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Sample records for cells including primary

  1. Pancreatic-type Acinar Cell Carcinoma of the Stomach Included in Multiple Primary Carcinomas.

    PubMed

    Yonenaga, Yoshikuni; Kurosawa, Manabu; Mise, Masahiro; Yamagishi, Miki; Higashide, Shunichi

    2016-06-01

    Pancreatic-type acinar cell carcinoma (ACC) in the stomach is extraordinarily rare. We pathologically examined two cases with multiple primary carcinomas, including gastric tumors. Gastric cancer specimens were examined by immunostaining and electron microscopy. Both cases had cancer cells with acinar patterns, resembling pancreatic ACC. The cancer cells in the first case were positive for exocrine markers, including chymotrypsin, lipase and alpha-1 antichymotrypsin (ACT), as well as neuroendocrine markers, including chromogranin A and synaptophysin. The cancer cells in the second case were positive for chymotrypsin and alpha-1 ACT, while being slightly positive for chromogranin A and synaptophysin. Ultrastructurally, cancer cells contained zymogen granules in both cases. The final diagnosis was pancreatic mixed acinar-neuroendocrine carcinoma and pure pancreatic ACC, respectively. We confirmed two cases with gastric pancreatic-type ACC included in multiple primary carcinomas. This type of double cancer has not been reported previously. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  2. Subretinal Pigment Epithelial Deposition of Drusen Components Including Hydroxyapatite in a Primary Cell Culture Model

    PubMed Central

    Pilgrim, Matthew G.; Lengyel, Imre; Lanzirotti, Antonio; Newville, Matt; Fearn, Sarah; Emri, Eszter; Knowles, Jonathan C.; Messinger, Jeffrey D.; Read, Russell W.; Guidry, Clyde; Curcio, Christine A.

    2017-01-01

    Purpose Extracellular deposits containing hydroxyapatite, lipids, proteins, and trace metals that form between the basal lamina of the RPE and the inner collagenous layer of Bruch's membrane are hallmarks of early AMD. We examined whether cultured RPE cells could produce extracellular deposits containing all of these molecular components. Methods Retinal pigment epithelium cells isolated from freshly enucleated porcine eyes were cultured on Transwell membranes for up to 6 months. Deposit composition and structure were characterized using light, fluorescence, and electron microscopy; synchrotron x-ray diffraction and x-ray fluorescence; secondary ion mass spectroscopy; and immunohistochemistry. Results Apparently functional primary RPE cells, when cultured on 10-μm-thick inserts with 0.4-μm-diameter pores, can produce sub-RPE deposits that contain hydroxyapatite, lipids, proteins, and trace elements, without outer segment supplementation, by 12 weeks. Conclusions The data suggest that sub-RPE deposit formation is initiated, and probably regulated, by the RPE, as well as the loss of permeability of the Bruch's membrane and choriocapillaris complex associated with age and early AMD. This cell culture model of early AMD lesions provides a novel system for testing new therapeutic interventions against sub-RPE deposit formation, an event occurring well in advance of the onset of vision loss. PMID:28146236

  3. Cathode for use in high energy primary thionyl chloride cell systems and high energy primary thionyl chloride cell systems including the cathode

    NASA Astrophysics Data System (ADS)

    Walker, C. W., Jr.; Wade, W. L., Jr.; Binder, M.; Gilman, S.

    1985-08-01

    A cathode is provided for use in high energy primary lithium-thionyl chloride cell systems or calcium-thionyl chloride cell systems. The cathode comprises an expanded metallic current collector screen into which has been pasted a mixture of a low surface area conductive carbon black and a high surface area conductive carbon black previously mixed with a binder.

  4. Mechanisms of growth inhibition of primary prostate epithelial cells following gamma irradiation or photodynamic therapy include senescence, necrosis, and autophagy, but not apoptosis.

    PubMed

    Frame, Fiona M; Savoie, Huguette; Bryden, Francesca; Giuntini, Francesca; Mann, Vincent M; Simms, Matthew S; Boyle, Ross W; Maitland, Norman J

    2016-01-01

    In comparison to more differentiated cells, prostate cancer stem-like cells are radioresistant, which could explain radio-recurrent prostate cancer. Improvement of radiotherapeutic efficacy may therefore require combination therapy. We have investigated the consequences of treating primary prostate epithelial cells with gamma irradiation and photodynamic therapy (PDT), both of which act through production of reactive oxygen species (ROS). Primary prostate epithelial cells were cultured from patient samples of benign prostatic hyperplasia and prostate cancer prior to treatment with PDT or gamma irradiation. Cell viability was measured using MTT and alamar blue assay, and cell recovery by colony-forming assays. Immunofluorescence of gamma-H2AX foci was used to quantify DNA damage, and autophagy and apoptosis were assessed using Western blots. Necrosis and senescence were measured by propidium iodide staining and beta-galactosidase staining, respectively. Both PDT and gamma irradiation reduced the colony-forming ability of primary prostate epithelial cells. PDT reduced the viability of all types of cells in the cultures, including stem-like cells and more differentiated cells. PDT induced necrosis and autophagy, whereas gamma irradiation induced senescence, but neither treatment induced apoptosis. PDT and gamma irradiation therefore inhibit cell growth by different mechanisms. We suggest these treatments would be suitable for use in combination as sequential treatments against prostate cancer. © 2015 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  5. Senescence-associated β-galactosidase staining in fish cell lines and primary cultures from several tissues and species, including rainbow trout coelomic fluid and milt.

    PubMed

    Vo, Nguyen T K; Mikhaeil, Michael S; Lee, Lucy E J; Pham, Phuc H; Bols, Niels C

    2015-04-01

    Cell lines and primary cultures from several teleost tissues and species were stained for senescence-associated β-galactosidase (SA β-Gal), revealing four general outcomes. (1) For long-standing fish cell lines that can be considered immortal, little or no SA β-Gal staining was observed, regardless of the culture conditions. (2) For a new walleye cell line from the bulbus arteriosus (WEBA), most cells stained for SA β-Gal even after 40 passages. This suggested that high SA β-Gal activity was a unique property of WEBA, perhaps reflecting their endothelial character, rather than cellular senescence. (3) For cell lines developed from the walleye caudal fin and from somatic cells in rainbow trout coelomic fluid, no SA β-Gal staining was observed in the earliest cultures to over 70 passages later. This suggested that cells from these anatomical sites do not undergo senescence in vitro. (4) By contrast, for cell lines developed from the walleye brain and from somatic cells in rainbow trout milt, most cells in the early-stage cultures stained for SA β-Gal, but as these were developed into cell lines, SA β-Gal-negative cells became dominant. This suggested that if cellular senescence occurred in vitro, this happened early in these cultures and subsequently a few SA β-Gal-negative cells went onto to form the cell line. Overall, the presence of SA β-Gal-positive cells in cultures could be interpreted in several ways, whereas their absence predicted that in these cultures, cells would proliferate indefinitely.

  6. Multimodality therapy including salvage surgical resection and intraoperative radiotherapy for patients with squamous-cell carcinoma of the anus with residual or recurrent disease after primary chemoradiotherapy.

    PubMed

    Hallemeier, Christopher L; You, Y Nancy; Larson, David W; Dozois, Eric J; Nelson, Heidi; Klein, Kristi A; Miller, Robert C; Haddock, Michael G

    2014-04-01

    For patients with residual or recurrent squamous-cell carcinoma of the anus after primary chemoradiotherapy, the standard treatment is surgical salvage. Patients with unresectable or borderline unresectable disease have poor outcomes, thus adjunctive treatments should be explored. The aim of this study is to report outcomes for patients with residual/recurrent anal cancer treated with multimodality therapy including salvage surgical resection and intraoperative radiotherapy. This is an observational study. This study was conducted at a tertiary referral center. Thirty-two patients were treated between 1993 and 2012. Median age was 53 years (range, 34-87). Salvage treatment was performed for residual disease (n = 9), first recurrence (n = 17), or second recurrence (n = 6) after primary chemoradiotherapy. Patients with recurrent disease received preoperative external beam reirradiation with concurrent chemotherapy. All patients underwent salvage surgical resection and intraoperative radiotherapy. Extent of surgical resection was R0 (negative margins, n = 16), R1 (microscopic residual, n = 13), or R2 (macroscopic residual, n = 3). The median intraoperative radiotherapy dose was 12.5 Gy. Treatment-related adverse events were classified according to the National Cancer Institute - Common Toxicity Criteria. Overall and disease-free survival were estimated by using the Kaplan-Meier technique. Central, local-regional, and distant failure were estimated by the use of the cumulative incidence method. Median length of hospital stay was 9 days. Mortality at 30 days after surgery and intraoperative radiotherapy was 0%. Fifteen patients (47%) experienced a total of 16 grade 3 treatment-related adverse events (wound complication (n = 6), bowel obstruction (n = 5), and ureteral obstruction (n = 3)). The 5-year estimates of overall and disease-free survival were 23% and 17%. The 5-year estimates of central, local-regional, and distant failure were 21%, 51%, and 40%. This was a

  7. Tropism and Infectivity of Influenza Virus, Including Highly Pathogenic Avian H5N1 Virus, in Ferret Tracheal Differentiated Primary Epithelial Cell Cultures

    PubMed Central

    Zeng, Hui; Goldsmith, Cynthia S.; Maines, Taronna R.; Belser, Jessica A.; Gustin, Kortney M.; Pekosz, Andrew; Zaki, Sherif R.; Katz, Jacqueline M.

    2013-01-01

    Tropism and adaptation of influenza viruses to new hosts is partly dependent on the distribution of the sialic acid (SA) receptors to which the viral hemagglutinin (HA) binds. Ferrets have been established as a valuable in vivo model of influenza virus pathogenesis and transmission because of similarities to humans in the distribution of HA receptors and in clinical signs of infection. In this study, we developed a ferret tracheal differentiated primary epithelial cell culture model that consisted of a layered epithelium structure with ciliated and nonciliated cells on its apical surface. We found that human-like (α2,6-linked) receptors predominated on ciliated cells, whereas avian-like (α2,3-linked) receptors, which were less abundant, were presented on nonciliated cells. When we compared the tropism and infectivity of three human (H1 and H3) and two avian (H1 and H5) influenza viruses, we observed that the human influenza viruses primarily infected ciliated cells and replicated efficiently, whereas a highly pathogenic avian H5N1 virus (A/Vietnam/1203/2004) replicated efficiently within nonciliated cells despite a low initial infection rate. Furthermore, compared to other influenza viruses tested, VN/1203 virus replicated more efficiently in cells isolated from the lower trachea and at a higher temperature (37°C) compared to a lower temperature (33°C). VN/1203 virus infection also induced higher levels of immune mediator genes and cell death, and virus was recovered from the basolateral side of the cell monolayer. This ferret tracheal differentiated primary epithelial cell culture system provides a valuable in vitro model for studying cellular tropism, infectivity, and the pathogenesis of influenza viruses. PMID:23255802

  8. Epigenetics targeted protein-vorinostat nanomedicine inducing apoptosis in heterogeneous population of primary acute myeloid leukemia cells including refractory and relapsed cases.

    PubMed

    Chandran, Parwathy; Kavalakatt, Anu; Malarvizhi, Giridharan Loghanathan; Vasanthakumari, Divya Rani Vikraman Nair; Retnakumari, Archana Payickattu; Sidharthan, Neeraj; Pavithran, Keechilat; Nair, Shantikumar; Koyakutty, Manzoor

    2014-05-01

    Aberrant epigenetics play a key role in the onset and progression of acute myeloid leukemia (AML). Herein we report in silico modelling based development of a novel, protein-vorinostat nanomedicine exhibiting selective and superior anti-leukemic activity against heterogeneous population of AML patient samples (n=9), including refractory and relapsed cases, and three representative cell lines expressing CD34(+)/CD38(-) stem cell phenotype (KG-1a), promyelocytic phenotype (HL-60) and FLT3-ITD mutation (MV4-11). Nano-vorinostat having ~100nm size exhibited enhanced cellular uptake rendering significantly lower IC50 in AML cell lines and patient samples, and induced enhanced HDAC inhibition, oxidative injury, cell cycle arrest and apoptosis compared to free vorinostat. Most importantly, nanomedicine showed exceptional single-agent activity against the clonogenic proliferative capability of bone marrow derived leukemic progenitors, while remaining non-toxic to healthy bone marrow cells. Collectively, this epigenetics targeted nanomedicine appears to be a promising therapeutic strategy against various French-American-British (FAB) classes of AML. Through the use of a protein-vorinostat agent, exceptional single-agent activity was demonstrated against the clonogenic proliferative capability of bone marrow derived leukemic progenitors, while remaining non-toxic to healthy bone marrow cells. The studied epigenetics targeted nanomedicine approach is a promising therapeutic strategy against various French-American-British classes of acute myeloid leukemia. © 2014 Elsevier Inc. All rights reserved.

  9. Photoelectrochemical cells including chalcogenophosphate photoelectrodes

    NASA Technical Reports Server (NTRS)

    Reichman, B.; Byvik, C. E. (Inventor)

    1984-01-01

    Photoelectrochemical cells employing chalcogenophosphate (MPX3) photoelectrodes are described where M is selected from the group of transition metal series of elements beginning with scandium (atomic number 21) through germanium (atomic number 32) yttrium (atomic number 39) through antimony (atomic number 51) and lanthanum (atomic number 57) through polonium (atomic number 84); P is phosphorus; and X is selected from the chalogenide series consisting of sulfur, selenium, and tellurium. These compounds have bandgaps in the desirable range from 2.0 eV to 2.2 eV for the photoelectrolysis of water and are stable when used as photoelectrodes for the same.

  10. Photoelectrochemical cells including chalcogenophosphate photoelectrodes

    NASA Astrophysics Data System (ADS)

    Reichman, B.; Byvik, C. E.

    1984-03-01

    Photoelectrochemical cells employing chalcogenophosphate (MPX3) photoelectrodes are described where M is selected from the group of transition metal series of elements beginning with scandium (atomic number 21) through germanium (atomic number 32) yttrium (atomic number 39) through antimony (atomic number 51) and lanthanum (atomic number 57) through polonium (atomic number 84); P is phosphorus; and X is selected from the chalogenide series consisting of sulfur, selenium, and tellurium. These compounds have bandgaps in the desirable range from 2.0 eV to 2.2 eV for the photoelectrolysis of water and are stable when used as photoelectrodes for the same.

  11. Electrochemical cell structure including an ionomeric barrier

    DOEpatents

    Lambert, Timothy N.; Hibbs, Michael

    2017-06-20

    An apparatus includes an electrochemical half-cell comprising: an electrolyte, an anode; and an ionomeric barrier positioned between the electrolyte and the anode. The anode may comprise a multi-electron vanadium phosphorous alloy, such as VP.sub.x, wherein x is 1-5. The electrochemical half-cell is configured to oxidize the vanadium and phosphorous alloy to release electrons. A method of mitigating corrosion in an electrochemical cell includes disposing an ionomeric barrier in a path of electrolyte or ion flow to an anode and mitigating anion accumulation on the surface of the anode.

  12. [Primary orbital squamous cell carcinoma].

    PubMed

    Campos Arbulú, Ana L; Sadava, Emmanuel E; Sánchez Ruiz, Alejandro; Fernández Vila, Juan M; Dillon, Horacio S; Mezzadri, Norberto A

    2017-01-01

    Primary orbital squamous cell carcinoma is a rare entity. There is little published literature. We report a case of primary squamous cell carcinoma of the orbital soft tissues. Surgical resection offered the best treatment for the patient. Complete resection of the lesion was achieved. The patient received adjuvant radiotherapy due to the proximity of the lesion to the surgical margins. Surgical treatment is feasible and should be considered as part of the surgeon's arsenal. However, therapeutic decisions must be made on a case-by-case basis.

  13. Primary gastric mantle cell lymphoma

    PubMed Central

    Petranovic, Duska; Pilcic, Gorazd; Peitl, Milena; Cubranic, Aleksandar; Valkovic, Toni; Nacinovic, Antica Duletic; Lucin, Ksenija; Jonjic, Nives

    2012-01-01

    Mantle cell lymphoma represents 2.5–7% all of non Hodgkin's lymphomas. Stomach is the most common site of extranodal lymphoma. However, that is not the case with mantle cell lymphoma, which is extremely rare. We present a case of 71-year-old woman admitted to the Internal Clinic of the University Clinical Hospital Center Rijeka, because of stomach discomfort and melena. Endoscopy and computed tomography revealed a polyp in gastric antrum. Histopathologic, immunohistochemic and genetic methods were also performed and the results were consistent with primary gastric mantle cell lymphoma without periepigastric and/or local or distant abdominal lymph node involvement. PMID:22567215

  14. Primary retroperitoneal acinar cell cystadenoma.

    PubMed

    Pesci, Anna; Castelli, Paola; Facci, Enrico; Romano, Luigi; Zamboni, Giuseppe

    2012-03-01

    In this report, we describe a case of hitherto unreported primary retroperitoneal acinar cell cystadenoma that morphologically and immunophenotypically resembled pancreatic acinar cell cystadenoma. Pancreatic acinar cell cystadenoma is a very uncommon benign lesion characterized by acinar cell differentiation, the evidence of pancreatic exocrine enzyme production, and the absence of cellular atypia. Our case occurred in a 55-year-old woman presenting a 10-cm multilocular cystic lesion in the retroperitoneum thought to be a mucinous cystic neoplasm. At laparotomy, the cystic mass, which showed no connection with any organ, was completely resected with a clinical diagnosis of cystic lymphangioma. The diagnosis of retroperitoneal acinar cell cystadenoma was based on the recognition of morphological acinar differentiation, the immunohistochemical demonstration of the acinar marker trypsin, and the absence of cellular atypia. These peculiar features can be used in the differential diagnosis with all the other cystic lesions of the retroperitoneum.

