeitos genéticos têm sido descritos. Recentemente, produtos de imunoglobulina, com aprimoramento em sua composição e para uso por via subcutânea, tornaram-se disponíveis em nosso meio. Com o objetivo de orientar o médico no uso da imunoglobulina humana para o tratamento das imunodeficiências primárias, os membros do Grupo de Assessoria em Imunodeficiências da Associação Brasileira de Alergia e Imunologia produziram um documento que teve por base uma revisão narrativa da literatura e sua experiência profissional, atualizando o I Consenso Brasileiro publicado em 2010. Apresentam-se novos conhecimentos sobre indicações e eficácia do tratamento com imunoglobulina nas imunodeficiências primárias, aspectos relevantes sobre a produção, forma de utilização (vias de administração, farmacocinética, doses e intervalos), efeitos adversos (principais efeitos, prevenção, tratamento e notificação), monitorização do

  15. Human herpesvirus 8 in primary effusion lymphoma in an HIV-seronegative male. A case report.

    Science.gov (United States)

    Munichor, Mariana; Cohen, Hector; Sarid, Ronit; Manov, Irena; Iancu, Theodore C

    2004-01-01

    AIDS-related body cavity-based lymphoma, or primary effusion lymphoma (PEL), is a distinct clinicopathologic entity that occurs predominantly in immunosuppressed patients infected with human herpesvirus 8 (HHV-8), also known as Kaposi's sarcoma-associated herpesvirus. Although it rarely occurs in human immunodeficiency virus (HIV)-negative patients, we report such a case here. A 74-year-old male, who was HIV and Epstein-Barr virus (EBV) negative, was admitted to the hospital with dyspnea and chest pain. Chest radiography and computed tomography showed right pleural effusion. Cytologic analysis of the pleural effusion revealed a high grade lymphoma with round nuclei, prominent nucleoli and abundant cytoplasm. Polymerase chain reaction performed on the pleural effusion was positive for HHV-8 and negative for EBV. On molecular studies, the immunoglobulin heavy and kappa light chains were rearranged. Flow cytometry revealed a hyperploid fraction with DNA index of 1.29 expressing CD30. Immunostaining for HHV-8 from a cell block was positive. Electron microscopy revealed lymphomalike cells, many in various stages of apoptosis, with large nucleoli and clusters of viruslike particles in the nucleoplasm. A firm diagnosis of PEL can be established by the examination of cells from the lymphomatous effusion by a combination of cytology, molecular genetics, phenotypic features, immunostaining and electron microscopy. To our knowledge, this is the first case in which immunostaining for anti-HHV-8 monoclonal antibodies was used to support the diagnosis.

  16. Next-generation sequencing of microRNAs in primary human polarized macrophages

    Directory of Open Access Journals (Sweden)

    Viviana Cobos Jiménez

    2014-12-01

    Full Text Available Macrophages are important for mounting inflammatory responses to tissue damage or infection by invading pathogens, and therefore modulation of their cellular functions is essential for the success of the immune system as well as for maintaining tissue homeostasis. Small non-coding RNAs are important regulatory elements of gene expression and microRNAs are the most widely known to be fundamental for the proper development of cells of the immune system. Macrophages can exhibit different phenotypes, depending on the cytokine environment they encounter in the affected tissues. We have analyzed the microRNA expression profiles during maturation of human primary monocytes into macrophages and polarization by pro- or anti-inflammatory cytokines. Here we describe the analysis of next-generation sequencing data deposited in EMBL–EBI ArrayExpress under accession number E-MTAB-1969 and associated with the study published by Cobos Jiménez and collaborators in Physiological Genomics in 2014 (1. The data presented here contributes to our understanding of microRNA expression profiles in human monocytes and macrophages and will also serve as a resource for novel microRNAs and other small RNA species expressed in these cells.

  17. Cellular Mechanics of Primary Human Cervical Fibroblasts: Influence of Progesterone and a Pro-inflammatory Cytokine.

    Science.gov (United States)

    Shukla, Vasudha; Barnhouse, Victoria; Ackerman, William E; Summerfield, Taryn L; Powell, Heather M; Leight, Jennifer L; Kniss, Douglas A; Ghadiali, Samir N

    2018-01-01

    The leading cause of neonatal mortality, pre-term birth, is often caused by pre-mature ripening/opening of the uterine cervix. Although cervical fibroblasts play an important role in modulating the cervix's extracellular matrix (ECM) and mechanical properties, it is not known how hormones, i.e., progesterone, and pro-inflammatory insults alter fibroblast mechanics, fibroblast-ECM interactions and the resulting changes in tissue mechanics. Here we investigate how progesterone and a pro-inflammatory cytokine, IL-1β, alter the biomechanical properties of human cervical fibroblasts and the fibroblast-ECM interactions that govern tissue-scale mechanics. Primary human fibroblasts were isolated from non-pregnant cervix and treated with estrogen/progesterone, IL-1β or both. The resulting changes in ECM gene expression, matrix remodeling, traction force generation, cell-ECM adhesion and tissue contractility were monitored. Results indicate that IL-1β induces a significant reduction in traction force and ECM adhesion independent of pre-treatment with progesterone. These cell level effects altered tissue-scale mechanics where IL-1β inhibited the contraction of a collagen gel over 6 days. Interestingly, progesterone treatment alone did not modulate traction forces or gel contraction but did result in a dramatic increase in cell-ECM adhesion. Therefore, the protective effect of progesterone may be due to altered adhesion dynamics as opposed to altered ECM remodeling.

  18. Development of a hemicornea from human primary cell cultures for pharmacotoxicology testing.

    Science.gov (United States)

    Builles, N; Bechetoille, N; Justin, V; André, V; Barbaro, V; Di Iorio, E; Auxenfans, C; Hulmes, D J S; Damour, O

    2007-07-01

    We report the reconstruction and characterization of a hemicornea (epithelialized stroma), using primary human cells, for use in research and as an alternative to the use of animals in pharmacotoxicology testing. To create a stromal equivalent, keratocytes from human corneas were cultured in collagen-glycosaminoglycan-chitosan foams. Limbal stem cell-derived epithelial cells were seeded on top of these, giving rise to hemi-corneas. The epithelium appeared morphologically similar to its physiological counterpart, as shown by the basal cell expression of p63 isoforms including, in some cases, the stem cell marker p63DeltaNalpha, and the expression of keratin 3 and 14-3-3sigma in the upper cell layers. In addition, the cuboidal basal epithelial cells were anchored to a basement membrane containing collagen IV, laminin 5, and hemidesmosomes. In the stromal part, the keratocytes colonized the porous scaffold, formed a network of interconnecting cells, and synthesized an ultrastructurally organized extracellular matrix (ECM) containing collagen types I, V, and VI. Electron microscopy showed the newly synthesized collagen fibrils to have characteristic periodic striations, with diameters and interfibril spacings similar to those found in natural corneas. Compared to existing models for corneal pharmacotoxicology testing, this new model more closely approaches physiological conditions by including the inducing effects of mesenchyme and cell-matrix interactions on epithelial cell morphogenesis.

  19. Primary cilia are sensors of electrical field stimulation to induce osteogenesis of human adipose-derived stem cells.

    Science.gov (United States)

    Cai, Shaobo; Bodle, Josephine C; Mathieu, Pattie S; Amos, Alison; Hamouda, Mehdi; Bernacki, Susan; McCarty, Greg; Loboa, Elizabeth G

    2017-01-01

    In this study, we report for the first time that the primary cilium acts as a crucial sensor for electrical field stimulation (EFS)-enhanced osteogenic response in osteoprogenitor cells. In addition, primary cilia seem to functionally modulate effects of EFS-induced cellular calcium oscillations. Primary cilia are organelles that have recently been implicated to play a crucial sensor role for many mechanical and chemical stimuli on stem cells. Here, we investigate the role of primary cilia in EFS-enhanced osteogenic response of human adipose-derived stem cells (hASCs) by knocking down 2 primary cilia structural proteins, polycystin-1 and intraflagellar protein-88. Our results indicate that structurally integrated primary cilia are required for detection of electrical field signals in hASCs. Furthermore, by measuring changes of cytoplasmic calcium concentration in hASCs during EFS, our findings also suggest that primary cilia may potentially function as a crucial calcium-signaling nexus in hASCs during EFS.-Cai, S., Bodle, J. C., Mathieu, P. S., Amos, A., Hamouda, M., Bernacki, S., McCarty, G., Loboa, E. G. Primary cilia are sensors of electrical field stimulation to induce osteogenesis of human adipose-derived stem cells. © FASEB.

  20. Foxp3-dependent transformation of human primary CD4+ T lymphocytes by the retroviral protein tax.

    Science.gov (United States)

    Chen, Li; Liu, Dan; Zhang, Yang; Zhang, Huan; Cheng, Hua

    2015-10-23

    The retroviral Tax proteins of human T cell leukemia virus type 1 and 2 (HTLV-1 and -2) are highly homologous viral transactivators. Both viral proteins can immortalize human primary CD4+ memory T cells, but when expressed alone they rarely transform T cells. In the present study, we found that the Tax proteins displayed a differential ability to immortalize human CD4+Foxp3+ T cells with characteristic expression of CTLA-4 and GITR. Because epidermal growth factor receptor (EGFR) was reportedly expressed and activated in a subset of CD4+Foxp3+ T cells, we introduced an activated EGFR into Tax-immortalized CD4+Foxp3+ T cells. We observed that these modified cells were grown independently of exogenous IL-2, correlating with a T cell transformation phenotype. In Tax-immortalized CD4+Foxp3- T cells, ectopic expression of Foxp3 was a prerequisite for Tax transformation of T cells. Accordingly, treatment of the transformed T cells with erlotinib, a selective inhibitor of EGFR, induced degradation of EGFR in lysosome, consequently causing T cell growth inhibition. Further, we identified autophagy as a crucial cellular survival pathway for the transformed T cells. Silencing key autophagy molecules including Beclin1, Atg5 and PI3 kinase class III (PI3KC3) resulted in drastic impairment of T cell growth. Our data, therefore, unveiled a previously unidentified role of Foxp3 in T cell transformation, providing a molecular basis for HTLV-1 transformation of CD4+Foxp3+ T cells. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. Effects of Rosmarinus officinalis extract on human primary omental preadipocytes and adipocytes.

    Science.gov (United States)

    Stefanon, Bruno; Pomari, Elena; Colitti, Monica

    2015-07-01

    The prevalence of obesity is increasing all over the world. Although it has been shown that natural substances influence fat metabolism, little is known about the effect on cellular and molecular mechanisms in human. In this in vitro study, the activity of Rosmarinus officinalis (RO) standardized extract in modulating human primary visceral preadipocytes differentiation, lipolysis, and apoptosis was investigated. Moreover, gene expression of key adipogenesis modulators and microRNAs-seq were evaluated. Preadipocytes treated with RO extract significantly reduced triglyceride incorporation during maturation in a dose-dependent manner without affecting cell viability. In addition, RO extract stimulated lipolytic activity in differentiating preadipocytes and mature adipocytes in treated cells compared to controls. Differentiating preadipocytes incubated in the presence of RO extract showed a decreased expression of cell cycle genes such as cyclin D1, cyclin-dependent kinase 4, cyclin-dependent kinase inhibitor 1A (p21, Cip1) and an increased expression of GATA binding protein 3, wingless-type MMTV integration site family, member 3A mRNA levels. Recent studies have demonstrated that some phytochemicals alter the expression of specific genes and microRNAs that play a fundamental role in the pathogenesis of obesity and related diseases. Interestingly, genes modulated in RO-treated cells were found to be validated miRNAs targets, such as let-7f-1, miR-17, and miR-143. The results indicated that RO extract modulates human adipocyte differentiation and significantly interferes with adipogenesis and lipid metabolism, supporting its interest as dietary supplement. © 2015 by the Society for Experimental Biology and Medicine.

  2. Elucidating mechanisms of toxicity using phenotypic data from primary human cell systems--a chemical biology approach for thrombosis-related side effects.

    Science.gov (United States)

    Berg, Ellen L; Polokoff, Mark A; O'Mahony, Alison; Nguyen, Dat; Li, Xitong

    2015-01-05

    Here we describe a chemical biology approach for elucidating potential toxicity mechanisms for thrombosis-related side effects. This work takes advantage of a large chemical biology data set comprising the effects of known, well-characterized reference agents on the cell surface levels of tissue factor (TF) in a primary human endothelial cell-based model of vascular inflammation, the BioMAP® 3C system. In previous work with the Environmental Protection Agency (EPA) for the ToxCast™ program, aryl hydrocarbon receptor (AhR) agonists and estrogen receptor (ER) antagonists were found to share an usual activity, that of increasing TF levels in this system. Since human exposure to compounds in both chemical classes is associated with increased incidence of thrombosis-related side effects, we expanded this analysis with a large number of well-characterized reference compounds in order to better understand the underlying mechanisms. As a result, mechanisms for increasing (AhR, histamine H1 receptor, histone deacetylase or HDAC, hsp90, nuclear factor kappa B or NFκB, MEK, oncostatin M receptor, Jak kinase, and p38 MAPK) and decreasing (vacuolar ATPase or V-ATPase) and mTOR) TF expression levels were uncovered. These data identify the nutrient, lipid, bacterial, and hypoxia sensing functions of autophagy as potential key regulatory points controlling cell surface TF levels in endothelial cells and support the mechanistic hypothesis that these functions are associated with thrombosis-related side effects in vivo.

  3. Exploiting high-throughput screens to optimize Adeno-Associated Viral Vectors for gene transfer into primary human keratinocytes

    OpenAIRE

    Sallach, Jessica

    2014-01-01

    Chronic non-healing wounds such as diabetic ulcers or burns represent a devastating health problem with significant clinical, physical and social implications. The healing can be frustrating and painful for patients. The difficult healing process requires advanced therapeutic strategies such as the use of primary human keratinocytes (HK) as autologous transplants, which may be considered for clinical use. To improve engraftment or to introduce therapeutic genes into primary HK, efficient and ...

  4. Efficacy of Various Intracanal Medicaments in Human Primary Teeth with Necrotic Pulp against Candida Biofilms: An in vivo Study

    OpenAIRE

    Paikkatt, Jophie V; Sreedharan, Sheela; Philomina, Beena; Kannan, VP; Santhakumar, Madhu; Kumar, TV Anupam

    2017-01-01

    Background Candida has been associated with cases of secondary and persistent root canal infections. The purpose of this study was to evaluate and compare the effectiveness of commonly used intracanal medicament against Candida biofilms found in root canals of human primary teeth with necrotic pulp. Materials and methods Pulp canals of 45 single-rooted primary maxillary anterior teeth with pulp necrosis in 34 children were included in the study. They were divided into three groups of 15 sampl...

  5. Silicon microfabricated beam expander

    Energy Technology Data Exchange (ETDEWEB)

    Othman, A., E-mail: aliman@ppinang.uitm.edu.my; Ibrahim, M. N.; Hamzah, I. H.; Sulaiman, A. A. [Faculty of Electrical Engineering, Universiti Teknologi MARA Malaysia, 40450, Shah Alam, Selangor (Malaysia); Ain, M. F. [School of Electrical and Electronic Engineering, Engineering Campus, Universiti Sains Malaysia, Seri Ampangan, 14300,Nibong Tebal, Pulau Pinang (Malaysia)

    2015-03-30

    The feasibility design and development methods of silicon microfabricated beam expander are described. Silicon bulk micromachining fabrication technology is used in producing features of the structure. A high-precision complex 3-D shape of the expander can be formed by exploiting the predictable anisotropic wet etching characteristics of single-crystal silicon in aqueous Potassium-Hydroxide (KOH) solution. The beam-expander consist of two elements, a micromachined silicon reflector chamber and micro-Fresnel zone plate. The micro-Fresnel element is patterned using lithographic methods. The reflector chamber element has a depth of 40 µm, a diameter of 15 mm and gold-coated surfaces. The impact on the depth, diameter of the chamber and absorption for improved performance are discussed.

  6. TRIPLE ACTION PALATE EXPANDER

    Directory of Open Access Journals (Sweden)

    Svetlana Yordanova

    2011-09-01

    Full Text Available Malocclusion correction essentially involves expansion of the maxilla, protrusion of anterior teeth and opening the bite. Expansion is often the stage preceding the treatment with fixed appliances. The elevation of the occlusion using accomplished with different devices (bite planes -fixed or removable, composite material on the occlusall surface of molars carries the risk of breaking or debonding them.The present article proposes an expanding appliance with triple action as a therapeutic means of choice in an orthodontic treatment with fixed appliances. The expander can simultaneously be used to protrude upper teeth, to expand the upper jaw and disarticulate the occlusion. It can be easily fabricated in clinical conditions, causes no discomfort and does not hamper oral hygiene because it can be removed and cleaned.

  7. In situ identification of CD44+/CD24- cancer cells in primary human breast carcinomas.

    Directory of Open Access Journals (Sweden)

    Giuseppe Perrone

    Full Text Available Breast cancer cells with the CD44+/CD24- phenotype have been reported to be tumourigenic due to their enhanced capacity for cancer development and their self-renewal potential. The identification of human tumourigenic breast cancer cells in surgical samples has recently received increased attention due to the implications for prognosis and treatment, although limitations exist in the interpretation of these studies. To better identify the CD44+/CD24- cells in routine surgical specimens, 56 primary breast carcinoma cases were analysed by immunofluorescence and confocal microscopy, and the results were compared using flow cytometry analysis to correlate the amount and distribution of the CD44+/CD24- population with clinicopathological features. Using these methods, we showed that the breast carcinoma cells displayed four distinct sub-populations based on the expression pattern of CD44 and CD24. The CD44+/CD24- cells were found in 91% of breast tumours and constituted an average of 6.12% (range, 0.11%-21.23% of the tumour. A strong correlation was found between the percentage of CD44+/CD24- cells in primary tumours and distant metastasis development (p = 0.0001; in addition, there was an inverse significant association with ER and PGR status (p = 0.002 and p = 0.001, respectively. No relationship was evident with tumour size (T and regional lymph node (N status, differentiation grade, proliferative index or HER2 status. In a multivariate analysis, the percentage of CD44+/CD24- cancer cells was an independent factor related to metastasis development (p = 0.004. Our results indicate that confocal analysis of fluorescence-labelled breast cancer samples obtained at surgery is a reliable method to identify the CD44+/CD24- tumourigenic cell population, allowing for the stratification of breast cancer patients into two groups with substantially different relapse rates on the basis of CD44+/CD24- cell percentage.

  8. Low-level laser therapy as an alternative for pulpotomy in human primary teeth.

    Science.gov (United States)

    Marques, Nádia Carolina Teixeira; Neto, Natalino Lourenço; Rodini, Camila de Oliveira; Fernandes, Ana Paula; Sakai, Vivien Thiemy; Machado, Maria Aparecida Andrade Moreira; Oliveira, Thais Marchini

    2015-09-01

    This study aimed to evaluate the effects of low-level laser therapy (LLLT) on pulpal response of primary teeth. Twenty mandibular primary molars were randomly divided into the following groups: group I Buckley's formocresol (diluted at 1:5), group II calcium hydroxide, group III LLLT + zinc oxide/eugenol, and group IV LLLT + calcium hydroxide. LLLT parameters were set at 660-nm wavelength, 10-mW power output, and 2.5 J/cm(2) energy density for 10 s in continuous mode (InGaAlP laser, Twin Laser®, MMOptics, Sao Carlos, Sao Paulo, Brazil). The teeth were extracted at the regular exfoliation period. The dentin-pulp complex was graded by an established histopathological score system. Statistical analysis was performed by Kruskal-Wallis and chi-square test. The histopathological assessment revealed statistically significant differences among groups (P < 0.05). The lowest degree of pulpal inflammation was present in LLLT + calcium hydroxide (P = 0.0296). Calcium hydroxide showed the highest rate of hard tissue barrier (P = 0.0033), odontoblastic layer (P = 0.0033), and dense collagen fibers (P = 0.0095). On the other hand, formocresol showed the highest incidence of internal resorption (P = 0.0142). Based on this study, low-level laser therapy preceding the use of calcium hydroxide exhibited satisfactory results on pulp tissue healing. However, further clinical studies on human teeth with long-term follow-up are needed to test the low-level laser therapy efficacy.

  9. Efficient Transduction of Human and Rhesus Macaque Primary T Cells by a Modified Human Immunodeficiency Virus Type 1-Based Lentiviral Vector.

    Science.gov (United States)

    He, Huan; Xue, Jing; Wang, Weiming; Liu, Lihong; Ye, Chaobaihui; Cong, Zhe; Kimata, Jason T; Qin, Chuan; Zhou, Paul

    2017-03-01

    Human immunodeficiency virus type 1 (HIV-1)-based lentiviral vectors efficiently transduce genes to human, but not rhesus, primary T cells and hematopoietic stem cells (HSCs). The poor transduction of HIV-1 vectors to rhesus cells is mainly due to species-specific restriction factors such as rhesus TRIM5α. Previously, several strategies to modify HIV-1 vectors were developed to overcome rhesus TRIM5α restriction. While the modified HIV-1 vectors efficiently transduce rhesus HSCs, they remain suboptimal for rhesus primary T cells. Recently, HIV-1 variants that encode combinations of LNEIE mutations in capsid (CA) protein and SIVmac239 Vif were found to replicate efficiently in rhesus primary T cells. Thus, the present study tested whether HIV-1 vectors packaged by a packaging construct containing these CA substitutions could efficiently transduce both human and rhesus primary CD4 T cells. To accomplish this, LNEIE mutations were made in the packaging construct CEMΔ8.9, and recombinant HIV-1 vectors packaged by Δ8.9 WT or Δ8.9 LNEIE were generated. Transduction rates, CA stability, and vector integration in CEMss-CCR5 and CEMss-CCR5-rhTRIM5α/green fluorescent protein cells, as well as transduction rates in human and rhesus primary CD4 T cells by Δ8.9 WT or Δ8.9 LNEIE-packaged HIV-1 vectors, were compared. Finally, the influence of rhesus TRIM5α variations in transduction rates to primary CD4 T cells from a cohort of 37 Chinese rhesus macaques was studied. While it maintains efficient transduction for human T-cell line and primary CD4 T cells, Δ8.9 LNEIE-packaged HIV-1 vector overcomes rhesus TRIM5α-mediated CA degradation, resulting in significantly higher transduction efficiency of rhesus primary CD4 T cells than Δ8.9 WT-packaged HIV-1 vector. Rhesus TRIM5α variations strongly influence transduction efficiency of rhesus primary CD4 T cells by both Δ8.9 WT or Δ8.9 LNEIE-packaged HIV-1 vectors. Thus, it is concluded that Δ8.9 LNEIE-packaged HIV-1

  10. Primary mesenchymal stem cells in human transplanted lungs are CD90/CD105 perivascularly located tissue-resident cells

    DEFF Research Database (Denmark)

    Rolandsson, Sara; Andersson Sjöland, Annika; Brune, Jan C

    2014-01-01

    BACKGROUND: Mesenchymal stem cells (MSC) have not only been implicated in the development of lung diseases, but they have also been proposed as a future cell-based therapy for lung diseases. However, the cellular identity of the primary MSC in human lung tissues has not yet been reported. This st......BACKGROUND: Mesenchymal stem cells (MSC) have not only been implicated in the development of lung diseases, but they have also been proposed as a future cell-based therapy for lung diseases. However, the cellular identity of the primary MSC in human lung tissues has not yet been reported...

  11. Differential susceptibility of primary cultured human skin cells to hypericin PDT in an in vitro model.

    Science.gov (United States)

    Popovic, A; Wiggins, T; Davids, L M

    2015-08-01

    Skin cancer is the most common cancer worldwide, and its incidence rate in South Africa is increasing. Photodynamic therapy (PDT) has been shown to be an effective treatment modality, through topical administration, for treatment of non-melanoma skin cancers. Our group investigates hypericin-induced PDT (HYP-PDT) for the treatment of both non-melanoma and melanoma skin cancers. However, a prerequisite for effective cancer treatments is efficient and selective targeting of the tumoral cells with minimal collateral damage to the surrounding normal cells, as it is well established that cancer therapies have bystander effects on normal cells in the body, often causing undesirable side effects. The aim of this study was to investigate the cellular and molecular effects of HYP-PDT on normal primary human keratinocytes (Kc), melanocytes (Mc) and fibroblasts (Fb) in an in vitro tissue culture model which represented both the epidermal and dermal cellular compartments of human skin. Cell viability analysis revealed a differential cytotoxic response to a range of HYP-PDT doses in all the human skin cell types, showing that Fb (LD50=1.75μM) were the most susceptible to HYP-PDT, followed by Mc (LD50=3.5μM) and Kc (LD50>4μM HYP-PDT) These results correlated with the morphological analysis which displayed distinct morphological changes in Fb and Mc, 24h post treatment with non-lethal (1μM) and lethal (3μM) doses of HYP-PDT, but the highest HYP-PDT doses had no effect on Kc morphology. Fluorescent microscopy displayed cytoplasmic localization of HYP in all the 3 skin cell types and additionally, HYP was excluded from the nuclei in all the cell types. Intracellular ROS levels measured in Fb at 3μM HYP-PDT, displayed a significant 3.8 fold (pPDT. These results depict a differential response to HYP-PDT by different human skin cells thus highlighting the efficacy and indeed, the potential bystander effect of if administered in vivo. This study contributes toward our knowledge

  12. Expanding specificity of class I restricted CD8+ T cells for viral epitopes following multiple inoculations of swine with a human adenovirus vectored foot-and-mouth disease virus (FMDV) vaccine

    DEFF Research Database (Denmark)

    Pedersen, Lasse E.; Patch, Jared R; Kenney, Mary

    2016-01-01

    class I major histocompatibility complex (MHC) tetramer staining. We also showed that a modified replication defective human adenovirus 5 vector expressing the FMDV structural proteins (Ad5-FMDV-T vaccine) targets the induction of a CD8(+) CTL response with a minimal humoral response. In this report, we...... show that the specificity of the CD8(+) T cell response to Ad5-FMDV-T varies between cohorts of genetically identical animals. Further, we demonstrate epitope specificity of CD8(+) T cells expands following multiple immunizations with this vaccine....

  13. P2X7Rs are involved in cell death, growth and cellular signaling in primary human osteoblasts

    DEFF Research Database (Denmark)

    Agrawal, Ankita; Henriksen, Zanne; Syberg, Susanne

    2017-01-01

    The ionotropic ATP-gated P2X7 receptor (P2X7R) is involved in the regulation of many physiological functions including bone metabolism. Several studies on osteoblasts from rodents and human osteoblast-like cell lines have addressed the expression and function of P2X7R on these bone-forming cells...... however; its role in human primary osteoblasts has not yet been reported. The aim of this study was to assess the expression of the P2X7R in bone marrow-derived stromal cells and in primary human trabecular osteoblasts and to determine the function in bone formation and cell signaling. We report...... that osteoblasts derived from human trabecular explants express a functional P2X7R capable of agonist-induced increase in intracellular calcium concentration and a positive permeability to fluorescent dyes. These osteoblasts are fully differentiated cells with alkaline phosphatase activity and the ability to form...

  14. Human Pol II promoter recognition based on primary sequences and free energy of dinucleotides

    Directory of Open Access Journals (Sweden)

    Yu Zu-Guo

    2008-02-01

    Full Text Available Abstract Background Promoter region plays an important role in determining where the transcription of a particular gene should be initiated. Computational prediction of eukaryotic Pol II promoter sequences is one of the most significant problems in sequence analysis. Existing promoter prediction methods are still far from being satisfactory. Results We attempt to recognize the human Pol II promoter sequences from the non-promoter sequences which are made up of exon and intron sequences. Four methods are used: two kinds of multifractal analysis performed on the numeric sequences obtained from the dinucleotide free energy, Z curve analysis and global descriptor of the promoter/non-promoter primary sequences. A total of 141 parameters are extracted from these methods and categorized into seven groups (methods. They are used to generate certain spaces and then each promoter/non-promoter sequence is represented by a point in the corresponding space. All the 120 possible combinations of the seven methods are tested. Based on Fisher's linear discriminant algorithm, with a relatively smaller number of parameters (96 and 117, we get satisfactory discriminant accuracies. Particularly, in the case of 117 parameters, the accuracies for the training and test sets reach 90.43% and 89.79%, respectively. A comparison with five other existing methods indicates that our methods have a better performance. Using the global descriptor method (36 parameters, 17 of the 18 experimentally verified promoter sequences of human chromosome 22 are correctly identified. Conclusion The high accuracies achieved suggest that the methods of this paper are useful for understanding the difficult problem of promoter prediction.