  15. Certain Adenylated Non-Coding RNAs, Including 5′ Leader Sequences of Primary MicroRNA Transcripts, Accumulate in Mouse Cells following Depletion of the RNA Helicase MTR4

    PubMed Central

    Dorweiler, Jane E.; Ni, Ting; Zhu, Jun; Munroe, Stephen H.; Anderson, James T.

    2014-01-01

    RNA surveillance plays an important role in posttranscriptional regulation. Seminal work in this field has largely focused on yeast as a model system, whereas exploration of RNA surveillance in mammals is only recently begun. The increased transcriptional complexity of mammalian systems provides a wider array of targets for RNA surveillance, and, while many questions remain unanswered, emerging data suggest the nuclear RNA surveillance machinery exhibits increased complexity as well. We have used a small interfering RNA in mouse N2A cells to target the homolog of a yeast protein that functions in RNA surveillance (Mtr4p). We used high-throughput sequencing of polyadenylated RNAs (PA-seq) to quantify the effects of the mMtr4 knockdown (KD) on RNA surveillance. We demonstrate that overall abundance of polyadenylated protein coding mRNAs is not affected, but several targets of RNA surveillance predicted from work in yeast accumulate as adenylated RNAs in the mMtr4KD. microRNAs are an added layer of transcriptional complexity not found in yeast. After Drosha cleavage separates the pre-miRNA from the microRNA's primary transcript, the byproducts of that transcript are generally thought to be degraded. We have identified the 5′ leading segments of pri-miRNAs as novel targets of mMtr4 dependent RNA surveillance. PMID:24926684

  16. Primary lithium cell life studies

    NASA Technical Reports Server (NTRS)

    Capulli, John; Donley, Sam; Deligiannis, Frank; Shen, David

    1990-01-01

    One solution for providing a truly independent power source is to package, within the critical subsystem element, a primary battery that can remain dormant for time periods as long as the mission life, which can be 10-15 years, maximum. When primary power from the spacecraft solar array/battery system is interrupted, the backup battery system, which is connected through a diode to the power input line, would automatically support the load to avoid a power interruption to the critical load for a time period long enough to ensure that ground control could access the satellite and correct the anomaly by sending appropriate commands to the spacecraft. Critical subsystems identified for the application are telemetry and command circuits, volatile computer memory, attitude control circuits, and some critical payloads. Due to volume packaging and weight restrictions that exist on most spacecraft, coupled with the long storage periods required, lithium cell technology was selected for the backup power source. Because of the high energy density (200-400 Wh/kg), long shelf life, and load capability, soluble cathode primary lithium technology was chosen. The most important lithium cell properties that require detail characterization for this application are capacity loss, shelf life, and the voltage delay mechanism. These are functions of storage time and temperature. During storage, a passive film builds up on the lithium electrode. The film protects the lithium electrode from progressive capacity decay but requires time to break down when a load is applied. This phenomenon results in a depressed voltage during the period of film breakdown which can last from fractions of a second to minutes.

  17. 42 CFR 81.25 - Guidelines for claims including two or more primary cancers.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... cancers. 81.25 Section 81.25 Public Health PUBLIC HEALTH SERVICE, DEPARTMENT OF HEALTH AND HUMAN SERVICES... Estimate Probability of Causation § 81.25 Guidelines for claims including two or more primary cancers. For claims including two or more primary cancers, DOL will use NIOSH-IREP to calculate the estimated...

  18. 42 CFR 81.25 - Guidelines for claims including two or more primary cancers.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... cancers. 81.25 Section 81.25 Public Health PUBLIC HEALTH SERVICE, DEPARTMENT OF HEALTH AND HUMAN SERVICES... Estimate Probability of Causation § 81.25 Guidelines for claims including two or more primary cancers. For claims including two or more primary cancers, DOL will use NIOSH-IREP to calculate the estimated...

  19. 42 CFR 81.25 - Guidelines for claims including two or more primary cancers.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... cancers. 81.25 Section 81.25 Public Health PUBLIC HEALTH SERVICE, DEPARTMENT OF HEALTH AND HUMAN SERVICES... Estimate Probability of Causation § 81.25 Guidelines for claims including two or more primary cancers. For claims including two or more primary cancers, DOL will use NIOSH-IREP to calculate the estimated...

  20. 42 CFR 81.25 - Guidelines for claims including two or more primary cancers.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... cancers. 81.25 Section 81.25 Public Health PUBLIC HEALTH SERVICE, DEPARTMENT OF HEALTH AND HUMAN SERVICES... Estimate Probability of Causation § 81.25 Guidelines for claims including two or more primary cancers. For claims including two or more primary cancers, DOL will use NIOSH-IREP to calculate the estimated...

  1. 42 CFR 81.25 - Guidelines for claims including two or more primary cancers.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... cancers. 81.25 Section 81.25 Public Health PUBLIC HEALTH SERVICE, DEPARTMENT OF HEALTH AND HUMAN SERVICES... Estimate Probability of Causation § 81.25 Guidelines for claims including two or more primary cancers. For claims including two or more primary cancers, DOL will use NIOSH-IREP to calculate the estimated...

  2. Tumor patients in psychodynamic psychotherapy including daydreaming: can imagery enhance primary process and positive emotions?

    PubMed

    Frick, Eckhard; Stigler, Michael; Georg, Hildegunde; Fischer, Norbert; Bumeder, Irmgard; Pokorny, Dan

    2008-07-01

    This therapy process study investigates the use of guided affective imagery for tumor patients. The therapeutic access to tumor patients is generally described as complex and challenging because of a disturbed emotion regulation and a defensive focus on reality. After autologous blood stem cell transplantation, 29 patients were treated with psychotherapy, including two daydreaming imagery sessions. Three text-analytical measures--Affective Dictionary Ulm, Regressive Imagery Dictionary, and Computerized Referential Activity for verbatim session transcripts--as well as the Quality of Life Questionnaire and the Karnofsky Performance Status were administered. Results show that guided affective imagery was able to enhance the psychotherapeutic process in tumor patients by activating the primary process, decreasing anxiety, and increasing referential activity. The positive emotional shift during imagery was achieved by the patients irrespective of their oncological severity status. Study limitations and future directions for research are discussed.

  3. Molluscan cells in culture: primary cell cultures and cell lines

    PubMed Central

    Yoshino, T. P.; Bickham, U.; Bayne, C. J.

    2013-01-01

    In vitro cell culture systems from molluscs have significantly contributed to our basic understanding of complex physiological processes occurring within or between tissue-specific cells, yielding information unattainable using intact animal models. In vitro cultures of neuronal cells from gastropods show how simplified cell models can inform our understanding of complex networks in intact organisms. Primary cell cultures from marine and freshwater bivalve and gastropod species are used as biomonitors for environmental contaminants, as models for gene transfer technologies, and for studies of innate immunity and neoplastic disease. Despite efforts to isolate proliferative cell lines from molluscs, the snail Biomphalaria glabrata Say, 1818 embryonic (Bge) cell line is the only existing cell line originating from any molluscan species. Taking an organ systems approach, this review summarizes efforts to establish molluscan cell cultures and describes the varied applications of primary cell cultures in research. Because of the unique status of the Bge cell line, an account is presented of the establishment of this cell line, and of how these cells have contributed to our understanding of snail host-parasite interactions. Finally, we detail the difficulties commonly encountered in efforts to establish cell lines from molluscs and discuss how these difficulties might be overcome. PMID:24198436

  4. Towards Optimal Education Including Self-Regulated Learning in Technology-Enhanced Preschools and Primary Schools

    ERIC Educational Resources Information Center

    Mooij, Ton; Dijkstra, Elma M.; Walraven, Amber; Kirschner, Paul A.

    2014-01-01

    At the start of preschool, four-year-old pupils differ in their development, including in their capacity to self-regulate their playing and learning. In preschool and primary school, educational processes are generally adapted to the mean age of the pupils in the class. The same may apply to pupil-monitoring systems based on information and…

  5. Towards Optimal Education Including Self-Regulated Learning in Technology-Enhanced Preschools and Primary Schools

    ERIC Educational Resources Information Center

    Mooij, Ton; Dijkstra, Elma M.; Walraven, Amber; Kirschner, Paul A.

    2014-01-01

    At the start of preschool, four-year-old pupils differ in their development, including in their capacity to self-regulate their playing and learning. In preschool and primary school, educational processes are generally adapted to the mean age of the pupils in the class. The same may apply to pupil-monitoring systems based on information and…

  6. Helping Teachers to Include Children with Special Educational Needs in the Primary Language Classroom

    ERIC Educational Resources Information Center

    Shield, Margaret

    2005-01-01

    One of the challenges facing primary school language teachers is the adaptation of the language syllabus to suit the needs of all of their learners, including those with special educational needs. Research into inclusion of such learners in the regular classroom has shown that adequate support for both child and teacher is essential if a child is…

  7. Thin film solar cell including a spatially modulated intrinsic layer

    DOEpatents

    Guha, Subhendu; Yang, Chi-Chung; Ovshinsky, Stanford R.

    1989-03-28

    One or more thin film solar cells in which the intrinsic layer of substantially amorphous semiconductor alloy material thereof includes at least a first band gap portion and a narrower band gap portion. The band gap of the intrinsic layer is spatially graded through a portion of the bulk thickness, said graded portion including a region removed from the intrinsic layer-dopant layer interfaces. The band gap of the intrinsic layer is always less than the band gap of the doped layers. The gradation of the intrinsic layer is effected such that the open circuit voltage and/or the fill factor of the one or plural solar cell structure is enhanced.

  8. Relationship of sea level muon charge ratio to primary composition including nuclear target effects

    NASA Technical Reports Server (NTRS)

    Goned, A.; Shalaby, M.; Salem, A. M.; Roushdy, M.

    1985-01-01

    The discrepancy between the muon charge ratio observed at low energies and that calculated using pp data is removed by including nuclear target effects. Calculations at high energies show that the primary iron spectrum is expected to change slope from 2 to 2.2 to 2.4 to 2.5 for energies approx. 4 x 10 to the 3 GeV/nucleon if scaling features continue to the highest energies.

  9. COST Action "EuroTelepath": digital pathology integration in electronic health record, including primary care centres.

    PubMed

    Rojo, Marcial García; Castro, Ana Morillo; Gonçalves, Luis

    2011-03-30

    Digital pathology includes the information technology that allows for the management of information, including data and images, generated in an anatomic pathology department. COST ACTION IC0604: The integration of digital slides in the electronic health record is one of the main objectives of COST Action IC0604 "Telepathology Network in Europe" (EURO-TELEPATH). Fostering use of medical informatics standards and adapting them to current needs is needed to manage efficiently extremely large medical images, like digital slide files. Digital slides can play a role in disease prevention, primary diagnosis, and second opinion. In all these tasks, automated image analysis can also be a most valuable tool. In order to achieve an efficient interoperability between pathology information systems with other clinical information systems, obtaining a seamless integration of pathology images (gross pictures and digital slides) with LIS-Pathology Information system in a web environment is an important task. Primary care information systems should also be included in the integration, since primary care centres play an essential role in the generation of clinical information and specimen collection. A common terminology, based in SNOMED CT is also needed. Main barrier in the integration of digital slides in pathology workflow and eHealth record is the cost of current digital slide scanners. Pathology information system vendors should participate in standardization bodies.

  10. Dendritic cell analysis in primary immunodeficiency

    PubMed Central

    Bigley, Venetia; Barge, Dawn; Collin, Matthew

    2016-01-01

    Purpose of review Dendritic cells are specialized antigen-presenting cells which link innate and adaptive immunity, through recognition and presentation of antigen to T cells. Although the importance of dendritic cells has been demonstrated in many animal models, their contribution to human immunity remains relatively unexplored in vivo. Given their central role in infection, autoimmunity, and malignancy, dendritic cell deficiency or dysfunction would be expected to have clinical consequences. Recent findings Human dendritic cell deficiency disorders, related to GATA binding protein 2 (GATA2) and interferon regulatory factor 8 (IRF8) mutations, have highlighted the importance of dendritic cells and monocytes in primary immunodeficiency diseases and begun to shed light on their nonredundant roles in host defense and immune regulation in vivo. The contribution of dendritic cell and monocyte dysfunction to the pathogenesis of primary immunodeficiency disease phenotypes is becoming increasingly apparent. However, dendritic cell analysis is not yet a routine part of primary immunodeficiency disease workup. Summary Widespread uptake of dendritic cell/monocyte screening in clinical practice will facilitate the discovery of novel dendritic cell and monocyte disorders as well as advancing our understanding of human dendritic cell biology in health and disease. PMID:27755182

  11. Primary Lithium-Thionyl Chloride Cell Evaluation

    DTIC Science & Technology

    1980-08-01

    AD A09466 0 AFWAL-TR-80-2076 PRIMARY LITHIUM THIONYL - CHLORIDE CELL EVALUATION Dr. A.E. Zolla R.R. Waterhouse D.J. DeBiccari G.L. Griffin, Jr. Altus...dS.,_b,I ......... S TYPE OF REPORT A PERIOD COVERED Primary Lithium - Thionyl Chloride Final 9/79 - 4/80 Cell Evaluation, 6 PERFORMING ORG. REPORT...the high performance characteristics of the Altus lithium - thionyl chloride cell. In particular features such as the inherent high energy density, the

  12. Analysis of primary cilia in directional cell migration in fibroblasts.

    PubMed

    Christensen, Søren T; Veland, Iben R; Schwab, Albrecht; Cammer, Michael; Satir, Peter

    2013-01-01

    Early studies of migrating fibroblasts showed that primary cilia orient in front of the nucleus and point toward the leading edge. Recent work has shown that primary cilia coordinate a series of signaling pathways critical to fibroblast cell migration during development and in wound healing. In particular, platelet-derived growth factor receptor alpha (PDGFRα) is compartmentalized to the primary cilium to activate signaling pathways that regulate reorganization of the cytoskeleton required for lamellipodium formation and directional migration in the presence of a specific ligand gradient. We summarize selected methods in analyzing ciliary function in directional cell migration, including immunofluorescence microscopy, scratch assay, and chemotaxis assay by micropipette addition of PDGFRα ligands to cultures of fibroblasts. These methods should be useful not only in studying cell migration but also more generally in delineating response pathways in cells with primary cilia. Copyright © 2013 Elsevier Inc. All rights reserved.

  13. Fuel cell repeater unit including frame and separator plate

    SciTech Connect

    Yamanis, Jean; Hawkes, Justin R; Chiapetta, Jr., Louis; Bird, Connie E; Sun, Ellen Y; Croteau, Paul F

    2013-11-05

    An example fuel cell repeater includes a separator plate and a frame establishing at least a portion of a flow path that is operative to communicate fuel to or from at least one fuel cell held by the frame relative to the separator plate. The flow path has a perimeter and any fuel within the perimeter flow across the at least one fuel cell in a first direction. The separator plate, the frame, or both establish at least one conduit positioned outside the flow path perimeter. The conduit is outside of the flow path perimeter and is configured to direct flow in a second, different direction. The conduit is fluidly coupled with the flow path.

  14. Modelling of an activated primary settling tank including the fermentation process and VFA elutriation.

    PubMed

    Ribes, J; Ferrer, J; Bouzas, A; Seco, A

    2002-10-01

    A complete model of a primary settler including both sedimentation and biological processes is presented. It is a one-dimensional model based on the solids flux concept and the conservation of mass that uses the Takács model for the settling velocity, which is corrected by a compression function in the lower layers. The biological model is based on the ASM2 and enlarged with the fermentation model proposed by this research group. The settler was split in ten layers and the flux terms in the mass balance for each layer is obtained by means of the settling model. A pilot plant has been operated to study the primary sludge fermentation and volatile fatty acids (VFA) elutriation in a primary settler tank. The model has been tested with pilot plant experimental data with very good results. It has been able to simulate the VFA production in the settler and their elutriation with the influent wastewater for all the studied experiments. The developed model is easily applicable to secondary settlers and thickeners, also taking into account biological activity inside them.

  15. Electrolytes including fluorinated solvents for use in electrochemical cells

    DOEpatents

    Tikhonov, Konstantin; Yip, Ka Ki; Lin, Tzu-Yuan

    2015-07-07

    Provided are electrochemical cells and electrolytes used to build such cells. The electrolytes include ion-supplying salts and fluorinated solvents capable of maintaining single phase solutions with the salts at between about -30.degree. C. to about 80.degree. C. The fluorinated solvents, such as fluorinated carbonates, fluorinated esters, and fluorinated esters, are less flammable than their non-fluorinated counterparts and increase safety characteristics of cells containing these solvents. The amount of fluorinated solvents in electrolytes may be between about 30% and 80% by weight not accounting weight of the salts. Fluorinated salts, such as fluoroalkyl-substituted LiPF.sub.6, fluoroalkyl-substituted LiBF.sub.4 salts, linear and cyclic imide salts as well as methide salts including fluorinated alkyl groups, may be used due to their solubility in the fluorinated solvents. In some embodiments, the electrolyte may also include a flame retardant, such as a phosphazene or, more specifically, a cyclic phosphazene and/or one or more ionic liquids.