  15. Human Pol II promoter recognition based on primary sequences and free energy of dinucleotides.

    Science.gov (United States)

    Yang, Jian-Yi; Zhou, Yu; Yu, Zu-Guo; Anh, Vo; Zhou, Li-Qian

    2008-02-24

    Promoter region plays an important role in determining where the transcription of a particular gene should be initiated. Computational prediction of eukaryotic Pol II promoter sequences is one of the most significant problems in sequence analysis. Existing promoter prediction methods are still far from being satisfactory. We attempt to recognize the human Pol II promoter sequences from the non-promoter sequences which are made up of exon and intron sequences. Four methods are used: two kinds of multifractal analysis performed on the numeric sequences obtained from the dinucleotide free energy, Z curve analysis and global descriptor of the promoter/non-promoter primary sequences. A total of 141 parameters are extracted from these methods and categorized into seven groups (methods). They are used to generate certain spaces and then each promoter/non-promoter sequence is represented by a point in the corresponding space. All the 120 possible combinations of the seven methods are tested. Based on Fisher's linear discriminant algorithm, with a relatively smaller number of parameters (96 and 117), we get satisfactory discriminant accuracies. Particularly, in the case of 117 parameters, the accuracies for the training and test sets reach 90.43% and 89.79%, respectively. A comparison with five other existing methods indicates that our methods have a better performance. Using the global descriptor method (36 parameters), 17 of the 18 experimentally verified promoter sequences of human chromosome 22 are correctly identified. The high accuracies achieved suggest that the methods of this paper are useful for understanding the difficult problem of promoter prediction.

  16. Zika Virus Persistently Infects and Is Basolaterally Released from Primary Human Brain Microvascular Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Megan C. Mladinich

    2017-07-01

    Full Text Available Zika virus (ZIKV is a mosquito-borne Flavivirus that has emerged as the cause of encephalitis and fetal microencephaly in the Americas. ZIKV uniquely persists in human bodily fluids for up to 6 months, is sexually transmitted, and traverses the placenta and the blood-brain barrier (BBB to damage neurons. Cells that support persistent ZIKV replication and mechanisms by which ZIKV establishes persistence remain enigmatic but central to ZIKV entry into protected neuronal compartments. The endothelial cell (EC lining of capillaries normally constrains transplacental transmission and forms the BBB, which selectively restricts access of blood constituents to neurons. We found that ZIKV (strain PRVABC59 persistently infects and continuously replicates in primary human brain microvascular ECs (hBMECs, without cytopathology, for >9 days and following hBMEC passage. ZIKV did not permeabilize hBMECs but was released basolaterally from polarized hBMECs, suggesting a direct mechanism for ZIKV to cross the BBB. ZIKV-infected hBMECs were rapidly resistant to alpha interferon (IFN-α and transiently induced, but failed to secrete, IFN-β and IFN-λ. Global transcriptome analysis determined that ZIKV constitutively induced IFN regulatory factor 7 (IRF7, IRF9, and IFN-stimulated genes (ISGs 1 to 9 days postinfection, despite persistently replicating in hBMECs. ZIKV constitutively induced ISG15, HERC5, and USP18, which are linked to hepatitis C virus (HCV persistence and IFN regulation, chemokine CCL5, which is associated with immunopathogenesis, as well as cell survival factors. Our results reveal that hBMECs act as a reservoir of persistent ZIKV replication, suggest routes for ZIKV to cross hBMECs into neuronal compartments, and define novel mechanisms of ZIKV persistence that can be targeted to restrict ZIKV spread.

  17. Differential Expression Profile of lncRNAs from Primary Human Hepatocytes Following DEET and Fipronil Exposure

    Directory of Open Access Journals (Sweden)

    Robert D. Mitchell III

    2017-10-01

    Full Text Available While the synthesis and use of new chemical compounds is at an all-time high, the study of their potential impact on human health is quickly falling behind, and new methods are needed to assess their impact. We chose to examine the effects of two common environmental chemicals, the insect repellent N,N-diethyl-m-toluamide (DEET and the insecticide fluocyanobenpyrazole (fipronil, on transcript levels of long non-protein coding RNAs (lncRNAs in primary human hepatocytes using a global RNA-Seq approach. While lncRNAs are believed to play a critical role in numerous important biological processes, many still remain uncharacterized, and their functions and modes of action remain largely unclear, especially in relation to environmental chemicals. RNA-Seq showed that 100 µM DEET significantly increased transcript levels for 2 lncRNAs and lowered transcript levels for 18 lncRNAs, while fipronil at 10 µM increased transcript levels for 76 lncRNAs and decreased levels for 193 lncRNAs. A mixture of 100 µM DEET and 10 µM fipronil increased transcript levels for 75 lncRNAs and lowered transcript levels for 258 lncRNAs. This indicates a more-than-additive effect on lncRNA transcript expression when the two chemicals were presented in combination versus each chemical alone. Differentially expressed lncRNA genes were mapped to chromosomes, analyzed by proximity to neighboring protein-coding genes, and functionally characterized via gene ontology and molecular mapping algorithms. While further testing is required to assess the organismal impact of changes in transcript levels, this initial analysis links several of the dysregulated lncRNAs to processes and pathways critical to proper cellular function, such as the innate and adaptive immune response and the p53 signaling pathway.

  18. Improving Primary Care with Human-Centered Design and Partnership-Based Leadership

    Directory of Open Access Journals (Sweden)

    May-Lynn Andresen

    2017-06-01

    Full Text Available Objective: The purpose of this quality improvement project was to empower and activate first-line staff (FLS to improve the six-month depression remission rate in a primary care clinic. Background: Lack of workforce engagement has been identified as an emerging national problem in health care and health care leaders have urged practice redesign to foster the Triple Aim of improved population health, improved care experience, and reduced cost of care (Berwick et al., 2008. Depression is difficult to manage and often exacerbates chronic illnesses and shortens lifespans, yet despite known effective treatments, six-month remission rates are low and care practices are often inadequate. Engaging in empowering leadership behaviors has demonstrated improvement in motivation, work outcomes, and empowerment in various industry settings across the world. Core approaches include: enhancing staff self-determination, encouraging participation in decision-making, and ensuring that staff have the knowledge and tools to achieve their performance goals, in addition to leadership communications that increase confidence in staff’s potential to perform at high levels, and their recognition that their efforts have an impact on improving organizational effectiveness. Methods: In this outpatient setting, care was siloed, staff were disengaged and a hierarchical paradigm was evident. Human-centered design principles were employed to intensively explore stakeholders’ experiences and to deeply engage end users in improving depression remission rates by creating, participating, and partnering in solutions. Leadership was educated in and deployed empowering leadership behaviors, which were synergistic with design thinking, and fostered empowerment. Results: Pre- and post-surveys demonstrated statistically significant improvement in empowerment. The six-month depression remission rate increased 167%, from 7.3% (N=261 to 19.4% (N=247. Conclusion: The convergence of

  19. α1-Antitrypsin reduces rhinovirus infection in primary human airway epithelial cells exposed to cigarette smoke.

    Science.gov (United States)

    Berman, Reena; Jiang, Di; Wu, Qun; Chu, Hong Wei

    2016-01-01

    Human rhinovirus (HRV) infections target airway epithelium and are the leading cause of acute exacerbations of COPD. Cigarette smoke (CS) increases the severity of viral infections, but there is no effective therapy for HRV infection. We determined whether α1-antitrypsin (A1AT) reduces HRV-16 infection in CS-exposed primary human airway epithelial cells. Brushed bronchial epithelial cells from normal subjects and patients diagnosed with COPD were cultured at air-liquid interface to induce mucociliary differentiation. These cells were treated with A1AT or bovine serum albumin for 2 hours and then exposed to air or whole cigarette smoke (WCS) with or without HRV-16 (5×10(4) 50% Tissue Culture Infective Dose [TCID50]/transwell) infection for 24 hours. WCS exposure significantly increased viral load by an average of fivefold and decreased the expression of antiviral genes interferon-λ1, OAS1, and MX1. When A1AT was added to WCS-exposed cells, viral load significantly decreased by an average of 29-fold. HRV-16 infection significantly increased HRV-16 receptor intercellular adhesion molecule-1 messenger RNA expression in air-exposed cells, which was decreased by A1AT. A1AT-mediated reduction of viral load was not accompanied by increased epithelial antiviral gene expression or by inhibiting the activity of 3C protease involved in viral replication or maturation. Our findings demonstrate that A1AT treatment prevents a WCS-induced increase in viral load and for the first time suggest a therapeutic effect of A1AT on HRV infection.

  20. Direction of movement is encoded in the human primary motor cortex.

    Directory of Open Access Journals (Sweden)

    Carolien M Toxopeus

    Full Text Available The present study investigated how direction of hand movement, which is a well-described parameter in cerebral organization of motor control, is incorporated in the somatotopic representation of the manual effector system in the human primary motor cortex (M1. Using functional magnetic resonance imaging (fMRI and a manual step-tracking task we found that activation patterns related to movement in different directions were spatially disjoint within the representation area of the hand on M1. Foci of activation related to specific movement directions were segregated within the M1 hand area; activation related to direction 0° (right was located most laterally/superficially, whereas directions 180° (left and 270° (down elicited activation more medially within the hand area. Activation related to direction 90° was located between the other directions. Moreover, by investigating differences between activations related to movement along the horizontal (0°+180° and vertical (90°+270° axis, we found that activation related to the horizontal axis was located more anterolaterally/dorsally in M1 than for the vertical axis, supporting that activations related to individual movement directions are direction- and not muscle related. Our results of spatially segregated direction-related activations in M1 are in accordance with findings of recent fMRI studies on neural encoding of direction in human M1. Our results thus provide further evidence for a direct link between direction as an organizational principle in sensorimotor transformation and movement execution coded by effector representations in M1.

  1. Zika Virus Persistently Infects and Is Basolaterally Released from Primary Human Brain Microvascular Endothelial Cells

    Science.gov (United States)

    Mladinich, Megan C.; Schwedes, John

    2017-01-01

    ABSTRACT Zika virus (ZIKV) is a mosquito-borne Flavivirus that has emerged as the cause of encephalitis and fetal microencephaly in the Americas. ZIKV uniquely persists in human bodily fluids for up to 6 months, is sexually transmitted, and traverses the placenta and the blood-brain barrier (BBB) to damage neurons. Cells that support persistent ZIKV replication and mechanisms by which ZIKV establishes persistence remain enigmatic but central to ZIKV entry into protected neuronal compartments. The endothelial cell (EC) lining of capillaries normally constrains transplacental transmission and forms the BBB, which selectively restricts access of blood constituents to neurons. We found that ZIKV (strain PRVABC59) persistently infects and continuously replicates in primary human brain microvascular ECs (hBMECs), without cytopathology, for >9 days and following hBMEC passage. ZIKV did not permeabilize hBMECs but was released basolaterally from polarized hBMECs, suggesting a direct mechanism for ZIKV to cross the BBB. ZIKV-infected hBMECs were rapidly resistant to alpha interferon (IFN-α) and transiently induced, but failed to secrete, IFN-β and IFN-λ. Global transcriptome analysis determined that ZIKV constitutively induced IFN regulatory factor 7 (IRF7), IRF9, and IFN-stimulated genes (ISGs) 1 to 9 days postinfection, despite persistently replicating in hBMECs. ZIKV constitutively induced ISG15, HERC5, and USP18, which are linked to hepatitis C virus (HCV) persistence and IFN regulation, chemokine CCL5, which is associated with immunopathogenesis, as well as cell survival factors. Our results reveal that hBMECs act as a reservoir of persistent ZIKV replication, suggest routes for ZIKV to cross hBMECs into neuronal compartments, and define novel mechanisms of ZIKV persistence that can be targeted to restrict ZIKV spread. PMID:28698279

  2. Differential Expression Profile of lncRNAs from Primary Human Hepatocytes Following DEET and Fipronil Exposure.

    Science.gov (United States)

    Mitchell Iii, Robert D; Wallace, Andrew D; Hodgson, Ernest; Roe, R Michael

    2017-10-07

    While the synthesis and use of new chemical compounds is at an all-time high, the study of their potential impact on human health is quickly falling behind, and new methods are needed to assess their impact. We chose to examine the effects of two common environmental chemicals, the insect repellent N,N-diethyl-m-toluamide (DEET) and the insecticide fluocyanobenpyrazole (fipronil), on transcript levels of long non-protein coding RNAs (lncRNAs) in primary human hepatocytes using a global RNA-Seq approach. While lncRNAs are believed to play a critical role in numerous important biological processes, many still remain uncharacterized, and their functions and modes of action remain largely unclear, especially in relation to environmental chemicals. RNA-Seq showed that 100 µM DEET significantly increased transcript levels for 2 lncRNAs and lowered transcript levels for 18 lncRNAs, while fipronil at 10 µM increased transcript levels for 76 lncRNAs and decreased levels for 193 lncRNAs. A mixture of 100 µM DEET and 10 µM fipronil increased transcript levels for 75 lncRNAs and lowered transcript levels for 258 lncRNAs. This indicates a more-than-additive effect on lncRNA transcript expression when the two chemicals were presented in combination versus each chemical alone. Differentially expressed lncRNA genes were mapped to chromosomes, analyzed by proximity to neighboring protein-coding genes, and functionally characterized via gene ontology and molecular mapping algorithms. While further testing is required to assess the organismal impact of changes in transcript levels, this initial analysis links several of the dysregulated lncRNAs to processes and pathways critical to proper cellular function, such as the innate and adaptive immune response and the p53 signaling pathway.

  3. α1-Antitrypsin reduces rhinovirus infection in primary human airway epithelial cells exposed to cigarette smoke

    Directory of Open Access Journals (Sweden)

    Berman R

    2016-06-01

    Full Text Available Reena Berman, Di Jiang, Qun Wu, Hong Wei Chu Department of Medicine, National Jewish Health, Denver, CO, USA Abstract: Human rhinovirus (HRV infections target airway epithelium and are the leading cause of acute exacerbations of COPD. Cigarette smoke (CS increases the severity of viral infections, but there is no effective therapy for HRV infection. We determined whether α1-antitrypsin (A1AT reduces HRV-16 infection in CS-exposed primary human airway epithelial cells. Brushed bronchial epithelial cells from normal subjects and patients diagnosed with COPD were cultured at air–liquid interface to induce mucociliary differentiation. These cells were treated with A1AT or bovine serum albumin for 2 hours and then exposed to air or whole cigarette smoke (WCS with or without HRV-16 (5×104 50% Tissue Culture Infective Dose [TCID50]/transwell infection for 24 hours. WCS exposure significantly increased viral load by an average of fivefold and decreased the expression of antiviral genes interferon-λ1, OAS1, and MX1. When A1AT was added to WCS-exposed cells, viral load significantly decreased by an average of 29-fold. HRV-16 infection significantly increased HRV-16 receptor intercellular adhesion molecule-1 messenger RNA expression in air-exposed cells, which was decreased by A1AT. A1AT-mediated reduction of viral load was not accompanied by increased epithelial antiviral gene expression or by inhibiting the activity of 3C protease involved in viral replication or maturation. Our findings demonstrate that A1AT treatment prevents a WCS-induced increase in viral load and for the first time suggest a therapeutic effect of A1AT on HRV infection. Keywords: α1-antitrypsin, rhinovirus, COPD, cigarette smoke, ICAM-1

  4. Simulation of Human Plasma Concentrations of Thalidomide and Primary 5-Hydroxylated Metabolites Explored with Pharmacokinetic Data in Humanized TK-NOG Mice.

    Science.gov (United States)

    Nishiyama, Sayako; Suemizu, Hiroshi; Shibata, Norio; Guengerich, F Peter; Yamazaki, Hiroshi

    2015-11-16

    Plasma concentrations of thalidomide and primary 5-hydroxylated metabolites including 5,6-dihydroxythalidomide and glutathione (GSH) conjugate(s) were investigated in chimeric mice with highly "humanized" liver cells harboring cytochrome P450 3A5*1. Following oral administration of thalidomide (100 mg/kg), plasma concentrations of GSH conjugate(s) of 5-hydroxythalidomide were higher in humanized mice than in controls. Simulation of human plasma concentrations of thalidomide were achieved with a simplified physiologically based pharmacokinetic model in accordance with reported thalidomide concentrations. The results indicate that the pharmacokinetics in humans of GSH conjugate and/or catechol primary 5-hydroxylated thalidomide contributing in vivo activation can be estimated for the first time.

  5. Novel approach for transient protein expression in primary cultures of human dental pulp-derived cells.

    Science.gov (United States)

    Suguro, Hisashi; Mikami, Yoshikazu; Koshi, Rieko; Ogiso, Bunnai; Watanabe, Eri; Watanabe, Nobukazu; Honda, Masaki J; Asano, Masatake; Komiyama, Kazuo

    2011-08-01

    Transfection is a powerful method for investigating variable biological functions of desired genes. However, the efficiency of transfection into primary cultures of dental pulp-derived cells (DPDC) is low. Therefore, using a recombinant vaccinia virus (vTF7-3), which contains T7 RNA polymerase, we have established a transient protein expression system in DPDCs. In this study, we used the human polymeric immunoglobulin receptor (pIgR) cDNA as a model gene. pIgR expression by the vTF7-3 expression system was confirmed by flow cytometry analysis and Western blotting. Furthermore, exogenous pIgR protein localized at the cell surface in DPDCs and formed a secretory component (SC). This suggests that exogenous pIgR protein expressed by the vTF7-3 expression system acts like endogenous pIgR protein. These results indicate the applicability of the method for cells outgrown from dental pulp tissue. In addition, as protein expression could be detected shortly after transfection (approximately 5h), this experimental system has been used intensely for experiments examining very early steps in protein exocytosis. Copyright © 2011 Elsevier Inc. All rights reserved.

  6. Prophylactic human papillomavirus vaccination and primary prevention of cervical cancer: issues and challenges.

    Science.gov (United States)

    Poljak, M

    2012-10-01

    Two prophylactic human papillomavirus (HPV) vaccines have been recently approved: one quadrivalent and the other a bivalent vaccine. When administered in a three-dose course to HPV-naive individuals, both vaccines exhibited excellent safety profiles and were highly efficacious against targeted clinical endpoints in large-scale international phase III clinical trials. Where coverage has been high for the appropriate target population, a reduction of HPV-related diseases with the shortest incubation periods has already been seen. By March 2012, universal HPV vaccination had been introduced into national vaccination programmes in more than 40 countries, but only in a few low-income and middle-income countries. With the growing market for HPV vaccines and competition between manufacturers, negotiated prices are already beginning to decline although they still remain out of reach of many countries. The great majority of countries are struggling to reach a level of coverage that will have the most impact on cervical cancer rates. Increasing coverage and improving completion of the HPV vaccine schedule, particularly of sexually naive females, is now the most important public-health issue in HPV vaccine efforts. A clear strategy for integrating primary (HPV vaccination) and secondary (screening) cervical cancer prevention must be agreed as soon as possible. Several second-generation prophylactic vaccines are being developed with the aim of resolving some of the limitations of the two current HPV prophylactic vaccines. © 2012 The Author. Clinical Microbiology and Infection © 2012 European Society of Clinical Microbiology and Infectious Diseases.

  7. Enamel matrix derivative promote primary human pulp cell differentiation and mineralization.

    Science.gov (United States)

    Riksen, Elisabeth Aurstad; Landin, Maria A; Reppe, Sjur; Nakamura, Yukio; Lyngstadaas, Ståle Petter; Reseland, Janne E

    2014-05-05

    Enamel matrix derivative (EMD) has been found to induce reactive dentin formation; however the molecular mechanisms involved are unclear. The effect of EMD (5-50 μg/mL) on primary human pulp cells were compared to untreated cells and cells incubated with 10⁻⁸ M dexamethasone (DEX) for 1, 2, 3, 7, and 14 days in culture. Expression analysis using Affymetrix microchips demonstrated that 10 μg/mL EMD regulated several hundred genes and stimulated the gene expression of proteins involved in mesenchymal proliferation and differentiation. Both EMD and DEX enhanced the expression of amelogenin (amel), and the dentinogenic markers dentin sialophosphoprotein (DSSP) and dentin matrix acidic phosphoprotein 1 (DMP1), as well as the osteogenic markers osteocalcin (OC, BGLAP) and collagen type 1 (COL1A1). Whereas, only EMD had effect on alkaline phosphatase (ALP) mRNA expression, the stimulatory effect were verified by enhanced secretion of OC and COL1A from EMD treated cells, and increased ALP activity in cell culture medium after EMD treatment. Increased levels of interleukin-6 (IL-6), interleukin-8 (IL-8), and monocyte chemoattractant proteins (MCP-1) in the cell culture medium were also found. Consequently, the suggested effect of EMD is to promote differentiation of pulp cells and increases the potential for pulpal mineralization to favor reactive dentine formation.

  8. Exposure to ambient ultrafine particulate matter alters the expression of genes in primary human neurons.

    Science.gov (United States)

    Solaimani, Parrisa; Saffari, Arian; Sioutas, Constantinos; Bondy, Stephen C; Campbell, Arezoo

    2017-01-01

    Exposure to ambient particulate matter (PM) has been associated with the onset of neurodevelopmental and neurodegenerative disorders, but the mechanism of toxicity remains unclear. To gain insight into this neurotoxicity, this study sought to examine global gene expression changes caused by exposure to ambient ultrafine PM. Microarray analysis was performed on primary human neurons derived from fetal brain tissue after a 24h exposure to 20μg/mL of ambient ultrafine particles. We found a majority of the changes in noncoding RNAs, which are involved in epigenetic regulation of gene expression, and thereby could impact the expression of several other protein coding gene targets. Although neurons from biologically different lot numbers were used, we found a significant increase in the expression of metallothionein 1A and 1F in all samples after exposure to particulate matter as confirmed by quantitative PCR. These metallothionein 1 proteins are responsible for neuroprotection after exposure to environmental insult but prolonged induction can be toxic. Epidemiological studies have reported that in utero exposure to ultrafine PM not only leads to neurodevelopmental and behavioral abnormalities, but may also predispose the progeny to neurodegenerative disease later in life by genetic imprinting. Our results pinpoint some of the PM-induced genetic changes that may underlie these findings. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. Regulation of Xylosyltransferase I Gene Expression by Interleukin 1β in Human Primary Chondrocyte Cells

    Science.gov (United States)

    Khair, Mostafa; Bourhim, Mustapha; Barré, Lydia; Li, Dong; Netter, Patrick; Magdalou, Jacques; Fournel-Gigleux, Sylvie; Ouzzine, Mohamed

    2013-01-01

    Xylosyltransferase I (XT-I) is an essential enzyme of proteoglycan (PG) biosynthesis pathway catalyzing the initial and rate-limiting step in glycosaminoglycan chain assembly. It plays a critical role in the regulation of PG synthesis in cartilage; however, little is known about underlying mechanism. Here, we provide evidence that, in human primary chondrocytes, IL-1β regulates XT-I gene expression into an early phase of induction and a late phase of down-regulation. Based on promoter deletions, the region up to −850 bp was defined as a major element of XT-I gene displaying both constitutive and IL-1β-regulated promoter activity. Point mutation and signaling analyses revealed that IL-1β-induced promoter activity is achieved through AP-1 response elements and mediated by SAP/JNK and p38 signaling pathways. Transactivation and chromatin immunoprecipitation assays indicated that AP-1 is a potent transactivator of XT-I promoter and that IL-1β-induced activity is mediated through increased recruitment of AP-1 to the promoter. Finally, we show that Sp3 is a repressor of XT-I promoter and bring evidence that the repressive effect of IL-1β during the late phase is mediated through Sp3 recruitment to the promoter. This suggests that modulation of Sp3 in cartilage could prevent IL-1β inhibition of PG synthesis and limit tissue degradation. PMID:23223231

  10. In vitro effects of fungi isolated from equine hooves on primary human keratinocytes.

    Science.gov (United States)

    Apprich, Veronika; Spergser, Joachim; Rosengarten, Renate; Stanek, Christian

    2006-12-01

    The effects of two dermatophytes (Microsporum gypseum and Trichophyton mentagrophytes) and four moulds (Scopulariopsis brevicaulis, Alternaria alternata, Geotrichum candidum and Penicillium spp.) on living keratinocyte cultures were examined in vitro using primary human keratinocytes. Rates of apoptosis of infected cells were determined using a colorimetric TUNEL system which detects the characteristic nuclear DNA fragmentation of apoptotic cells. The cytotoxicity of the individual fungi was tested by quantitatively measuring cytosolic enzyme lactate dehydrogenase, released upon cell lysis, in culture supernatants. Additionally, the cell structures within the infected keratinocytes in cultures were examined by scanning electron microscopy. All of the fungi exhibited high cytotoxicity, whereas the development of only the two dermatophytes and the mould Scopulariopsis brevicaulis resulted in distinctly increased apoptosis. Electron microscopy showed that all fungi studied caused similar alterations in the cell structure, with Microsporum gypseum being the most harmful. Increasing loss of cell adhesion as a consequence of a decreasing number of reticulating cell appendices and a reduced cell plasticity were the most evident alterations.

  11. A qualitative study on human resources for primary health care in Iran.

    Science.gov (United States)

    Nekoei Moghadam, Mahmood; Amiresmaili, Mohammadreza; Sadeghi, Vahideh; Zeinalzadeh, Ali-Hossein; Tupchi, Mahmud; Parva, Sahar

    2017-02-03

    According to World Bank Group report, while Primary Health Care (PHC) services in Iran were appropriate to the needs of the population in the late 1970s and 1980s, the changing burden of diseases and shifting demand patterns have rendered the existing PHC system no longer suitable to meet current and emergent needs. Based on previous studies, one of the main PHC challenges in Iran relates to human resources issues. This study was conducted in 2012 at 3 scales of local (Tabriz city), provincial (East Azerbaijan), and national levels. Two qualitative methods were used in the study: free-focus group discussions and in-depth interviews. Framework analysis was used to analyze collected data. Categories of analysis were developed using framework analysis approach, and main themes were emerged. Four themes were developed and finalized out of focus group discussions and interviews: availability of health workers in PHC, competency, PHC health workers' motivation, and PHC managerial issues. Based on findings and issues discovered in this study, several suggestions can be made, including development, implementation, and evaluation of needs assessment strategies for various vocational posts, promotion of educational courses and PHC-based training, strengthening of relationships and coordination between practical and scientific bodies, application of incentive programs in PHC, and strengthening of system management capacity through use of qualified managers. Copyright © 2017 John Wiley & Sons, Ltd.