  16. Primary cutaneous T-cell lymphomas.

    PubMed

    Rosen, Steven T; Querfeld, Christiane

    2006-01-01

    Primary cutaneous T-cell lymphomas (CTCLs) encompass a clinically and biologically heterogeneous group of non-Hodgkin lymphomas (NHLs) defined by clonal proliferation of skin-homing malignant T lymphocytes and natural killer cells. They account for up to 75% to 80% of all cutaneous lymphomas. The current WHO-EORTC classification of cutaneous lymphomas with primary cutaneous manifestations lists 13 entities. The most common subtypes-mycosis fungoides, Sézary syndrome, primary cutaneous anaplastic large cell lymphoma, and lymphomatoid papulosis-which represent approximately 95% of CTCLs, will be discussed in the following review. Each entity has unique biological characteristics and clinical course. Topical and/or systemic therapies are employed based on the stage of the disease and the tempo of progression.

  17. 75 FR 66798 - Ormet Primary Aluminum Corporation Including On-Site Temporary Workers, Hannibal, OH; Notice of...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-29

    ... Employment and Training Administration Ormet Primary Aluminum Corporation Including On-Site Temporary Workers... regarding eligibility for workers and former workers of Ormet Primary Aluminum Corporation, including on... United States production, consumption, and importation of primary and secondary aluminum to supplement...

  18. Association of types of dyspnea including 'bendopnea' with cardiopulmonary disease in primary care.

    PubMed

    Martínez Cerón, Diana María; Garcia Rosa, Maria Luiza; Lagoeiro Jorge, Antônio Jose; de Andrade Martins, Wolney; Tinoco Mesquita, Evandro; Di Calafriori Freire, Monica; da Silva Correia, Dayse Mary; Kang, Hye Chung

    2017-03-01

    Dyspnea is the symptom most commonly reported by patients with heart failure (HF) and/or pulmonary disease, the obese and the elderly. Recently 'bendopnea' (shortness of breath when bending forward) has been described in patients with HF. To determine the association of exertional dyspnea, orthopnea, paroxysmal nocturnal dyspnea and bendopnea with chronic disease, especially heart failure, and their phenotypes in primary care. This cross-sectional study included 633 individuals aged between 45 and 99 years enrolled in a primary care program in Niteroi, Brazil. Participants underwent clinical assessment and laboratory tests and completed a questionnaire, all on the same day. Paroxysmal nocturnal dyspnea and bendopnea were associated with HF (unadjusted OR 2.42, 95% CI 1.10-5.29 and OR 2.59, 95% CI 1.52-4.44, respectively). In multivariate models, chronic obstructive pulmonary disease, coronary heart disease and myocardial infarction were not associated with bendopnea. Bendopnea was the only type of dyspnea not linked to respiratory disease or coronary heart disease. Even after adjusting for depression and body mass index, the association remained with HF with or without preserved ejection fraction, and bendopnea thus appears to be a promising symptom to differentiate HF from the other two disease groups. Copyright © 2016 Sociedade Portuguesa de Cardiologia. Publicado por Elsevier España, S.L.U. All rights reserved.

  19. Treatment Option Overview (Plasma Cell Neoplasms Including Multiple Myeloma)

    MedlinePlus

    ... Neoplasms for more information. High-dose chemotherapy with stem cell transplant This treatment is a way of giving ... blood -forming cells destroyed by the cancer treatment. Stem cells (immature blood cells) are removed from the blood ...

  20. General Information about Plasma Cell Neoplasms (Including Multiple Myeloma)

    MedlinePlus

    ... Neoplasms for more information. High-dose chemotherapy with stem cell transplant This treatment is a way of giving ... blood -forming cells destroyed by the cancer treatment. Stem cells (immature blood cells) are removed from the blood ...

  1. Treatment Options for Plasma Cell Neoplasms (Including Multiple Myeloma)

    MedlinePlus

    ... Neoplasms for more information. High-dose chemotherapy with stem cell transplant This treatment is a way of giving ... blood -forming cells destroyed by the cancer treatment. Stem cells (immature blood cells) are removed from the blood ...

  2. Stages of Plasma Cell Neoplasms (Including Multiple Myeloma)

    MedlinePlus

    ... Neoplasms for more information. High-dose chemotherapy with stem cell transplant This treatment is a way of giving ... blood -forming cells destroyed by the cancer treatment. Stem cells (immature blood cells) are removed from the blood ...

  3. Primary processes in sensory cells: current advances.

    PubMed

    Frings, Stephan

    2009-01-01

    In the course of evolution, the strong and unremitting selective pressure on sensory performance has driven the acuity of sensory organs to its physical limits. As a consequence, the study of primary sensory processes illustrates impressively how far a physiological function can be improved if the survival of a species depends on it. Sensory cells that detect single-photons, single molecules, mechanical motions on a nanometer scale, or incredibly small fluctuations of electromagnetic fields have fascinated physiologists for a long time. It is a great challenge to understand the primary sensory processes on a molecular level. This review points out some important recent developments in the search for primary processes in sensory cells that mediate touch perception, hearing, vision, taste, olfaction, as well as the analysis of light polarization and the orientation in the Earth's magnetic field. The data are screened for common transduction strategies and common transduction molecules, an aspect that may be helpful for researchers in the field.

  4. Allogeneic transplantation strategies including haploidentical transplantation in sickle cell disease.

    PubMed

    Gluckman, Eliane

    2013-01-01

    Sickle cell disease (SCD) is the most common inherited hemoglobinopathy. Despite antenatal counseling and neonatal screening programs implemented in higher income countries, SCD is still associated with multiple morbidities and early mortality. To date, the only curative approach to SCD is hematopoietic stem cell transplantation, but this therapy is not yet established worldwide. The registries of the European Blood and Marrow Transplant (EBMT) and the Centre for International Blood and Marrow Transplant Research (CIBMTR) account, respectively, for 611 and 627 patients receiving transplantations for SCD. Most of these patients were transplanted with grafts from an HLA-identical sibling donor. The main obstacles to increasing the number of transplantations are a lack of awareness on the part of physicians and families, the absence of reliable prognostic factors for severity, and the perceived risk that transplantation complications may outweigh the benefits of early transplantation. Results show that more than 90% of patients having undergone an HLA-identical sibling transplantation after myeloablative conditioning are cured, with very limited complications. Major improvement is expected from the use of new reduced-toxicity conditioning regimens and the use of alternative donors, including unrelated cord blood transplantations and related haploidentical bone marrow or peripheral blood stem cell transplantations.

  5. Primary cell culture of human adenocarcinomas--practical considerations.

    PubMed

    Lerescu, Lucian; Tucureanu, Cătălin; Caraş, Iuliana; Neagu, Stefan; Melinceanu, Laura; Sălăgeanu, Aurora

    2008-01-01

    Cell culture is one of the major tools for oncology research, being an excellent system in which to study the biochemistry and molecular biology associated with individual cancer types and to understand cancer cell physiology. Progress in understanding the biology of any type of carcinoma has been impeded by the inability to culture adequately malignant cells from most epithelial tissues. The ultimate in vitro tumor model would completely reflect the in vivo tumor microenvironment in function and mechanism. Unfortunately, such a model does not currently exist. Homogeneous cell lines that can be continuously propagated on plastic surfaces have been extensively used as a surrogate for tumor environment; however they are very different from the in vivo tumor cells. Model systems involving primary culture represent the situation most closely related to the original tissue although they have a number of disadvantages over cell lines, such as the limited ability to repeat studies with a well characterized culture system that can be used in multiple laboratories. The primary culture may contain many types of stromal and infiltrating cell types potentially complicating the interpretation of data. Yet, their properties better reflect the cellular interactions present in intact tissue. The present article reviews the critical steps in obtaining, routine maintenance and cryopreservation of primary tumor cell cultures, based on information from literature and personal experience on the subject. The article also includes an updated protocol for primary tumor cell isolation and culture.

  6. T cell-B cell interactions in primary immunodeficiencies.

    PubMed

    Tangye, Stuart G; Deenick, Elissa K; Palendira, Umaimainthan; Ma, Cindy S

    2012-02-01

    Regulated interactions between cells of the immune system facilitate the generation of successful immune responses, thereby enabling efficient neutralization and clearance of pathogens and the establishment of both cell- and humoral-mediated immunological memory. The corollary of this is that impediments to efficient cell-cell interactions, normally necessary for differentiation and effector functions of immune cells, underly the clinical features and disease pathogenesis of primary immunodeficiencies. In affected individuals, these defects manifest as impaired long-term humoral immunity and susceptibility to infection by specific pathogens. In this review, we discuss the importance of, and requirements for, effective interactions between B cells and T cells during the formation of CD4(+) T follicular helper cells and the elicitation of cytotoxic function of virus-specific CD8(+) T cells, as well as how these processes are abrogated in primary immunodeficiencies due to loss-of-function mutations in defined genes. © 2012 New York Academy of Sciences.

  7. Failure of cell cleavage induces senescence in tetraploid primary cells.

    PubMed

    Panopoulos, Andreas; Pacios-Bras, Cristina; Choi, Justin; Yenjerla, Mythili; Sussman, Mark A; Fotedar, Rati; Margolis, Robert L

    2014-10-15

    Tetraploidy can arise from various mitotic or cleavage defects in mammalian cells, and inheritance of multiple centrosomes induces aneuploidy when tetraploid cells continue to cycle. Arrest of the tetraploid cell cycle is therefore potentially a critical cellular control. We report here that primary rat embryo fibroblasts (REF52) and human foreskin fibroblasts become senescent in tetraploid G1 after drug- or small interfering RNA (siRNA)-induced failure of cell cleavage. In contrast, T-antigen-transformed REF52 and p53+/+ HCT116 tumor cells rapidly become aneuploid by continuing to cycle after cleavage failure. Tetraploid primary cells quickly become quiescent, as determined by loss of the Ki-67 proliferation marker and of the fluorescent ubiquitination-based cell cycle indicator/late cell cycle marker geminin. Arrest is not due to DNA damage, as the γ-H2AX DNA damage marker remains at control levels after tetraploidy induction. Arrested tetraploid cells finally become senescent, as determined by SA-β-galactosidase activity. Tetraploid arrest is dependent on p16INK4a expression, as siRNA suppression of p16INK4a bypasses tetraploid arrest, permitting primary cells to become aneuploid. We conclude that tetraploid primary cells can become senescent without DNA damage and that induction of senescence is critical to tetraploidy arrest.

  8. Battery cell for a primary battery

    SciTech Connect

    Hakkinen, A.

    1984-12-11

    A battery cell for a primary battery, particularly a flat cell battery to be activated on being taken into use, e.g., when submerged into water. The battery cell comprises a positive current collector and a negative electrode. A separator layer which, being in contact with the negative electrode, is disposed between said negative electrode and the positive current collector. A depolarizing layer containing a depolarizing agent is disposed between the positive current collector and the separate layer. An intermediate layer of a porous, electrically insulating, and water-absorbing material is disposed next to the positive current collector and arranged in contact with the depolarizing agent.

  9. Mantle cell lymphoma: primary oral presentation.

    PubMed

    Ainscough, S; Power, A M; Brown, A N

    2017-01-01

    Mantle-cell lymphoma is an uncommon lymphoid malignancy of B-cells. It is often aggressive and prognosis is poor. A 69-year-old gentleman with a history of ischaemic heart disease was referred from primary care with a painless right floor of mouth swelling that had been present for 1 month. He otherwise completely asymptomatic. Incisional biopsy of the lesion was undertaken and marker studies demonstrated mantle cell lymphoma. Positron emission tomography-computed tomography and bone marrow biopsy showed widespread but low volume involvement. The patient was referred to the haematology multidisciplinary team for further assessment and treatment.

  10. Primary cells and stem cells in drug discovery: emerging tools for high-throughput screening.

    PubMed

    Eglen, Richard; Reisine, Terry

    2011-04-01

    Many drug discovery screening programs employ immortalized cells, recombinantly engineered to express a defined molecular target. Several technologies are now emerging that render it feasible to employ more physiologically, and clinically relevant, cell phenotypes. Consequently, numerous approaches use primary cells, which retain many functions seen in vivo, as well as endogenously expressing the target of interest. Furthermore, stem cells, of either embryonic or adult origin, as well as those derived from differentiated cells, are now finding a place in drug discovery. Collectively, these cells are expanding the utility of authentic human cells, either as screening tools or as therapeutics, as well as providing cells derived directly from patients. Nonetheless, the growing use of phenotypically relevant cells (including primary cells or stem cells) is not without technical difficulties, particularly when their envisioned use lies in high-throughput screening (HTS) protocols. In particular, the limited availability of homogeneous primary or stem cell populations for HTS mandates that novel technologies be developed to accelerate their adoption. These technologies include detection of responses with very few cells as well as protocols to generate cell lines in abundant, homogeneous populations. In parallel, the growing use of changes in cell phenotype as the assay readout is driving greater use of high-throughput imaging techniques in screening. Taken together, the greater availability of novel primary and stem cell phenotypes as well as new detection technologies is heralding a new era of cellular screening. This convergence offers unique opportunities to identify drug candidates for disorders at which few therapeutics are presently available.

  11. Cutaneous primary B-cell lymphomas: from diagnosis to treatment*

    PubMed Central

    Lima, Margarida

    2015-01-01

    Primary cutaneous B-cell lymphomas are a heterogeneous group of mature B-cells neoplasms with tropism for the skin, whose biology and clinical course differ significantly from the equivalent nodal lymphomas. The most indolent forms comprise the primary cutaneous marginal zone and follicle center B-cell lymphomas that despite the excellent prognosis have cutaneous recurrences very commonly. The most aggressive forms include the primary cutaneous large B-cell lymphomas, consisting in two major groups: the leg type, with poor prognosis, and others, the latter representing a heterogeneous group of lymphomas from which specific entities are supposed to be individualized over time, such as intravascular large B-cell lymphomas. Treatment may include surgical excision, radiotherapy, antibiotics, corticosteroids, interferon, monoclonal antibodies and chemotherapy, depending on the type of lymphoma and on the type and location of the skin lesions. In subtypes with good prognosis is contraindicated overtreatment and in those associated with a worse prognosis the recommended therapy relies on CHOP-like regimens associated with rituximab, assisted or not with local radiotherapy. We review the primary cutaneous B-cell lymphomas, remembering the diagnostic criteria, differential diagnosis, classification, and prognostic factors and presenting the available therapies. PMID:26560215

  12. Standardized cryopreservation of human primary cells.

    PubMed

    Ramos, Thomas V; Mathew, Aby J; Thompson, Maria L; Ehrhardt, Rolf O

    2014-09-02

    Cryopreservation is the use of low temperatures to preserve structurally intact living cells. The cells that survive the thermodynamic journey from the 37 °C incubator to the -196 °C liquid nitrogen storage tank are free from the influences of time. Thus, cryopreservation is a critical component of cell culture and cell manufacturing protocols. Successful cryopreservation of human cells requires that the cells be derived from patient samples that are collected in a standardized manner, and carefully handled from blood draw through cell isolation. Furthermore, proper equipment must be in place to ensure consistency, reproducibility, and sterility. In addition, the correct choice and amount of cryoprotectant agent must be added at the correct temperature, and a controlled rate of freezing (most commonly 1 °C/min) must be applied prior to a standardized method of cryogenic storage. This appendix describes how human primary cells can be frozen for long-term storage and thawed for growth in a tissue culture vessel.

  13. Primary cell culture method for the honeybee Apis mellifera.

    PubMed

    Ju, Hyunhee; Ghil, Sungho

    2015-10-01

    Honeybees are among the most important pollinators in nature, and honeybee-associated products are useful in various areas, including the food industry. However, honeybees may be infected by various types of pathogens. The study of honeybee-associated diseases would greatly benefit from a successful cell culture system, but although some honeybee cell culture techniques have been introduced, these methods have not yet been fully established. Here, we describe a primary cell culture method for the honeybee, Apis mellifera. We isolated, sterilized, and seeded egg cells into non-coated cell culture dishes to generate cell aggregates. After approximately 10 d, aggregates were dissociated and seeded to cell culture dishes. Cell passages were continuously performed, with sub-culturing every 3-4 d. The cells expressed non-adherent phenotypes. Their growth increased with the passage number when they were cultured in growth medium based on L-15 insect medium but not Schneider's insect medium. Finally, polymerase chain reaction confirmed that the cells originated from A. mellifera. Our results suggest that the culturing methods described herein are appropriate for isolating primary cells from honeybee eggs. These methods could thus facilitate the study of honeybee-associated pathogenesis, development, and toxicology.

  14. B cell suppression in primary glomerular disease.

    PubMed

    Rood, Ilse M; Hofstra, Julia M; Deegens, Jeroen K J; Wetzels, Jack F M

    2014-03-01

    Membranous nephropathy, focal segmental glomerulosclerosis (FSGS), and minimal change disease (MCD) are the most common causes of idiopathic nephrotic syndrome. For many years prednisone, alkylating agents, and calcineurin inhibitors have been the standard of therapy for these patients. More effective or better tolerated treatment modalities are needed. B cell targeted therapy was recently introduced in clinical practice. In this review, we briefly summarize the current standard therapy and discuss the efficacy of B cell targeted therapy in primary glomerular diseases. Observational, short-term studies suggest that rituximab is effective and comparable to standard therapy in maintaining remissions in patients with frequently relapsing or steroid-dependent MCD or FSGS. In contrast, response is limited in patients with steroid-resistant nephrotic syndrome. Rituximab also induces remissions in patients with membranous nephropathy. Controlled clinical trials on kidney endpoints are urgently needed to position B cell targeted therapy in clinical practice.

  15. Inflammatory Cell Distribution in Primary Merkel Cell Carcinoma

    PubMed Central

    Wheat, Rachel; Roberts, Claudia; Waterboer, Tim; Steele, Jane; Marsden, Jerry; Steven, Neil M.; Blackbourn, David J.

    2014-01-01

    Merkel cell carcinoma (MCC) is an aggressive poorly differentiated neuroendocrine cutaneous carcinoma associated with older age, immunodeficiency and Merkel cell polyomavirus (MCPyV) integrated within malignant cells. The presence of intra-tumoural CD8+ lymphocytes reportedly predicts better MCC-specific survival. In this study, the distribution of inflammatory cells and properties of CD8+ T lymphocytes within 20 primary MCC specimens were characterised using immunohistochemistry and multicolour immunofluorescent staining coupled to confocal microscopy. CD8+ cells and CD68+ macrophages were identified in 19/20 primary MCC. CD20+ B cells were present in 5/10, CD4+ cells in 10/10 and FoxP3+ cells in 7/10 specimens. Only two specimens had almost no inflammatory cells. Within specimens, inflammatory cells followed the same patchy distribution, focused at the edge of sheets and nodules and, in some cases, more intense in trabecular areas. CD8+ cells were outside vessels on the edge of tumour. Those few within malignant sheets typically lined up in fine septa not contacting MCC cells expressing MCPyV large T antigen. The homeostatic chemokine CXCL12 was expressed outside malignant nodules whereas its receptor CXCR4 was identified within tumour but not on CD8+ cells. CD8+ cells lacked CXCR3 and granzyme B expression irrespective of location within stroma versus malignant nodules or of the intensity of the intra-tumoural infiltrate. In summary, diverse inflammatory cells were organised around the margin of malignant deposits suggesting response to aberrant signaling, but were unable to penetrate the tumour microenvironment itself to enable an immune response against malignant cells or their polyomavirus. PMID:24961933

  16. Flexible organic solar cells including efficiency enhancing grating structures.

    PubMed

    de Oliveira Hansen, Roana Melina; Liu, Yinghui; Madsen, Morten; Rubahn, Horst-Günter

    2013-04-12

    In this work, a new method for the fabrication of organic solar cells containing functional light-trapping nanostructures on flexible substrates is presented. Polyimide is spin-coated on silicon support substrates, enabling standard micro- and nanotechnology fabrication techniques, such as photolithography and electron-beam lithography, besides the steps required for the bulk-heterojunction organic solar cell fabrication. After the production steps, the solar cells on polyimide are peeled off the silicon support substrates, resulting in flexible devices containing nanostructures for light absorption enhancement. Since the solar cells avoid using brittle electrodes, the performance of the flexible devices is not affected by the peeling process. We have investigated three different nanostructured grating designs and conclude that gratings with a 500 nm pitch distance have the highest light-trapping efficiency for the selected active layer material (P3HT:PCBM), resulting in an enhancement of about 34% on the solar cell efficiency. The presented method can be applied to a large variety of flexible nanostructured devices in future applications.

  17. Flexible organic solar cells including efficiency enhancing grating structures

    NASA Astrophysics Data System (ADS)

    Melina de Oliveira Hansen, Roana; Liu, Yinghui; Madsen, Morten; Rubahn, Horst-Günter

    2013-04-01

    In this work, a new method for the fabrication of organic solar cells containing functional light-trapping nanostructures on flexible substrates is presented. Polyimide is spin-coated on silicon support substrates, enabling standard micro- and nanotechnology fabrication techniques, such as photolithography and electron-beam lithography, besides the steps required for the bulk-heterojunction organic solar cell fabrication. After the production steps, the solar cells on polyimide are peeled off the silicon support substrates, resulting in flexible devices containing nanostructures for light absorption enhancement. Since the solar cells avoid using brittle electrodes, the performance of the flexible devices is not affected by the peeling process. We have investigated three different nanostructured grating designs and conclude that gratings with a 500 nm pitch distance have the highest light-trapping efficiency for the selected active layer material (P3HT:PCBM), resulting in an enhancement of about 34% on the solar cell efficiency. The presented method can be applied to a large variety of flexible nanostructured devices in future applications.

  18. Primary Mediastinal B-Cell Lymphoma

    PubMed Central

    Pileri, Stefano A.; Gaidano, Gianluca; Zinzani, Pier Luigi; Falini, Brunangelo; Gaulard, Philippe; Zucca, Emanuele; Pieri, Federica; Berra, Eva; Sabattini, Elena; Ascani, Stefano; Piccioli, Milena; Johnson, Peter W. M.; Giardini, Roberto; Pescarmona, Edoardo; Novero, Domenico; Piccaluga, Pier Paolo; Marafioti, Teresa; Alonso, Miguel A.; Cavalli, Franco

    2003-01-01

    Although primary mediastinal (thymic) large B-cell lymphoma has been primarily studied, its precise phenotype, molecular characteristics, and histogenesis are still a matter of debate. The International Extranodal Lymphoma Study Group collected 137 such cases for extensive pathological review. Histologically, the lymphomatous growth was predominantly diffuse with fibrosis that induced compartmentalized cell aggregation. It consisted of large cells with varying degrees of nuclear polymorphism and clear to basophilic cytoplasm. On immunohistochemistry, the following phenotype was observed: CD45+, CD20+, CD79a+, PAX5/BSAP+, BOB.1+, Oct-2+, PU.1+, Bcl-2+, CD30+, HLA-DR+, MAL protein+/−, Bcl-6+/−, MUM1/IRF4+/−, CD10−/+, CD21−, CD15−, CD138−, CD68−, and CD3−. Immunoglobulins were negative both at immunohistochemistry and in situ hybridization. Molecular analysis, performed in 45 cases, showed novel findings. More than half of the cases displayed BCL-6 gene mutations, which usually occurred along with functioning somatic IgVH gene mutations and Bcl-6 and/or MUM1/IRF4 expression. The present study supports the concept that a sizable fraction of cases of this lymphoma are from activated germinal center or postgerminal center cells. However, it differs from other aggressive B-cell lymphomas in that it shows defective immunoglobulin production despite the expression of OCT-2, BOB.1, and PU.1 transcription factors and the lack of IgVH gene crippling mutations. PMID:12507907

  19. Immortalization of primary human smooth muscle cells.

    PubMed Central

    Perez-Reyes, N; Halbert, C L; Smith, P P; Benditt, E P; McDougall, J K

    1992-01-01

    Primary human aortic and myometrial smooth muscle cells (SMCs) were immortalized using an amphotropic recombinant retroviral construct containing the E6 and E7 open reading frames (ORFs) of human papillomavirus type 16. The SMCs expressing the E6/E7 ORFs have considerably elevated growth rates when compared with nonimmortalized control cells and show no signs of senescence with long-term passage. The first SMC line derived in this study has been maintained in continuous tissue culture for greater than 1 year (greater than 180 population doublings). The immortalized SMCs have decreased cell size and decreased content of muscle-specific alpha-actin filaments as determined by indirect immunofluorescence. Southern blot analysis has demonstrated the stable integration of the E6/E7 ORFs in the retrovirally infected cells, and radioimmunoprecipitation has confirmed the continued expression of the E6 and E7 genes. Cytogenetic studies of the SMC lines have revealed essentially diploid populations except for the myometrial clonal line, which became aneuploid at late passage (greater than 125 doublings). These cell lines were not tumorigenic in nude mice. Images PMID:1311088

  20. Effective activity of cytokine-induced killer cells against autologous metastatic melanoma including cells with stemness features.

    PubMed

    Gammaitoni, Loretta; Giraudo, Lidia; Leuci, Valeria; Todorovic, Maja; Mesiano, Giulia; Picciotto, Franco; Pisacane, Alberto; Zaccagna, Alessandro; Volpe, Maria Giuseppa; Gallo, Susanna; Caravelli, Daniela; Giacone, Elena; Venesio, Tiziana; Balsamo, Antonella; Pignochino, Ymera; Grignani, Giovanni; Carnevale-Schianca, Fabrizio; Aglietta, Massimo; Sangiolo, Dario

    2013-08-15

    We investigate the unknown tumor-killing activity of cytokine-induced killer (CIK) cells against autologous metastatic melanoma and the elusive subset of putative cancer stem cells (mCSC). We developed a preclinical autologous model using same patient-generated CIK cells and tumor targets to consider the unique biology of each patient/tumor pairing. In primary tumor cell cultures, we visualized and immunophenotypically defined a putative mCSC subset using a novel gene transfer strategy that exploited their exclusive ability to activate the promoter of stemness gene Oct4. The CIK cells from 10 patients with metastatic melanoma were successfully expanded (median, 23-fold; range, 11-117). Primary tumor cell cultures established and characterized from the same patients were used as autologous targets. Patient-derived CIK cells efficiently killed autologous metastatic melanoma [up to 71% specific killing (n = 26)]. CIK cells were active in vivo against autologous melanoma, resulting in delayed tumor growth, increased necrotic areas, and lymphocyte infiltration at tumor sites. The metastatic melanoma cultures presented an average of 11.5% ± 2.5% putative mCSCs, which was assessed by Oct4 promoter activity and stemness marker expression (Oct4, ABCG2, ALDH, MITF). Expression was confirmed on mCSC target molecules recognized by CIK cells (MIC A/B; ULBPs). CIK tumor killing activity against mCSCs was intense (up to 71%, n = 4) and comparable with results reported against differentiated metastatic melanoma cells (P = 0.8). For the first time, the intense killing activity of CIK cells against autologous metastatic melanoma, including mCSCs, has been shown. These findings move clinical investigation of a new immunotherapy for metastatic melanoma, including mCSCs, closer. ©2013 AACR.

  1. Thymoquinone causes multiple effects, including cell death, on dividing plant cells.

    PubMed

    Hassanien, Sameh E; Ramadan, Ahmed M; Azeiz, Ahmed Z Abdel; Mohammed, Rasha A; Hassan, Sabah M; Shokry, Ahmed M; Atef, Ahmed; Kamal, Khalid B H; Rabah, Samar; Sabir, Jamal S M; Abuzinadah, Osama A; El-Domyati, Fotouh M; Martin, Gregory B; Bahieldin, Ahmed

    2013-01-01

    Thymoquinone (TQ) is a major constituent of Nigella sativa oil with reported anti-oxidative activity and anti-inflammatory activity in animal cells. It also inhibits proliferation and induces programmed cell death (apoptosis) in human skin cancer cells. The present study sought to detect the influence of TQ on dividing cells of three plant systems and on expression of Bcl2-associated athanogene-like (BAG-like) genes that might be involved during the process of cell death. BAG genes are known for the regulation of diverse physiological processes in animals, including apoptosis, tumorigenesis, stress responses, and cell division. Synthetic TQ at 0.1mg/mL greatly reduced wheat seed germination rate, whereas 0.2mg/mL completely inhibited germination. An Evans blue assay revealed moderate cell death in the meristematic zone of Glycine max roots after 1h of TQ treatment (0.2mg/mL), with severe cell death occurring in this zone after 2h of treatment. Light microscopy of TQ-treated (0.2mg/mL) onion hairy root tips for 1h revealed anti-mitotic activity and also cell death-associated changes, including nuclear membrane disruption and nuclear fragmentation. Transmission electron microscopy of TQ-treated cells (0.2mg/mL) for 1h revealed shrinkage of the plasma membrane, leakage of cell lysate, degradation of cell walls, enlargement of vacuoles and condensation of nuclei. Expression of one BAG-like gene, previously associated with cell death, was induced 20 min after TQ treatment in Glycine max root tip cells. Thus, TQ has multiple effects, including cell death, on dividing plant cells and plants may serve as a useful system to further investigate the mechanisms underlying the response of eukaryotic cells to TQ.

  2. Inferring time derivatives including cell growth rates using Gaussian processes

    PubMed Central

    Swain, Peter S.; Stevenson, Keiran; Leary, Allen; Montano-Gutierrez, Luis F.; Clark, Ivan B.N.; Vogel, Jackie; Pilizota, Teuta

    2016-01-01

    Often the time derivative of a measured variable is of as much interest as the variable itself. For a growing population of biological cells, for example, the population's growth rate is typically more important than its size. Here we introduce a non-parametric method to infer first and second time derivatives as a function of time from time-series data. Our approach is based on Gaussian processes and applies to a wide range of data. In tests, the method is at least as accurate as others, but has several advantages: it estimates errors both in the inference and in any summary statistics, such as lag times, and allows interpolation with the corresponding error estimation. As illustrations, we infer growth rates of microbial cells, the rate of assembly of an amyloid fibril and both the speed and acceleration of two separating spindle pole bodies. Our algorithm should thus be broadly applicable. PMID:27941811

  3. Inferring time derivatives including cell growth rates using Gaussian processes

    NASA Astrophysics Data System (ADS)

    Swain, Peter S.; Stevenson, Keiran; Leary, Allen; Montano-Gutierrez, Luis F.; Clark, Ivan B. N.; Vogel, Jackie; Pilizota, Teuta

    2016-12-01

    Often the time derivative of a measured variable is of as much interest as the variable itself. For a growing population of biological cells, for example, the population's growth rate is typically more important than its size. Here we introduce a non-parametric method to infer first and second time derivatives as a function of time from time-series data. Our approach is based on Gaussian processes and applies to a wide range of data. In tests, the method is at least as accurate as others, but has several advantages: it estimates errors both in the inference and in any summary statistics, such as lag times, and allows interpolation with the corresponding error estimation. As illustrations, we infer growth rates of microbial cells, the rate of assembly of an amyloid fibril and both the speed and acceleration of two separating spindle pole bodies. Our algorithm should thus be broadly applicable.

  4. High-throughput RNAi screening in vitro: From cell lines to primary cells

    PubMed Central

    OVCHARENKO, DMITRIY; JARVIS, RICHARD; HUNICKE-SMITH, SCOTT; KELNAR, KEVIN; BROWN, DAVID

    2005-01-01

    Small interfering RNAs (siRNAs) are being used to induce sequence-specific gene silencing in cultured cells to study mammalian gene function. Libraries of siRNAs targeting entire human gene classes can be used to identify genes with specific cellular functions. Here we describe high-throughput siRNA delivery methods to facilitate siRNA library screening experiments with both immortalized and primary cells. We adapted chemical reverse transfection for immortalized adherent cell lines in a 96-well format. The method is fast, robust, and exceptionally effective for many cell types. For primary cells and immortalized cells that are recalcitrant to lipofection-based methods, we developed electropermeabilization (electroporation) conditions that facilitate siRNA delivery to a broad range of cell types, including primary human T-cells, hMSC, NHA, NDHF-Neo, HUVEC, DI TNC1, RPTEC, PC12, and K562 cells. To enable high-throughput electropermeabilization of primary cells, we developed a novel 96-well electroporation device that provides highly efficient and reproducible delivery of siRNAs. The combination of high-throughput chemical reverse transfection and electroporation makes it possible to deliver libraries of siRNAs to virtually any cell type, enabling gene function analysis and discovery on a genome scale. PMID:15923380

  5. High-throughput RNAi screening in vitro: from cell lines to primary cells.

    PubMed

    Ovcharenko, Dmitriy; Jarvis, Richard; Hunicke-Smith, Scott; Kelnar, Kevin; Brown, David

    2005-06-01

    Small interfering RNAs (siRNAs) are being used to induce sequence-specific gene silencing in cultured cells to study mammalian gene function. Libraries of siRNAs targeting entire human gene classes can be used to identify genes with specific cellular functions. Here we describe high-throughput siRNA delivery methods to facilitate siRNA library screening experiments with both immortalized and primary cells. We adapted chemical reverse transfection for immortalized adherent cell lines in a 96-well format. The method is fast, robust, and exceptionally effective for many cell types. For primary cells and immortalized cells that are recalcitrant to lipofection-based methods, we developed electropermeabilization (electroporation) conditions that facilitate siRNA delivery to a broad range of cell types, including primary human T-cells, hMSC, NHA, NDHF-Neo, HUVEC, DI TNC1, RPTEC, PC12, and K562 cells. To enable high-throughput electropermeabilization of primary cells, we developed a novel 96-well electroporation device that provides highly efficient and reproducible delivery of siRNAs. The combination of high-throughput chemical reverse transfection and electroporation makes it possible to deliver libraries of siRNAs to virtually any cell type, enabling gene function analysis and discovery on a genome scale.