  12. Pupil size directly modulates the feedforward response in human primary visual cortex independently of attention.

    Science.gov (United States)

    Bombeke, Klaas; Duthoo, Wout; Mueller, Sven C; Hopf, Jens-Max; Boehler, C Nico

    2016-02-15

    Controversy revolves around the question of whether psychological factors like attention and emotion can influence the initial feedforward response in primary visual cortex (V1). Although traditionally, the electrophysiological correlate of this response in humans (the C1 component) has been found to be unaltered by psychological influences, a number of recent studies have described attentional and emotional modulations. Yet, research into psychological effects on the feedforward V1 response has neglected possible direct contributions of concomitant pupil-size modulations, which are known to also occur under various conditions of attentional load and emotional state. Here we tested the hypothesis that such pupil-size differences themselves directly affect the feedforward V1 response. We report data from two complementary experiments, in which we used procedures that modulate pupil size without differences in attentional load or emotion while simultaneously recording pupil-size and EEG data. Our results confirm that pupil size indeed directly influences the feedforward V1 response, showing an inverse relationship between pupil size and early V1 activity. While it is unclear in how far this effect represents a functionally-relevant adaptation, it identifies pupil-size differences as an important modulating factor of the feedforward response of V1 and could hence represent a confounding variable in research investigating the neural influence of psychological factors on early visual processing. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Effect of Cyclosporin A and Angiotensin II on cytosolic calcium levels in primary human gingival fibroblasts

    Directory of Open Access Journals (Sweden)

    Ajitkumar Supraja

    2016-01-01

    Full Text Available Background: To evaluate the effect of Cyclosporin A (CsA and angiotensin II (Ang II on cytosolic calcium levels in cultured human gingival fibroblasts (HGFs. Materials and Methods: Healthy gingival samples from six volunteers were obtained, and primary HGFs were cultured. Cell viability and proliferation assay were performed to identify the ideal concentrations of CsA and Ang II. Cytosolic calcium levels in cultured gingival fibroblasts treated with CsA and Ang II were studied using colorimetric assay, confocal and fluorescence imaging. Statistical analyses were done using SPSS software and GraphPad Prism. Results: Higher levels of cytosolic levels were evident in cells treated with CsA and Ang II when compared to control group and was statistically significant (P < 0.05 in both colorimetric assay and confocal imaging. Fluorescent images of the cultured HGFs revealed the same. Conclusion: Thus calcium being a key player in major cellular functions, plays a major role in the pathogenesis of drug-induced gingival overgrowth.

  14. Immunoproteasome overexpression underlies the pathogenesis of thyroid oncocytes and primary hypothyroidism: studies in humans and mice.

    Directory of Open Access Journals (Sweden)

    Hiroaki J Kimura

    2009-11-01

    Full Text Available Oncocytes of the thyroid gland (Hürthle cells are found in tumors and autoimmune diseases. They have a unique appearance characterized by abundant granular eosinophilic cytoplasm and hyperchromatic nucleus. Their pathogenesis has remained, thus far, unknown.Using transgenic mice chronically expressing IFNgamma in thyroid gland, we showed changes in the thyroid follicular epithelium reminiscent of the human oncocyte. Transcriptome analysis comparing transgenic to wild type thyrocytes revealed increased levels of immunoproteasome subunits like LMP2 in transgenics, suggesting an important role of the immunoproteasome in oncocyte pathogenesis. Pharmacologic blockade of the proteasome, in fact, ameliorated the oncocytic phenotype. Genetic deletion of LMP2 subunit prevented the development of the oncocytic phenotype and primary hypothyroidism. LMP2 was also found expressed in oncocytes from patients with Hashimoto thyroiditis and Hürthle cell tumors.In summary, we report that oncocytes are the result of an increased immunoproteasome expression secondary to a chronic inflammatory milieu, and suggest LMP2 as a novel therapeutic target for the treatment of oncocytic lesions and autoimmune hypothyroidism.

  15. In vitro toxicity of propolis in comparison with other primary teeth pulpotomy agents on human fibroblasts.

    Science.gov (United States)

    Al-Haj Ali, Sanaa Najeh

    2016-08-01

    The aim of this study was to assess and compare the in vitro toxicity of propolis with other primary teeth pulpotomy medicaments. Human periodontal ligament (PDL) cells were subjected to different concentrations of propolis, formocresol, ferric sulfate, and gray mineral trioxide aggregate (MTA) (0.05, 0.5, 5, 50, and 100 μg/mL) for 24 h at 37°C. Cells that were not exposed to the tested materials served as the negative control. In vitro toxicity was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Statistical analysis of the data was accomplished using anova and Tukey statistical tests (P propolis and gray MTA had comparable cell viability to the negative control group. Almost all remaining concentrations of tested materials were significantly inferior to the negative control after 24 h of exposure (P Propolis and MTA are more biocompatible than formocresol and ferric sulfate since they were both able to preserve PDL fibroblasts for up to 24 h. © 2015 Wiley Publishing Asia Pty Ltd.

  16. Ambroxol inhibits rhinovirus infection in primary cultures of human tracheal epithelial cells.

    Science.gov (United States)

    Yamaya, Mutsuo; Nishimura, Hidekazu; Nadine, Lusamba Kalonji; Ota, Chiharu; Kubo, Hiroshi; Nagatomi, Ryoichi

    2014-04-01

    The mucolytic drug ambroxol hydrochloride reduces the production of pro-inflammatory cytokines and the frequency of exacerbation in patients with chronic obstructive pulmonary disease (COPD). However, the inhibitory effects of ambroxol on rhinovirus infection, the major cause of COPD exacerbations, have not been studied. We examined the effects of ambroxol on type 14 rhinovirus (RV14) infection, a major RV group, in primary cultures of human tracheal epithelial cells. RV14 infection increased virus titers and cytokine content in the supernatants and RV14 RNA in the cells. Ambroxol (100 nM) reduced RV14 titers and cytokine concentrations of interleukin (IL)-1β, IL-6 and IL-8 in the supernatants and RV14 RNA in the cells after RV14 infection, in addition to reducing susceptibility to RV14 infection. Ambroxol also reduced the expression of intercellular adhesion molecule-1 (ICAM-1), the receptor for RV14, and the number of acidic endosomes from which RV14 RNA enters the cytoplasm. In addition, ambroxol reduced the activation of the transcription factor nuclear factor kappa B (NF-κB) in the nucleus. These results suggest that ambroxol inhibits RV14 infection partly by reducing ICAM-1 and acidic endosomes via the inhibition of NF-κB activation. Ambroxol may modulate airway inflammation by reducing the production of cytokines in rhinovirus infection.

  17. Rapid detection of new and expanding human settlements in the Limpopo province of South Africa using a spatio-temporal change detection method

    CSIR Research Space (South Africa)

    Kleynhans, W

    2015-08-01

    Full Text Available Recent development has identified the benefits of using hyper-temporal satellite time series data for land cover change detection and classification in South Africa. In particular, the monitoring of human settlement expansion in the Limpopo province...

  18. Transcriptome patterns from primary cutaneous Leishmania braziliensis infections associate with eventual development of mucosal disease in humans.

    Science.gov (United States)

    Maretti-Mira, Ana Claudia; Bittner, Jaime; Oliveira-Neto, Manoel Paes; Liu, Minghsun; Kang, Dezhi; Li, Huiying; Pirmez, Claude; Craft, Noah

    2012-01-01

    Localized Cutaneous Leishmaniasis (LCL) and Mucosal Leishmaniasis (ML) are two extreme clinical forms of American Tegumentary Leishmaniasis that usually begin as solitary primary cutaneous lesions. Host and parasite factors that influence the progression of LCL to ML are not completely understood. In this manuscript, we compare the gene expression profiles of primary cutaneous lesions from patients who eventually developed ML to those that did not. Using RNA-seq, we analyzed both the human and Leishmania transcriptomes in primary cutaneous lesions. Limited number of reads mapping to Leishmania transcripts were obtained. For human transcripts, compared to ML patients, lesions from LCL patients displayed a general multi-polarization of the adaptive immune response and showed up-regulation of genes involved in chemoattraction of innate immune cells and in antigen presentation. We also identified a potential transcriptional signature in the primary lesions that may predict long-term disease outcome. We were able to simultaneously sequence both human and Leishmania mRNA transcripts in primary cutaneous leishmaniasis lesions. Our results suggest an intrinsic difference in the immune capacity of LCL and ML patients. The findings correlate the complete cure of L. braziliensis infection with a controlled inflammatory response and a balanced activation of innate and adaptive immunity.

  19. Transcriptome patterns from primary cutaneous Leishmania braziliensis infections associate with eventual development of mucosal disease in humans.

    Directory of Open Access Journals (Sweden)

    Ana Claudia Maretti-Mira

    Full Text Available INTRODUCTION: Localized Cutaneous Leishmaniasis (LCL and Mucosal Leishmaniasis (ML are two extreme clinical forms of American Tegumentary Leishmaniasis that usually begin as solitary primary cutaneous lesions. Host and parasite factors that influence the progression of LCL to ML are not completely understood. In this manuscript, we compare the gene expression profiles of primary cutaneous lesions from patients who eventually developed ML to those that did not. METHODS: Using RNA-seq, we analyzed both the human and Leishmania transcriptomes in primary cutaneous lesions. RESULTS: Limited number of reads mapping to Leishmania transcripts were obtained. For human transcripts, compared to ML patients, lesions from LCL patients displayed a general multi-polarization of the adaptive immune response and showed up-regulation of genes involved in chemoattraction of innate immune cells and in antigen presentation. We also identified a potential transcriptional signature in the primary lesions that may predict long-term disease outcome. CONCLUSIONS: We were able to simultaneously sequence both human and Leishmania mRNA transcripts in primary cutaneous leishmaniasis lesions. Our results suggest an intrinsic difference in the immune capacity of LCL and ML patients. The findings correlate the complete cure of L. braziliensis infection with a controlled inflammatory response and a balanced activation of innate and adaptive immunity.

  20. The genomic profile of human malignant glioma is altered early in primary cell culture and preserved in spheroids

    NARCIS (Netherlands)

    de Witt Hamer, P. C.; van Tilborg, A. A. G.; Eijk, P. P.; Sminia, P.; Troost, D.; van Noorden, C. J. F.; Ylstra, B.; Leenstra, S.

    2008-01-01

    Screening of therapeutics relies on representative cancer models. The representation of human glioblastoma by in vitro cell culture models is questionable. We obtained genomic profiles by array comparative genomic hybridization of both short-and long-term primary cell and spheroid cultures, derived

  1. The awareness of primary caregivers in South Africa of the human rights of their children with intellectual disabilities.

    Science.gov (United States)

    Huus, K; Dada, S; Bornman, J; Lygnegård, F

    2016-11-01

    Besides the right to freedom, human rights can be seen as a basic requirement also for the maintenance of human dignity and the opportunity to thrive - particularly in the case of children with disabilities. It is imperative to explore primary caregivers' awareness of the human rights of their children with intellectual disabilities in view of the role they may play in either facilitating or restricting these rights. This paper explores the awareness of 219 primary caregivers of the human rights of their children with intellectual disabilities. A descriptive survey design was used with a custom-designed questionnaire that employed a deductive content analysis based on the articles of the United Nations Convention on the Rights of a Child. Comparisons were drawn between the awareness of primary caregivers from urban and those from rural areas. The majority (85.5%) of participants agreed that their child with intellectual disability had rights. Three broad kinds of right were mentioned (in descending order): provision rights, protection rights and participation rights. Participants from both urban and rural areas mentioned education (a provision right) most frequently. However, participants from urban areas were more aware of the different rights that existed than were their counterparts from rural areas. Primary caregivers in both rural and urban areas are aware of the rights of their children with disabilities, although there are significant differences between them. © 2016 John Wiley & Sons Ltd.

  2. DIFFERENCES IN PROPIONATE-INDUCED INHIBITION OF CHOLESTEROL AND TRIACYLGLYCEROL SYNTHESIS BETWEEN HUMAN AND RAT HEPATOCYTES IN PRIMARY CULTURE

    NARCIS (Netherlands)

    LIN, YG; VONK, RJ; SLOOFF, MJH; KUIPERS, F; SMIT, MJ

    Propionate is a short-chain fatty acid formed in the colon and supposedly involved in the cholesterol-lowering effect of soluble fibre. To explore the underlying mechanism(s) of this fibre action, we have used human hepatocytes in primary culture to study the effects of propionate on hepatic lipid

  3. Characterization of connective tissue growth factor expression in primary cultures of human tubular epithelial cells: modulation by hypoxia

    NARCIS (Netherlands)

    Kroening, Sven; Neubauer, Emily; Wullich, Bernd; Aten, Jan; Goppelt-Struebe, Margarete

    2010-01-01

    Kroening S, Neubauer E, Wullich B, Aten J, Goppelt-Struebe M. Characterization of connective tissue growth factor expression in primary cultures of human tubular epithelial cells: modulation by hypoxia. Am J Physiol Renal Physiol 298:F796-F806, 2010. First published December 23, 2009;

  4. RNAi mediated acute depletion of Retinoblastoma protein (pRb promotes aneuploidy in human primary cells via micronuclei formation

    Directory of Open Access Journals (Sweden)

    Iovino Flora

    2009-11-01

    Full Text Available Abstract Background Changes in chromosome number or structure as well as supernumerary centrosomes and multipolar mitoses are commonly observed in human tumors. Thus, centrosome amplification and mitotic checkpoint dysfunctions are believed possible causes of chromosomal instability. The Retinoblastoma tumor suppressor (RB participates in the regulation of synchrony between DNA synthesis and centrosome duplication and it is involved in transcription regulation of some mitotic genes. Primary human fibroblasts were transfected transiently with short interfering RNA (siRNA specific for human pRb to investigate the effects of pRb acute loss on chromosomal stability. Results Acutely pRb-depleted fibroblasts showed altered expression of genes necessary for cell cycle progression, centrosome homeostasis, kinetochore and mitotic checkpoint proteins. Despite altered expression of genes involved in the Spindle Assembly Checkpoint (SAC the checkpoint seemed to function properly in pRb-depleted fibroblasts. In particular AURORA-A and PLK1 overexpression suggested that these two genes might have a role in the observed genomic instability. However, when they were post-transcriptionally silenced in pRb-depleted fibroblasts we did not observe reduction in the number of aneuploid cells. This finding suggests that overexpression of these two genes did not contribute to genomic instability triggered by RB acute loss although it affected cell proliferation. Acutely pRb-depleted human fibroblasts showed the presence of micronuclei containing whole chromosomes besides the presence of supernumerary centrosomes and aneuploidy. Conclusion Here we show for the first time that RB acute loss triggers centrosome amplification and aneuploidy in human primary fibroblasts. Altogether, our results suggest that pRb-depleted primary human fibroblasts possess an intact spindle checkpoint and that micronuclei, likely caused by mis-attached kinetochores that in turn trigger

  5. Radiation response of chemically derived mitochondrial DNA-deficient AG01522 human primary fibroblasts.

    Science.gov (United States)

    Nieri, D; Fioramonti, M; Berardinelli, F; Leone, S; Cherubini, R; De Nadal, V; Gerardi, S; Moreno, S; Nardacci, R; Tanzarella, C; Antoccia, A

    2013-08-30

    Mitochondria are the main cellular source of Reactive Oxygen Species (ROS). Alterations of mitochondrial metabolism and consequent loss of mitochondrial membrane potential may lead to redox imbalance and in turn to DNA damage, chromosomal instability and apoptosis. On the other hand, impaired mitochondrial functions may either exacerbate the detrimental effects of geno- and cytotoxic agents or may bring beneficial cellular responses. To study the role of mitochondria within this framework, AG01522 human primary fibroblasts were incubated with the mitochondrial polymerase γ inhibitor 2',3'-dideoxycytidine (ddC), leading to mitochondrial DNA (mtDNA) depletion and to mitochondrial dysfunctions. The successful treatment toward mtDNA depletion was confirmed by Complex-IV subunit I (COX-I) immunofluorescence and western blot assays. mtDNA-depleted cells and their counterparts were ultrastructurally characterized by transmission electron microscopy. mtDNA-depleted cells showed dramatic mitochondrial alterations such as fragmentation and cristae disruption along with a reduction of the mitochondrial membrane potential and elevated levels of ROS. Despite increased ROS levels, we did not find any difference in telomere length between ddC-treated and untreated cells. The spontaneous rate of DNA double-strand breaks (DSBs) and chromosome aberrations was significantly enhanced in mtDNA-depleted cells whereas the induction of DSBs by low-Linear Energy Transfer (LET) (X-rays; 7.7keV/μm protons) and high-LET radiations (28.5keV/μm protons) did not differ when compared with normal cells. However, in irradiated cells impaired mitochondrial functions seemed to bring beneficial cellular responses to the detrimental effect of radiations. In fact, after X-irradiation mtDNA-depleted cells show less remaining unrejoined DSBs than normal cells and furthermore a lower induction of cytogenetic damage. Overall, these data show that active mitochondrial functions are required for the proper

  6. Expandable LED array interconnect

    Science.gov (United States)

    Yuan, Thomas Cheng-Hsin; Keller, Bernd

    2011-03-01

    A light emitting device that can function as an array element in an expandable array of such devices. The light emitting device comprises a substrate that has a top surface and a plurality of edges. Input and output terminals are mounted to the top surface of the substrate. Both terminals comprise a plurality of contact pads disposed proximate to the edges of the substrate, allowing for easy access to both terminals from multiple edges of the substrate. A lighting element is mounted to the top surface of the substrate. The lighting element is connected between the input and output terminals. The contact pads provide multiple access points to the terminals which allow for greater flexibility in design when the devices are used as array elements in an expandable array.

  7. Grazing incidence beam expander

    Energy Technology Data Exchange (ETDEWEB)

    Akkapeddi, P.R.; Glenn, P.; Fuschetto, A.; Appert, Q.; Viswanathan, V.K.

    1985-01-01

    A Grazing Incidence Beam Expander (GIBE) telescope is being designed and fabricated to be used as an equivalent end mirror in a long laser resonator cavity. The design requirements for this GIBE flow down from a generic Free Electron Laser (FEL) resonator. The nature of the FEL gain volume (a thin, pencil-like, on-axis region) dictates that the output beam be very small. Such a thin beam with the high power levels characteristic of FELs would have to travel perhaps hundreds of meters or more before expanding enough to allow reflection from cooled mirrors. A GIBE, on the other hand, would allow placing these optics closer to the gain region and thus reduces the cavity lengths substantially. Results are presented relating to optical and mechanical design, alignment sensitivity analysis, radius of curvature analysis, laser cavity stability analysis of a linear stable concentric laser cavity with a GIBE. Fabrication details of the GIBE are also given.

  8. Expanding the HAWC Observatory

    Energy Technology Data Exchange (ETDEWEB)

    Mori, Johanna [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2016-08-17

    The High Altitude Water Cherenkov Gamma-Ray Observatory is expanding its current array of 300 water tanks to include 350 outrigger tanks to increase sensitivity to gamma rays above 10 TeV. This involves creating and testing hardware with which to build the new tanks, including photomultiplier tubes, high voltage supply units, and flash analog to digital converters. My responsibilities this summer included preparing, testing and calibrating that equipment.

  9. Bioinformatics analysis of proteomics profiles in senescent human primary proximal tubule epithelial cells.

    Science.gov (United States)

    Lu, Yang; Wang, Jingchao; Dapeng, Chen; Wu, Di; Cai, Guangyan; Chen, Xiangmei

    2016-04-01

    Dysfunction of renal tubule epithelial cells is associated with renal tubulointerstitial fibrosis. Exploration of the proteomic profiles of senesced tubule epithelial cells is essential to elucidate the mechanism of tubulointerstitium development. Primary human proximal tubule epithelial cells from passage 3 (P3) and passage 6 (P6) were selected for evaluation. EdU and SA-β-galactosidase staining were used to detect cell senescence. p53, p21, and p16 were detected by Western blot analysis. Liquid chromatography mass spectrometry (LC-MS) was used to examine differentially expressed proteins (DEPs) between P6 and P3 cells. The expression of DEPs was examined by Western blot analysis. Bioinformatics analysis was performed by protein-protein interaction and gene ontology analyses. The majority of tubule cells from passage 6 (P6) stained positive for SA-β-galactosidase, whereas passage 3 (P3) cells were negative. Senescence biomarkers, including p53, p21, and p16, were upregulated in P6 cells relative to P3 cells. EdU staining results showed a lower rate of EdU positive cells in P6 cells than in P3 cells. LC-MS was used to examine DEPs between P6 and P3 cells. These DEPs are involved in glycolysis, response to stress, cytoskeleton regulation, oxidative reduction, ATP binding, and oxidative stress. Using Western blot analysis, we validated the down-regulation of AKR1B1, EEF2, EEF1A1, and HSP90 and the up-regulation of VIM in P6 cells seen in the LC-MS data. More importantly, we built the molecular network based on biological functions and protein-protein interactions and found that the DEPs are involved in translation elongation, stress, and glycolysis, and that they are all associated with cytoskeleton regulation, which regulates senescent cell activities such as apoptosis and EMT in tubule epithelial cells. We explored proteomic profile changes in cell culture-induced senescent cells and built senescence-associated molecular networks, which will help to elucidate the

  10. Ex vivo expanded human regulatory T cells delay islet allograft rejection via inhibiting islet-derived monocyte chemoattractant protein-1 production in CD34+ stem cells-reconstituted NOD-scid IL2rγnull mice.

    Directory of Open Access Journals (Sweden)

    Fang Xiao

    Full Text Available Type 1 diabetes mellitus (T1DM is an autoimmune disease caused by immune-mediated destruction of insulin-secreting β cells of the pancreas. Near complete dependence on exogenous insulin makes T1DM very difficult to control, with the result that patients are exposed to high blood glucose and risk of diabetic complications and/or intermittent low blood glucose that can cause unconsciousness, fits and even death. Allograft transplantation of pancreatic islets restores normoglycemia with a low risk of surgical complications. However, although successful immediately after transplantation, islets are progressively lost, with most of the patients requiring exogenous insulin within 2 years post-transplant. Therefore, there is an urgent requirement for the development of new strategies to prevent islet rejection. In this study, we explored the importance of human regulatory T cells in the control of islets allograft rejection. We developed a pre-clinical model of human islet transplantation by reconstituting NOD-scid IL2rγnull mice with cord blood-derived human CD34+ stem cells and demonstrated that although the engrafted human immune system mediated the rejection of human islets, their survival was significantly prolonged following adoptive transfer of ex vivo expanded human Tregs. Mechanistically, Tregs inhibited the infiltration of innate immune cells and CD4+ T cells into the graft by down-regulating the islet graft-derived monocyte chemoattractant protein-1. Our findings might contribute to the development of clinical strategies for Treg therapy to control human islet rejection. We also show for the first time that CD34+ cells-reconstituted NOD-scid IL2rγnull mouse model could be beneficial for investigating human innate immunity in vivo.

  11. Rotavirus structural proteins and dsRNA are required for the human primary plasmacytoid dendritic cell IFNalpha response.

    Directory of Open Access Journals (Sweden)

    Emily M Deal

    2010-06-01

    Full Text Available Rotaviruses are the leading cause of severe dehydrating diarrhea in children worldwide. Rotavirus-induced immune responses, especially the T and B cell responses, have been extensively characterized; however, little is known about innate immune mechanisms involved in the control of rotavirus infection. Although increased levels of systemic type I interferon (IFNalpha and beta correlate with accelerated resolution of rotavirus disease, multiple rotavirus strains, including rhesus rotavirus (RRV, have been demonstrated to antagonize type I IFN production in a variety of epithelial and fibroblast cell types through several mechanisms, including degradation of multiple interferon regulatory factors by a viral nonstructural protein. This report demonstrates that stimulation of highly purified primary human peripheral plasmacytoid dendritic cells (pDCs with either live or inactivated RRV induces substantial IFNalpha production by a subset of pDCs in which RRV does not replicate. Characterization of pDC responses to viral stimulus by flow cytometry and Luminex revealed that RRV replicates in a small subset of human primary pDCs and, in this RRV-permissive small subset, IFNalpha production is diminished. pDC activation and maturation were observed independently of viral replication and were enhanced in cells in which virus replicates. Production of IFNalpha by pDCs following RRV exposure required viral dsRNA and surface proteins, but neither viral replication nor activation by trypsin cleavage of VP4. These results demonstrate that a minor subset of purified primary human peripheral pDCs are permissive to RRV infection, and that pDCs retain functionality following RRV stimulus. Additionally, this study demonstrates trypsin-independent infection of primary peripheral cells by rotavirus, which may allow for the establishment of extraintestinal viremia and antigenemia. Importantly, these data provide the first evidence of IFNalpha induction in primary

  12. Rotavirus structural proteins and dsRNA are required for the human primary plasmacytoid dendritic cell IFNalpha response.

    Science.gov (United States)

    Deal, Emily M; Jaimes, Maria C; Crawford, Sue E; Estes, Mary K; Greenberg, Harry B

    2010-06-03

    Rotaviruses are the leading cause of severe dehydrating diarrhea in children worldwide. Rotavirus-induced immune responses, especially the T and B cell responses, have been extensively characterized; however, little is known about innate immune mechanisms involved in the control of rotavirus infection. Although increased levels of systemic type I interferon (IFNalpha and beta) correlate with accelerated resolution of rotavirus disease, multiple rotavirus strains, including rhesus rotavirus (RRV), have been demonstrated to antagonize type I IFN production in a variety of epithelial and fibroblast cell types through several mechanisms, including degradation of multiple interferon regulatory factors by a viral nonstructural protein. This report demonstrates that stimulation of highly purified primary human peripheral plasmacytoid dendritic cells (pDCs) with either live or inactivated RRV induces substantial IFNalpha production by a subset of pDCs in which RRV does not replicate. Characterization of pDC responses to viral stimulus by flow cytometry and Luminex revealed that RRV replicates in a small subset of human primary pDCs and, in this RRV-permissive small subset, IFNalpha production is diminished. pDC activation and maturation were observed independently of viral replication and were enhanced in cells in which virus replicates. Production of IFNalpha by pDCs following RRV exposure required viral dsRNA and surface proteins, but neither viral replication nor activation by trypsin cleavage of VP4. These results demonstrate that a minor subset of purified primary human peripheral pDCs are permissive to RRV infection, and that pDCs retain functionality following RRV stimulus. Additionally, this study demonstrates trypsin-independent infection of primary peripheral cells by rotavirus, which may allow for the establishment of extraintestinal viremia and antigenemia. Importantly, these data provide the first evidence of IFNalpha induction in primary human pDCs by a ds

  13. Complex nature of SNP genotype effects on gene expression in primary human leucocytes

    Directory of Open Access Journals (Sweden)

    Dinesen Lotte C

    2009-01-01

    Full Text Available Abstract Background Genome wide association studies have been hugely successful in identifying disease risk variants, yet most variants do not lead to coding changes and how variants influence biological function is usually unknown. Methods We correlated gene expression and genetic variation in untouched primary leucocytes (n = 110 from individuals with celiac disease – a common condition with multiple risk variants identified. We compared our observations with an EBV-transformed HapMap B cell line dataset (n = 90, and performed a meta-analysis to increase power to detect non-tissue specific effects. Results In celiac peripheral blood, 2,315 SNP variants influenced gene expression at 765 different transcripts (cis expression quantitative trait loci, eQTLs. 135 of the detected SNP-probe effects (reflecting 51 unique probes were also detected in a HapMap B cell line published dataset, all with effects in the same allelic direction. Overall gene expression differences within the two datasets predominantly explain the limited overlap in observed cis-eQTLs. Celiac associated risk variants from two regions, containing genes IL18RAP and CCR3, showed significant cis genotype-expression correlations in the peripheral blood but not in the B cell line datasets. We identified 14 genes where a SNP affected the expression of different probes within the same gene, but in opposite allelic directions. By incorporating genetic variation in co-expression analyses, functional relationships between genes can be more significantly detected. Conclusion In conclusion, the complex nature of genotypic effects in human populations makes the use of a relevant tissue, large datasets, and analysis of different exons essential to enable the identification of the function for many genetic risk variants in common diseases.

  14. Pharmacological inhibition of dynamin II reduces constitutive protein secretion from primary human macrophages.

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    Maaike Kockx

    Full Text Available Dynamins are fission proteins that mediate endocytic and exocytic membrane events and are pharmacological therapeutic targets. These studies investigate whether dynamin II regulates constitutive protein secretion and show for the first time that pharmacological inhibition of dynamin decreases secretion of apolipoprotein E (apoE and several other proteins constitutively secreted from primary human macrophages. Inhibitors that target recruitment of dynamin to membranes (MiTMABs or directly target the GTPase domain (Dyngo or Dynole series, dose- and time- dependently reduced the secretion of apoE. SiRNA oligo's targeting all isoforms of dynamin II confirmed the involvement of dynamin II in apoE secretion. Inhibition of secretion was not mediated via effects on mRNA or protein synthesis. 2D-gel electrophoresis showed that inhibition occurred after apoE was processed and glycosylated in the Golgi and live cell imaging showed that inhibited secretion was associated with reduced post-Golgi movement of apoE-GFP-containing vesicles. The effect was not restricted to macrophages, and was not mediated by the effects of the inhibitors on microtubules. Inhibition of dynamin also altered the constitutive secretion of other proteins, decreasing the secretion of fibronectin, matrix metalloproteinase 9, Chitinase-3-like protein 1 and lysozyme but unexpectedly increasing the secretion of the inflammatory mediator cyclophilin A. We conclude that pharmacological inhibitors of dynamin II modulate the constitutive secretion of macrophage apoE as a class effect, and that their capacity to modulate protein secretion may affect a range of biological processes.

  15. Biocompatibility of magnesium implants in primary human reaming debris-derived cells stem cells in vitro.

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    Charyeva, Olga; Dakischew, Olga; Sommer, Ursula; Heiss, Christian; Schnettler, Reinhard; Lips, Katrin Susanne

    2016-03-01

    Use of magnesium for resorbable metal implants is a new concept in orthopaedic and dental medicine. The majority of studies on magnesium's biocompatibility in vitro have assessed the short-term effect of magnesium extract on cells. The aim of this study was to evaluate the influence of direct exposure to magnesium alloys on the bioactivity of primary human reaming debris-derived (HRD) cells. Pure Mg, Mg2Ag, WE43 and Mg10Gd were tested for biocompatibility. The study consisted of assessment of cell viability by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test, evaluation of alkaline phosphatase (ALP) content, and study of cell morphology under light microscopy, scanning electron microscopy (SEM) and transmission electron microscopy (TEM), along with determination of calcification and pH changes induced by magnesium. The number of viable cells in the presence of Mg2Ag was high over the entire observation period. Inhibition of ALP content in osteogenic differentiating HRD was caused by pure Mg at day 14 and 28. All other magnesium alloys did not affect the ALP content. Exposure of HRD to magnesium increased the amount of lysosomes and endocytotic vesicles. Cellular attachment was generally the best for those crystals that formed on the surface of all materials. A decrease was observed in Ca(2+) in the medium from day 1 to day 14. In terms of cell morphology, cell viability and differentiation, cell density and the effect on the surrounding pH, Mg2Ag showed the most promising results. All magnesium materials induced calcification, which is beneficial for orthopaedic and dental applications.

  16. Clonal mutations in primary human glial tumors: evidence in support of the mutator hypothesis

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    Sarkar Chitra

    2007-10-01

    Full Text Available Abstract Background A verifiable consequence of the mutator hypothesis is that even low grade neoplasms would accumulate a large number of mutations that do not influence the tumor phenotype (clonal mutations. In this study, we have attempted to quantify the number of clonal mutations in primary human gliomas of astrocytic cell origin. These alterations were identified in tumor tissue, microscopically confirmed to have over 70% neoplastic cells. Methods Random Amplified Polymorphic DNA (RAPD analysis was performed using a set of fifteen 10-mer primers of arbitrary but definite sequences in 17 WHO grade II astrocytomas (low grade diffuse astrocytoma or DA and 16 WHO grade IV astrocytomas (Glioblastoma Multiforme or GBM. The RAPD profile of the tumor tissue was compared with that of the leucocyte DNA of the same patient and alteration(s scored. A quantitative estimate of the overall genomic changes in these tumors was obtained by 2 different modes of calculation. Results The overall change in the tumors was estimated to be 4.24% in DA and 2.29% in GBM by one method and 11.96% and 6.03% in DA and GBM respectively by the other. The difference between high and lower grade tumors was statistically significant by both methods. Conclusion This study demonstrates the presence of extensive clonal mutations in gliomas, more in lower grade. This is consistent with our earlier work demonstrating that technique like RAPD analysis, unbiased for locus, is able to demonstrate more intra-tumor genetic heterogeneity in lower grade gliomas compared to higher grade. The results support the mutator hypothesis proposed by Loeb.

  17. Measuring Connectivity in the Primary Visual Pathway in Human Albinism Using Diffusion Tensor Imaging and Tractography.

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    Grigorian, Anahit; McKetton, Larissa; Schneider, Keith A

    2016-08-11

    In albinism, the number of ipsilaterally projecting retinal ganglion cells (RGCs) is significantly reduced. The retina and optic chiasm have been proposed as candidate sites for misrouting. Since a correlation between the number of lateral geniculate nucleus (LGN) relay neurons and LGN size has been shown, and based on previously reported reductions in LGN volumes in human albinism, we suggest that fiber projections from LGN to the primary visual cortex (V1) are also reduced. Studying structural differences in the visual system of albinism can improve the understanding of the mechanism of misrouting and subsequent clinical applications. Diffusion data and tractography are useful for mapping the OR (optic radiation). This manuscript describes two algorithms for OR reconstruction in order to compare brain connectivity in albinism and controls.An MRI scanner with a 32-channel head coil was used to acquire structural scans. A T1-weighted 3D-MPRAGE sequence with 1 mm(3) isotropic voxel size was used to generate high-resolution images for V1 segmentation. Multiple proton density (PD) weighted images were acquired coronally for right and left LGN localization. Diffusion tensor imaging (DTI) scans were acquired with 64 diffusion directions. Both deterministic and probabilistic tracking methods were run and compared, with LGN as the seed mask and V1 as the target mask. Though DTI provides relatively poor spatial resolution, and accurate delineation of OR may be challenging due to its low fiber density, tractography has been shown to be advantageous both in research and clinically. Tract based spatial statistics (TBSS) revealed areas of significantly reduced white matter integrity within the OR in patients with albinism compared to controls. Pairwise comparisons revealed a significant reduction in LGN to V1 connectivity in albinism compared to controls. Comparing both tracking algorithms revealed common findings, strengthening the reliability of the technique.

  18. Spatio-temporal correlates of taste processing in the human primary gustatory cortex.

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    Iannilli, E; Noennig, N; Hummel, T; Schoenfeld, A M

    2014-07-25

    In humans the identification of the primary gustatory cortex (PGC) is still under debate. Neuroimaging studies indicate insula and overlying opercula as the best candidates but the exact position of the PGC within this region is not entirely clear. Moreover, inconsistencies appear when comparing results from studies using functional magnetic resonance imaging (fMRI), and gustatory event-related potentials (gERP), or gustatory event-related magnetic fields (gERMF). fMRI indicates activations in the anterior part of the insula and frontal operculum, while gERP and/or gERMF indicate activations at the transition between the parietal operculum and insula in its posterior part. Here it is important to note that for gERP and gERMF temporal and spatial characteristics of the stimulus must be well controlled to evoke a useful brain response. In the present study gERMF and gERP were recorded simultaneously using a whole-head system with 249 magnetometers and 32 electrodes, respectively; taste stimuli were applied using a stimulator providing excellent temporal and spatial control of the stimulus. Separate ERP and ERMF averaged waveforms were derived time-locked to the onset of the taste stimuli. The source analysis for the early time range revealed activity in the left and right anterior and mid part of the insula, where in the later time range the sources were located more in the posterior part of the insula. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

  19. Sensory-motor interactions for vocal pitch monitoring in non-primary human auditory cortex.

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    Jeremy D W Greenlee

    Full Text Available The neural mechanisms underlying processing of auditory feedback during self-vocalization are poorly understood. One technique used to study the role of auditory feedback involves shifting the pitch of the feedback that a speaker receives, known as pitch-shifted feedback. We utilized a pitch shift self-vocalization and playback paradigm to investigate the underlying neural mechanisms of audio-vocal interaction. High-resolution electrocorticography (ECoG signals were recorded directly from auditory cortex of 10 human subjects while they vocalized and received brief downward (-100 cents pitch perturbations in their voice auditory feedback (speaking task. ECoG was also recorded when subjects passively listened to playback of their own pitch-shifted vocalizations. Feedback pitch perturbations elicited average evoked potential (AEP and event-related band power (ERBP responses, primarily in the high gamma (70-150 Hz range, in focal areas of non-primary auditory cortex on superior temporal gyrus (STG. The AEPs and high gamma responses were both modulated by speaking compared with playback in a subset of STG contacts. From these contacts, a majority showed significant enhancement of high gamma power and AEP responses during speaking while the remaining contacts showed attenuated response amplitudes. The speaking-induced enhancement effect suggests that engaging the vocal motor system can modulate auditory cortical processing of self-produced sounds in such a way as to increase neural sensitivity for feedback pitch error detection. It is likely that mechanisms such as efference copies may be involved in this process, and modulation of AEP and high gamma responses imply that such modulatory effects may affect different cortical generators within distinctive functional networks that drive voice production and control.

  20. The altered landscape of the human skin microbiome in patients with primary immunodeficiencies.

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    Oh, Julia; Freeman, Alexandra F; Park, Morgan; Sokolic, Robert; Candotti, Fabio; Holland, Steven M; Segre, Julia A; Kong, Heidi H

    2013-12-01

    While landmark studies have shown that microbiota activate and educate host immunity, how immune systems shape microbiomes and contribute to disease is incompletely characterized. Primary immunodeficiency (PID) patients suffer recurrent microbial infections, providing a unique opportunity to address this issue. To investigate the potential influence of host immunity on the skin microbiome, we examined skin microbiomes in patients with rare monogenic PIDs: hyper-IgE (STAT3-deficient), Wiskott-Aldrich, and dedicator of cytokinesis 8 syndromes. While specific immunologic defects differ, a shared hallmark is atopic dermatitis (AD)-like eczema. We compared bacterial and fungal skin microbiomes (41 PID, 13 AD, 49 healthy controls) at four clinically relevant sites representing the major skin microenvironments. PID skin displayed increased ecological permissiveness with altered population structures, decreased site specificity and temporal stability, and colonization with microbial species not observed in controls, including Clostridium species and Serratia marcescens. Elevated fungal diversity and increased representation of opportunistic fungi (Candida, Aspergillus) supported increased PID skin permissiveness, suggesting that skin may serve as a reservoir for the recurrent fungal infections observed in these patients. The overarching theme of increased ecological permissiveness in PID skin was counterbalanced by the maintenance of a phylum barrier in which colonization remained restricted to typical human-associated phyla. Clinical parameters, including markers of disease severity, were positively correlated with prevalence of Staphylococcus, Corynebacterium, and other less abundant taxa. This study examines differences in microbial colonization and community stability in PID skin and informs our understanding of host-microbiome interactions, suggesting a bidirectional dialogue between skin commensals and the host organism.

  1. Cytotoxicity and bioactivity of various pulpotomy materials on stem cells from human exfoliated primary teeth.

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    Collado-González, M; García-Bernal, D; Oñate-Sánchez, R E; Ortolani-Seltenerich, P S; Álvarez-Muro, T; Lozano, A; Forner, L; Llena, C; Moraleda, J M; Rodríguez-Lozano, F J

    2017-12-01

    To investigate the cytotoxicity and bioactivity of several pulpotomy materials: Biodentine (Septodont, Saint-Maur-des-Fosses, France) MTA (Angelus, Londrina, PR, Brazil), Theracal LC (Bisco Inc., Schamburg, IL, USA) and IRM (Dentsply DeTrey GmbH, Konstanz, Germany), after contact with stem cells isolated from human exfoliated primary teeth (SHEDs). SHEDs were cultured in the presence of the eluates of various pulpotomy materials for 24, 48 and 72 h. Cell viability was determined by mitochondrial dehydrogenase enzymatic (MTT) assay. Apoptosis and changes in cell phenotype were evaluated by flow cytometry. Also, an in vitro scratch wound-healing assay was used to determine their effects on cell migration. To assess cell morphology and attachment to the different pulpotomy materials, SHEDs were directly seeded onto the material surfaces and analysed by scanning electron microscopy (SEM). Finally, the deposition of a calcified matrix in presence of these materials was verified by Alizarin Red staining. Statistical analysis was performed with analysis of variance and Bonferroni or Tukey post-test (α = 0.05). Cell viability in the presence of Biodentine eluates was significantly higher to that obtained using complete medium alone (control; P Biodentine. SEM studies revealed a suitable proliferation rate, cell spreading and attachment, especially when using Biodentine and MTA Angelus discs. Finally, Biodentine eluates significantly induced calcified matrix deposition from 7 days of culture (P Biodentine exhibited better cytocompatibility and bioactivity than MTA Angelus, Theracal LC and IRM. © 2017 International Endodontic Journal. Published by John Wiley & Sons Ltd.

  2. Adiponectin gene expression in human primary adipocyte culture treated with uremic serum

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    Sultan Alouffi

    2015-01-01

    Full Text Available End-stage renal disease (ESRD is accompanied by an increased rate of morbidity and mortality due to cardiovascular disease (CVD. Although renal replacement therapy is required at this stage, it is associated with additional complications such as inflammation and dyslipidemia. It has been suggested that adiponectin has anti-inflammatory properties. We studied the potential role of uremic mileu on the adiponectin expression in human primary adipocyte culture. A cohort of 18 patients with ESRD (hemo-and peritoneal dialysis and nine healthy controls were analyzed in a prospective cross-sectional study. Single blood samples were taken pre-and post-hemodialysis and in peritoneal dialysis patients. Serum concentrations of total adiponectin (7.95 ± 1.44 μg/mL; 6.73 ± 1.2 μg/mL; 13.7 ± 3.04 μg/mL, respectively and high molecular weight adiponectin (3.03 ± 1.95 μg/mL; 3.57 ± 2.44 14 μg/mL; 8.02 ± 5 μg/mL respectively were measured. Other biochemical parameters (cholesterol, low-density lipoprotein cholesterol and triglycerides were assessed in all groups of patients. Adiponectin gene expression was determined using real-time polymerase chain reaction, and was found to be lower in ESRD patients compared with healthy controls with low dose but not with high-dose treatments. Serum concentrations of total adiponectin and high molecular weight adiponectin were significantly higher in the ESRD versus control group. These results provide an initial insight into understanding the putative role of adipose tissue in contributing to the association of CVD risk in patients with chronic kidney disease.

  3. Arsenic compromises conducting airway epithelial barrier properties in primary mouse and immortalized human cell cultures.

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    Cara L Sherwood

    Full Text Available Arsenic is a lung toxicant that can lead to respiratory illness through inhalation and ingestion, although the most common exposure is through contaminated drinking water. Lung effects reported from arsenic exposure include lung cancer and obstructive lung disease, as well as reductions in lung function and immune response. As part of their role in innate immune function, airway epithelial cells provide a barrier that protects underlying tissue from inhaled particulates, pathogens, and toxicants frequently found in inspired air. We evaluated the effects of a five-day exposure to environmentally relevant levels of arsenic {<4μM [~300 μg/L (ppb] as NaAsO2} on airway epithelial barrier function and structure. In a primary mouse tracheal epithelial (MTE cell model we found that both micromolar (3.9 μM and submicromolar (0.8 μM arsenic concentrations reduced transepithelial resistance, a measure of barrier function. Immunofluorescent staining of arsenic-treated MTE cells showed altered patterns of localization of the transmembrane tight junction proteins claudin (Cl Cl-1, Cl-4, Cl-7 and occludin at cell-cell contacts when compared with untreated controls. To better quantify arsenic-induced changes in tight junction transmembrane proteins we conducted arsenic exposure experiments with an immortalized human bronchial epithelial cell line (16HBE14o-. We found that arsenic exposure significantly increased the protein expression of Cl-4 and occludin as well as the mRNA levels of Cl-4 and Cl-7 in these cells. Additionally, arsenic exposure resulted in altered phosphorylation of occludin. In summary, exposure to environmentally relevant levels of arsenic can alter both the function and structure of airway epithelial barrier constituents. These changes likely contribute to the observed arsenic-induced loss in basic innate immune defense and increased infection in the airway.

  4. In Vitro Experimental Model of Trained Innate Immunity in Human Primary Monocytes.

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    Bekkering, Siroon; Blok, Bastiaan A; Joosten, Leo A B; Riksen, Niels P; van Crevel, Reinout; Netea, Mihai G

    2016-12-01

    Innate immune memory, or trained immunity, has recently been described to be an important property of cells of the innate immune system. Due to the increased interest in this important new field of immunological investigation, we sought to determine the optimal conditions for an in vitro experimental protocol of monocyte training using three of the most commonly used training stimuli from the literature: β-glucan, the bacillus Calmette-Guérin (BCG) vaccine, and oxidized low-density lipoprotein (oxLDL). We investigated and optimized a protocol of monocyte trained immunity induced by an initial training period with β-glucan, BCG, or oxLDL, followed by washing and resting of the cells and, thereafter, restimulation with secondary bacterial stimuli. The training and resting time intervals were varied to identify the optimal setting for the long-term induction of trained immunity. Trained immunity was assessed in terms of the secondary cytokine response, the production of reactive oxygen species, cell morphology, and induction of glycolysis. Monocytes primed with β-glucan, BCG, and oxLDL showed increased pro- and anti-inflammatory cytokine responses upon restimulation with nonrelated stimuli. Also, all three stimuli induced a switch to glycolysis (the Warburg effect). These effects were most pronounced when the training interval was 24 h and the resting time interval was 6 days. Training with BCG and oxLDL also led to the increased production of reactive oxygen species, whereas training with β-glucan led to the decreased production of reactive oxygen species. We describe the optimal conditions for an in vitro experimental model with human primary monocytes for study of the induction of trained innate immunity by microbial and metabolic stimuli. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  5. Utilization of replication-competent XMRV reporter-viruses reveals severe viral restriction in primary human cells.

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    Christina Martina Stürzel

    Full Text Available The gammaretrovirus termed xenotropic murine leukemia virus-related virus (XMRV was described to be isolated from prostate cancer tissue biopsies and from blood of patients suffering from chronic fatigue syndrome. However, many studies failed to detect XMRV and to verify these disease associations. Data suggesting the contamination of specimens in particular by PCR-based methods and recent reports demonstrating XMRV generation via recombination of two murine leukemia virus precursors raised serious doubts about XMRV being a genuine human pathogen. To elucidate cell tropism of XMRV, we generated replication competent XMRV reporter viruses encoding a green fluorescent protein or a secretable luciferase as tools to analyze virus infection of human cell lines or primary human cells. Transfection of proviral DNAs into LNCaP prostate cancer cells resulted in readily detectably reporter gene expression and production of progeny virus. Inoculation of known XMRV susceptible target cells revealed that these virions were infectious and expressed the reporter gene, allowing for a fast and highly sensitive quantification of XMRV infection. Both reporter viruses were capable of establishing a spreading infection in LNCaP and Raji B cells and could be easily passaged. However, after inoculation of primary human blood cells such as CD4 T cells, macrophages or dendritic cells, infection rates were very low, and a spreading infection was never established. In line with these results we found that supernatants derived from these XMRV infected primary cell types did not contain infectious virus. Thus, although XMRV efficiently replicated in some human cell lines, all tested primary cells were largely refractory to XMRV infection and did not support viral spread. Our results provide further evidence that XMRV is not a human pathogen.

  6. Expanding the Scope of Anatomical Sciences: The Case of "Human Evolution--The Fossil Evidence" Course at the Sackler School of Medicine, Tel-Aviv University

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    Notzer, Netta; Abramovitz, Ruth

    2012-01-01

    The Anatomy Department at Tel-Aviv University Medical School offers its students an elective course of 26 didactic hours on human evolution. The course is open to students from all faculties, who must fulfill all academic requirements, without a prerequisite of a background in anatomy. Approximately 120 students attend annually, a third of them…

  7. Molecular Validation of Chondrogenic Differentiation and Hypoxia Responsiveness of Platelet-Lysate Expanded Adipose Tissue–Derived Human Mesenchymal Stromal Cells

    NARCIS (Netherlands)

    Galeano-Garces, Catalina; Camilleri, Emily T.; Riester, Scott M.; Dudakovic, Amel; Larson, Dirk R.; Qu, Wenchun; Smith, Jay; Dietz, Allan B.; Im, Hee-Jeong; Krych, Aaron J.; Larson, A. Noelle; Karperien, Marcel; van Wijnen, Andre J.

    2017-01-01

    Objective: To determine the optimal environmental conditions for chondrogenic differentiation of human adipose tissue–derived mesenchymal stromal/stem cells (AMSCs). In this investigation we specifically investigate the role of oxygen tension and 3-dimensional (3D) culture systems. Design: Both

  8. Evaluating a Human Rights-Based Advocacy Approach to Expanding Access to Pain Medicines and Palliative Care: Global Advocacy and Case Studies from India, Kenya, and Ukraine.

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    Lohman, Diederik; Amon, Joseph J

    2015-12-10

    Palliative care has been defined as care that is person-centered and attentive to physical symptoms and psychological, social, and existential distress in patients with severe or life-threatening illness. The identification of access to palliative care and pain treatment as a human rights issue first emerged among palliative care advocates, physicians, and lawyers in the 1990s, with a basis in the right to health and the right to be free from cruel, inhuman, and degrading treatment. Using a case study approach, we evaluate the results of a human rights-based advocacy approach on access to pain medicine and palliative care in India, Kenya, and Ukraine. In each country, human rights advocacy helped raise awareness of the issue, identify structural barriers to care, define government obligations, and contribute to the reform of laws, policies, and practices impeding the availability of palliative care services. In addition, advocacy efforts stimulated civil society engagement and high-level political leadership that fostered the implementation of human rights-based palliative care programs. Globally, access to palliative care was increasingly recognized by human rights bodies and within global health and drug policy organizations as a government obligation central to the right to health. Copyright © 2015 Lohman, Amon. This is an open access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original author and source are credited.

  9. Prevalence and factors associated with breast milk donation in banks that receive human milk in primary health care units,

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    Tatiana Mota Xavier de Meneses

    Full Text Available Abstract Objective: To estimate the prevalence and to analyze factors associated with breast milk donation at primary health care units in order to increase the human milk bank reserves. Methods: Cross-sectional study carried out in 2013 in Rio de Janeiro, Brazil. A representative sample of 695 mothers of children younger than 1 year attended to at the nine primary health care units with human milk donation services were interviewed. A hierarchical approach was used to obtain adjusted prevalence ratios (APR by Poisson regression with robust variance. The final model included the variables associated with breast milk donation (p ≤ 0.05. Results: 7.3% of the mothers had donated breast milk. Having been encouraged to donate breast milk by healthcare professionals, relatives, or friends (APR = 7.06, receiving information on breast milk expression by the primary health care unit (APR = 3.65, and receiving help from the unit professionals to breastfeed (APR = 2.24 were associated with a higher prevalence of donation. Admission of the newborn to the neonatal unit was associated with a lower prevalence of donation (APR = 0.09. Conclusions: Encouragement to breast milk donation, and information and help provided by primary health care unit professionals to breastfeeding were shown to be important for the practice of human milk donation.

  10. Prevalence and factors associated with breast milk donation in banks that receive human milk in primary health care units.

    Science.gov (United States)

    Meneses, Tatiana Mota Xavier de; Oliveira, Maria Inês Couto de; Boccolini, Cristiano Siqueira

    To estimate the prevalence and to analyze factors associated with breast milk donation at primary health care units in order to increase the human milk bank reserves. Cross-sectional study carried out in 2013 in Rio de Janeiro, Brazil. A representative sample of 695 mothers of children younger than 1 year attended to at the nine primary health care units with human milk donation services were interviewed. A hierarchical approach was used to obtain adjusted prevalence ratios (APR) by Poisson regression with robust variance. The final model included the variables associated with breast milk donation (p≤0.05). 7.3% of the mothers had donated breast milk. Having been encouraged to donate breast milk by healthcare professionals, relatives, or friends (APR=7.06), receiving information on breast milk expression by the primary health care unit (APR=3.65), and receiving help from the unit professionals to breastfeed (APR=2.24) were associated with a higher prevalence of donation. Admission of the newborn to the neonatal unit was associated with a lower prevalence of donation (APR=0.09). Encouragement to breast milk donation, and information and help provided by primary health care unit professionals to breastfeeding were shown to be important for the practice of human milk donation. Copyright © 2017. Published by Elsevier Editora Ltda.

  11. Metabolic activation of diesel exhaust carcinogens in primary and immortalized human TP53 knock-in (Hupki) mouse embryo fibroblasts.

    Science.gov (United States)

    Kucab, Jill E; Phillips, David H; Arlt, Volker M

    2012-04-01

    Approximately 50% of human tumors have a mutation in TP53. The pattern and spectra of TP53 mutations often differ between cancer types, perhaps due to different etiological factors. The Hupki (human TP53 knock-in) mouse embryo fibroblast (HUF) immortalization assay is useful for studying mutagenesis in the human TP53 gene by environmental carcinogens. Prior to initiating an immortalization assay, carcinogen treatment conditions must be optimized, which can require a large number of cells. As primary HUF cultures senesce within 2 weeks, restricting their use, we investigated whether immortalized HUFs retaining wild-type TP53 can be surrogates for primary HUFs in initial treatment optimization. DNA damage by eight compounds found in diesel exhaust, benzo[a]pyrene, 3-nitrobenzanthrone, 1-nitropyrene, 1,3-dinitropyrene, 1,6-dinitropyrene, 1,8-dinitropyrene, 6-nitrochrysene, and 3-nitrofluorene, was assessed by (32) P-postlabeling and the alkaline comet assay in primary HUFs and in an immortal HUF cell line J201. For most compounds, higher levels of DNA adducts accumulated in J201 cells than in primary HUFs. This difference was not reflected in the comet assay or by cell viability changes. Experiments in three additional immortal HUF cell lines (AAI49, U56, and E2-143) confirmed strong differences in DNA adduct levels compared with primary HUFs. However, these did not correlate with the protein expression of Nqo1 or Nat1/2, or with gene expression of Cyp1a1 or Cyp1b1. Our results show that using immortal HUFs as surrogates for primary HUFs in genotoxicity screening has limitations and that DNA adduct formation is the best measure of genotoxicity of the nitro-polycyclic aromatic hydrocarbons tested in HUFs. Copyright © 2011 Wiley Periodicals, Inc.

  12. Efficacy of Various Intracanal Medicaments in Human Primary Teeth with Necrotic Pulp against Candida Biofilms: An in vivo Study.

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    Paikkatt, Jophie V; Sreedharan, Sheela; Philomina, Beena; Kannan, V P; Santhakumar, Madhu; Kumar, Tv Anupam

    2017-01-01

    Candida has been associated with cases of secondary and persistent root canal infections. The purpose of this study was to evaluate and compare the effectiveness of commonly used intracanal medicament against Candida biofilms found in root canals of human primary teeth with necrotic pulp. Pulp canals of 45 single-rooted primary maxillary anterior teeth with pulp necrosis in 34 children were included in the study. They were divided into three groups of 15 samples each - group I: Ca(OH)2 (calcium hydroxide); group II: 1% chlorhexidine gel (CHX); and group III: 1% metronidazole gel. Bacterial count was obtained from each tooth at two different stages: (1) after instrumentation, and (2) after placement of the medication. Statistical analysis using the Statistical Package for the Social Sciences version 10.0 software program (Inc., Chicago, IL, USA) with Wilcoxon signed rank test after grouping the samples was performed. Ca(OH)2, 1% CHX gel, and 1% metronidazole gel were ineffective in completely eliminating Candida biofilms from root canal of human primary teeth with necrotic pulp. None of the commonly used intracanal medicaments, i.e., Ca(OH)2, 1% CHX gel, and 1% metronidazole gel, was effective in completely eliminating Candida biofilm from root canal of human primary teeth with necrotic pulp. Ineffectiveness of these medicaments against Candida has opened new door of research regarding the use of suitable intracanal medicaments against single and multispecies biofilms. Paikkatt JV, Sreedharan S, Philomina B, Kannan VP, Santhakumar M, Kumar TVA. Efficacy of Various Intracanal Medicaments in Human Primary Teeth with Necrotic Pulp against Candida Biofilms: An in vivo Study. Int J Clin Pediatr Dent 2017;10(1):45-48.

  13. Sex differences in liver toxicity-do female and male human primary hepatocytes react differently to toxicants in vitro?