  6. Establishing and maintaining primary cell cultures derived from the ctenophore Mnemiopsis leidyi.

    PubMed

    Vandepas, Lauren E; Warren, Kaitlyn J; Amemiya, Chris T; Browne, William E

    2017-04-01

    We have developed an efficient method for the preparation and maintenance of primary cell cultures isolated from adult Mnemiopsis leidyi, a lobate ctenophore. Our primary cell cultures are derived from tissue explants or enzymatically dissociated cells, and maintained in a complex undefined ctenophore mesogleal serum. These methods can be used to isolate, maintain and visually monitor ctenophore cells to assess proliferation, cellular morphology and cell differentiation in future studies. Exemplar cell types that can be easily isolated from primary cultures include proliferative ectodermal and endodermal cells, motile amebocyte-like cells, and giant smooth muscle cells that exhibit inducible contractile properties. We have also derived 'tissue envelopes' containing sections of endodermal canal surrounded by mesoglea and ectoderm that can be used to monitor targeted cell types in an in vivo context. Access to efficient and reliably generated primary cell cultures will facilitate the analysis of ctenophore development, physiology and morphology from a cell biological perspective.

  7. Distinct speed dependence of entorhinal island and ocean cells, including respective grid cells

    PubMed Central

    Sun, Chen; Kitamura, Takashi; Yamamoto, Jun; Martin, Jared; Pignatelli, Michele; Kitch, Lacey J.; Schnitzer, Mark J.; Tonegawa, Susumu

    2015-01-01

    Entorhinal–hippocampal circuits in the mammalian brain are crucial for an animal’s spatial and episodic experience, but the neural basis for different spatial computations remain unknown. Medial entorhinal cortex layer II contains pyramidal island and stellate ocean cells. Here, we performed cell type-specific Ca2+ imaging in freely exploring mice using cellular markers and a miniature head-mounted fluorescence microscope. We found that both oceans and islands contain grid cells in similar proportions, but island cell activity, including activity in a proportion of grid cells, is significantly more speed modulated than ocean cell activity. We speculate that this differential property reflects island cells’ and ocean cells’ contribution to different downstream functions: island cells may contribute more to spatial path integration, whereas ocean cells may facilitate contextual representation in downstream circuits. PMID:26170279

  8. Primary prevention of latex allergy in healthcare-spectrum of strategies including the European glove standardization.

    PubMed

    Wrangsjö, Karin; Boman, Anders; Lidén, Carola; Meding, Birgitta

    2012-04-01

    IgE-mediated allergy to natural rubber latex was first noted from rubber gloves in 1979. The initial reports in dermatological journals described contact urticarial reactions; later, severe generalized allergic reactions and asthma were documented. A considerable proportion of the people involved in medical and dental care were found to be sensitized to latex. This article describes and surveys a broad range of primary prevention measures at the local and national levels. Examples are given from paediatrics, dental education, and medical care. National strategies and position papers on latex allergy are presented in which medical professionals, manufacturers and public authorities have cooperated. Special reference is paid to the European work to standardize medical gloves, which led to document EN 455:3.

  9. Gene therapy of primary T cell immunodeficiencies.

    PubMed

    Fischer, Alain; Hacein-Bey-Abina, Salima; Cavazzana-Calvo, Marina

    2013-08-10

    Gene therapy of severe combined immunodeficiencies has been proven to be effective to provide sustained correction of the T cell immunodeficiencies. This has been achieved for 2 forms of SCID, i.e SCID-X1 (γc deficiency) and adenosine deaminase deficiency. Occurrence of gene toxicity generated by integration of first generation retroviral vectors, as observed in the SCID-X1 trials has led to replace these vectors by self inactivated (SIN) retro(or lenti) viruses that may provide equivalent efficacy with a better safety profile. Results of ongoing clinical studies in SCID as well as in other primary immunodeficiencies, such as the Wiskott Aldrich syndrome, will be thus very informative.

  10. [A methodological proposal to include nutrition education in primary schools. Experience in Chile].

    PubMed

    Olivares, Sonia; Morón, Cecilio; Kain, Juliana; Zacarías, Isabel; Andrade, Margarita; Lera, Lydia; Díaz, Nora; Vio, Fernando

    2004-06-01

    This article presents the methodology to incorporate nutrition education in Chilean primary schools. In 2001, nutritional status, food and nutrition knowledge, attitudes and practices of 1701 school children from ten urban and rural public schools (3rd to 7th grade) were evaluated. This information was necessary to design the nutrition education strategy. The prevalence of obesity was 15.4% and overweight 19.6%. Daily intake of dairy products, fruits and vegetables was low, while the consumption of energy dense snacks was very high. Because the Ministry of Education does not allow the incorporation of new programs to the curriculum, the educational strategy was based on the development of a text book, a teacher's guide, five practical guides for students from third to eight grade and a CD-Rom. These materials were validated by 36 teachers through an educational intervention during 5 months in six schools (intervention groups). The teachers reported that the educational materials were useful, motivational and easy to understand for both, teachers and students. Preliminary results showed a significant increase in food and nutrition knowledge, in the consumption of dairy products and a decrease in the consumption of bread among the intervention groups. Intake of snacks increased in both groups, but it was significantly higher in the control group. These results indicate that nutrition education will only produce significant changes in food habits if health and educational authorities establish regulations for food advertisement oriented to children and also to food items sold in the schools.

  11. Second primary malignancies after radiotherapy including HDR (252)Cf brachytherapy for cervical cancer.

    PubMed

    Samerdokiene, Vitalija; Valuckas, Konstantinas Povilas; Janulionis, Ernestas; Atkocius, Vydmantas; Rivard, Mark J

    2015-01-01

    Second primary malignancies (SPMs) are among the most serious late adverse effects after radiotherapy experienced over time by the increasing population of cancer survivors worldwide. The study aim was to determine the rate and distribution of SPMs for neutron- and photon-emitting brachytherapy (BT) sources for patients treated for cervical cancer. The cohort comprised 662 patients with invasive cervical cancer (Stages IIB and IIIB) and contributed 5,224 patient-years (PY) of observation. These patients were treated by radiotherapy during the 1989-1999 year period with cobalt-60 source ((60)Co) teletherapy. The first group of patients (N = 375; 3,154 PY) received high-dose-rate (HDR) californium-252 source ((252)Cf) BT, whereas the second group (N = 287; 2,070 PY) received HDR (60)Co BT. Over a 25-year period, 35 SPMs were observed, amounting to 5.3% of all observed patients: in 16 (2.4%) heavily, 2 (0.3%) moderately, 14 (2.1%) lightly irradiated body sites, and 3 (0.5%) other sites. Of these, 21 cases (5.6%) were observed in the HDR (252)Cf BT group, whereas 14 cases (4.9%) were observed in the HDR (60)Co BT group. Exposures received during (60)Co teletherapy and HDR BT with either (252)Cf or (60)Co had statistically equivalent (p = 0.68) effects on SPM development. Cure rates are improving, and therefore, there are more long-term survivors from cervical cancer. This study shows no significant difference in rates or distribution of SPMs in women treated with neutron BT compared with photon BT (p = 0.68). After reviewing related literature and our research results, it is evident that a detailed investigation of SPM frequency, localization, and dose to adjacent organs is a suitable topic for further research. Copyright © 2015 American Brachytherapy Society. Published by Elsevier Inc. All rights reserved.

  12. Primary oat cell carcinoma of the larynx

    SciTech Connect

    Aguilar, E.A. III; Robbins, K.T.; Stephens, J.; Dimery, I.W.; Batsakis, J.G.

    1987-02-01

    The aggressiveness of small (oat) cell carcinoma of the larynx presents a therapeutic challenge to the oncologist. Since the first description of this type of carcinoma in 1972, 52 patients have been reported in the literature and a variety of treatment regimens have been used. The purpose of this study was to report two new cases and review all previous reports to determine the disease's biological behavior, clinical manifestations, and optimum treatment. Thirty-five percent of the tumors were transglottic, and 27% were supraglottic. Fifty-four percent of patients had regional metastases at initial presentation and 17.6% had distant metastases. The median survival was 10 months for all patients. Patients who were treated with chemotherapy with or without other modalities had the best 2-year survival rates (52.2%). Forty-one percent of patients had regional recurrence only, 12.5% had regional recurrence and distant metastases, and 2% developed distant metastases only. We conclude that patients with oat cell carcinoma of the larynx should be treated with combination chemotherapy and radiation therapy. Surgery is best reserved for persistent and recurrent disease at the primary site and neck.

  13. Assessment of cell viability in primary neuronal cultures.

    PubMed

    Aras, Mandar A; Hartnett, Karen A; Aizenman, Elias

    2008-07-01

    This unit contains five protocols for assaying cell viability in vitro using primary neuronal cultures, including a novel method for use with transfected neurons. Three of the assays are based on the principle that cell death cascades alter membrane permeability. The lactate dehydrogenase (LDH) release assay measures the amount of the cytoplasmic enzyme released into the bathing medium, while the trypan blue and propidium iodide assays measure the ability of cells to exclude dye from their cytoplasm. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay measures the mitochondrial activity of viable cells by quantifying the conversion of the tetrazolium salt to its formazan product. Finally, the fifth assay details the measurement of luciferase expression as an indication of neuronal viability within a relatively small population of transfected neurons.

  14. Emergency response facilities including primary and secondary prevention strategies across 79 professional football clubs in England.

    PubMed

    Malhotra, Aneil; Dhutia, Harshil; Gati, Sabiha; Yeo, Tee-Joo; Finnochiaro, Gherardo; Keteepe-Arachi, Tracey; Richards, Thomas; Walker, Mike; Birt, Robin; Stuckey, David; Robinson, Laurence; Tome, Maite; Beasley, Ian; Papadakis, Michael; Sharma, Sanjay

    2017-06-14

    To assess the emergency response planning and prevention strategies for sudden cardiac arrest (SCA) across a wide range of professional football clubs in England. A written survey was sent to all professional clubs in the English football league, namely the Premiership, Championship, League 1 and League 2. Outcomes included: (1) number of clubs performing cardiac screening and frequency of screening; (2) emergency planning and documentation; (3) automated external defibrillator (AED) training and availability; and (4) provision of emergency services at sporting venues. 79 clubs (86%) responded to the survey. 100% clubs participated in cardiac screening. All clubs had AEDs available on match days and during training sessions. 100% Premiership clubs provided AED training to designated staff. In contrast, 30% of lower division clubs with AEDs available did not provide formal training. Most clubs (n=66; 83%) reported the existence of an emergency action plan for SCA but formal documentation was variable. All clubs in the Premiership and League 1 provided an ambulance equipped for medical emergencies on match days compared with 75% of clubs in the Championship and 66% in League 2. The majority of football clubs in England have satisfactory prevention strategies and emergency response planning in line with European recommendations. Additional improvements such as increasing awareness of European guidelines for emergency planning, AED training and mentorship with financial support to lower division clubs are necessary to further enhance cardiovascular safety of athletes and spectators and close the gap between the highest and lower divisions. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  15. Highly efficient baculovirus-mediated multigene delivery in primary cells

    PubMed Central

    Mansouri, Maysam; Bellon-Echeverria, Itxaso; Rizk, Aurélien; Ehsaei, Zahra; Cianciolo Cosentino, Chiara; Silva, Catarina S.; Xie, Ye; Boyce, Frederick M.; Davis, M. Wayne; Neuhauss, Stephan C. F.; Taylor, Verdon; Ballmer-Hofer, Kurt; Berger, Imre; Berger, Philipp

    2016-01-01

    Multigene delivery and subsequent cellular expression is emerging as a key technology required in diverse research fields including, synthetic and structural biology, cellular reprogramming and functional pharmaceutical screening. Current viral delivery systems such as retro- and adenoviruses suffer from limited DNA cargo capacity, thus impeding unrestricted multigene expression. We developed MultiPrime, a modular, non-cytotoxic, non-integrating, baculovirus-based vector system expediting highly efficient transient multigene expression from a variety of promoters. MultiPrime viruses efficiently transduce a wide range of cell types, including non-dividing primary neurons and induced-pluripotent stem cells (iPS). We show that MultiPrime can be used for reprogramming, and for genome editing and engineering by CRISPR/Cas9. Moreover, we implemented dual-host-specific cassettes enabling multiprotein expression in insect and mammalian cells using a single reagent. Our experiments establish MultiPrime as a powerful and highly efficient tool, to deliver multiple genes for a wide range of applications in primary and established mammalian cells. PMID:27143231

  16. Measuring energy metabolism in cultured cells, including human pluripotent stem cells and differentiated cells

    PubMed Central

    Zhang, Jin; Nuebel, Esther; Wisidagama, Dona R R; Setoguchi, Kiyoko; Hong, Jason S; Van Horn, Christine M; Imam, Sarah S; Vergnes, Laurent; Malone, Cindy S; Koehler, Carla M; Teitell, Michael A

    2013-01-01

    Measurements of glycolysis and mitochondrial function are required to quantify energy metabolism in a wide variety of cellular contexts. In human pluripotent stem cells (hPSCs) and their differentiated progeny, this analysis can be challenging because of the unique cell properties, growth conditions and expense required to maintain these cell types. Here we provide protocols for analyzing energy metabolism in hPSCs and their early differentiated progenies that are generally applicable to mature cell types as well. Our approach has revealed distinct energy metabolism profiles used by hPSCs, differentiated cells, a variety of cancer cells and Rho-null cells. The protocols measure or estimate glycolysis on the basis of the extracellular acidification rate, and they measure or estimate oxidative phosphorylation on the basis of the oxygen consumption rate. Assays typically require 3 h after overnight sample preparation. Companion methods are also discussed and provided to aid researchers in developing more sophisticated experimental regimens for extended analyses of cellular bioenergetics. PMID:22576106

  17. Sealed Primary Lithium-Inorganic Electrolyte Cell.

    DTIC Science & Technology

    Primary batteries , Reliability(Electronics), Lithium compounds, Aluminum compounds, Chlorides , Thionyl chloride , Battery components, Storage, Life tests, Explosions, Hazards, Temperature, Ventilation

  18. Interaction between arsenic trioxide and human primary cells: emphasis on human cells of myeloid origin.

    PubMed

    Binet, François; Antoine, Francis; Girard, Denis

    2009-03-01

    Arsenic trioxide (As(2)O(3); ATO) is considered to be one of the most potent drugs in cancer chemotherapy and is highly effective in the treatment of acute promyelocytic leukemia (APL). It is well established that treatment of APL patients with ATO is associated with the disappearance of the PML-RARalpha fusion transcript, the characteristic APL gene product of the chromosomal translocation t(15;17). Although its mode of action is still not fully understood, ATO is known to induce cell apoptosis via generation of reactive oxygen species and activation of caspases. Several reports have indicated that ATO acts principally by inducing cell apoptosis not only in APL, but in a variety of non-APL cells including myeloma cells, chronic myeloid leukemia cells and cells of immune origin, including B or T lymphocytes, macrophages and, more recently, neutrophils. There is an increasing amount of data, including some from our laboratory, concerning the interaction between ATO and human primary cells. The focus of this review will be to cover the role of ATO in human immune primary cells with special emphasis on cells of myeloid origin.

  19. Low temperature cell pausing: an alternative short-term preservation method for use in cell therapies including stem cell applications.

    PubMed

    Robinson, Nathalie J; Picken, Andrew; Coopman, Karen

    2014-02-01

    Encouraging advances in cell therapies have produced a requirement for an effective short-term cell preservation method, enabling time for quality assurance testing and transport to their clinical destination. Low temperature pausing of cells offers many advantages over cryopreservation, including the ability to store cells at scale, reduced cost and a simplified procedure with increased reliability. This review will focus on the importance of developing a short-term cell preservation platform as well highlighting the major successes of cell pausing and the key challenges which need addressing, to enable application of the process to therapeutically relevant cells.

  20. Collective cell migration of primary zebrafish keratocytes.

    PubMed

    Rapanan, Jose L; Cooper, Kimbal E; Leyva, Kathryn J; Hull, Elizabeth E

    2014-08-01

    Fish keratocytes are an established model in single cell motility but little is known about their collective migration. Initially, sheets migrate from the scale at ~145 μm/h but over the course of 24h the rate of leading edge advance decreases to ~23 μm/h. During this period, leader cells retain their ability to migrate rapidly when released from the sheet and follower cell area increases. After the addition of RGD peptide, leader cell lamellae are lost, altering migratory forces within the sheet, resulting in rapid retraction. Leader and follower cell states interconvert within minutes with changes in cell-cell adhesions. Leader cells migrate as single cells when they detach from the leading edge and single cells appear to become leader cells if they rejoin the sheet. Follower cells rapidly establish leader cell morphology during closing of holes formed during sheet expansion and revert to follower cell morphology after hole-closure. Inhibition of Rho associated kinase releases leader cells and halts advancement of the leading edge suggesting an important role for the intercellular actomyosin cable at the leading edge. In addition, the presence of the stationary scale orients direction of sheet migration which is characterized by a more uniform advance of the leading edge than in some cell line systems. These data establish fish keratocyte explant cultures as a collective cell migration system and suggest that cell-cell interactions determine the role of keratocytes within the migrating sheet. Copyright © 2014 Elsevier Inc. All rights reserved.