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    Milena Mennecozzi

    Full Text Available There is increasing amount of evidence for sex variation in drug efficiency and toxicity profiles. Women are more susceptible than men to acute liver injury from xenobiotics. In general, this is attributed to sex differences at a physiological level as well as differences in pharmacokinetics and pharmacodynamics, but neither of these can give a sufficient explanation for the diverse responses to xenobiotics. Existing data are mainly based on animal models and limited data exist on in vitro sex differences relevant to humans. To date, male and female human hepatocytes have not yet been compared in terms of their responses to hepatotoxic drugs. We investigated whether sex-specific differences in acute hepatotoxicity can be observed in vitro by comparing hepatotoxic drug effects in male and female primary human hepatocytes. Significant sex-related differences were found for certain parameters and individual drugs, showing an overall higher sensitivity of female primary hepatocytes to hepatotoxicants. Moreover, our work demonstrated that high content screening is feasible with pooled primary human hepatocytes in suspension.

  14. Escape is a more common mechanism than avidity reduction for evasion of CD8+ T cell responses in primary human immunodeficiency virus type 1 infection

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    Williams Ian

    2011-06-01

    Full Text Available Abstract Background CD8+ T cells play an important role in control of viral replication during acute and early human immunodeficiency virus type 1 (HIV-1 infection, contributing to containment of the acute viral burst and establishment of the prognostically-important persisting viral load. Understanding mechanisms that impair CD8+ T cell-mediated control of HIV replication in primary infection is thus of importance. This study addressed the relative extent to which HIV-specific T cell responses are impacted by viral mutational escape versus reduction in response avidity during the first year of infection. Results 18 patients presenting with symptomatic primary HIV-1 infection, most of whom subsequently established moderate-high persisting viral loads, were studied. HIV-specific T cell responses were mapped in each individual and responses to a subset of optimally-defined CD8+ T cell epitopes were followed from acute infection onwards to determine whether they were escaped or declined in avidity over time. During the first year of infection, sequence variation occurred in/around 26/33 epitopes studied (79%. In 82% of cases of intra-epitopic sequence variation, the mutation was confirmed to confer escape, although T cell responses were subsequently expanded to variant sequences in some cases. In contrast, Conclusions Escape appears to constitute a much more important means of viral evasion of CD8+ T cell responses in acute and early HIV infection than decline in functional avidity of epitope-specific T cells. These findings support the design of vaccines to elicit T cell responses that are difficult for the virus to escape.

  15. The expanding universe

    CERN Document Server

    Lew, Kristi

    2011-01-01

    People have always been fascinated with the stars above and the universe that contains them. Over the years, astronomers have developed numerous theories to explain how the universe began, how it works, and what its ultimate fate will be. But all of the scientists' questions are far from answered. The Expanding Universe goes beyond the creation of the universe to explain how scientists think the universe works, grows, and changes, including what great thinkers Isaac Newton and Albert Einstein had to say about its fate. Readers will also learn about how researchers are slowly shedding light on

  16. Expanding Your Horizon 2015

    CERN Multimedia

    Kaltenhauser, Kristin

    2015-01-01

    Expanding your horizons is a bi-annual “Science Day” for girls aged 11 to 14, held at the University of Geneva on 14 November. The girls had the opportunity to take part in hands-on workshops held by local professional women in the field of science, mathematics, engineering and technology. For the fourth time, CERN was part of this event, offering three workshops as well as a booth at the Discovery Fair, including Higgnite, an interactive visualization of the Higgs Field.

  17. Quantifying Human Appropriated Net Primary Productivity (HANPP) in a Ghanaian Cocoa System

    Science.gov (United States)

    Morel, A.; Adu-Bredu, S.; Adu Sasu, M.; Ashley Asare, R.; Boyd, E.; Hirons, M. A.; Malhi, Y.; Mason, J.; Norris, K.; Robinson, E. J. Z.; McDermott, C. L.

    2015-12-01

    Ghana is the second largest producer of cocoa (Theobroma cacoa), exporting approximately 18 percent of global volumes. These cocoa farms are predominantly small-scale, ranging in size from 2-4 hectares (ha). Traditionally, the model of cocoa expansion in Ghana relied on clearing new areas of forest and establishing a farm under remnant forest trees. This is increasingly less practical due to few unprotected forest areas remaining and management practices favoring close to full sun cocoa to maximize short-term yields. This study is part of a larger project, ECOLMITS, which is an interdisciplinary, ESPA-funded[1] initiative exploring the ecological limits of ecosystem system services (ESS) for alleviating poverty in small-scale agroforestry systems. The ecological study plots are situated within and around the Kakum National Forest, a well-protected, moist-evergreen forest of the Lower Guinea Forest region. Net primary productivity (NPP) is a measure of the rate at which carbon dioxide (CO2) is incorporated into plant tissues (e.g. canopy, stem and root). For this study, NPP was monitored in situ using methods developed by the Global Environmental Monitoring Network (GEM, http://gem.tropicalforests.ox.ac.uk/). By comparing NPP measured in intact forest and farms, the human appropriated NPP (HANPP) of this system can be estimated. The forest measures provide the "potential" NPP of the region, and then the reduction in NPP for farm plots is calculated for both land-cover change (HANPPLUC) and cocoa harvesting (HANPPHARV). The results presented are of the first year of NPP measurements across the cocoa landscape, including measurements from intact forest, logged forest and cocoa farms across a shade gradient and located at varying distances from the forest edge (e.g. 100 m, 500 m, 1 km and 5 km). These measures will have implications for carbon sequestration potential over the region and long-term sustainability of the Ghanaian cocoa sector. [1] Ecosystem Services for

  18. Prognostic implication of human papillomavirus types and species in cervical cancer patients undergoing primary treatment.

    Directory of Open Access Journals (Sweden)

    Yat Ming Lau

    Full Text Available High-risk human papillomavirus (HPV types are associated with cervical cancer. It is well established that individual HPV types vary in oncogenicity, but current data on their prognostic implication remain controversial. We examined the association between HPV types/species and the survival of 236 Chinese women aged 26-87 (mean 54.4 years after receiving primary treatment for cervical cancer. Overall, 45.8% were of FIGO stage I, 41.9% stage II, and 12.3% stage III. The four most prevalent types found were HPV-16 (60.2%, HPV-18 (21.6%, HPV-52 (11.9%, and HPV-58 (9.3%. Overall, 19.5% of patients had multiple-type infections, 78.4% harboured one or more alpha-9 species, and 28.8% harboured one or more alpha-7 species. After a median follow-up of 8.0 years, 156 (66.1% patients survived. The 3-year overall survival rate was 75.5%. Factors independently associated with a poorer 3-year overall survival were age >60 years, tumour size >4 cm, lymph node involvement and treatment with radiotherapy+/-chemotherapy. Univariate analysis showed HPV-16 single-type infection was associated with a marginally poorer disease-specific survival (71.6% vs. 87.0%, HR: 1.71, 95% CI = 1.01-2.90, whereas non-HPV-16 alpha-9 species was associated with a better disease-specific survival (90.0% vs. 76.2%, HR: 0.36, 95% CI = 0.16-0.79. However, on multivariate analysis, HPV infection status irrespective of different grouping methods, including individual types, species, single-type or co-infection, did not carry any significant prognostic significance. In conclusion, we did not observe any association between infection with a particular HPV type/species and survival. An HPV type-based stratification in treatment and follow-up plan could not be recommended.

  19. Prognostic implication of human papillomavirus types and species in cervical cancer patients undergoing primary treatment.

    Science.gov (United States)

    Lau, Yat Ming; Cheung, Tak Hong; Yeo, Winnie; Mo, Frankie; Yu, Mei Yung; Lee, Kun Min; Ho, Wendy C S; Yeung, Apple C M; Law, Priscilla T Y; Chan, Paul K S

    2015-01-01

    High-risk human papillomavirus (HPV) types are associated with cervical cancer. It is well established that individual HPV types vary in oncogenicity, but current data on their prognostic implication remain controversial. We examined the association between HPV types/species and the survival of 236 Chinese women aged 26-87 (mean 54.4) years after receiving primary treatment for cervical cancer. Overall, 45.8% were of FIGO stage I, 41.9% stage II, and 12.3% stage III. The four most prevalent types found were HPV-16 (60.2%), HPV-18 (21.6%), HPV-52 (11.9%), and HPV-58 (9.3%). Overall, 19.5% of patients had multiple-type infections, 78.4% harboured one or more alpha-9 species, and 28.8% harboured one or more alpha-7 species. After a median follow-up of 8.0 years, 156 (66.1%) patients survived. The 3-year overall survival rate was 75.5%. Factors independently associated with a poorer 3-year overall survival were age >60 years, tumour size >4 cm, lymph node involvement and treatment with radiotherapy+/-chemotherapy. Univariate analysis showed HPV-16 single-type infection was associated with a marginally poorer disease-specific survival (71.6% vs. 87.0%, HR: 1.71, 95% CI = 1.01-2.90), whereas non-HPV-16 alpha-9 species was associated with a better disease-specific survival (90.0% vs. 76.2%, HR: 0.36, 95% CI = 0.16-0.79). However, on multivariate analysis, HPV infection status irrespective of different grouping methods, including individual types, species, single-type or co-infection, did not carry any significant prognostic significance. In conclusion, we did not observe any association between infection with a particular HPV type/species and survival. An HPV type-based stratification in treatment and follow-up plan could not be recommended.

  20. CD133-enriched Xeno-Free human embryonic-derived neural stem cells expand rapidly in culture and do not form teratomas in immunodeficient mice

    Directory of Open Access Journals (Sweden)

    Daniel L. Haus

    2014-09-01

    Full Text Available Common methods for the generation of human embryonic-derived neural stem cells (hNSCs result in cells with potentially compromised safety profiles due to maintenance of cells in conditions containing non-human proteins (e.g. in bovine serum or on mouse fibroblast feeders. Additionally, sufficient expansion of resulting hNSCs for scaling out or up in a clinically relevant time frame has proven to be difficult. Here, we report a strategy that produces hNSCs in completely “Xeno-Free” culture conditions. Furthermore, we have enriched the hNSCs for the cell surface marker CD133 via magnetic sorting, which has led to an increase in the expansion rate and neuronal fate specification of the hNSCs in vitro. Critically, we have also confirmed neural lineage specificity upon sorted hNSC transplantation into the immunodeficient NOD-scid mouse brain. The future use or adaptation of these protocols has the potential to better facilitate the advancement of pre-clinical strategies from the bench to the bedside.

  1. Repression of GW/P body components and the RNAi microprocessor impacts primary ciliogenesis in human astrocytes

    Directory of Open Access Journals (Sweden)

    Rattner Jerome B

    2011-08-01

    Full Text Available Abstract Background In most cells, the centriolar component of the centrosome can function as a basal body supporting the formation of a primary cilium, a non-motile sensory organelle that monitors information from the extracellular matrix and relays stimuli into the cell via associated signaling pathways. Defects in the formation and function of primary cilia underlie multiple human diseases and are hallmarks of malignancy. The RNA silencing pathway is involved in the post-transcriptional silencing of > 50% of mRNA that occurs within GW/P bodies. GW/P bodies are found throughout the cytoplasm and previously published live cell imaging data suggested that in a malignant cell type (U2OS, two GW/P bodies reside at the centrosome during interphase. This led us to investigate if a similar relationship exists in primary cells and if the inhibition of the miRNA pathway impairs primary cilium formation. Results Two GW/P bodies as marked by GW182 and hAgo2 colocalized to the basal body of primary human astrocytes as well as human synoviocytes during interphase and specifically with the distal end of the basal body in the pericentriolar region. Since it is technically challenging to examine the two centrosomal GW/P bodies in isolation, we investigated the potential relationship between the global population of GW/P bodies and primary ciliogenesis. Astrocytes were transfected with siRNA directed to GW182 and hAgo2 and unlike control astrocytes, a primary cilium was no longer associated with the centrosome as detected in indirect immunofluorescence assays. Ultrastructural analysis of siRNA transfected astrocytes revealed that knock down of GW182, hAgo2, Drosha and DGCR8 mRNA did not affect the appearance of the earliest stage of ciliogenesis but did prevent the formation and elongation of the ciliary axoneme. Conclusions This study confirms and extends a previously published report that GW/P bodies reside at the centrosome in U2OS cells and documents that

  2. Expanding the scope of anatomical sciences: the case of "Human evolution: The fossil evidence" course at the Sackler School of Medicine, Tel-Aviv University.

    Science.gov (United States)

    Notzer, Netta; Abramovitz, Ruth

    2012-01-01

    The Anatomy Department at Tel-Aviv University Medical School offers its students an elective course of 26 didactic hours on human evolution. The course is open to students from all faculties, who must fulfill all academic requirements, without a prerequisite of a background in anatomy. Approximately 120 students attend annually, a third of them are nonmedical students who major in philosophy, archeology, and sociology. This article discusses the course's contributions to students' understanding of a scientific concept that a scientific theory can be contradicted by new evidence, because facts govern science. Also, research methods of applying scientific principles establish the understanding of the human body, which evidently contributes to health and medicine. In the classes, the students are divided into mini-groups of 2-3 students, while the lecturer moves among students to examine fossils. In addition, analogies, open-discussions, and explanations accompany the tangible experiences. The lecturer of the course is an experienced anthropologist-anatomist researcher. He is a role-model and a mentor, sharing with the students his belief that a scientist should be persistent in his research to overcome difficult circumstances. Students, regardless of their backgrounds, express high appreciation of the course in their feedback questionnaires. The message conveyed by this course is that not only knowledge counts but also its integration with scientific principles. This course teaches us that science can bring students from different areas to study together and share ideas. In conclusion, this is a unique course in the eyes of the faculty and students alike. Copyright © 2012 American Association of Anatomists.

  3. Fourier Descriptors Based on the Structure of the Human Primary Visual Cortex with Applications to Object Recognition

    OpenAIRE

    Bohi, Amine; Prandi, Dario; Guis, Vincente; Bouchara, Frédéric; Gauthier, Jean-Paul

    2016-01-01

    International audience; In this paper we propose a supervised object recognition method using new global features and inspired by the model of the human primary visual cortex V1 as the semidiscrete roto-translation group $SE(2,N)=\\mathbb Z_N\\rtimes \\mathbb{R}^2$. The proposed technique is based on generalized Fourier descriptors on the latter group, which are invariant to natural geometric transformations (rotations, translations). These descriptors are then used to feed an SVM classifier. We...

  4. A Review of the Human Clinical Studies Involving Citrus aurantium (Bitter Orange) Extract and its Primary Protoalkaloid p-Synephrine

    OpenAIRE

    Stohs, Sidney J.; Preuss, Harry G.; Shara, Mohd

    2012-01-01

    This review summarizes the published as well as unpublished human studies involving Citrus aurantium (bitter orange) extract and its primary protoalkaloid p-synephrine, providing information and an assessment of the safety and efficacy of these widely used products. The results of over 20 studies involving a total of approximately 360 subjects that consumed p-synephrine alone or in combination with other ingredients are reviewed and critiqued. Over 50 % of the subjects involved in these studi...

  5. Human resources requirements for diabetic patients healthcare in primary care clinics of the Mexican Institute of Social Security

    OpenAIRE

    Doubova, Svetlana V; Claudine Ramírez-Sánchez; Alejandro Figueroa-Lara; Ricardo Pérez-Cuevas

    2013-01-01

    Objective. To estimate the requirements of human resources (HR) of two models of care for diabetes patients: conventional and specific, also called DiabetIMSS, which are provided in primary care clinics of the Mexican Institute of Social Security (IMSS). Materials and methods. An evaluative research was conducted. An expert group identified the HR activities and time required to provide healthcare consistent with the best clinical practices for diabetic patients. HR were estimated by using th...

  6. The Multifaceted Responses of Primary Human Astrocytes and Brain Microvascular Endothelial Cells to the Lyme Disease Spirochete, Borrelia Burgdorferi

    Directory of Open Access Journals (Sweden)

    Catherine A. Brissette

    2013-07-01

    Full Text Available The vector-borne pathogen, Borrelia burgdorferi, causes a multi-system disorder including neurological complications. These neurological disorders, collectively termed neuroborreliosis, can occur in up to 15% of untreated patients. The neurological symptoms are probably a result of a glial-driven, host inflammatory response to the bacterium. However, the specific contributions of individual glial and other support cell types to the pathogenesis of neuroborreliosis are relatively unexplored. The goal of this project was to characterize specific astrocyte and endothelial cell responses to B. burgdorferi. Primary human astrocytes and primary HBMEC (human brain microvascular endothelial cells were incubated with B. burgdorferi over a 72-h period and the transcriptional responses to the bacterium were analyzed by real-time PCR arrays. There was a robust increase in several surveyed chemokine and related genes, including IL (interleukin-8, for both primary astrocytes and HBMEC. Array results were confirmed with individual sets of PCR primers. The production of specific chemokines by both astrocytes and HBMEC in response to B. burgdorferi, including IL-8, CXCL-1, and CXCL-10, were confirmed by ELISA. These results demonstrate that primary astrocytes and HBMEC respond to virulent B. burgdorferi by producing a number of chemokines. These data suggest that infiltrating phagocytic cells, particularly neutrophils, attracted by chemokines expressed at the BBB (blood–brain barrier may be important contributors to the early inflammatory events associated with neuroborreliosis.

  7. Expanding the Trilinos developer community.

    Energy Technology Data Exchange (ETDEWEB)

    Heroux, Michael Allen

    2010-10-01

    The Trilinos Project started approximately nine years ago as a small effort to enable research, development and ongoing support of small, related solver software efforts. The 'Tri' in Trilinos was intended to indicate the eventual three packages we planned to develop. In 2007 the project expanded its scope to include any package that was an enabling technology for technical computing. Presently the Trilinos repository contains over 55 packages covering a broad spectrum of reusable tools for constructing full-featured scalable scientific and engineering applications. Trilinos usage is now worldwide, and many applications have an explicit dependence on Trilinos for essential capabilities. Users come from other US laboratories, universities, industry and international research groups. Awareness and use of Trilinos is growing rapidly outside of Sandia. Members of the external research community are becoming more familiar with Trilinos, its design and collaborative nature. As a result, the Trilinos project is receiving an increasing number of requests from external community members who want to contribute to Trilinos as developers. To-date we have worked with external developers in an ad hoc fashion. Going forward, we want to develop a set of policies, procedures, tools and infrastructure to simplify interactions with external developers. As we go forward with multi-laboratory efforts such as CASL and X-Stack, and international projects such as IESP, we will need a more streamlined and explicit process for making external developers 'first-class citizens' in the Trilinos development community. This document is intended to frame the discussion for expanding the Trilinos community to all strategically important external members, while at the same time preserving Sandia's primary leadership role in the project.

  8. Expanding hollow metal rings

    Science.gov (United States)

    Peacock, Harold B [Evans, GA; Imrich, Kenneth J [Grovetown, GA

    2009-03-17

    A sealing device that may expand more planar dimensions due to internal thermal expansion of a filler material. The sealing material is of a composition such that when desired environment temperatures and internal actuating pressures are reached, the sealing materials undergoes a permanent deformation. For metallic compounds, this permanent deformation occurs when the material enters the plastic deformation phase. Polymers, and other materials, may be using a sealing mechanism depending on the temperatures and corrosivity of the use. Internal pressures are generated by either rapid thermal expansion or material phase change and may include either liquid or solid to gas phase change, or in the gaseous state with significant pressure generation in accordance with the gas laws. Sealing material thickness and material composition may be used to selectively control geometric expansion of the seal such that expansion is limited to a specific facing and or geometric plane.

  9. An unusual case of primary human immunodeficiency virus infection presenting as mononucleosis-like syndrome and acute aseptic meningoencephalitis. Report of a case and review of the literature

    Directory of Open Access Journals (Sweden)

    Marcelo Corti

    2014-01-01

    Full Text Available Clinical presentation of primary human immunodeficiency virus (HIV infection includes a wide spectrum of manifestations from asymptomatic infection to a symptomatic and severe illness. Central nervous system involvement should be always considered as a severe clinical form of primary HIV infection. Physicians should be aware to the broad clinical spectrum of primary HIV infection. We report a case of a female with diagnosis of mononucleosis-like syndrome and acute aseptic meningoencephalitis during primary HIV infection.

  10. CPM Is a Useful Cell Surface Marker to Isolate Expandable Bi-Potential Liver Progenitor Cells Derived from Human iPS Cells

    Directory of Open Access Journals (Sweden)

    Taketomo Kido

    2015-10-01

    Full Text Available To develop a culture system for large-scale production of mature hepatocytes, liver progenitor cells (LPCs with a high proliferation potential would be advantageous. We have found that carboxypeptidase M (CPM is highly expressed in embryonic LPCs, hepatoblasts, while its expression is decreased along with hepatic maturation. Consistently, CPM expression was transiently induced during hepatic specification from human-induced pluripotent stem cells (hiPSCs. CPM+ cells isolated from differentiated hiPSCs at the immature hepatocyte stage proliferated extensively in vitro and expressed a set of genes that were typical of hepatoblasts. Moreover, the CPM+ cells exhibited a mature hepatocyte phenotype after induction of hepatic maturation and also underwent cholangiocytic differentiation in a three-dimensional culture system. These results indicated that hiPSC-derived CPM+ cells share the characteristics of LPCs, with the potential to proliferate and differentiate bi-directionally. Thus, CPM is a useful marker for isolating hiPSC-derived LPCs, which allows development of a large-scale culture system for producing hepatocytes and cholangiocytes.

  11. "Voice Forum" The Human Voice as Primary Instrument in Music Therapy

    DEFF Research Database (Denmark)

    Pedersen, Inge Nygaard; Storm, Sanne

    2009-01-01

    Aspects will be drawn on the human voice as tool for embodying our psychological and physiological state, and attempting integration of feelings. Presentations and dialogues on different methods and techniques in "Therapy related body-and voice work.", as well as the human voice as a tool for non...... for nonverbal orientation and information both to our selves and others. Focus on training on the voice instrument, the effect and impact of the human voice, and listening perspectives...

  12. Nanostructural effect of acid-etching and fluoride application on human primary and permanent tooth enamels

    Energy Technology Data Exchange (ETDEWEB)

    Cheong, Youjin [Department of Biomedical Engineering and Healthcare Industry Research Institute, College of Medicine, Kyung Hee University, Seoul (Korea, Republic of); Choi, Samjin [Department of Biomedical Engineering and Healthcare Industry Research Institute, College of Medicine, Kyung Hee University, Seoul (Korea, Republic of); Department of Orthodontics, College of Dental Medicine, Kyung Hee University, Seoul (Korea, Republic of); Kim, So Jung [Department of Pediatric Dentistry, College of Dental Medicine, Kyung Hee University, Seoul (Korea, Republic of); Park, Hun-Kuk, E-mail: sigmoidus@khu.ac.kr [Department of Biomedical Engineering and Healthcare Industry Research Institute, College of Medicine, Kyung Hee University, Seoul (Korea, Republic of); Program of Medical Engineering, Kyung Hee University, Seoul (Korea, Republic of)

    2012-07-01

    This study examined the nanostructural effects of fluoride application and the acid-etching time with respect to the time elapsed after fluoride application on the primary and permanent tooth enamel layers using atomic force microscopy (AFM) and scanning electron microscopy (SEM). 192 non-carious teeth were assigned to sixteen experimental groups (n = 12) including primary (1 to 8) and permanent (9 to 16) teeth, based on the timing of acid-etching with 37% phosphoric acid after an acidulated phosphate fluoride (APF) pre-treatment. The APF pre-treatment led to a decrease in surface roughness in both the primary and permanent teeth. After the APF treatment, the roughness in both primary and permanent teeth increased with the time elapsed. An acid-etching time of 40 s led to increased nanostructural changes in the enamel surfaces compared to the conventional acid-etching time of 20 s. This acid-etching process led to a higher roughness changes in the primary teeth than in the permanent teeth. To obtain proper enamel adhesion of a sealant after APF pre-treatment, it is important to apply acid-etching two weeks after pre-treatment. In addition, the acid-etching time should be prolonged to apply etching more quickly than two weeks, regardless of the primary and permanent teeth. Highlights: Black-Right-Pointing-Pointer APF pre-treatment led to decreased surface roughness in the enamel. Black-Right-Pointing-Pointer After APF treatment, the more roughness increased with increasing time elapsed. Black-Right-Pointing-Pointer Acid-etching should be performed two weeks after fluoride application.

  13. Genomic and phenotypic profiles of two Brazilian breast cancer cell lines derived from primary human tumors

    DEFF Research Database (Denmark)

    Corrêa, Natássia C R; Kuasne, Hellen; Faria, Jerusa A Q A

    2013-01-01

    Breast cancer is the most common type of cancer among women worldwide. Research using breast cancer cell lines derived from primary tumors may provide valuable additional knowledge regarding this type of cancer. Therefore, the aim of this study was to investigate the phenotypic profiles of MACL-1...... and MGSO-3, the only Brazilian breast cancer cell lines available for comparative studies. We evaluated the presence of hormone receptors, proliferation, differentiation and stem cell markers, using immunohistochemical staining of the primary tumor, cultured cells and xenografts implanted...

  14. Human embryonic stem cells in culture possess primary cilia with hedgehog signaling machinery

    DEFF Research Database (Denmark)

    Kiprilov, Enko N; Awan, Aashir; Desprat, Romain

    2008-01-01

    are present in three undifferentiated hESC lines. EM reveals the characteristic 9 + 0 axoneme. The number and length of cilia increase after serum starvation. Important components of the hedgehog (Hh) pathway, including smoothened, patched 1 (Ptc1), and Gli1 and 2, are present in the cilia. Stimulation...... of the pathway results in the concerted movement of Ptc1 out of, and smoothened into, the primary cilium as well as up-regulation of GLI1 and PTC1. These findings show that hESCs contain primary cilia associated with working Hh machinery. Udgivelsesdato: 2008-Mar-10...

  15. Expression of CD44 splice variants in human primary brain tumors

    NARCIS (Netherlands)

    Kaaijk, P.; Troost, D.; Morsink, F.; Keehnen, R. M.; Leenstra, S.; Bosch, D. A.; Pals, S. T.

    1995-01-01

    Expression of CD44, particularly of certain splice variants, has been linked to tumor progression and metastatic potential in a number of different animal and human cancers. Although differential expression of CD44 standard epitopes (CD44s) in human brain tumors has been reported, the expression of

  16. The human beta 2-microglobulin gene. Primary structure and definition of the transcriptional unit

    NARCIS (Netherlands)

    Güssow, D.; REIN, R.; GINJAAR, I.; Hochstenbach, F.; SEEMANN, G.; KOTTMAN, A.; Ploegh, H. L.

    1987-01-01

    The human genomic clone pb2m13 contains a functional beta 2-microglobulin (B2m) gene, which upon transfection is readily expressed in murine fibroblasts. Here we report the nucleotide sequence of the human beta 2m gene and of a nearly full length cDNA clone. A comparison with the murine beta 2m gene

  17. Biokinetics of chlorpromazine in primary rat and human hepatocytes and human HepaRG cells after repeated exposure

    NARCIS (Netherlands)

    Broeders, Jessica J W; Parmentier, Céline; Truisi, Germaine L; Jossé, Rozenn; Alexandre, Eliane; Savary, Camille C; Hewitt, Philip G; Mueller, Stefan O; Guillouzo, André; Richert, Lysiane; van Eijkeren, Jan C H; Hermens, Joop L M; Blaauboer, Bas J

    2015-01-01

    Since drug induced liver injury is difficult to predict in animal models, more representative tests are needed to better evaluate these effects in humans. Existing in vitro systems hold great potential to detect hepatotoxicity of pharmaceuticals. In this study, the in vitro biokinetics of the model

  18. Expression and regulation of Schlafen (SLFN family members in primary human monocytes, monocyte-derived dendritic cells and T cells

    Directory of Open Access Journals (Sweden)

    Alexander Puck

    2015-01-01

    Full Text Available Schlafen (SLFN/Slfn family members have been investigated for their involvement in fundamental cellular processes including growth regulation, differentiation and control of viral replication. However, most research has been focused on the characterization of Slfns within the murine system or in human cell lines. Since little is known about SLFNs in primary human immune cells, we set out to analyze the expression and regulation of the six human SLFN genes in monocytes, monocyte-derived dendritic cells (moDCs and T cells. Comparison of SLFN gene expression across these three cell types showed high mRNA expression of SLFN11 in monocytes and moDCs and high SLFN5 expression in T cells, indicating functional importance within these cell types. Differentiation of monocytes to moDCs leads to the gradual upregulation of SLFN12L and SLFN13 while SLFN12 levels were decreased by differentiation stimuli. Stimulation of moDCs via human rhinovirus, lipopolysaccharide, or IFN-α lead to strong upregulation of SLFN gene expression, while peptidoglycan poorly stimulated regulation of both SLFNs and the classical interferon-stimulated gene MxA. T cell activation was found to downregulate the expression of SLFN5, SLFN12 and SLFN12L, which was reversible upon addition of exogenous IFN-α. In conclusion, we demonstrate, that SLFN gene upregulation is mainly dependent on autocrine type I interferon signaling in primary human immune cells. Rapid decrease of SLFN expression levels following T cell receptor stimulation indicates a role of SLFNs in the regulation of human T cell quiescence.

  19. Concordant testing results between various Human Papillomavirus assays in primary cervical cancer screening

    DEFF Research Database (Denmark)

    de Thurah, Lena; Bonde, Jesper; Hoa Lam, Janni Uyen

    2018-01-01

    (HC2) and other assays. METHODS: We searched PubMed, Embase and Scopus for studies of primary screening with HC2 and ≥one more assay, with cross-tabulated testing results for the assays. Two authors applied inclusion criteria and three authors extracted data from included studies. For each inter...

  20. Differential heat shock response of primary human cell cultures and established cell lines

    DEFF Research Database (Denmark)

    Richter, W W; Issinger, O G

    1986-01-01

    degrees C treatment, whereas in immortalized cell lines usually 90% of the cells were found in suspension. Enhanced expression of the major heat shock protein (hsp 70) was found in all heat-treated cells. In contrast to the primary cell cultures, established and transformed cell lines synthesized...

  1. Identification of genetic disparity between primary and metastatic melanoma in human patients.

    Science.gov (United States)

    Swoboda, Alexander; Rasin-Streden, Doris; Schanab, Oliver; Okamoto, Ichiro; Pehamberger, Hubert; Petzelbauer, Peter; Mikula, Mario

    2011-09-01

    It is commonly accepted that cancer cell progression is accompanied by accumulation of genetic changes. Here we searched for copy number variations in melanoma and asked whether homozygous losses always cumulate during tumor cell progression. Therefore we investigated either melanoma cell lines or tissue derived from the primary lesion and from the lymph node metastasis of the same individual patient. In vitro studies of melanoma cell lines revealed high migratory and anchorage independent growth of metastasis-derived cells. Surprisingly, whole genome DNA analysis of a primum-derived cell line revealed a total of 10 homozygous losses, whereas the matched metastasis-derived cell line only shared five of those losses. We further tested these cells in a mouse model for intradermal melanoma growth and detected fast growth of the metastasis-derived cell line and no growth of primum-derived cells. Additionally, we screened matched pairs of patient-derived melanoma primum and metastasis samples and we could also identify a case with homozygous deletions exclusively present in the primary lesion. Therefore, we suggest that tumor cell progression at the metastatic niche can occur parallel and independently from the primary tumor. We propose that for mutation-targeted therapy genotyping should be performed not only from primary, but also from metastatic melanoma. Copyright © 2011 Wiley-Liss, Inc.

  2. Arsenite Effects on Mitochondrial Bioenergetics in Human and Mouse Primary Hepatocytes Follow a Nonlinear Dose Response

    Directory of Open Access Journals (Sweden)

    Hemantkumar Chavan

    2017-01-01

    Full Text Available Arsenite is a known carcinogen and its exposure has been implicated in a variety of noncarcinogenic health concerns. Increased oxidative stress is thought to be the primary cause of arsenite toxicity and the toxic effect is thought to be linear with detrimental effects reported at all concentrations of arsenite. But the paradigm of linear dose response in arsenite toxicity is shifting. In the present study we demonstrate that arsenite effects on mitochondrial respiration in primary hepatocytes follow a nonlinear dose response. In vitro exposure of primary hepatocytes to an environmentally relevant, moderate level of arsenite results in increased oxidant production that appears to arise from changes in the expression and activity of respiratory Complex I of the mitochondrial proton circuit. In primary hepatocytes the excess oxidant production appears to elicit adaptive responses that promote resistance to oxidative stress and a propensity to increased proliferation. Taken together, these results suggest a nonlinear dose-response characteristic of arsenite with low-dose arsenite promoting adaptive responses in a process known as mitohormesis, with transient increase in ROS levels acting as transducers of arsenite-induced mitohormesis.

  3. In vitro growth characteristics of human lymphoid malignancies in primary cell culture

    NARCIS (Netherlands)

    I.P. Touw (Ivo)

    1986-01-01

    textabstractThe experimental work presented in this thesis deals with the analysis of the in vitro growth and differentiation characteristics of acute lymphoblastic leukemia ( T end non-T), T cell non Hodgkin's lymphoma and B cell chronic lymphocytic leukemia in primary cell culture. For

  4. Effects of air-polishing devices with different abrasives on bovine primary and second teeth and deciduous human teeth.

    Science.gov (United States)

    Khalefa, Mohammad; Finke, Christian; Jost-Brinkmann, Paul-Georg

    2013-09-01

    The present study investigates the effect of air polishing using different combinations of devices and abrasive powders on bovine secondary and primary dentition and on human deciduous teeth. Lower incisors of freshly slaughtered calves and cows were partially embedded in polyurethane and polished flat. Human deciduous incisors and second molars were subjected to the same procedure. These various tooth types (bovine secondary, bovine primary, human deciduous) were then randomly assigned to 22 groups containing 10 teeth each. The specimens in each group were treated by a specific combination of an air-polishing device (n=2; PROPHYflex 3®, Air-Flow® Handy 2+) and an abrasive (n=3; Air-Flow® Pulver Classic, ClinPro™ Prophy Powder, PROPHYpearls®) applied from a distance of 5±0.5 mm at maximum setting for 60 s. Additional groups of specimens were polished with CCS® 40 or Cleanic® pastes applied with a rotating brush at low speed. A Perthometer PCV profilometer was used to analyze the degrees of surface roughness and enamel reduction in each group. PROPHYpearls® created significantly (pteeth than on bovine primary or human deciduous teeth, with no significant difference between the latter two. The degrees of surface roughness induced by air polishing surpassed the effect of CCS® 40 paste but resembled the effect of Cleanic® paste. Both Air-Flow® Handy 2+ and PROPHYflex 3® are appropriate devices to remove plaque and discoloration from the surface of deciduous teeth. PROPHYpearls® powder is excessively aggressive.

  5. Predictivity of dog co-culture model, primary human hepatocytes and HepG2 cells for the detection of hepatotoxic drugs in humans

    Energy Technology Data Exchange (ETDEWEB)

    Atienzar, Franck A., E-mail: franck.atienzar@ucb.com [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium); Novik, Eric I. [H mu rel Corporation, 675 U.S. Highway 1, North Brunswick, NJ 08902 (United States); Gerets, Helga H. [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium); Parekh, Amit [H mu rel Corporation, 675 U.S. Highway 1, North Brunswick, NJ 08902 (United States); Delatour, Claude; Cardenas, Alvaro [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium); MacDonald, James [Chrysalis Pharma Consulting, LLC, 385 Route 24, Suite 1G, Chester, NJ 07930 (United States); Yarmush, Martin L. [Department of Biomedical Engineering, Rutgers University, Piscataway, NJ 08854 (United States); Dhalluin, Stéphane [UCB Pharma SA, Non-Clinical Development, Chemin du Foriest, 1420 Braine-l' Alleud (Belgium)

    2014-02-15

    Drug Induced Liver Injury (DILI) is a major cause of attrition during early and late stage drug development. Consequently, there is a need to develop better in vitro primary hepatocyte models from different species for predicting hepatotoxicity in both animals and humans early in drug development. Dog is often chosen as the non-rodent species for toxicology studies. Unfortunately, dog in vitro models allowing long term cultures are not available. The objective of the present manuscript is to describe the development of a co-culture dog model for predicting hepatotoxic drugs in humans and to compare the predictivity of the canine model along with primary human hepatocytes and HepG2 cells. After rigorous optimization, the dog co-culture model displayed metabolic capacities that were maintained up to 2 weeks which indicates that such model could be also used for long term metabolism studies. Most of the human hepatotoxic drugs were detected with a sensitivity of approximately 80% (n = 40) for the three cellular models. Nevertheless, the specificity was low approximately 40% for the HepG2 cells and hepatocytes compared to 72.7% for the canine model (n = 11). Furthermore, the dog co-culture model showed a higher superiority for the classification of 5 pairs of close structural analogs with different DILI concerns in comparison to both human cellular models. Finally, the reproducibility of the canine system was also satisfactory with a coefficient of correlation of 75.2% (n = 14). Overall, the present manuscript indicates that the dog co-culture model may represent a relevant tool to perform chronic hepatotoxicity and metabolism studies. - Highlights: • Importance of species differences in drug development. • Relevance of dog co-culture model for metabolism and toxicology studies. • Hepatotoxicity: higher predictivity of dog co-culture vs HepG2 and human hepatocytes.

  6. siRNA delivery into cultured primary human myoblasts--optimization of electroporation parameters and theoretical analysis.

    Science.gov (United States)

    Lojk, Jasna; Mis, Katarina; Pirkmajer, Sergej; Pavlin, Mojca

    2015-12-01

    Introduction of genetic material into muscle tissue has been extensively researched, including isolation and in vitro expansion of primary myoblasts as a potential source of cells for skeletal and heart muscle tissue engineering applications. In this study, we optimized the electroporation protocol for introduction of short interfering ribonucleic acid (siRNA) against messenger RNA for Hypoxia Inducible Factor 1α (HIF-1α) into cultured primary human myoblasts. We established optimal pulsing protocol for siRNA electro transfection, and theoretically analyzed the effect of electric field and pulse duration on silencing efficiency and electrophoretic displacement of siRNA. Silencing of HIF-1α was determined with quantitative polymerase chain reaction and Western Blot. The most efficient silencing (71% knockdown) was achieved with 8 × 2 ms pulses, E = 0.6 kV/cm. Viability was determined immediately, 1 h and 48 h after electroporation. In general, there was a trade-off between efficient silencing and preserved viability. Electric field and pulse duration are crucial parameters for silencing, since both increase membrane permeabilization and electrophoretic transfer of siRNA. Short-term viability showed immediate toxicity of pulses due to membrane damage, while indirect effects on cell proliferation were observed after 48 h. Presented results are important for faster optimization of electroporation parameters for ex vivo electrotransfer of short RNA molecules into primary human myoblasts. © 2015 Wiley Periodicals, Inc.

  7. Expander chunked codes

    Science.gov (United States)

    Tang, Bin; Yang, Shenghao; Ye, Baoliu; Yin, Yitong; Lu, Sanglu

    2015-12-01

    Chunked codes are efficient random linear network coding (RLNC) schemes with low computational cost, where the input packets are encoded into small chunks (i.e., subsets of the coded packets). During the network transmission, RLNC is performed within each chunk. In this paper, we first introduce a simple transfer matrix model to characterize the transmission of chunks and derive some basic properties of the model to facilitate the performance analysis. We then focus on the design of overlapped chunked codes, a class of chunked codes whose chunks are non-disjoint subsets of input packets, which are of special interest since they can be encoded with negligible computational cost and in a causal fashion. We propose expander chunked (EC) codes, the first class of overlapped chunked codes that have an analyzable performance, where the construction of the chunks makes use of regular graphs. Numerical and simulation results show that in some practical settings, EC codes can achieve rates within 91 to 97 % of the optimum and outperform the state-of-the-art overlapped chunked codes significantly.

  8. Are "Human Factors" the Primary Cause of Complications in the Field of Implant Dentistry?

    Science.gov (United States)

    Renouard, Franck; Amalberti, René; Renouard, Erell

    Complications in medicine and dentistry are usually analyzed from a purely technical point of view. Rarely is the role of human behavior or judgment considered as a reason for adverse outcomes. When the role of human factors is considered, these are usually described in general terms rather than specifically identifying the factors responsible for an adverse event. The impact of cognitive and behavioral factors in the explanation of adverse events has been studied in other high-stakes areas such as aviation and nuclear power. Specific protocols have been developed to reduce rates of human error, and, where human error is unavoidable, to lessen its impact. This approach has dramatically reduced the incidence of accidents in these fields. This article aims to review how a similar approach may prove valuable in the reduction of complications in implant dentistry.

  9. Analysis of adequacy levels for human resources improvement within primary health care framework in Africa

    National Research Council Canada - National Science Library

    Parent, Florence; Fromageot, Audrey; Coppieters, Yves; Lejeune, Colette; Lemenu, Dominique; Garant, Michèle; Piette, Danielle; Levêque, Alain; De Ketele, Jean-Marie

    2005-01-01

    Human resources in health care system in sub-Saharan Africa are generally picturing a lack of adequacy between expected skills from the professionals and health care needs expressed by the populations...

  10. A Primary Human Critical Success Factors Model for the ERP System Implementation

    National Research Council Canada - National Science Library

    Aleksander Jenko; Matjaž Roblek

    2016-01-01

    ...) and the different causes of ERP implementation project failures. Despite a detailed literature preview, we were unable to find an appropriate research with a comprehensive overview of the true causes behind CSFs, observed from a human factors perspective...

  11. Cytopathic effect of Human cosavirus (HCoSV) on primary cell cultures of human embryonic lung MRC5.

    Science.gov (United States)

    Rezig, Dorra; Touzi, Henda; Meddeb, Zina; Triki, Henda

    2014-10-01

    Human cosaviruses (HCoSVs) are newly discovered viruses in Picornaviridae family. Until now, most published studies reported HCoSV detection using molecular techniques and genetic characterization of the virus. Nevertheless, no laboratory has yet reported the replication of these viruses in cultured cell lines. In the present work, the propagation of HCoSV strains isolated from human fecal specimens in MRC5 cell line and their induced cytopathic effects (CPE) was studied. The first sign of virus growth was observed 24-48h after inoculation. The cells rounded up and clumped together rapidly; empty areas became visible and, on the third day of CPE, a remarkable decrease in live cells was observed. This represents the first report on in vitro model of HCoSV replication which opens up opportunities for future investigations of these new viruses. Copyright © 2014 Elsevier B.V. All rights reserved.

  12. Direct Phenotypical and Functional Dysregulation of Primary Human B Cells by Human Immunodeficiency Virus (HIV) Type 1 In Vitro

    OpenAIRE

    Ana Judith Perisé-Barrios; María Ángeles Muñoz-Fernandez; Marjorie Pion

    2012-01-01

    BACKGROUND: Human immunodeficiency virus type 1 (HIV-1) induces a general dysregulation of immune system. Dysregulation of B cell compartment is generally thought to be induced by HIV-related immune activation and lymphopenia. However, a direct influence of HIV-1 particles on B cells was recently proposed as the third pathway of B cells dysregulation. METHODS/PRINCIPAL FINDINGS: We evaluated the direct and specific consequences of HIV-1 contact on activation, survival, proliferation and pheno...

  13. Effect of milk and soy-based infant formulas on in situ demineralization of human primary enamel.

    Science.gov (United States)

    de Mazer Papa, Anna Maria Cia; Tabchoury, Cínthia Pereira Machado; Del Bel Cury, Altair Antoninha; Tenuta, Livia Maria Andaló; Arthur, Rodrigo Alex; Cury, Jaime Aparecido

    2010-01-01

    The effect of infant formulas sweetened or not with sucrose on primary enamel demineralization and dental biofilm is not known. This study's purpose was to evaluate the effect of infant formula and sucrose association on demineralization of primary enamel and dental biofilm formed. A cross-over, split-mouth, and blind in situ study was conducted during 3 phases, each of 10 days duration, during which 11 volunteers wore palatal appliances containing slabs of human primary enamel. The dental slabs were extraorally subjected 8 times a day to 6 groups of treatment: distilled and deionized water; 10% sucrose solution; and milk-based and soy-based formula with or without 10% sucrose added. After each phase, the acidogenicity, biochemical and microbiological composition of dental biofilm formed was analyzed, and enamel demineralization was assessed by microhardness. Both formulas induced significant enamel mineral loss, which increased when sucrose was added. Both formulas were fermented, resulting in a decrease of biofilm pH, irrespective of sucrose addition. Also, lactobacilli counts were higher in biofilm formed in the presence of both formulas when compared to the water group. Milk and soy-based formulas present potential to induce demineralization in primary enamel, which increased when sweetened with sucrose.

  14. Primary human cytomegalovirus infection induces the expansion of virus-specific activated and atypical memory B cells.

    Science.gov (United States)

    Dauby, Nicolas; Kummert, Caroline; Lecomte, Sandra; Liesnard, Corinne; Delforge, Marie-Luce; Donner, Catherine; Marchant, Arnaud

    2014-10-15

    Although neutralizing antibodies play a central role in the control of cytomegalovirus (CMV) dissemination, little is known about the response of B lymphocytes to primary human CMV infection. The proportion, phenotype, specificity, and functionality of B-cell subsets were studied in a cohort of pregnant women with primary CMV infection. CMV-seronegative pregnant women, as well as CMV-seronegative and CMV-seropositive healthy adults, were included as controls. Primary CMV infection was associated with a sustained expansion of activated (CD27(+)CD20(+)CD21(low)) and atypical (CD27(-)CD20(+)CD21(low)) memory B cells (MBCs). Both subsets expressed an effector phenotype, and their proportions were correlated with viremia. Activated MBCs expressed high levels of activation markers and included high frequencies of tumor necrosis α (TNF-α)-producing cells, whereas atypical MBCs expressed high levels of inhibitory receptors and had low TNF-α responses. Fluorescent-labeled antigen experiments indicated that activated and atypical MBCs were enriched in CMV-specific cells. Primary CMV infection mobilizes a large pool of memory B cells that includes activated and atypical MBCs. The functional regulation of CMV-specific MBCs may limit the production of antibodies and the control of viral dissemination. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  15. Pulp tissue response to Portland cement associated with different radio pacifying agents on pulpotomy of human primary molars.

    Science.gov (United States)

    Marques, N; Lourenço Neto, N; Fernandes, A P; Rodini, C; Hungaro Duarte, M; Rios, D; Machado, M A; Oliveira, T

    2015-12-01

    The objective of this research was to evaluate the response of Portland cement associated with different radio pacifying agents on pulp treatment of human primary teeth by clinical and radiographic exams and microscopic analysis. Thirty mandibular primary molars were randomly divided into the following groups: Group I - Portland cement; Group II - Portland cement with iodoform (Portland cement + CHI3 ); Group III - Portland cement with zirconium oxide (Portland cement + ZrO2 ); and treated by pulpotomy technique (removal of a portion of the pulp aiming to maintain the vitally of the remaining radicular pulp tissue using a therapeutic dressing). Clinical and radiographic evaluations were recorded at 6, 12 and 24 months follow-up. The teeth at the regular exfoliation period were extracted and processed for histological analysis. Data were tested using statistical analysis with a significance level of 5%. The microscopic findings were descriptively analysed. All treated teeth were clinically and radiographically successful at follow-up appointments. The microscopic analysis revealed positive response to pulp repair with hard tissue barrier formation and pulp calcification in the remaining roots of all available teeth. The findings of this study suggest that primary teeth pulp tissue exhibited satisfactory biological response to Portland cement associated with radio pacifying agents. However, further studies with long-term follow-up are needed to determine the safe clinical indication of this alternative material for pulp therapy of primary teeth. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

  16. Expression of SHH signaling molecules in the developing human primary dentition

    Science.gov (United States)

    2013-01-01

    Background Our current knowledge on tooth development derives primarily from studies in mice. Very little is known about gene expression and function during human odontogenesis. Sonic Hedgehog (SHH) signaling has been demonstrated to play crucial roles in the development of multiple organs in mice, including the tooth. However, if SHH signaling molecules are expressed and function in the developing human embryonic tooth remain unknown. Results We conducted microarray assay to reveal the expression profile of SHH signaling pathway molecules. We then used in situ hybridization to validate and reveal spatial and temporal expression patterns of a number of selected molecules, including SHH, PTC1, SMO, GLI1, GLI2, and GLI3, in the developing human embryonic tooth germs, and compared them with that in mice. We found that all these genes exhibit similar but slightly distinct expression patterns in the human and mouse tooth germ at the cap and bell stages. Conclusions Our results demonstrate the operation of active SHH signaling in the developing human tooth and suggest a conserved function of SHH signaling pathway during human odontogenesis. PMID:23566240

  17. Quantitative Assessment of the Human Appropriation of Net Primary Production (HANPP in the Coastal Areas of Jiangsu, China

    Directory of Open Access Journals (Sweden)

    Fangyi Zhang

    2015-11-01

    Full Text Available Global increases in population and consumption have raised concerns regarding the sustainability of the current and future use of natural resources. The human appropriation of net primary production (HANPP provides a useful measure for determining human-derived alterations in the amount of biomass that is available in ecosystems each year. HANPP was calculated based on vegetation modelling, agricultural statistics, and remote sensing data on land use and land cover to assess the human impacts on ecosystems in the coastal areas of Jiangsu, China. The results showed that HANPP increased from 332 g·C/m2/year in 2000 to 442 g·C/m2/year in 2010, with an average annual increase of 2.9%. The proportion of appropriated net primary production increased from 50.3% to 71.0% of NPPpot, mainly driven by HANPPharv (harvested NPP with an increase from 45.2% to 61.3% of NPPpot. Additionally, the spatial variation in average HANPP was striking among counties in the observed period with the lowest and highest values of 21.8% and 63.8% of NPPpot, respectively. Further analysis showed that observed levels of HANPP are high due to a high level of biomass harvest from cropland and the increases in fertilizer use, farmland irrigation rate and population and economic growth explain the trends in HANPP in the coastal areas of Jiangsu.

  18. The research of morphological variations and sexual dimorphism of primary grooves on the medial side of brain hemispheres in humans

    Directory of Open Access Journals (Sweden)

    Spasojević Goran

    2012-01-01

    Full Text Available Morphological studies of the various parts of the brain show certain morphological and morphometric differences in correlation with sex, so-called sexual dimorphism of the brain. Our research has been done on the cerebral hemispheres, taken from cadavers of both sexes and different age without pathological processes in the brain. The sample comprised 26 male brains and 16 female brains. We studied three primary grooves (sulcus cinguli, sulcus parietooccipitalis and sulcus calcarinus of the medial surface of the human cerebral hemispheres. We conducted morphological typology of grooves and morphometric measurements of primary brain grooves length in relation to sex and side of hemisphere. The results showed a statistically significant sex difference in the cingulate sulcus length (p0,05. Determined morphometric sexual dimorphism in cingulate sulcus length is significant because it implies the correlation between morphology and function of the explored areas of the cerebral cortex.

  19. Individual differences on immunostimulatory activity of raw and black garlic extract in human primary immune cells.

    Science.gov (United States)

    Purev, Uranchimeg; Chung, Mi Ja; Oh, Deog-Hwan

    2012-08-01

    The immunostimulatory activities of garlic extract using a cell line or animal models have been reported; however, no previous studies have evaluated individual differences in regards to the immunostimulatory activities. The immunostimulatory activities such as cell proliferation, tumor necrosis factor (TNF-α) and nitric oxides (NO) production of raw garlic and black garlic extracts on individual primary lymphocytes or macrophages isolated from the blood of 21 volunteers were evaluated. The antioxidant and anticancer effects of raw garlic and black garlic ethanol extract was measured to determine the optimum conditions for extraction. The 70% ethanol black garlic extracts at 70°C for 12 h (70% BGE) showed the strongest antioxidant and anticancer activities. Immunostimulatory activities of garlic extracts extracted under optimal condition on primary immune cells obtained from 21 volunteers were analyzed. Results showed that the cell proliferation, TNF-α and NO production of primary immune cells treated with 70% raw garlic extract (70% RGE) were significantly different; however, little difference was observed for the 70% BGE treatment. BGE showed stronger immunostimulatory activities than RGE. These results indicate that the immunostimulatory activities of RGE and BGE can be strongly correlated with the antioxidant and anticancer activities. Determination of immunostimulatory activities of different types of garlic using immune cells isolated from volunteers was dependent on the individual constituents due to changes in the composition of garlic during processing. Individual primary immune cells might be used as important tools to determine individual differences in all food ingredients for the development of personalized immunostimulatory active foods.

  20. Neural systems underlying aversive conditioning in humans with primary and secondary reinforcers

    Directory of Open Access Journals (Sweden)

    Mauricio R Delgado

    2011-05-01

    Full Text Available Money is a secondary reinforcer commonly used across a range of disciplines in experimental paradigms investigating reward learning and decision-making. The effectiveness of monetary reinforcers during aversive learning and its neural basis, however, remains a topic of debate. Specifically, it is unclear if the initial acquisition of aversive representations of monetary losses depends on similar neural systems as more traditional aversive conditioning that involves primary reinforcers. This study contrasts the efficacy of a biologically defined primary reinforcer (shock and a socially defined secondary reinforcer (money during aversive learning and its associated neural circuitry. During a two-part experiment, participants first played a gambling game where wins and losses were based on performance to gain an experimental bank. Participants were then exposed to two separate aversive conditioning sessions. In one session, a primary reinforcer (mild shock served as an unconditioned stimulus (US and was paired with one of two colored squares, the conditioned stimuli (CS+ and CS-, respectively. In another session, a secondary reinforcer (loss of money served as the US and was paired with one of two different CS. Skin conductance responses were greater for CS+ compared to CS- trials irrespective of type of reinforcer. Neuroimaging results revealed that the striatum, a region typically linked with reward-related processing, was found to be involved in the acquisition of aversive conditioned response irrespective of reinforcer type. In contrast, the amygdala was involved during aversive conditioning with primary reinforcers, as suggested by both an exploratory fMRI analysis and a follow-up case study with a patient with bilateral amygdala damage. Taken together, these results suggest that learning about potential monetary losses may depend on reinforcement learning related systems, rather than on typical structures involved in more biologically based

  1. Optimizing Transfection of Primary Human Umbilical Vein Endothelial Cells Using Commercially Available Chemical Transfection Reagents

    OpenAIRE

    Hunt, Michelle A.; Currie, Margaret J.; Robinson, Bridget A.; Dachs, Gabi U.

    2010-01-01

    Primary cells, such as HUVEC, are notoriously difficult to transfect and are susceptible to the toxic effects of transfection reagents. A transfection reagent with a high transfection efficiency and low cytotoxicity was sought to retain sufficient viability of transfected HUVEC for subsequent assays. Nine chemical transfection reagents, currently commercially available, were compared for their ability to transfect HUVEC in vitro. A plasmid expressing the enhanced GFP (EGFP) was used for trans...

  2. Magnesium corrosion particles do not interfere with the immune function of primary human and murine macrophages

    OpenAIRE

    Roth, Isabelle; Schumacher, Stephan; Basler, Tina; Baumert, Kathrin; Seitz, Jan-Marten; Evertz, Florian; Mueller, Peter Paul; Baeumer, Wolfgang; Kietzmann, Manfred

    2015-01-01

    Magnesium is currently under investigation as a prospective biodegradable implant material. Biodegradation of magnesium causes a release of magnesium, hydroxide ions and hydrogen gas but it can also lead to the formation of particulate debris. Implant-derived particles may have immunotoxic effects. To investigate the influence of magnesium-derived particles on the immune functions of primary macrophages, up to 500 mu g/ml magnesium or magnesium corrosion particles were added to the cell cultu...

  3. Knowledge of primary health care and career choice at primary health care settings among final year medical students - challenges to human resources for health in Vietnam.