  1. T-cell primary leptomeningeal lymphoma in cerebellopontine angle

    PubMed Central

    Briongos-Figuero, Laisa Socorro; Gómez-Traveso, Tamara; Pérez-Castrillon, José Luis

    2015-01-01

    Primary meningeal lymphomas are very rare and those derived from T cells are even more infrequent (less than 5% of primary central nervous system lymphomas). Cerebellopontine angle involvement in the primary T-cell lymphoma is exceptional. Clinical presentation depends on the type of lesions, and histological diagnosis is needed. We present a rare case of a 50-year-old woman who presented with clinical cerebellar syndrome with posterior opsoclonus-myoclonus syndrome. Necropsy evaluation revealed primary diffuse leptomeningeal non-Hodgkin's T-cell lymphoma. PMID:25750225

  2. Pleomorphism and drug resistant cancer stem cells are characteristic of aggressive primary meningioma cell lines.

    PubMed

    Khan, Ishaq; Baeesa, Saleh; Bangash, Mohammed; Schulten, Hans-Juergen; Alghamdi, Fahad; Qashqari, Hanadi; Madkhali, Nawal; Carracedo, Angel; Saka, Mohamad; Jamal, Awatif; Al-Maghrabi, Jaudah; AlQahtani, Mohammed; Al-Karim, Saleh; Damanhouri, Ghazi; Saini, Kulvinder; Chaudhary, Adeel; Abuzenadah, Adel; Hussein, Deema

    2017-01-01

    Meningioma tumors arise in arachnoid membranes, and are the most reported central nervous system (CNS) tumors worldwide. Up to 20% of grade I meningioma tumors reoccur and currently predictive cancer stem cells (CSCs) markers for aggressive and drug resistant meningiomas are scarce. Meningioma tissues and primary cell lines were investigated using whole transcriptome microarray analysis, immunofluorescence staining of CSCs markers (including CD133, Sox2, Nestin, and Frizzled 9), and drug treatment with cisplatin or etoposide. Unsupervised hierarchical clustering of six meningioma samples separated tissues into two groups. Analysis identified stem cells related pathways to be differential between the two groups and indicated the de-regulation of the stem cell associated genes Reelin (RELN), Calbindin 1 (CALB1) and Anterior Gradient 2 Homolog (AGR2). Immunofluorescence staining for four tissues confirmed stemness variation in situ. Biological characterization of fifteen meningioma primary cell lines concordantly separated cells into two functionally distinct sub-groups. Pleomorphic cell lines (NG type) grew significantly faster than monomorphic cell lines (G type), had a higher number of cells that express Ki67, and were able to migrate aggressively in vitro. In addition, NG type cell lines had a lower expression of nuclear Caspase-3, and had a significantly higher number of CSCs co-positive for CD133+ Sox2+ or AGR2+ BMI1+. Importantly, these cells were more tolerant to cisplatin and etoposide treatment, showed a lower level of nuclear Caspase-3 in treated cells and harbored drug resistant CSCs. Collectively, analyses of tissues and primary cell lines revealed stem cell associated genes as potential targets for aggressive and drug resistant meningiomas.

  3. Efficient Gene Editing in Primary Human T Cells.

    PubMed

    Chen, Yvonne Y

    2015-11-01

    Recent advances in T-cell therapy for cancer, viral infections, and autoimmune diseases highlight the broad therapeutic potential of T-cell engineering. However, site-specific genetic manipulation in primary human T cells remains challenging. Two recent studies describe efficient genome editing in T cells using CRISPR and TALEN approaches.

  4. Primary Tumor and MEF Cell Isolation to Study Lung Metastasis.

    PubMed

    Dong, Shengli; Maziveyi, Mazvita; Alahari, Suresh K

    2015-05-20

    In breast tumorigenesis, the metastatic stage of the disease poses the greatest threat to the affected individual. Normal breast cells with altered genotypes now possess the ability to invade and survive in other tissues. In this protocol, mouse mammary tumors are removed and primary cells are prepared from tumors. The cells isolated from this procedure are then available for gene profiling experiments. For successful metastasis, these cells must be able to intravasate, survive in circulation, extravasate to distant organs, and survive in that new organ system. The lungs are the typical target of breast cancer metastasis. A set of genes have been discovered that mediates the selectivity of metastasis to the lung. Here we describe a method of studying lung metastasis from a genetically engineered mouse model.. Furthermore, another protocol for analyzing mouse embryonic fibroblasts (MEFs) from the mouse embryo is included. MEF cells from the same animal type provide a clue of non-cancer cell gene expression. Together, these techniques are useful in studying mouse mammary tumorigenesis, its associated signaling mechanisms and pathways of the abnormalities in embryos.

  5. Enhanced peripheral visual processing in congenitally deaf humans is supported by multiple brain regions, including primary auditory cortex.

    PubMed

    Scott, Gregory D; Karns, Christina M; Dow, Mark W; Stevens, Courtney; Neville, Helen J

    2014-01-01

    Brain reorganization associated with altered sensory experience clarifies the critical role of neuroplasticity in development. An example is enhanced peripheral visual processing associated with congenital deafness, but the neural systems supporting this have not been fully characterized. A gap in our understanding of deafness-enhanced peripheral vision is the contribution of primary auditory cortex. Previous studies of auditory cortex that use anatomical normalization across participants were limited by inter-subject variability of Heschl's gyrus. In addition to reorganized auditory cortex (cross-modal plasticity), a second gap in our understanding is the contribution of altered modality-specific cortices (visual intramodal plasticity in this case), as well as supramodal and multisensory cortices, especially when target detection is required across contrasts. Here we address these gaps by comparing fMRI signal change for peripheral vs. perifoveal visual stimulation (11-15° vs. 2-7°) in congenitally deaf and hearing participants in a blocked experimental design with two analytical approaches: a Heschl's gyrus region of interest analysis and a whole brain analysis. Our results using individually-defined primary auditory cortex (Heschl's gyrus) indicate that fMRI signal change for more peripheral stimuli was greater than perifoveal in deaf but not in hearing participants. Whole-brain analyses revealed differences between deaf and hearing participants for peripheral vs. perifoveal visual processing in extrastriate visual cortex including primary auditory cortex, MT+/V5, superior-temporal auditory, and multisensory and/or supramodal regions, such as posterior parietal cortex (PPC), frontal eye fields, anterior cingulate, and supplementary eye fields. Overall, these data demonstrate the contribution of neuroplasticity in multiple systems including primary auditory cortex, supramodal, and multisensory regions, to altered visual processing in congenitally deaf adults.

  6. In vitro methods to culture primary human breast epithelial cells.

    PubMed

    Raouf, Afshin; Sun, Yu Jia

    2013-01-01

    Current evidence suggests that much like leukemia, breast tumors are maintained by a small subpopulation of tumor cells that have stem cell properties. These cancer stem cells are envisaged to be responsible for tumor formation and relapse. Therefore, knowledge about their nature will provide a platform to develop therapies to eliminate these breast cancer stem cells. This concept highlights the need to understand the mechanisms that regulate the normal functions of the breast stem cells and their immediate progeny as alterations to these same mechanisms can cause these primitive cells to act as cancer stem cells. The study of the primitive cell functions relies on the ability to isolate them from primary sources of breast tissue. This chapter describes processing of discarded tissue from reduction mammoplasty samples as sources of normal primary human breast epithelial cells and describes cell culture systems to grow single-cell suspensions prepared from these reduction samples in vitro.

  7. Pure Primary Ovarian Squamous Cell Carcinoma Perforating the Rectum

    PubMed Central

    Okada, Aiko; Haraguchi, Naotsugu; Tomimatsu, Takuji; Kimura, Tadashi

    2017-01-01

    Rectal perforation is uncommon in ovarian cancer, even in advanced stages. Pure primary ovarian squamous cell carcinoma is a very rare subtype of ovarian cancer and has not been reported to cause rectal perforation. A 50-year-old woman presented with rectal bleeding. Rectosigmoidoscopy suggested perforation of a pelvic tumor into the rectum. Abdominopelvic magnetic resonance imaging revealed a 9 cm heterogeneous mass in the pouch of Douglas. We performed complete cytoreduction, including an en-bloc resection of the tumor and rectosigmoid colon. Histopathology showed squamous cell carcinoma of the left ovary penetrating the rectal wall. A common symptom of rectal bleeding was caused by a very rare entity of ovarian cancer penetrating the rectal wall, but thorough evaluation led to its accurate diagnosis and appropriate treatment. PMID:28316851

  8. Cultivate Primary Nasal Epithelial Cells from Children and Reprogram into Induced Pluripotent Stem Cells.

    PubMed

    Ulm, Ashley; Mayhew, Christopher N; Debley, Jason; Khurana Hershey, Gurjit K; Ji, Hong

    2016-03-10

    Nasal epithelial cells (NECs) are the part of the airways that respond to air pollutants and are the first cells infected with respiratory viruses. They are also involved in many airway diseases through their innate immune response and interaction with immune and airway stromal cells. NECs are of particular interest for studies in children due to their accessibility during clinical visits. Human induced pluripotent stem cells (iPSCs) have been generated from multiple cell types and are a powerful tool for modeling human development and disease, as well as for their potential applications in regenerative medicine. This is the first protocol to lay out methods for successful generation of iPSCs from NECs derived from pediatric participants for research purposes. It describes how to obtain nasal epithelial cells from children, how to generate primary NEC cultures from these samples, and how to reprogram primary NECs into well-characterized iPSCs. Nasal mucosa samples are useful in epidemiological studies related to the effects of air pollution in children, and provide an important tool for studying airway disease. Primary nasal cells and iPSCs derived from them can be a tool for providing unlimited material for patient-specific research in diverse areas of airway epithelial biology, including asthma and COPD research.

  9. Cultivate Primary Nasal Epithelial Cells from Children and Reprogram into Induced Pluripotent Stem Cells

    PubMed Central

    Ulm, Ashley; Mayhew, Christopher N.; Debley, Jason; Khurana Hershey, Gurjit K.; Ji, Hong

    2016-01-01

    Nasal epithelial cells (NECs) are the part of the airways that respond to air pollutants and are the first cells infected with respiratory viruses. They are also involved in many airway diseases through their innate immune response and interaction with immune and airway stromal cells. NECs are of particular interest for studies in children due to their accessibility during clinical visits. Human induced pluripotent stem cells (iPSCs) have been generated from multiple cell types and are a powerful tool for modeling human development and disease, as well as for their potential applications in regenerative medicine. This is the first protocol to lay out methods for successful generation of iPSCs from NECs derived from pediatric participants for research purposes. It describes how to obtain nasal epithelial cells from children, how to generate primary NEC cultures from these samples, and how to reprogram primary NECs into well-characterized iPSCs. Nasal mucosa samples are useful in epidemiological studies related to the effects of air pollution in children, and provide an important tool for studying airway disease. Primary nasal cells and iPSCs derived from them can be a tool for providing unlimited material for patient-specific research in diverse areas of airway epithelial biology, including asthma and COPD research. PMID:27022951

  10. Characterization of cancer stem cells and primary cilia in medulloblastoma.

    PubMed

    Gate, David; Danielpour, Moise; Bannykh, Serguei; Town, Terrence

    2015-01-01

    Medulloblastoma, a tumor of the cerebellum, is the most common pediatric central nervous system malignancy. These tumors are etiologically linked to mutations in the Sonic hedgehog (Shh) pathway, which signals through the primary, non-motile cilium. The growth of these aggressive tumors relies on self-renewal of tumor-propagating cells known as cancer stem cells (CSCs). Previous reports have implicated CD133-expressing cells as CSCs in brain tumors, while those expressing CD15 have been shown to propagate medulloblastoma. Here, we demonstrate that CD133+ and CD15+ cells are distinct medulloblastoma populations. CD15+ cells comprise approximately 0.5-1% of total human medulloblastoma cells, display CSC properties in culture and are detected in the Smoothened A1 transgenic mouse model of medulloblastoma. Additionally, we report on a medulloblastoma patient with enriched CD15+ cells in recurrent vs primary medulloblastoma. We also demonstrate that human medulloblastoma cells critically rely on establishment of primary cilia to drive Shh-mediated cell division. Primary cilia are found in external granule cells of human fetal cerebellum and in 12/14 medulloblastoma samples. Yet, CD15+ medulloblastoma cells lack primary cilia, suggesting that this CSC population signals independently of Shh. These results are important when considering the effects of current and prospective treatment modalities on medulloblastoma CSC populations.

  11. Primary cilium - antenna-like structure on the surface of most mammalian cell types

    NASA Astrophysics Data System (ADS)

    Dvorak, J.; Sitorova, V.; Hadzi Nikolov, D.; Mokry, J.; Richter, I.; Kasaova, L.; Filip, S.; Ryska, A.; Petera, J.

    2011-12-01

    The primary cilium is a sensory solitary non-motile microtubule-based organelle protruding in the quiescent phase of the cell cycle from the surface of the majority of human cells, including embryonic cells, stem cells and stromal cells of malignant tumors. The presence of a primary cilium on the surface of a cell is transient, limited to the quiescent G1(G0) phase and the beginning of the S phase of the cell cycle. The primary cilium is formed from the mother centriole. Primary cilia are key coordinators of signaling pathways during development and tissue homeostasis and, when deffective, they are a major cause of human diseases and developmental disorders, now commonly referred to as ciliopathies. Most cancer cells do not possess a primary cilium. The loss of the primary cilium is a regular feature of neoplastic transformation in the majority of solid tumors. The primary cilium could serve as a tumor suppressor organelle. The aim of this paper was to provide a review of the current knowledge of the primary cilium.

  12. BRCA1 haploinsufficiency for replication stress suppression in primary cells.

    PubMed

    Pathania, Shailja; Bade, Sangeeta; Le Guillou, Morwenna; Burke, Karly; Reed, Rachel; Bowman-Colin, Christian; Su, Ying; Ting, David T; Polyak, Kornelia; Richardson, Andrea L; Feunteun, Jean; Garber, Judy E; Livingston, David M

    2014-11-17

    BRCA1-a breast and ovarian cancer suppressor gene-promotes genome integrity. To study the functionality of BRCA1 in the heterozygous state, we established a collection of primary human BRCA1(+/+) and BRCA1(mut/+) mammary epithelial cells and fibroblasts. Here we report that all BRCA1(mut/+) cells exhibited multiple normal BRCA1 functions, including the support of homologous recombination- type double-strand break repair (HR-DSBR), checkpoint functions, centrosome number control, spindle pole formation, Slug expression and satellite RNA suppression. In contrast, the same cells were defective in stalled replication fork repair and/or suppression of fork collapse, that is, replication stress. These defects were rescued by reconstituting BRCA1(mut/+) cells with wt BRCA1. In addition, we observed 'conditional' haploinsufficiency for HR-DSBR in BRCA1(mut/+) cells in the face of replication stress. Given the importance of replication stress in epithelial cancer development and of an HR defect in breast cancer pathogenesis, both defects are candidate contributors to tumorigenesis in BRCA1-deficient mammary tissue.

  13. Primary cells - A forecast of performance

    NASA Astrophysics Data System (ADS)

    Hazkany, H.; Peled, E.; Raz, B.

    Of the several existing methods of exploratory forecasting the method of trend extrapolation has been chosen. This method is built upon the assumption that excepting technological revolution, technological characteristics develop in an orderly manner and that development trends are predictable. Attention is given to the Leclanche cell, the alkaline cell, the zinc silver oxide cell, the magnesium cell, the mercury cell, zinc-air cells, and Li cells. It is found that safe commercial high rate cells will have an energy density which is 10 to 20 percent lower than the value corresponding to the maximum capability of the technology. Cells with about 500-630 W-hr/kg are expected to penetrate the market at 1990. Low and high temperature performance and shelf life have almost reached the desirable levels and no vast improvement is expected or needed. Power density is expected to grow to the 500-1000 W/kg range (pulses), but safety problems must be resolved before commercialization.

  14. HYDROGEN-OXYGEN PRIMARY EXTRATERRESTRIAL (HOPE) FUEL CELL PROGRAM

    DTIC Science & Technology

    The HOPE (Hydrogen-Oxygen Primary Extraterrestrial) Fuel Cell Program is a multi-phase effort to advance the state-of-the-art of fuel cells by...configuration fuel cell module. The HOPE spacecraft, fuel supply tanks, pneumatics, and thermal systems were designed and fabricated to provide...verify water removal, thermal design, and 30-day shelf-life of the fuel cell . The 35-cell module was subjected to a series of performance tests

  15. Growth inhibitory activity of Ankaferd hemostat on primary melanoma cells and cell lines

    PubMed Central

    Turk, Seyhan; Malkan, Umit Yavuz; Ghasemi, Mehdi; Hocaoglu, Helin; Mutlu, Duygu; Gunes, Gursel; Aksu, Salih; Haznedaroglu, Ibrahim Celalettin

    2017-01-01

    Objective: Ankaferd hemostat is the first topical hemostatic agent about the red blood cell–fibrinogen relations tested in the clinical trials. Ankaferd hemostat consists of standardized plant extracts including Alpinia officinarum, Glycyrrhiza glabra, Thymus vulgaris, Urtica dioica, and Vitis vinifera. The aim of this study was to determine the effect of Ankaferd hemostat on viability of melanoma cell lines. Methods: Dissimilar melanoma cell lines and primary cells were used in this study. These cells were treated with different concentrations of Ankaferd hemostat to assess the impact of different dosages of the drug. All cells treated with different concentrations were incubated for different time intervals. After the data had been obtained, one-tailed T-test was used to determine whether the Ankaferd hemostat would have any significant inhibitory impact on cell growth. Results: We demonstrated in this study that cells treated with Ankaferd hemostat showed a significant decrease in cell viability compared to control groups. The cells showed different resistances against Ankaferd hemostat which depended on the dosage applied and the time treated cells had been incubated. We also demonstrated an inverse relationship between the concentration of the drug and the incubation time on one hand and the viability of the cells on the other hand, that is, increasing the concentration of the drug and the incubation time had a negative impact on cell viability. Conclusion: The findings in our study contribute to our knowledge about the anticancer impact of Ankaferd hemostat on different melanoma cells. PMID:28293423

  16. Primary lithium-thionyl chloride cell evaluation

    NASA Astrophysics Data System (ADS)

    Zolla, A. E.; Waterhouse, R.; Debiccari, D.; Griffin, G. L.