    Science.gov (United States)

    Giang, Kim Bao; Minh, Hoang Van; Hien, Nguyen Van; Ngoc, Nguyen Minh; Hinh, Nguyen Duc

    2015-01-01

    There is a shortage of medical doctors in primary health care (PHC) settings in Vietnam. Evidence about the knowledge medical students have about PHC and their career decision-making is important for making policy in human resources for health. The objective of this study was to analyse knowledge and attitudes about PHC among medical students in their final year and their choice to work in PHC after graduation. A cross-sectional study was conducted among 400 final year general medical students from Hanoi Medical University. Self-administered interviews were conducted. Key variables were knowledge, awareness of the importance of PHC and PHC career choices. Descriptive and analytic statistics were performed. Students had essential knowledge of the concept and elements of PHC and were well aware of its importance. However, only one-third to one half of them valued PHC with regard to their professional development or management opportunities. Less than 1% of students would work at commune or district health facilities after graduation. This study evidences challenges related to increasing the number of medical doctors working in PHC settings. Immediate and effective interventions are needed to make PHC settings more attractive and to encourage medical graduates to start and continue a career in PHC.

  4. CTCF and CohesinSA-1 Mark Active Promoters and Boundaries of Repressive Chromatin Domains in Primary Human Erythroid Cells.

    Directory of Open Access Journals (Sweden)

    Laurie A Steiner

    Full Text Available CTCF and cohesinSA-1 are regulatory proteins involved in a number of critical cellular processes including transcription, maintenance of chromatin domain architecture, and insulator function. To assess changes in the CTCF and cohesinSA-1 interactomes during erythropoiesis, chromatin immunoprecipitation coupled with high throughput sequencing and mRNA transcriptome analyses via RNA-seq were performed in primary human hematopoietic stem and progenitor cells (HSPC and primary human erythroid cells from single donors.Sites of CTCF and cohesinSA-1 co-occupancy were enriched in gene promoters in HSPC and erythroid cells compared to single CTCF or cohesin sites. Cell type-specific CTCF sites in erythroid cells were linked to highly expressed genes, with the opposite pattern observed in HSPCs. Chromatin domains were identified by ChIP-seq with antibodies against trimethylated lysine 27 histone H3, a modification associated with repressive chromatin. Repressive chromatin domains increased in both number and size during hematopoiesis, with many more repressive domains in erythroid cells than HSPCs. CTCF and cohesinSA-1 marked the boundaries of these repressive chromatin domains in a cell-type specific manner.These genome wide data, changes in sites of protein occupancy, chromatin architecture, and related gene expression, support the hypothesis that CTCF and cohesinSA-1 have multiple roles in the regulation of gene expression during erythropoiesis including transcriptional regulation at gene promoters and maintenance of chromatin architecture. These data from primary human erythroid cells provide a resource for studies of normal and perturbed erythropoiesis.

  5. Gene-chip studies of adipogenesis-regulated microRNAs in mouse primary adipocytes and human obesity

    Directory of Open Access Journals (Sweden)

    Gallagher Iain J

    2011-03-01

    Full Text Available Abstract Background Adipose tissue abundance relies partly on the factors that regulate adipogenesis, i.e. proliferation and differentiation of adipocytes. While components of the transcriptional program that initiates adipogenesis is well-known, the importance of microRNAs in adipogenesis is less well studied. We thus set out to investigate whether miRNAs would be actively modulated during adipogenesis and obesity. Methods Several models exist to study adipogenesis in vitro, of which the cell line 3T3-L1 is the most well known, albeit not the most physiologically appropriate. Thus, as an alternative, we produced EXIQON microarray of brown and white primary murine adipocytes (prior to and following differentiation to yield global profiles of miRNAs. Results We found 65 miRNAs regulated during in vitro adipogenesis in primary adipocytes. We evaluated the similarity of our responses to those found in non-primary cell models, through literature data-mining. When comparing primary adipocyte profiles, with those of cell lines reported in the literature, we found a high degree of difference in 'adipogenesis' regulated miRNAs suggesting that the model systems may not be accurately representing adipogenesis. The expression of 10 adipogenesis-regulated miRNAs were studied using real-time qPCR and then we selected 5 miRNAs, that showed robust expression, were profiled in subcutaneous adipose tissue obtained from 20 humans with a range of body mass indices (BMI, range = 21-48, and all samples have U133+2 Affymetrix profiles provided. Of the miRNAs tested, mir-21 was robustly expressed in human adipose tissue and positively correlated with BMI (R2 = 0.49, p Conclusion In conclusion, we provide a preliminary analysis of miRNAs associated with primary cell in vitro adipogenesis and demonstrate that the inflammation-associated miRNA, mir-21 is up-regulated in subcutaneous adipose tissue in human obesity. Further, we provide a novel transcriptomics database of

  6. Mutation Rate Variation is a Primary Determinant of the Distribution of Allele Frequencies in Humans.

    Directory of Open Access Journals (Sweden)

    Arbel Harpak

    2016-12-01

    Full Text Available The site frequency spectrum (SFS has long been used to study demographic history and natural selection. Here, we extend this summary by examining the SFS conditional on the alleles found at the same site in other species. We refer to this extension as the "phylogenetically-conditioned SFS" or cSFS. Using recent large-sample data from the Exome Aggregation Consortium (ExAC, combined with primate genome sequences, we find that human variants that occurred independently in closely related primate lineages are at higher frequencies in humans than variants with parallel substitutions in more distant primates. We show that this effect is largely due to sites with elevated mutation rates causing significant departures from the widely-used infinite sites mutation model. Our analysis also suggests substantial variation in mutation rates even among mutations involving the same nucleotide changes. In summary, we show that variable mutation rates are key determinants of the SFS in humans.

  7. Expression patterns of genes critical for BMP signaling pathway in developing human primary tooth germs.

    Science.gov (United States)

    Dong, Xiuqing; Shen, Bin; Ruan, Ningsheng; Guan, Zhen; Zhang, Yanding; Chen, YiPing; Hu, Xuefeng

    2014-12-01

    The developing murine tooth has been used as an excellent model system to study the molecular mechanism of organ development and regeneration. While the expression patterns of numerous regulatory genes have been examined and their roles have begun to be revealed in the developing murine tooth, little is known about gene expression and function in human tooth development. In order to unveil the molecular mechanisms that regulate human tooth morphogenesis, we examined the expression patterns of the major BMP signaling pathway molecules in the developing human tooth germ at the cap and bell stages by in situ hybridization, immunohistochemistry, and real-time RT-PCR. Expression of BMP ligands and antagonist, including BMP2, BMP3, BMP4, BMP7, and NOOGGIN, exhibited uniform patterns in the tooth germs of incisor and molar at the cap and bell stages with stronger expression in the inner dental epithelium than that in the dental mesenchyme. Both type I and type II BMP receptors were present in widespread expression pattern in the whole-enamel organ and the dental mesenchyme with the strongest expression in inner dental epithelium at the cap and bell stages. SMAD4 and SMAD1/5/8 showed an expression pattern similar to that of BMP ligands with more intensive signals in the inner dental epithelium. Despite some unique and distinct patterns as compared to the mouse, the intensive expression of BMP signaling pathway molecules in the developing human tooth strongly suggests conserved functions of BMP signaling during human odontogenesis, such as in mediating tissue interactions and regulating differentiation and organization of odontogenic tissues. Our results provide an important set of documents for studying molecular regulatory mechanisms underlying tooth development and regeneration in humans.

  8. Evaluation of E-Cigarette Liquid Vapor and Mainstream Cigarette Smoke after Direct Exposure of Primary Human Bronchial Epithelial Cells

    OpenAIRE

    Stefanie Scheffler; Hauke Dieken; Olaf Krischenowski; Christine Förster; Detlev Branscheid; Michaela Aufderheide

    2015-01-01

    E-cigarettes are emerging products, often described as “reduced-risk” nicotine products or alternatives to combustible cigarettes. Many smokers switch to e-cigarettes to quit or significantly reduce smoking. However, no regulations for e-cigarettes are currently into force, so that the quality and safety of e-liquids is not necessarily guaranteed. We exposed primary human bronchial epithelial cells of two different donors to vapor of e-cigarette liquid with or without nicotine, vapor of the c...

  9. Bringing transcranial mapping into shape: Sulcus-aligned mapping captures motor somatotopy in human primary motor hand area

    DEFF Research Database (Denmark)

    Raffin, Estelle; Pellegrino, Giovanni; Di Lazzaro, Vincenzo

    2015-01-01

    Motor representations express some degree of somatotopy in human primary motor hand area (M1HAND), but within-M1HAND corticomotor somatotopy has been difficult to study with transcranial magnetic stimulation (TMS). Here we introduce a “linear” TMS mapping approach based on the individual shape...... motor threshold. We pseudorandomly stimulated six targets located on a straight mediolateral line corresponding to the overall orientation of the central sulcus with a fixed coil orientation of 45° to the mid-sagittal line (STRAIGHT-450 FIX) or seven targets in the posterior part of the crown...

  10. Protective Effects of Diallyl Sulfide Against Ethanol-Induced Injury in Rat Adipose Tissue and Primary Human Adipocytes.

    Science.gov (United States)

    Kema, Venkata Harini; Khan, Imran; Jamal, Reshma; Vishwakarma, Sandeep Kumar; Lakki Reddy, Chandrakala; Parwani, Kirti; Patel, Farhin; Patel, Dhara; Khan, Aleem A; Mandal, Palash

    2017-06-01

    Alcohol consumption is the fourth leading cause of death and disability worldwide. Several cellular pathways contribute to alcohol-mediated tissue injury. Adipose tissue apart from functioning as an endocrine organ secretes several hormones and cytokines known as adipokines that are known to play a significant role in alcohol-induced tissue damage. This study was designed to test the efficacy of diallyl sulfide (DAS) in regulating the alcohol-induced outcomes on adipose tissue. Male Wistar rats were fed with 36% Lieber-DeCarli liquid diet containing ethanol (EtOH) for 4 weeks. Control rats were pair-fed with isocaloric diet containing maltodextrin instead of EtOH. During the last week of feeding protocol, the EtOH-fed rat group was given 200 mg/kg body weight of DAS through diet. We also studied DAS effect on isolated human primary adipocytes. Viability of human primary adipocytes on DAS treatment was assessed by MTT assay. Malondialdehyde (MDA), a marker of oxidative stress, was measured by HPLC and the thiobarbituric acid method. Expression of inflammatory genes and lipogenic genes was studied by qRT-PCR and Western blotting. Serum inflammatory gene expression was studied by ELISA. Our study results showed that DAS could alleviate EtOH-induced expression levels of proinflammatory and endoplasmic reticulum (ER) stress genes and improve adipose tissue mass and adipocyte morphology in male Wistar rats fed Lieber-DeCarli diet containing 6% EtOH. Further, we showed that DAS reduced the expression of lipogenic genes and improved lipid accumulation and adipocyte mass in human primary adipocytes treated with EtOH. Subsequently, we also showed that oxidative stress, as measured by the changes in MDA levels, was reduced in both male Wistar rats and human primary adipocytes treated with EtOH plus DAS. Our study results prove that DAS is effective in ameliorating EtOH-induced damage to adipose tissue as evidenced by the reduction brought about by DAS in oxidative stress

  11. Efficient gene disruption in cultured primary human endothelial cells by CRISPR/Cas9.

    Science.gov (United States)

    Abrahimi, Parwiz; Chang, William G; Kluger, Martin S; Qyang, Yibing; Tellides, George; Saltzman, W Mark; Pober, Jordan S

    2015-07-03

    The participation of endothelial cells (EC) in many physiological and pathological processes is widely modeled using human EC cultures, but genetic manipulation of these untransformed cells has been technically challenging. Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 nuclease (Cas9) technology offers a promising new approach. However, mutagenized cultured cells require cloning to yield homogeneous populations, and the limited replicative lifespan of well-differentiated human EC presents a barrier for doing so. To create a simple but highly efficient method using CRISPR/Cas9 to generate biallelic gene disruption in untransformed human EC. To demonstrate proof-of-principle, we used CRISPR/Cas9 to disrupt the gene for the class II transactivator. We used endothelial colony forming cell-derived EC and lentiviral vectors to deliver CRISPR/Cas9 elements to ablate EC expression of class II major histocompatibility complex molecules and with it, the capacity to activate allogeneic CD4(+) T cells. We show the observed loss-of-function arises from biallelic gene disruption in class II transactivator that leaves other essential properties of the cells intact, including self-assembly into blood vessels in vivo, and that the altered phenotype can be rescued by reintroduction of class II transactivator expression. CRISPR/Cas9-modified human EC provides a powerful platform for vascular research and for regenerative medicine/tissue engineering. © 2015 American Heart Association, Inc.

  12. Humans rather than climate the primary cause of Pleistocene megafaunal extinction in Australia

    NARCIS (Netherlands)

    van der Kaars, W.A.; Miller, Gifford; Turney, Chris; Cook, Ellyn; Nürnberg, Dirk; Schönfeld, Joachim; Kershaw, Peter; Lehman, Scott

    2017-01-01

    Environmental histories that span the last full glacial cycle and are representative of regional change in Australia are scarce, hampering assessment of environmental change preceding and concurrent with human dispersal on the continent ca. 47,000 years ago. Here we present a continuous 150,000-year

  13. Restriction of human papillomavirus DNA testing in primary cervical screening to women above age 30

    DEFF Research Database (Denmark)

    Rebolj, Matejka; Njor, Sisse H; Lynge, Elsebeth

    2012-01-01

    Cervical screening with human papillomavirus (HPV) testing is less specific for high-grade cervical intraepithelial neoplasia (=CIN3) than cytology. The aim of this systematic review was to determine whether a restriction of HPV testing to women aged at least 30 years would eliminate the problem...

  14. CO(2) laser vaporization as primary therapy for human papillomavirus lesions. A prospective observational study.

    Science.gov (United States)

    Savoca, S; Nardo, L G; Rosano, T F; D'Agosta, S; Nardo, F

    2001-12-01

    Human papillomavirus manifestations occur with increased frequency and severity amongst sexually active people. Several therapeutic approaches have been suggested to treat this viral disease. The aim of this prospective observational study was to assess the effectiveness of CO(2) laser vaporization for human papillomavirus warts. Eighty healthy sexually active women with cytologically, colposcopically and histologically diagnosed human papillomavirus urogenital and perianal warts were enrolled and then treated by CO(2) laser (16-18 W). Male partners were also investigated, and interferon-beta was eventually administered. All patients were then followed up for twelve months consecutively. At twelve-month follow-up, warts clearance was observed in 70 (87.5%) women. Recurrence was reported in ten (12.5%) women with multiple partners and affected by flat or endophytic condiloma of the cervix. Moreover, there were no complaints of pain, scar tissue deformity or other side effects. CO(2) laser vaporization is an effective, as well as safe and simple therapeutic approach for treatment of human papillomavirus warts. Its use should be encouraged for condyloma acuminata not associated with malignancy, as well as during pregnancy.

  15. Apoptosis in the human periodontal membrane evaluated in primary and permanent teeth

    DEFF Research Database (Denmark)

    Bille, Marie-Louise Bastholm; Thomsen, Bjarke; Kjær, Inger

    2011-01-01

    Abstract Objective. Recent studies revealed a highly innervated layer in close proximity to the root surface in the periodontal membrane of human teeth. Persistence of the epithelial cells of Malassez along root surfaces without resorption has also been demonstrated. It is hypothesized that resor...

  16. Pathogenesis of primary R5 human immunodeficiency virus type 1 clones in SCID-hu mice

    NARCIS (Netherlands)

    Scoggins, R. M.; Taylor, J. R.; Patrie, J.; van't Wout, A. B.; Schuitemaker, H.; Camerini, D.

    2000-01-01

    We studied the replication and cytopathicity in SCID-hu mice of R5 human immunodeficiency virus type 1 (HIV-1) biological clones from early and late stages of infection of three patients who never developed MT-2 cell syncytium-inducing (SI; R5X4 or X4) viruses. Several of the late-stage non-MT-2

  17. A feeder- and xeno-free human induced pluripotent stem cell line obtained from primary human dermal fibroblasts with epigenetic repression of reprogramming factors expression: GPCCi001-A

    Directory of Open Access Journals (Sweden)

    Michał Stefan Lach

    2017-04-01

    Full Text Available The primary human dermal fibroblasts (PHDFs from breast cancer patient were obtained to generate the human induced pluripotent stem cell line GPCCi001-A via lentiviral transfection. Thus, a modified EF1a-hSTEMCCA-loxP with tetO operator which regulates transgene expression was used. This method takes advantage of epigenetic regulation of transcription and allows for stable silencing of the reprogramming factors in obtained hiPS cells. To increase the potential utility of hiPSCs for clinical applications, they were adapted to feeder- and xeno-free conditions. The pluripotency of GPCCi001-A cell line and ability to differentiate into three germ layers was confirmed.

  18. IL-17 Enhances Chemotaxis of Primary Human B Cells during Asthma.

    Directory of Open Access Journals (Sweden)

    Rabih Halwani

    Full Text Available IL-17 is a pro-inflammatory mediator that is believed to play a critical role in regulating tissue inflammation during asthma, COPD, as well as other inflammatory disorders. The level of expression of IL-17 has been shown to be upregulated in lung bronchial tissue of asthmatic patients. Several reports have provided further evidence that this cytokine could play a key role in enhancing the migration of inflammatory as well as structural cells of the bronchial lung tissue during asthma and COPD. B cell infiltration to sites of inflammation during inflammatory disorders such as bowel disease, asthma and COPD has been reported. Accordingly, in this study we hypothesized that IL-17 may exert a chemotactic effect on primary B cells during asthma. We observed that B cells from asthmatic patients expressed significantly higher levels of IL-17RA and IL-17RC, compared to those of healthy subjects. Using an in-vitro migration assay, B cells were shown to migrate towards both IL-17A and IL-17F. Interestingly, blocking IL-17A and IL-17F signaling using either anti-IL-17R antibodies or MAP kinase inhibitors prevented in vitro migration of B cell towards IL-17. These observations indicate a direct chemotactic effect of IL-17 cytokines on primary peripheral blood B cells with higher effect being on asthmatic B cells. These findings revealed a key role for IL-17 in enhancing the migration of B cells to the lung tissue during asthma or COPD.

  19. Normal mammary fibroblasts induce reversion of the malignant phenotype in human primary breast cancer.

    Science.gov (United States)

    Römer, Anna-Maria; Lühr, Inke; Klein, Andreas; Friedl, Andreas; Sebens, Susanne; Rösel, Frank; Arnold, Norbert; Strauss, Alexander; Jonat, Walter; Bauer, Maret

    2013-04-01

    The tumor microenvironment plays a major role in tumor growth and progression. Its manipulation can lead to a reversion of the malignant phenotype. Here we explored the ability of normal mammary fibroblasts (HMFs) to induce reversion of the malignant phenotype of primary breast carcinoma cells (PBCs) in a three-dimensional (3D) context. PBCs were isolated from 13 primary breast carcinomas and cultured in 3D collagen-I gels as mono- or co-culture with HMFs. In five co-cultures, PBCs exhibited reversion of their malignant phenotype, whereas PBCs in matched monocultures exhibited disorganized growth. Reversion, defined as the restoration of the complete baso-apical polarity axis, was confirmed with established polarity markers. Secretion of the tissue-specific glycoprotein MAM-6 into the acinar lumens and deposition of basement membrane indicated functional differentiation. Gene expression analysis revealed a set of differentially regulated genes which possibly affect the reversion process. These included MAL, ELF5, MAP6, ZMYND11 and SQLE. These findings highlight the significant role of fibroblasts in regulating the carcinoma phenotype.

  20. Gene disruption of Plasmodium falciparum p52 results in attenuation of malaria liver stage development in cultured primary human hepatocytes.

    Directory of Open Access Journals (Sweden)

    Ben C L van Schaijk

    Full Text Available Difficulties with inducing sterile and long lasting protective immunity against malaria with subunit vaccines has renewed interest in vaccinations with attenuated Plasmodium parasites. Immunizations with sporozoites that are attenuated by radiation (RAS can induce strong protective immunity both in humans and rodent models of malaria. Recently, in rodent parasites it has been shown that through the deletion of a single gene, sporozoites can also become attenuated in liver stage development and, importantly, immunization with these sporozoites results in immune responses identical to RAS. The promise of vaccination using these genetically attenuated sporozoites (GAS depends on translating the results in rodent malaria models to human malaria. In this study, we perform the first essential step in this transition by disrupting, p52, in P. falciparum an ortholog of the rodent parasite gene, p36p, which we had previously shown can confer long lasting protective immunity in mice. These P. falciparum P52 deficient sporozoites demonstrate gliding motility, cell traversal and an invasion rate into primary human hepatocytes in vitro that is comparable to wild type sporozoites. However, inside the host hepatocyte development is arrested very soon after invasion. This study reveals, for the first time, that disrupting the equivalent gene in both P. falciparum and rodent malaria Plasmodium species generates parasites that become similarly arrested during liver stage development and these results pave the way for further development of GAS for human use.

  1. [EVALUATION OF THE HUMAN SENSITIVITY TO SMALLPOX VIRUS BY THE PRIMARY CULTURES OF THE MONOCYTE-MACROPHAGES].

    Science.gov (United States)

    Zamedyanskaya, A S; Titova, K A; Sergeev, Al A; Kabanov, A S; Bulychev, L E; Sergeev, Ar A; Galakhova, D O; Nesterov, A E; Nosareva, O V; Shishkina, L N; Taranov, O S; Omigov, V V; Agafonov, A P; Sergeev, A N

    2016-01-01

    Studies of the primary cultures of granulocytes, mononuclear, and monocyte-macrophage cells derived from human blood were performed using variola virus (VARV) in the doses of 0.001-0.021 PFU/cell (plaques-forming units per cell). Positive dynamics of the virus accumulation was observed only in the monocyte-macrophages with maximum values of virus concentration (5.0-5.5 Ig PFU/ml) mainly within six days after the infection. The fact of VARV replication in the monocyte-macrophages was confirmed by the data of electron microscopy. At the same time, virus vaccines when tested in doses 3.3 and 4.2 Ig PFU/ml did not show the ability to reproduce in these human cells. The people sensitivity to VARV as assessed from the data obtained on human monocyte-macrophages corresponded to -1 PFU (taking into account the smooth interaction of the virus in the body to the cells of this type), which is consistent to previously found theoretical data on the virus sensitivity. The human susceptibility to VARV assessed experimentally can be used to predict the adequacy of developed smallpox models (in vivo) based on susceptible animals. This is necessary for reliable assessment of the efficiency of development of drugs for treatment and prophylaxis of the smallpox.

  2. Outgrowth of human liver metastases after resection of the primary colorectal tumor: a shift in the balance between apoptosis and proliferation.

    NARCIS (Netherlands)

    Peeters, C.F.J.M.; Waal, R.M.W. de; Wobbes, Th.; Westphal, J.R.; Ruers, T.J.M.

    2006-01-01

    Rapid outgrowth of metastases after removal of the primary tumor has been described in several mouse models. Loss of primary tumor-induced inhibition of angiogenesis in the metastases has been suggested as the underlying cause. Accordingly, we recently demonstrated that vascular density in human

  3. Automated bioreactor system for cartilage tissue engineering of human primary nasal septal chondrocytes.

    Science.gov (United States)

    Princz, Sascha; Wenzel, Ulla; Tritschler, Hanna; Schwarz, Silke; Dettmann, Christian; Rotter, Nicole; Hessling, Martin

    2017-10-26

    An automated bioreactor system for three-dimensional (3D) cultivation of facial cartilage replacement matrices (e.g. whole human auricles) with automatised medium exchange, gas flow and temperature control was developed. The measurement of O2 saturation and pH value in the medium was performed with a non-invasive optical method. The whole system can be observed via remote monitoring worldwide. First results demonstrated that the complete system remained sterile throughout a period of 42 days. Human chondrocytes migrated into the employed cartilage replacement matrix consisting of decellularised porcine nasoseptal cartilage (pNSC). Furthermore, an improved migration and new synthesis of aggrecan was detected. A first evaluation of the system was conducted by comparison of the results from laboratory analysis with computational fluid dynamics (CFD).

  4. Analysis of adequacy levels for human resources improvement within primary health care framework in Africa

    Directory of Open Access Journals (Sweden)

    Piette Danielle

    2005-12-01

    Full Text Available Abstract Human resources in health care system in sub-Saharan Africa are generally picturing a lack of adequacy between expected skills from the professionals and health care needs expressed by the populations. It is, however, possible to analyse these various lacks of adequacy related to human resource management and their determinants to enhance the effectiveness of the health care system. From two projects focused on nurse professionals within the health care system in Central Africa, we present an analytic grid for adequacy levels looking into the following aspects: - adequacy between skills-based profiles for health system professionals, quality of care and service delivery (health care system /medical standards, needs and expectations from the populations, - adequacy between allocation of health system professionals, quality of care and services delivered (health care system /medical standards, needs and expectations from the populations, - adequacy between human resource management within health care system and medical standards, - adequacy between human resource management within education/teaching/training and needs from health care system and education sectors, - adequacy between basic and on-going education and realities of tasks expected and implemented by different categories of professionals within the health care system body, - adequacy between intentions for initial and on-going trainings and teaching programs in health sciences for trainers (teachers/supervisors/health care system professionals/ directors (teaching managers of schools.... This tool is necessary for decision-makers as well as for health care system professionals who share common objectives for changes at each level of intervention within the health system. Setting this adequacy implies interdisciplinary and participative approaches for concerned actors in order to provide an overall vision of a more broaden system than health district, small island with self

  5. Humans rather than climate the primary cause of Pleistocene megafaunal extinction in Australia

    OpenAIRE

    van der Kaars, S.; Miller, G.; Turney, C.; Cook, E. J.; Nürnberg, Dirk; Schönfeld, Joachim; Kershaw, A. P.; Lehman, S.

    2017-01-01

    Environmental histories that span the last full glacial cycle and are representative of regional change in Australia are scarce, hampering assessment of environmental change preceding and concurrent with human dispersal on the continent ca. 47,000 years ago. Here we present a continuous 150,000-year record offshore south-western Australia and identify the timing of two critical late Pleistocene events: wide-scale ecosystem change and regional megafaunal population collapse. We establish that ...

  6. Lysophosphatidic acid (LPA 18:1 transcriptional regulation of primary human gingival fibroblasts

    Directory of Open Access Journals (Sweden)

    D. Roselyn Cerutis

    2014-12-01

    Full Text Available The pleiotropic, bioactive lipid lysophosphatidic acid [(LPA, 1-acyl-sn-glycerol-3-phosphate] exerts critical regulatory actions in physiology and pathophysiology in many systems. It is present in normal bodily fluids, and is elevated in pathology (1. In vivo, “LPA” exists as distinct molecular species, each having a single fatty acid of varying chain length and degree of unsaturation covalently attached to the glycerol backbone via an acyl, alkyl, or alkenyl link. These species differ in affinities for the individual LPA receptors [(LPARs, LPA1-6] and coupling to G proteins (2. However, LPA 18:1 has been and continues to be the most commonly utilized species in reported studies. The actions of “LPA” remain poorly defined in oral biology and pathophysiology. Our laboratory has addressed this knowledge gap by studying in vitro the actions of the major human salivary LPA species [18:1, 18:0, and 16:0 (3] in human oral cells (4–7. This includes gingival fibroblasts (GF, which our flow cytometry data from multiple donors found that they express LPA1-5 (6. We have also reported that these species are ten-fold elevated to pharmacologic levels in the saliva and gingival crevicular fluid obtained from patients with moderate–severe periodontitis (8. As the potential of LPA to regulate transcriptional activity had not been examined in the oral system, this study used whole human genome microarray analysis to test the hypothesis that LPA 18:1-treated human GF would show significant changes in gene transcripts relevant to their biology, wound-healing, and inflammatory responses. LPA 18:1 was found to significantly regulate a large, complex set of genes critical to GF biology in these categories and to periodontal disease. The raw data has been deposited at NCBI's GEO database as record GSE57496.

  7. Lysophosphatidic acid (LPA) 18:1 transcriptional regulation of primary human gingival fibroblasts.

    Science.gov (United States)

    Cerutis, D Roselyn; Weston, Michael D; Ogunleye, Afolabi O; McVaney, Timothy P; Miyamoto, Takanari

    2014-12-01

    The pleiotropic, bioactive lipid lysophosphatidic acid [(LPA), 1-acyl-sn-glycerol-3-phosphate] exerts critical regulatory actions in physiology and pathophysiology in many systems. It is present in normal bodily fluids, and is elevated in pathology (1). In vivo, "LPA" exists as distinct molecular species, each having a single fatty acid of varying chain length and degree of unsaturation covalently attached to the glycerol backbone via an acyl, alkyl, or alkenyl link. These species differ in affinities for the individual LPA receptors [(LPARs), LPA1-6] and coupling to G proteins (2). However, LPA 18:1 has been and continues to be the most commonly utilized species in reported studies. The actions of "LPA" remain poorly defined in oral biology and pathophysiology. Our laboratory has addressed this knowledge gap by studying in vitro the actions of the major human salivary LPA species [18:1, 18:0, and 16:0 (3)] in human oral cells (4-7). This includes gingival fibroblasts (GF), which our flow cytometry data from multiple donors found that they express LPA1-5 (6). We have also reported that these species are ten-fold elevated to pharmacologic levels in the saliva and gingival crevicular fluid obtained from patients with moderate-severe periodontitis (8). As the potential of LPA to regulate transcriptional activity had not been examined in the oral system, this study used whole human genome microarray analysis to test the hypothesis that LPA 18:1-treated human GF would show significant changes in gene transcripts relevant to their biology, wound-healing, and inflammatory responses. LPA 18:1 was found to significantly regulate a large, complex set of genes critical to GF biology in these categories and to periodontal disease. The raw data has been deposited at NCBI's GEO database as record GSE57496.

  8. Advancing the Capabilities of an Authentic Ex Vivo Model of Primary Human Prostate Cancer

    Science.gov (United States)

    2014-10-01

    The lactate dehydrogenase (LDH) assay is one of the most widely used assays for quantifying cytotoxicity in organotypic cultures. We performed LDH...R. Keshari et al., Metabolic reprogramming and validation of hyperpolarized 13C lactate as a prostate cancer biomarker using a human prostate tissue...inherent drawbacks hinder their clinical relevance.1,2 Cell lines, often artificially immortalized, acquire mutations in culture and lack the

  9. Determinants of Human Immunodeficiency Virus Type 1 Escape from the Primary CD8+ Cytotoxic T Lymphocyte Response

    Science.gov (United States)

    Jones, Nicola A.; Wei, Xiping; Flower, Darren R.; Wong, MaiLee; Michor, Franziska; Saag, Michael S.; Hahn, Beatrice H.; Nowak, Martin A.; Shaw, George M.; Borrow, Persephone

    2004-01-01

    CD8+ cytotoxic T lymphocytes (CTLs) play an important role in containment of virus replication in primary human immunodeficiency virus (HIV) infection. HIV's ability to mutate to escape from CTL pressure is increasingly recognized; but comprehensive studies of escape from the CD8 T cell response in primary HIV infection are currently lacking. Here, we have fully characterized the primary CTL response to autologous virus Env, Gag, and Tat proteins in three patients, and investigated the extent, kinetics, and mechanisms of viral escape from epitope-specific components of the response. In all three individuals, we observed variation beginning within weeks of infection at epitope-containing sites in the viral quasispecies, which conferred escape by mechanisms including altered peptide presentation/recognition and altered antigen processing. The number of epitope-containing regions exhibiting evidence of early CTL escape ranged from 1 out of 21 in a subject who controlled viral replication effectively to 5 out of 7 in a subject who did not. Evaluation of the extent and kinetics of HIV-1 escape from >40 different epitope-specific CD8 T cell responses enabled analysis of factors determining escape and suggested that escape is restricted by costs to intrinsic viral fitness and by broad, codominant distribution of CTL-mediated pressure on viral replication. PMID:15545352

  10. The Expanding Universe: Dark Energy

    Energy Technology Data Exchange (ETDEWEB)

    Lincoln, Don [Fermilab; Nord, Brian [Fermilab

    2014-09-01

    In 1998, observations of distant supernovae led physicists that not only was the universe expanding, but the expansion was speeding up. In this article, we describe the evidence for an expanding universe and describe what physicists and cosmologists have learned in the intervening years. The target audience for this article is high school physics teachers and college physics professors at teaching institutions.

  11. Next-generation sequencing reveals low-dose effects of cationic dendrimers in primary human bronchial epithelial cells.

    Science.gov (United States)

    Feliu, Neus; Kohonen, Pekka; Ji, Jie; Zhang, Yuning; Karlsson, Hanna L; Palmberg, Lena; Nyström, Andreas; Fadeel, Bengt

    2015-01-27

    Gene expression profiling has developed rapidly in recent years with the advent of deep sequencing technologies such as RNA sequencing (RNA Seq) and could be harnessed to predict and define mechanisms of toxicity of chemicals and nanomaterials. However, the full potential of these technologies in (nano)toxicology is yet to be realized. Here, we show that systems biology approaches can uncover mechanisms underlying cellular responses to nanomaterials. Using RNA Seq and computational approaches, we found that cationic poly(amidoamine) dendrimers (PAMAM-NH2) are capable of triggering down-regulation of cell-cycle-related genes in primary human bronchial epithelial cells at doses that do not elicit acute cytotoxicity, as demonstrated using conventional cell viability assays, while gene transcription was not affected by neutral PAMAM-OH dendrimers. The PAMAMs were internalized in an active manner by lung cells and localized mainly in lysosomes; amine-terminated dendrimers were internalized more efficiently when compared to the hydroxyl-terminated dendrimers. Upstream regulator analysis implicated NF-κB as a putative transcriptional regulator, and subsequent cell-based assays confirmed that PAMAM-NH2 caused NF-κB-dependent cell cycle arrest. However, PAMAM-NH2 did not affect cell cycle progression in the human A549 adenocarcinoma cell line. These results demonstrate the feasibility of applying systems biology approaches to predict cellular responses to nanomaterials and highlight the importance of using relevant (primary) cell models.

  12. Primary Human Blood Dendritic Cells for Cancer Immunotherapy—Tailoring the Immune Response by Dendritic Cell Maturation

    Directory of Open Access Journals (Sweden)

    Simone P. Sittig

    2015-12-01

    Full Text Available Dendritic cell (DC-based cancer vaccines hold the great promise of tipping the balance from tolerance of the tumor to rejection. In the last two decades, we have gained tremendous knowledge about DC-based cancer vaccines. The maturation of DCs has proven indispensable to induce immunogenic T cell responses. We review the insights gained from the development of maturation cocktails in monocyte derived DC-based trials. More recently, we have also gained insights into the functional specialization of primary human blood DC subsets. In peripheral human blood, we can distinguish at least three primary DC subsets, namely CD1c+ and CD141+ myeloid DCs and plasmacytoid DCs. We reflect the current knowledge on maturation and T helper polarization by these blood DC subsets in the context of DC-based cancer vaccines. The maturation stimulus in combination with the DC subset will determine the type of T cell response that is induced. First trials with these natural DCs underline their excellent in vivo functioning and mark them as promising tools for future vaccination strategies.

  13. PRIMARY CUTANEOUS HISTOPLASMOSIS PADA PASIEN DENGAN INFEKSI HUMAN IMMUNODEFICIENCY VIRUS (HIV

    Directory of Open Access Journals (Sweden)

    Nyoman Upadana

    2014-09-01

    Full Text Available Histoplasmosis adalah penyakit granulomatosa disebabkan oleh jamur dimorphic Histoplasma capsulatum. Lesi kulit merupakan bentuk sekunder, tetapi dapat juga primer pada kasus self inoculation. Kasus adalah laki-laki 27 tahun dengan benjolan sejak 3 bulan yang lalu, diawali pada kedua kaki meluas ke tangan dan wajah. Pemeriksaan kulit didapatkan papul, nodul, dan ulkus dengan tepi landai, tertutup krusta tebal. Hasil pemeriksaan CD4 adalah 4/mm3. Pemeriksaan histopatologi pada stroma tampak sebaran sel histiosit mengandung mikroorganisme, bentuk bulat, berdinding tebal, berwarna eosinofilik, dan dikelilingi oleh clear halo. Positif dengan pewarnaan periodic acid schiff dan grocott methenamic silver, dan sesuai untuk Histoplasma capsulatum. Diagnosis pasien adalah primary cutaneous histoplasmosis dan diterapi flukonazol 200 mg IV dengan respon terapi yang baik. [MEDICINA 2013;44:113-117].

  14. Mechanotransduction in primary human osteoarthritic chondrocytes is mediated by metabolism of energy, lipids, and amino acids.

    Science.gov (United States)

    Zignego, Donald L; Hilmer, Jonathan K; June, Ronald K

    2015-12-16

    Chondrocytes are the sole cell type found in articular cartilage and are repeatedly subjected to mechanical loading in vivo. We hypothesized that physiological dynamic compression results in changes in energy metabolism to produce proteins for maintenance of the pericellular and extracellular matrices. The objective of this study was to develop an in-depth understanding for the short term (mechanotransduction with aging dependence. Targeted studies found a transient increase in the ratio of NADP+ to NADPH and an initial decrease in the ratio of GDP to GTP, suggesting a flux of energy into the TCA cycle. By characterizing metabolomics profiles of primary chondrocytes in response to applied dynamic compression, this study provides insight into how OA chondrocytes respond to mechanical load. These results are consistent with increases in glycolytic energy utilization by mechanically induced signaling, and add substantial new data to a complex picture of how chondrocytes transduce mechanical loads. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. [Biochemical, structural and biomechanical features of human scleral matrix in primary open-angle glaucoma].

    Science.gov (United States)

    Iomdina, E N; Ignat'eva, N Iu; Danilov, N A; Arutiunian, L L; Kiseleva, O A; Nazarenko, L A

    2011-01-01

    The goal of the investigation was to study the changes of scleral matrix collagen in progressing primary open-angle glaucoma (POAG). In scleral samples of 16 patients aged 52-81 years with different stages of POAG amino acid composition, collagen content and cross-linking ratio characterizing tissue rigidity were estimated. Collagen I was found to be the dominant type in sclera of glaucomatous and normal eyes, though as POAG progressing the content and cross-linking increase, that underlies the scleral rigidity increase and permeability decrease. Similar ratio of collagen cross-linking with age and glaucoma progression give evidence of specific pathologic changes of the dominant matrix component, that may be one of significant pathogenic factor of POAG.

  16. Establishment and characterization of a new cell line from primary human breast carcinoma.

    Science.gov (United States)

    Amadori, D; Bertoni, L; Flamigni, A; Savini, S; De Giovanni, C; Casanova, S; De Paola, F; Amadori, A; Giulotto, E; Zoli, W

    1993-12-01

    A new cell line (BRC-230) was established from surgical material of primary ductal infiltrating breast carcinoma. The epithelial nature of this cell line was confirmed by ultrastructural analysis and demonstrated the retention of structural properties characteristic of the original tumor. The BRC-230 cell line induced tumor in athymic Cr1:nu/nu(CD-1)BR nude mice, it possessed an abnormal karyotype with a modal chromosome number between 60-61 with eight recurrent marker chromosomes, and it presented a doubling time of 30.5 hr. Scatchard analysis demonstrated that both primary tumor and BRC-230 cells were estrogen and progesterone receptor negative. Immunoenzymatic and radioimmunoassays showed a production of marker antigens (CEA, TPA, CA125, CA15-3, CA19-9) which was similar in the patient's serum and BRC-230 cells. The in vitro drug sensitivity assay of the cell line and of the parental tumor tissue showed overlapping results to all tested antiblastic drugs. BRC-230 cells were resistant to 4-Idroperoxy-cyclophosphamide, Idarubicinol, Mitoxantrone, Etoposide, 4'Epidoxorubicin, and Doxorubicin, showing a multiple drug resistance phenotype. Amplification or rearrangement of Her-2neu, Ha-ras, and C-myc genes was observed neither in the original tumor nor in BRC-230 cells; the mdr-1 gene was also present in a single copy. We conclude from these studies that the BRC-230 cell line maintains the same characteristics as the original tumor and may provide us with a good model to study in vitro the biology of drug resistance of breast cancer.

  17. The influence of contralateral primary afferents on Ia inhibitory interneurones in humans.

    Science.gov (United States)

    Delwaide, P J; Pepin, J L

    1991-01-01

    1. Contralateral influences on short latency reciprocal inhibition between wrist extensor and flexor muscles were investigated in twenty-two healthy volunteers. Reciprocal inhibition, probably mediated through the Ia inhibitory interneurone, was measured by conditioning the flexor carpi radialis (FCR) H reflex by weak stimulation of the ipsilateral radial nerve. Maximum reciprocal inhibition occurring at a precise delay between conditioning and conditioned stimulations was taken as the test level of inhibition. 2. Contralateral median or radial nerves were stimulated at short intervals before the onset of reciprocal inhibition. The latter was increased by 8.6% after median nerve stimulation and decreased by 16.5% after radial nerve stimulation. 3. The contribution of sensory fibres in the two nerves to contralateral effects was investigated by stimulating purely sensory branches of the nerves. No clear modification of the contralateral reciprocal inhibition was observed. The effects produced by mixed nerve stimulation are thus likely to have been mediated by Ia fibres. 4. In three hemiplegic patients where reciprocal inhibition was reduced unilaterally, stimulation on the spastic side produced contralateral effects similar to those observed in normal subjects. This result indicates that contralateral effects are not mediated through the Ia inhibitory interneurone ipsilateral to the conditioning stimulus. 5. Since contralateral effects occur after short delays (2 ms, median nerve; 3 ms, radial nerve), we suggest a functional scheme in which the excitability of Ia inhibitory interneurones is modified by contralateral primary afferents via the interneurones activated by group I fibres, probably Ia fibres. The short delays indicate that the interneurone transmitting primary afferent influences to the contralateral side is probably excitatory. PMID:1895236

  18. Granulocytes affect double-strand break repair assays in primary human lymphocytes.

    Science.gov (United States)

    Lacoste, Sandrine; Bhatia, Ravi; Bhatia, Smita; O'Connor, Timothy R

    2014-01-01

    Patients who develop therapy-related myelodysplasia/acute myeloid leukemia after autologous-hematopoietic stem cell (aHCT) transplant show lower expression levels of DNA repair genes in their pre-aHCT CD34+ cells. To investigate whether this leads to functional differences in DNA repair abilities measurable in patients, we adapted two plasmid-based host-cell reactivation assays for use in primary lymphocytes. Prior to applying these assays to patients who underwent aHCT, we wanted first to verify whether sample preparation affected repair measurements, as patient samples were simply depleted of erythrocytes (with hetastarch) prior to freezing, which is not the classical way to prepare lymphocytes prior to DNA repair experiments (with a density gradient). We show here that lymphocytes from healthy donors freshly prepared with hetastarch show systematically a higher level of double-strand break repair as compared to when prepared with a density gradient, but that most of this difference disappears after samples were frozen. Several observations points to granulocytes as the source for this effect of sample preparation on repair: 1) removal of granulocytes makes the effect disappear, 2) DSB repair measurements for the same individual correlate to the percentage of granulocytes in the sample and 3) nucleofection in presence of granulocytes increases the level of reactive oxygen species (ROS) in neighboring lymphocytes in a dose-dependent manner (R2 of 0.95). These results indicate that co-purified granulocytes, possibly through the release of ROS at time of transfection, can lead to an enhanced repair in lymphocytes that obfuscates any evaluation of inter individual differences in repair as measured by host-cell reactivation. As a result, hetastarch-prepared samples are likely unsuitable for the assessment of DSB repair in primary cells with that type of assay. Granulocyte contamination that exists after a density gradient preparation, although much more limited, could

  19. Replication of different clones of human immunodeficiency virus type 1 in primary fetal human astrocytes: enhancement of viral gene expression by Nef.

    Science.gov (United States)

    Bencheikh, M; Bentsman, G; Sarkissian, N; Canki, M; Volsky, D J

    1999-04-01

    Dementia is a common complication of AIDS which is associated with human immunodeficiency virus type 1 (HIV-1) infection of brain macrophages and microglia. Recent studies have shown that astrocytes are also infected in the brain but HIV-1 replication in these cells is restricted. To determine virus specificity of this restriction we tested the expression of 15 HIV-1 molecular clones in primary human fetal astrocytes by infection and DNA transfection. Infection with cell-free viruses was poorly productive and revealed no clone-specific differences. In contrast, transfected cells produced transiently high levels of HIV-1 p24 core antigen, up to 50 nanograms per ml culture supernatant, and nanogram levels of p24 were detected 3-4 weeks after transfection of some viral clones. The average peak expression of HIV-1 in astrocytes varied as a function of viral clone used by a factor of 15 but the differences and the subsequent virus spread did not correlate with the tropism of the viral clones to T cells or macrophages. Functional vif, vpu, and vpr genes were dispensable for virus replication from transfected DNA, but intact nef provided a detectable enhancement of early viral gene expression and promoted maintenance of HIV-1 infection. We conclude that primary astrocytes present no fundamental barriers to moderate expression of different strains of HIV-1 and that the presence of functional Nef is advantageous to virus infection in these cells.

  20. TGF-β signaling is an effective target to impair survival and induce apoptosis of human cholangiocarcinoma cells: A study on human primary cell cultures.

    Directory of Open Access Journals (Sweden)

    Anna Maria Lustri

    Full Text Available Cholangiocarcinoma (CCA and its subtypes (mucin- and mixed-CCA arise from the neoplastic transformation of cholangiocytes, the epithelial cells lining the biliary tree. CCA has a high mortality rate owing to its aggressiveness, late diagnosis and high resistance to radiotherapy and chemotherapeutics. We have demonstrated that CCA is enriched for cancer stem cells which express epithelial to mesenchymal transition (EMT traits, with these features being associated with aggressiveness and drug resistance. TGF-β signaling is upregulated in CCA and involved in EMT. We have recently established primary cell cultures from human mucin- and mixed-intrahepatic CCA. In human CCA primary cultures with different levels of EMT trait expression, we evaluated the anticancer effects of: (i CX-4945, a casein kinase-2 (CK2 inhibitor that blocks TGF-β1-induced EMT; and (ii LY2157299, a TGF-β receptor I kinase inhibitor. We tested primary cell lines expressing EMT trait markers (vimentin, N-cadherin and nuclear catenin but negative for epithelial markers, and cell lines expressing epithelial markers (CK19-positive in association with EMT traits. Cell viability was evaluated by MTS assays, apoptosis by Annexin V FITC and cell migration by wound-healing assay.at a dose of 10 μM, CX4945 significantly decreased cell viability of primary human cell cultures from both mucin and mixed CCA, whereas in CK19-positive cell cultures, the effect of CX4945 on cell viability required higher concentrations (>30μM. At the same concentrations, CX4945 also induced apoptosis (3- fold increase vs controls which correlated with the expression level of CK2 in the different CCA cell lines (mucin- and mixed-CCA. Indeed, no apoptotic effects were observed in CK19-positive cells expressing lower CK2 levels. The effects of CX4945 on viability and apoptosis were associated with an increased number of γ-H2ax (biomarker for DNA double-strand breaks foci, suggesting the active role of CK2 as

  1. Rate of hepatitis C viral clearance by human livers in human patients: Liver transplantation modeling primary infection and implications for studying entry inhibition.

    Directory of Open Access Journals (Sweden)

    Michael G Hughes

    Full Text Available To better understand the dynamics of early hepatitis C virus (HCV infection, we determined how rapidly non-cirrhotic HCV-uninfected liver allografts clear HCV from the circulation of cirrhotic HCV-infected patients at the time of transplantation but before administration of immunosuppression. Specifically, we characterized serum HCV kinetics during the first 90 min of reperfusion for 19 chronically HCV-infected patients transplanted with an HCV-uninfected liver by measuring serum viral load immediately prior to reperfusion (t = 0 and then every 15 min for a total of 90 min (t = 90. Immunosuppression was withheld until all samples were taken to better model primary infection. During this period, rates of viral clearance varied more than 20-fold with a median rate constant of 0.0357 1/min, range 0.0089-0.2169; half-life (minutes median 19.4, range 3.2-77.8. The majority of viral clearance occurred within the first 60 min. The amount of blood transfused during this 90-min period (a potential confounding variable of this human liver transplant model of primary infection accounted for 53% and 59% of k (r = 0.53, p = 0.05 and half-life (r = 0.59, p = 0.03 variability, respectively. No other clinical variables tested (age, allograft weight, and degree of reperfusion injury as assessed by peak postoperative ALT or AST accounted for the remaining variability (p>0.05.In a human liver transplant model of primary infection, HCV rapidly clears the bloodstream. With approximately 90% of clearance occurring in the first 90 minutes of reperfusion, studies of HCV entry inhibition could utilize rate of clearance during this early period as an outcome measure.

  2. Human papillomavirus as prognostic marker with rising prevalence in neck squamous cell carcinoma of unknown primary: A retrospective multicentre study.

    Science.gov (United States)

    Schroeder, Lea; Boscolo-Rizzo, Paolo; Dal Cin, Elisa; Romeo, Salvatore; Baboci, Lorena; Dyckhoff, Gerhard; Hess, Jochen; Lucena-Porcel, Carlota; Byl, Anne; Becker, Nikolaus; Alemany, Laia; Castellsagué, Xavier; Quer, Miquel; León, Xavier; Wiesenfarth, Manuel; Pawlita, Michael; Holzinger, Dana

    2017-03-01

    Patients with neck squamous cell carcinomas of unknown primary tumour (NSCCUP) present with lymph node metastasis without evidence for a primary tumour. Most patients undergo an aggressive multimodal treatment, which induces severe, potentially unnecessary toxicity. Primary tumours of NSCCUP can be hidden in the oropharynx. Human papillomavirus (HPV) is causally involved in a subgroup of oropharyngeal squamous cell carcinomas (OPSCC) associated with early lymph node metastasis and good prognosis. Detection of markers for HPV transformation in NSCCUP could allow focussing on the oropharynx in primary tumour search and could be of value for choice and extent of treatment. In a retrospective multicentre study (Germany, Italy and Spain), we analysed metastatic lymph nodes from 180 NSCCUP patients for the presence of HPV DNA, HPV E6*I mRNA and cellular p16INK4a overexpression, a surrogate marker for HPV-induced transformation. HPV status, defined as positivity for viral mRNA with at least one additional marker, was correlated with clinical parameters and survival outcome. A substantial proportion (16%) of NSCCUP were HPV-driven, mainly by HPV16 (89%). HPV prevalence increased with year of diagnosis from 9% during 1998-2004 to 23% during 2005-2014 (p = 0.007). HPV-driven NSCCUP had significantly better overall and progression-free survival rates (p ≤ 0.008). Based on this survival benefit, it is contended that HPV RNA status should be included in NSCCUP diagnosis and in therapeutic decision-making. Deintensification of radiation in patients with HPV-driven NSCCUP, while concurrently concentrating on the oropharynx appears to be a promising therapeutic strategy, the efficacy of which should be assessed in prospective trials. To our knowledge, this is the largest study on HPV in NSCCUP. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. A novel in vitro model for studying quiescence and activation of primary isolated human myoblasts

    DEFF Research Database (Denmark)

    Sellathurai, Jeeva; Cheedipudi, Sirisha; Dhawan, Jyotsna

    2013-01-01

    term exponentially proliferating cultures normally used for in vitro studies. Human myoblasts cultured through many passages inevitably consist of a mixture of proliferating and non-proliferating cells, while cells activated from G0 are in a synchronously proliferating phase, and therefore may...... be a better model for in vivo proliferating satellite cells. Furthermore, the temporal propagation of proliferation in these synchronized cultures resembles the pattern seen in vivo during regeneration. We therefore present this culture model as a useful and novel condition for molecular analysis...

  4. Methods and theory in bone modeling drift: comparing spatial analyses of primary bone distributions in the human humerus.

    Science.gov (United States)

    Maggiano, Corey M; Maggiano, Isabel S; Tiesler, Vera G; Chi-Keb, Julio R; Stout, Sam D

    2016-01-01

    This study compares two novel methods quantifying bone shaft tissue distributions, and relates observations on human humeral growth patterns for applications in anthropological and anatomical research. Microstructural variation in compact bone occurs due to developmental and mechanically adaptive circumstances that are 'recorded' by forming bone and are important for interpretations of growth, health, physical activity, adaptation, and identity in the past and present. Those interpretations hinge on a detailed understanding of the modeling process by which bones achieve their diametric shape, diaphyseal curvature, and general position relative to other elements. Bone modeling is a complex aspect of growth, potentially causing the shaft to drift transversely through formation and resorption on opposing cortices. Unfortunately, the specifics of modeling drift are largely unknown for most skeletal elements. Moreover, bone modeling has seen little quantitative methodological development compared with secondary bone processes, such as intracortical remodeling. The techniques proposed here, starburst point-count and 45° cross-polarization hand-drawn histomorphometry, permit the statistical and populational analysis of human primary tissue distributions and provide similar results despite being suitable for different applications. This analysis of a pooled archaeological and modern skeletal sample confirms the importance of extreme asymmetry in bone modeling as a major determinant of microstructural variation in diaphyses. Specifically, humeral drift is posteromedial in the human humerus, accompanied by a significant rotational trend. In general, results encourage the usage of endocortical primary bone distributions as an indicator and summary of bone modeling drift, enabling quantitative analysis by direction and proportion in other elements and populations. © 2015 Anatomical Society.

  5. Genetics Home Reference: juvenile primary osteoporosis

    Science.gov (United States)

    ... Home Health Conditions Juvenile primary osteoporosis Juvenile primary osteoporosis Printable PDF Open All Close All Enable Javascript ... view the expand/collapse boxes. Description Juvenile primary osteoporosis is a skeletal disorder characterized by thinning of ...

  6. Genetics Home Reference: juvenile primary lateral sclerosis

    Science.gov (United States)

    ... Conditions Juvenile primary lateral sclerosis Juvenile primary lateral sclerosis Printable PDF Open All Close All Enable Javascript ... the expand/collapse boxes. Description Juvenile primary lateral sclerosis is a rare disorder characterized by progressive weakness ...

  7. COmplex coronary Bifurcation lesions: RAndomized comparison of a strategy using a dedicated self-expanding biolimus-eluting stent versus a culotte strategy using everolimus-eluting stents: primary results of the COBRA trial.

    Science.gov (United States)

    Dubois, Christophe; Bennett, Johan; Dens, Joseph; De Cock, Dries; Desmet, Walter; Belmans, Ann; Ughi, Giovanni J; Sinnaeve, Peter; Vrolix, Mathias; D'hooge, Jan; Adriaenssens, Tom

    2016-04-20

    We aimed to compare healing responses with optical coherence tomography, and clinical and angiographic outcome after treatment of coronary bifurcation lesions with a dedicated stent versus a conventional culotte technique. Forty patients with true and complex coronary bifurcation lesions were randomly assigned to treatment with the Axxess™ bifurcation stent in the proximal main vessel (MV) and additional BioMatrix™ stents in the branches (Biosensors Europe SA, Morges, Switzerland), versus a culotte technique using XIENCE™ stents (Abbott Vascular, Santa Clara, CA, USA). The primary endpoint of percentage of uncovered struts at nine months was similar with the dedicated strategy vs. culotte in the proximal MV (median 17.8 [IQR 3.3-24.7] vs. 6.8 [2.0-20.5]; p=0.19), bifurcation core (9.5 [5.7-19.5] vs. 4.0 [0.7-17.6]; p=0.17), distal MV (2.6 [2.3-18] vs. 2.2 [0.5-6.0]; p=0.09) and side branch (5.7 [1.5-11.5] vs. 1.9 [0-5.8]; p=0.14). As compared with culotte, a strategy using Axxess resulted in a significantly larger lumen in the proximal MV both acutely (minimum lumen diameter 3.03±0.51 vs. 2.71±0.44 mm, p=0.04) and at follow-up (mean lumen area 10.0±2.1 vs. 7.1±1.8 mm2, pstrategies resulted in good clinical outcomes at one year, and no stent thromboses. As compared with a culotte strategy with XIENCE stents, complex bifurcation stenting using a dedicated strategy combining Axxess and BioMatrix stents results in similar stent strut coverage at nine-month follow-up, and a significantly larger lumen and lower angiographic late lumen loss in the proximal MV.