    1980-08-01

    A test program was conducted to evaluate the Altus 1350AH cell performance against the Minuteman Survival Ground Power requirements. Twelve cells of the 17 inch diameter, 1-3/8 inch heights were fabricated and tested during this study. Under discharge rates varying from C/100 to C/400 at ambient temperature, the volumetric and gravimetric energy density performance requirements of 15 watt hours per cubic inch and 150 watt hours per pound were exceeded in all cases. All other performance requirements of voltage, current, configuration, capacity volume, weight, electrolyte leakage (none), and maintainability (none required), were met or exceeded. The abuse testing demonstrated the Altus Cell's ability to safely withstand short circuit by external shorting, short circuit by penetration with a conductive object, forced discharge, and forced charging of a cell. Disposal of discharged cells by incineration is an environmentally safe and efficient method of disposal.

  17. Increasing resource allocation and research into tobacco control activities: a comprehensive approach including primary prevention, treatment and brief intervention.

    PubMed

    Richmond, R

    1993-01-01

    The range of tobacco control activities should be viewed as essential parts of a complex multi-component puzzle. Intervention strategies designed to address tobacco control should be comprehensive and include both primary and secondary prevention activities and be multi-faceted and capable of bringing about change at both the individual and broader social and cultural levels. In this paper I argue for a mutually inclusive framework in which the various components contribute in important and different ways. I examine the prevalence of smoking and identify the high risk groups, then I examine the range of available strategies and present the evidence for their success. I discuss the primary prevention approaches such as warning labels, taxes, price increases, workplace bans, education in schools, mass media and self-help materials, as well as brief interventions and treatment strategies which are conducted at the worksite, general practice and specialized cessation clinics. The areas for future research are delineated for increased resource allocation and include: the best ways to disseminate brief interventions to smokers, methods to motivate smokers; training of health professionals to deliver brief interventions; enhancing quitting and access to existing treatment resources among specific disadvantaged minority groups, e.g. migrants, unemployed youth, the effect on smoking prevalence of warning labels on cigarette packets and price rises on cigarettes.

  18. Human prion protein-induced autophagy flux governs neuron cell damage in primary neuron cells.

    PubMed

    Moon, Ji-Hong; Lee, Ju-Hee; Nazim, Uddin Md; Lee, You-Jin; Seol, Jae-Won; Eo, Seong-Kug; Lee, John-Hwa; Park, Sang-Youel

    2016-05-24

    An unusual molecular structure of the prion protein, PrPsc is found only in mammals with transmissible prion diseases. Prion protein stands for either the infectious pathogen itself or a main component of it. Recent studies suggest that autophagy is one of the major functions that keep cells alive and has a protective effect against the neurodegeneration. In this study, we investigated that the effect of human prion protein on autophagy-lysosomal system of primary neuronal cells. The treatment of human prion protein induced primary neuron cell death and decreased both LC3-II and p62 protein amount indicating autophagy flux activation. Electron microscope pictures confirmed the autophagic flux activation in neuron cells treated with prion protein. Inhibition of autophagy flux using pharmacological and genetic tools prevented neuron cell death induced by human prion protein. Autophagy flux induced by prion protein is more activated in prpc expressing cells than in prpc silencing cells. These data demonstrated that prion protein-induced autophagy flux is involved in neuron cell death in prion disease and suggest that autophagy flux might play a critical role in neurodegenerative diseases including prion disease.

  19. Sealed Primary Lithium-Inorganic Electrolyte Cell

    DTIC Science & Technology

    1977-02-01

    Battery , Thionyl Chloride , Lithium , Lithium Aluminum Chloride , Hermetic Lithium Battery , D Cell, Voltage-Delay, Shelf Life, High Energy Density Battery ... lithium - thionyl chloride , inorganic electrclyte system is one of the highest energy density systems known to date (1-4). The cells contain an Li anoae, a...However, this is not tne case with te thionyl chloride system. A completely discharged battery , while sitting on

  20. Sealed Primary Lithium-Inorganic Electrolyte Cell

    DTIC Science & Technology

    1977-11-01

    Thionyl Chloride , Lithium , Lithium Aluminum Chloride , Hermetic Lithium Battery, D Cell I Voltage-Delay 1 Shelf Life 1 High_ Energy Density...and the propagation of the thermal runaway en- countered in the thionyl cells. For our initial studies we restricted ourselves to the stable...types of sulfur e.g. rhombic (^) and monoclinic (A ). 3. Thionyl Chloride (SOCI2) The thermogram of SOCI2 (0.161 gm) at 50C/min heating rate is

  1. Primary cell uses neither liquid nor fused electrolytes

    NASA Technical Reports Server (NTRS)

    Gutmann, F.; Herman, A. M.; Rembaum, A.

    1967-01-01

    Dry, solid state primary battery cell establishes an electrode reaction by a charge transfer mechanism without liquid phase ionization of electrolyte compounds. The charge transfer complex is sufficiently conductive to permit the passage of useful current.

  2. Primary Central Nervous System Anaplastic Large T-cell Lymphoma

    PubMed Central

    Splavski, Bruno; Muzevic, Dario; Ladenhauser-Palijan, Tatjana; Jr, Brano Splavski

    2016-01-01

    Introduction: Primary central nervous system lymphoma (PCNSL) of T-cell origin is an exceptionally rare, highly malignant intracranial neoplasm. Although such a tumor typically presents with a focal mass lesion. Case report: Past medical history of a 26-year-old male patient with a PCNS lymphoma of T-cell origin was not suggestive of intracranial pathology or any disorder of other organs and organic systems. To achieve a gross total tumor resection, surgery was performed via osteoplastic craniotomy using the left frontal transcortical transventricular approach. Histological and immunohistochemical analyses of the tissue removed described tumor as anaplastic large cell lymphoma of T-cells (T-ALCL). Postoperative and neurological recovery was complete, while control imaging of the brain showed no signs of residual tumor at a six-month follow-up. The patient, who did not appear immunocompromized, was referred to a hematologist and an oncologist where corticosteroids, the particular chemotherapeutic protocol and irradiation therapy were applied. Conclusion: Since PCNS lymphoma is a potentially curable brain tumor, we believe that proper selection of the management options, including early radical tumor resection for solitary PCNS lymphoma, may be proposed as a major treatment of such a tumor in selected patients, resulting in a satisfactory outcome. PMID:27703297

  3. Primary Cutaneous Mantle Cell Lymphoma: A Case Report

    PubMed Central

    Mazzuoccolo, L. D.; Castro Perez, G. A.; Sorin, I.; Bravo, A. I.

    2013-01-01

    Primary cutaneous mantle cell lymphoma (MCL) is a rare cutaneous proliferation of naive pregerminal CD-5 positive B cells in the skin with no extracutaneous involvement. Overexpression of cyclin D1 is pathognomonic of this condition, and surgery and radiation therapy are the most common therapeutic options. In this case, we describe the clinical, histopathological, immunohistochemical, and molecular characteristics of a new case of primary cutaneous MCL. PMID:23762653

  4. The Correlation between NK Cell and Liver Function in Patients with Primary Hepatocellular Carcinoma

    PubMed Central

    Zeng, Xiao-Hui; Min, Lu

    2014-01-01

    Background/Aims This study aimed to detect the expression of natural killer (NK) cell receptor natural killer group 2D (NKG2D) in the peripheral blood of patients with primary hepatocellular carcinoma and to discuss the correlation between NK cell cytotoxicity and liver function. Methods The number of NK cells and the expression of NK cell receptor NKG2D in peripheral blood were determined by flow cytometry in patients with primary hepatocellular carcinoma, hepatitis B cirrhosis, chronic hepatitis B, and healthy controls. Results When compared with patients in the healthy and the chronic hepatitis B groups, the primary hepatocellular carcinoma group showed significant decreases in all parameters, including the cytotoxicity of NK cells on K562 cells, expression rate of NKG2D in NK cells, number of NKG2D+ NK cells, expression level of NKG2D, and number of NK cells (p<0.05). The activity of NK cells showed a positive correlation, whereas the Child-Pugh scores in the primary hepatocellular carcinoma and the hepatitis B cirrhosis groups showed a negative correlation with all parameters detected above. Conclusions The decrease of NK cell activity in patients with primary hepatocellular carcinoma is closely related to their lower expression of NKG2D. Liver function affects the expression of NKG2D and the activity of NK cells. PMID:24827627

  5. Metabolic Profiling of Developing Pear Fruits Reveals Dynamic Variation in Primary and Secondary Metabolites, Including Plant Hormones.

    PubMed

    Oikawa, Akira; Otsuka, Takao; Nakabayashi, Ryo; Jikumaru, Yusuke; Isuzugawa, Kanji; Murayama, Hideki; Saito, Kazuki; Shiratake, Katsuhiro

    2015-01-01

    Metabolites in the fruits of edible plants include sweet sugars, visually appealing pigments, various products with human nutritional value, and biologically active plant hormones. Although quantities of these metabolites vary during fruit development and ripening because of cell division and enlargement, there are few reports describing the actual dynamics of these changes. Therefore, we applied multiple metabolomic techniques to identify the changes in metabolite levels during the development and ripening of pear fruits (Pyrus communis L. 'La France'). We quantified and classified over 250 metabolites into six groups depending on their specific patterns of variation during development and ripening. Approximately half the total number of metabolites, including histidine and malate, accumulated transiently around the blooming period, during which cells are actively dividing, and then decreased either rapidly or slowly. Furthermore, the amounts of sulfur-containing amino acids also increased in pear fruits around 3-4 months after the blooming period, when fruit cells are enlarging, but virtually disappeared from ripened fruits. Some metabolites, including the plant hormone abscisic acid, accumulated particularly in the receptacle prior to blooming and/or fruit ripening. Our results show several patterns of variation in metabolite levels in developing and ripening pear fruits, and provide fundamental metabolomic data that is useful for understanding pear fruit physiology and enhancing the nutritional traits of new cultivars.

  6. Metabolic Profiling of Developing Pear Fruits Reveals Dynamic Variation in Primary and Secondary Metabolites, Including Plant Hormones

    PubMed Central

    Oikawa, Akira; Otsuka, Takao; Nakabayashi, Ryo; Jikumaru, Yusuke; Isuzugawa, Kanji; Murayama, Hideki; Saito, Kazuki; Shiratake, Katsuhiro

    2015-01-01

    Metabolites in the fruits of edible plants include sweet sugars, visually appealing pigments, various products with human nutritional value, and biologically active plant hormones. Although quantities of these metabolites vary during fruit development and ripening because of cell division and enlargement, there are few reports describing the actual dynamics of these changes. Therefore, we applied multiple metabolomic techniques to identify the changes in metabolite levels during the development and ripening of pear fruits (Pyrus communis L. ‘La France’). We quantified and classified over 250 metabolites into six groups depending on their specific patterns of variation during development and ripening. Approximately half the total number of metabolites, including histidine and malate, accumulated transiently around the blooming period, during which cells are actively dividing, and then decreased either rapidly or slowly. Furthermore, the amounts of sulfur-containing amino acids also increased in pear fruits around 3–4 months after the blooming period, when fruit cells are enlarging, but virtually disappeared from ripened fruits. Some metabolites, including the plant hormone abscisic acid, accumulated particularly in the receptacle prior to blooming and/or fruit ripening. Our results show several patterns of variation in metabolite levels in developing and ripening pear fruits, and provide fundamental metabolomic data that is useful for understanding pear fruit physiology and enhancing the nutritional traits of new cultivars. PMID:26168247

  7. Immune cells in primary and metastatic gastrointestinal stromal tumors (GIST).

    PubMed

    Cameron, Silke; Gieselmann, Marieke; Blaschke, Martina; Ramadori, Giuliano; Füzesi, Laszlo

    2014-01-01

    We have previously described immune cells in untreated primary gastrointestinal stromal tumors (GIST). Here we compare immune cells in metastatic and primary GIST, and describe their chemoattractants. For this purpose, tissue microarrays from 196 patients, 188 primary and 51 metastasized GIST were constructed for paraffin staining. Quantitative analysis was performed for cells of macrophage lineage (Ki-M1P, CD68), T-cells (CD3, CD56) and B-cells (CD20). Chemokine gene-expression was evaluated by real-time RT-PCR. Immuno-localisation was verified by immunofluorescence. Ki-M1P+ cells were the predominant immune cells in both primary and metastatic GIST (2 8.8% ± 7.1, vs. 26.7% ± 6.3). CD68+ macrophages were significantly fewer, with no significant difference between primary GIST (3.6% ± 2.1) and metastases (4.6% ± 1.5). CD3+ T-cells were the most dominant lymphocytes with a significant increase in metastases (7.3% ± 2.3 vs. 2.2% ± 1.8 in primary GIST, P < 0.01). The percentage of CD56+ NK-cells was 1.1% ± 0.9 in the primary, and 2.4 ± 0.7 (P < 0.05) in the metastases. The number of CD20+ B-cells was generally low with 0.6% ± 0.7 in the primary and 1.8% ± 0.3 (P < 0.05) in the metastases. Analysis of the metastases showed significantly more Ki-M1P+ cells in peritoneal metastases (31.8% ± 7.4 vs. 18.2% ± 3.7, P < 0.01), whilst CD3+ T-cells were more common in liver metastases (11.7% ± 1.8 vs. 4.4% ± 2.6, P < 0.01). The highest transcript expression was seen for monocyte chemotactic protein 1 (MCP1/CCL2), macrophage inflammatory protein 1α (MIP-1α/CCL3) and the pro-angiogenic growth-related oncoprotein 1 (Gro-α/CXCL-1). Whilst the ligands were predominantly expressed in tumor cells, their receptors were mostly present in immune cells. This locally specific microenvironment might influence neoplastic progression of GIST at the different metastatic sites.

  8. Immune cells in primary and metastatic gastrointestinal stromal tumors (GIST)

    PubMed Central

    Cameron, Silke; Gieselmann, Marieke; Blaschke, Martina; Ramadori, Giuliano; Füzesi, Laszlo

    2014-01-01

    We have previously described immune cells in untreated primary gastrointestinal stromal tumors (GIST). Here we compare immune cells in metastatic and primary GIST, and describe their chemoattractants. For this purpose, tissue microarrays from 196 patients, 188 primary and 51 metastasized GIST were constructed for paraffin staining. Quantitative analysis was performed for cells of macrophage lineage (Ki-M1P, CD68), T-cells (CD3, CD56) and B-cells (CD20). Chemokine gene-expression was evaluated by real-time RT-PCR. Immuno-localisation was verified by immunofluorescence. Ki-M1P+ cells were the predominant immune cells in both primary and metastatic GIST (2 8.8% ± 7.1, vs. 26.7% ± 6.3). CD68+ macrophages were significantly fewer, with no significant difference between primary GIST (3.6% ± 2.1) and metastases (4.6% ± 1.5). CD3+ T-cells were the most dominant lymphocytes with a significant increase in metastases (7.3% ± 2.3 vs. 2.2% ± 1.8 in primary GIST, P < 0.01). The percentage of CD56+ NK-cells was 1.1% ± 0.9 in the primary, and 2.4 ± 0.7 (P < 0.05) in the metastases. The number of CD20+ B-cells was generally low with 0.6% ± 0.7 in the primary and 1.8% ± 0.3 (P < 0.05) in the metastases. Analysis of the metastases showed significantly more Ki-M1P+ cells in peritoneal metastases (31.8% ± 7.4 vs. 18.2% ± 3.7, P < 0.01), whilst CD3+ T-cells were more common in liver metastases (11.7% ± 1.8 vs. 4.4% ± 2.6, P < 0.01). The highest transcript expression was seen for monocyte chemotactic protein 1 (MCP1/CCL2), macrophage inflammatory protein 1α (MIP-1α/CCL3) and the pro-angiogenic growth-related oncoprotein 1 (Gro-α/CXCL-1). Whilst the ligands were predominantly expressed in tumor cells, their receptors were mostly present in immune cells. This locally specific microenvironment might influence neoplastic progression of GIST at the different metastatic sites. PMID:25120735

  9. Role for suppressor T cells in the pathogenesis of autoimmune diseases (including rheumatoid arthritis). Facts and hypotheses.

    PubMed

    Berthelot, Jean-Marie; Maugars, Yves

    2004-09-01

    Although uncontrolled clones of autoreactive T cells play a central role in the pathogenesis of autoimmunity, another mechanism potentially involved in many autoimmune diseases is deficiency of suppressor T cells, most notably those belonging to the antiidiopeptide TH3/Tr1 TCD4+CD25+(high) subset. Failure of suppressor mechanisms may be in part primary, due to defective positive selection of suppressor T cells in the thymus, and in part acquired, secondary to chronic infections promoted by deficiencies in innate immunity. Renewed interest in suppressor TCD4+ cells has generated plausible explanations for many events including paradoxical induction of autoimmune disorders by immunosuppressive agents or thymectomy. Insights into the physiology of these regulatory T-cell clones might suggest new treatment options, although many currently used drugs (including anti-TNF alpha agents) enhance the activity of several suppressor T-cell clones. Investigation of these suppressor clones in rheumatoid arthritis is still in its infancy and faces obstacles such as the need for identifying key clones in each individual patient and the presence of T-cell repertoire contraction. This last phenomenon exists at disease onset and may stem from early thymus dysfunction, which may also lead to a reduction in suppressor TCD4+ cell counts. Thus, although restoring deficient suppressor clones may provide a full recovery in animals, the high prevalence of T-cell repertoire contraction in humans with rheumatoid arthritis may severely limit the beneficial effects of this therapeutic approach.

  10. Effect of antifreeze proteins on frozen primary prostatic adenocarcinoma cells.

    PubMed

    Koushafar, H; Rubinsky, B

    1997-03-01

    Recent studies show that prostate adenocarcinoma cells can survive cryosurgery and that cell destruction depends on the specific thermal parameters used during freezing. The goal of this preliminary study is to determine whether certain chemical compounds, known as antifreeze proteins, can induce complete human primary prostatic adenocarcinoma cell destruction by freezing, regardless of the thermal parameters used. The study also examines the mechanism by which antifreeze proteins bring about cell destruction. Antifreeze proteins were added to solutions containing human primary prostatic adenocarcinoma cells. The cells were frozen with controlled thermal parameters using a directional solidification apparatus attached to a light microscope. Cell viability was determined after thawing as a function of antifreeze protein concentration and cooling rate during freezing. The dose response study shows that for all the cooling rates tested, 10-mg/mL solutions of antifreeze protein cause the complete destruction of human primary prostatic adenocarcinoma cells frozen to a temperature at which, without these proteins, the cells survive freezing. Light microscopy shows that the lethal effect of the antifreeze proteins is related to the formation of intracellular ice in the frozen cells. CONCLUSIONS; This preliminary study has demonstrated that antifreeze proteins have the ability to generate complete destruction of prostatic adenocarcinoma cells frozen to high subzero temperatures irrespective of the cooling rates used during freezing. This suggests that introducing antifreeze proteins into undesirable tissues prior to freezing may increase the efficacy and the control over tissue destruction by cryosurgery.

  11. B Cell Depletion Therapy Normalizes Circulating Follicular Th Cells in Primary Sjögren Syndrome.

    PubMed

    Verstappen, Gwenny M; Kroese, Frans G M; Meiners, Petra M; Corneth, Odilia B; Huitema, Minke G; Haacke, Erlin A; van der Vegt, Bert; Arends, Suzanne; Vissink, Arjan; Bootsma, Hendrika; Abdulahad, Wayel H

    2017-01-01

    To assess the effect of B cell depletion therapy on effector CD4+ T cell homeostasis and its relation to objective measures of disease activity in patients with primary Sjögren syndrome (pSS). Twenty-four patients with pSS treated with rituximab (RTX) and 24 healthy controls (HC) were included. Frequencies of circulating effector CD4+ T cell subsets were examined by flow cytometry at baseline and 16, 24, 36, and 48 weeks after the first RTX infusion. Th1, Th2, follicular Th (TFH), and Th17 cells were discerned based on surface marker expression patterns. Additionally, intracellular cytokine staining was performed for interferon-γ, interleukin (IL)-4, IL-21, and IL-17 and serum levels of these cytokines were analyzed. In patients with pSS, frequencies of circulating TFH cells and Th17 cells were increased at baseline compared with HC, whereas frequencies of Th1 and Th2 cells were unchanged. B cell depletion therapy resulted in a pronounced decrease in circulating TFH cells, whereas Th17 cells were only slightly lowered. Frequencies of IL-21-producing and IL-17-producing CD4+ T cells and serum levels of IL-21 and IL-17 were also reduced. Importantly, the decrease in circulating TFH cells was associated with lower systemic disease activity over time, as measured by the European League Against Rheumatism Sjögren's Syndrome Disease Activity Index scores and serum IgG levels. B cell depletion therapy in patients with pSS results in normalization of the elevated levels of circulating TFH cells. This reduction is associated with improved objective clinical disease activity measures. Our observations illustrate the pivotal role of the crosstalk between B cells and TFH cells in the pathogenesis of pSS.

  12. Clinicopathological features of five unusual cases of intraosseous myoepithelial carcinomas, mimicking conventional primary bone tumours, including EWSR1 rearrangement in one case.

    PubMed

    Rekhi, Bharat; Joshi, Sujit; Panchwagh, Yogesh; Gulia, Ashish; Borges, Anita; Bajpai, Jyoti; Jambehekar, Nirmala A; Pant, Vinita; Mandholkar, Mahesh; Byregowda, Suman; Puri, Ajay

    2016-04-01

    Primary intraosseous myoepithelial tumours, including carcinomas are rare tumours. The concept of histopathological spectrum of these tumours is evolving. We describe clinicopathological and immunohistochemical features of five myoepithelial carcinomas, including molecular cytogenetic results in one case. There were five male patients within age-range of 8-40 years (median = 26). Four tumours occurred in the long bones, including two tumours, each, in the femur and fibula, respectively, while a single tumour occurred in the proximal phalanges. Tumour size (n = 3 cases) varied from 5.6 to 8.6 cm. On radiological imaging, most tumours appeared as expansile, lytic and destructive lesions. Two tumours appeared as sclerotic lesions. Two cases were referred with diagnoses of chondrosarcomas and a single case was referred with two different diagnoses, including an adamantinoma and an osteosarcoma. Histopathological examination in all these cases showed multinodular tumours comprising mostly polygonal cells, exhibiting moderate nuclear atypia and interspersed mitotic figures within a stroma containing variable amount of myxoid, chondroid, hyalinised and osteoid-like material. Three tumours revealed prominent squamous differentiation. By immunohistochemistry, tumour cells were positive for EMA (5/5), pan CK (AE1/AE3) (3/3), CK5/6 (4/4), CK MNF116 (1/1), S100 protein (5/5) and GFAP (3/5). The first tumour revealed EWSR1 rearrangement. The first patient, 10 months after tumour resection and a simultaneous lung metastatectomy, is free-of-disease (FOD). The second patient, 11 months after tumour resection is FOD. The third and fourth patients underwent wide resections and are on follow-up. The fifth patient underwent resections, including a lung metastatectomy. Primary intraosseous myoepithelial carcinomas are rare and mimic conventional primary bone tumours. Some primary intraosseous myoepithelial carcinomas display EWSR1 rearrangement. Squamous differentiation may be

  13. Inactivation of Ricin Toxin by Nanosecond Pulsed Electric Fields Including Evidences from Cell and Animal Toxicity.

    PubMed

    Wei, Kai; Li, Wei; Gao, Shan; Ji, Bin; Zang, Yating; Su, Bo; Wang, Kaile; Yao, Maosheng; Zhang, Jue; Wang, Jinglin

    2016-01-05

    Ricin is one of the most toxic and easily produced plant protein toxin extracted from the castor oil plant, and it has been classified as a chemical warfare agent. Here, nanosecond pulsed electric fields (nsPEFs) at 30 kV/cm (pulse durations: 10 ns, 100 ns, and 300 ns) were applied to inactivating ricin up to 4.2 μg/mL. To investigate the efficacy, cells and mice were tested against the ricin treated by the nsPEFs via direct intraperitoneal injection and inhalation exposure. Results showed that nsPEFs treatments can effectively reduce the toxicity of the ricin. Without the nsPEFs treatment, 100% of mice were killed upon the 4 μg ricin injection on the first day, however 40% of the mice survived the ricin treated by the nsPEFs. Compared to injection, inhalation exposure even with higher ricin dose required longer time to observe mice fatality. Pathological observations revealed damages to heart, lung, kidney, and stomach after the ricin exposure, more pronounced for lung and kidney including severe bleeding. Sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE) and circular dichroism (CD) analyses revealed that although the primary structure of ricin was not altered, its secondary structures (beta-sheet and beta-turn) underwent transition upon the nsPEFs treatment.

  14. Inactivation of Ricin Toxin by Nanosecond Pulsed Electric Fields Including Evidences from Cell and Animal Toxicity

    PubMed Central

    Wei, Kai; Li, Wei; Gao, Shan; Ji, Bin; Zang, Yating; Su, Bo; Wang, Kaile; Yao, Maosheng; Zhang, Jue; Wang, Jinglin

    2016-01-01

    Ricin is one of the most toxic and easily produced plant protein toxin extracted from the castor oil plant, and it has been classified as a chemical warfare agent. Here, nanosecond pulsed electric fields (nsPEFs) at 30 kV/cm (pulse durations: 10 ns, 100 ns, and 300 ns) were applied to inactivating ricin up to 4.2 μg/mL. To investigate the efficacy, cells and mice were tested against the ricin treated by the nsPEFs via direct intraperitoneal injection and inhalation exposure. Results showed that nsPEFs treatments can effectively reduce the toxicity of the ricin. Without the nsPEFs treatment, 100% of mice were killed upon the 4 μg ricin injection on the first day, however 40% of the mice survived the ricin treated by the nsPEFs. Compared to injection, inhalation exposure even with higher ricin dose required longer time to observe mice fatality. Pathological observations revealed damages to heart, lung, kidney, and stomach after the ricin exposure, more pronounced for lung and kidney including severe bleeding. Sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE) and circular dichroism (CD) analyses revealed that although the primary structure of ricin was not altered, its secondary structures (beta-sheet and beta-turn) underwent transition upon the nsPEFs treatment. PMID:26728251

  15. Inactivation of Ricin Toxin by Nanosecond Pulsed Electric Fields Including Evidences from Cell and Animal Toxicity

    NASA Astrophysics Data System (ADS)

    Wei, Kai; Li, Wei; Gao, Shan; Ji, Bin; Zang, Yating; Su, Bo; Wang, Kaile; Yao, Maosheng; Zhang, Jue; Wang, Jinglin

    2016-01-01

    Ricin is one of the most toxic and easily produced plant protein toxin extracted from the castor oil plant, and it has been classified as a chemical warfare agent. Here, nanosecond pulsed electric fields (nsPEFs) at 30 kV/cm (pulse durations: 10 ns, 100 ns, and 300 ns) were applied to inactivating ricin up to 4.2 μg/mL. To investigate the efficacy, cells and mice were tested against the ricin treated by the nsPEFs via direct intraperitoneal injection and inhalation exposure. Results showed that nsPEFs treatments can effectively reduce the toxicity of the ricin. Without the nsPEFs treatment, 100% of mice were killed upon the 4 μg ricin injection on the first day, however 40% of the mice survived the ricin treated by the nsPEFs. Compared to injection, inhalation exposure even with higher ricin dose required longer time to observe mice fatality. Pathological observations revealed damages to heart, lung, kidney, and stomach after the ricin exposure, more pronounced for lung and kidney including severe bleeding. Sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE) and circular dichroism (CD) analyses revealed that although the primary structure of ricin was not altered, its secondary structures (beta-sheet and beta-turn) underwent transition upon the nsPEFs treatment.

  16. Primary biliary cirrhosis: an orchestrated immune response against epithelial cells.

    PubMed

    Gershwin, M E; Ansari, A A; Mackay, I R; Nakanuma, Y; Nishio, A; Rowley, M J; Coppel, R L

    2000-04-01

    Primary biliary cirrhosis (PBC) is an organ-specific autoimmune disease that predominantly affects women and is characterized by chronic progressive destruction of small intrahepatic bile ducts with portal inflammation and ultimately fibrosis. The serologic hallmark of PBC is the presence of antibodies to mitochondria, especially to the E2 component of the pyruvate dehydrogenase complex. The mechanisms by which (and if) such antibodies produce liver tissue injury are unknown. However, the presence of these antibodies has allowed detailed immunological definition of the antigenic epitopes, the nature of reactive autoantibodies and the characterization of T-cell responses. Several mechanisms may now be proposed regarding the immune-mediated bile duct damage in PBC, including the possible role of T-cell-mediated cytotoxicity and intracellular interaction between the IgA class of antimitochondrial antibodies and mitochondrial autoantigens. There are major questions which remain unanswered, including, of course, etiology, but also the reasons for female predominance, the absence of PBC in children, the relative ineffectiveness of immunosuppressive drugs, and the specific role of mitochondrial antigens. The data so far provide suggestive evidence that PBC is a mucosal disease; this thesis provides a basis for discussion of etiology via the enterohepatic circulation of toxins and/or infection.

  17. The exocyst localizes to the primary cilium in MDCK cells.

    PubMed

    Rogers, Katherine K; Wilson, Patricia D; Snyder, Richard W; Zhang, Xiaoyu; Guo, Wei; Burrow, Christopher R; Lipschutz, Joshua H

    2004-06-18

    Primary cilia play a role in the maintenance of tubular epithelial differentiation and ciliary dysfunction can result in abnormal cyst formation, such as occurs in autosomal dominant polycystic kidney disease (ADPKD). We previously showed that the exocyst, an eight-protein complex involved in the biogenesis of polarity from yeast to mammals, is centrally involved in cyst formation [Mol. Biol. Cell. 11 (2000) 4259]. Here we show that the exocyst complex localizes to the primary cilium in Madin-Darby canine kidney (MDCK) tubular epithelial cells. We further show that the exocyst is overexpressed in both cell lines and primary cell cultures of ADPKD origin, suggesting that the exocyst may be involved in the pathogenesis of ADPKD.

  18. Cathode catalyst for primary phosphoric fuel cells

    NASA Technical Reports Server (NTRS)

    Walsh, F.

    1980-01-01

    Alkylation of Vulcan XC-72 provided the most stable bond type for linking CoTAA to the surface of the carbon; this result is based on data obtained by cyclic voltammetry, pulse voltammetry and by release of 14C from bonded CoTAA. Half-cell tests at 100 C in 85% phosphoric acid showed that CoTAA bonded to the surface of carbon (Vulcan XC-72) via an alkylation procedure is a more active catalyst than is platinum based on a factor of two improvement in Tafel slope; dimeric CoTAA has catalytic activity equal to platinum. Half-cell tests also showed that bonded CoTAA catalysts do not suffer a loss in potential when air is used as a fuel rather than oxygen. Commercially available PTFE was shown to be stable for four months in 200 C 85% phosphoric acid based on lack of change in surface wetting properties, IR and physical characteristics. When stressed electrochemically in 150 C 85% phosphoric acid, PTFE also showed no changes after one month.

  19. Advances in the theory and application of BSF cells. [including electrical resistivity and photovoltaic cells

    NASA Technical Reports Server (NTRS)

    Mandelkorn, J.; Lamneck, J. H.

    1975-01-01

    The characteristics and behavior of p(+), p solar cells were investigated. The p(+), p cells were made by the removal of the n(+) surface layers from n(+), p p(+), BSF cells followed by application of a suitable contact to the resultant p(+), p structures. The open circuit voltage of p(+), p cells was found to increase with increasing 'p' bulk resistivity. The measured open circuit velocity-temperature coefficients were positive and increased with increasing resistivity. An outline of prior limitations in solar cell design is presented, and the removal of these limitations through use of BSF effects is pointed out. The study of BSF effects made feasible production of very thin high efficiency silicon cells as well as high resistivity-high efficiency cells, two desirable types of silicon cells which were previously impossible to make.

  20. A rare case of primary cardiac B cell lymphoma

    PubMed Central

    2014-01-01

    Primary cardiac lymphomas represent an extremely rare entity of extranodal lymphomas and should be distinguished from secondary cardiac involvement of disseminated lymphomas belonging to the non-Hodgkin’s classification of blood cancers. Only 90 cases have been reported in literature. Presentation of cardiac lymphomas on imaging studies may not be unambiguous since they potentially mimic other cardiac neoplasms including myxomas, angiosarcoma or rhadomyomas and therefore require multimodality cardiac imaging, endomyocardial biopsy, excisional intraoperative biopsy and pericardial fluid cytological evaluation to establish final diagnosis. Herein we report the case of a 70 y/o immunocompetent Caucasian female with a rapidly progressing superior vena cava syndrome secondary to a large primary cardiac diffuse large B cell lymphoma (NHL lymphoma) almost completely obstructing the right atrium, right ventricle and affecting both mitral and tricuspid valve. The patient had no clinical evidence of disseminated disease and was successfully treated with extensive debulking during open-heart surgery on cardiopulmonary bypass and 6 cycles of rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone chemotherapy (R-CHOP). PMID:24422789

  1. Chromosome Conformation Capture in Primary Human Cells.

    PubMed

    Cortesi, Alice; Bodega, Beatrice

    2016-01-01

    3D organization of the genome, its structural and regulatory function of cell identity, is acquiring prominent features in epigenetics studies; more efforts have been done to develop techniques that allow studying nuclear structure. Chromosome conformation capture (3C) has been set up in 2002 from Dekker and from that moment great investments were made to develop genomics variants of 3C technology (4C, 5C, Hi-C) providing new tools to investigate the shape of the genome in a more systematic and unbiased manner. 3C method allows scientists to fix dynamic and variable 3D interactions in nuclear space, and consequently to study which sequences interact, how a gene is regulated by different and distant enhancer, or how a set of enhancer could regulate transcriptional units; to follow the conformation that mediates regulation change in development; and to evaluate if this fine epigenetic mechanism is impaired in disease condition.

  2. Colon-targeted delivery of live bacterial cell biotherapeutics including microencapsulated live bacterial cells

    PubMed Central

    Prakash, Satya; Malgorzata Urbanska, Aleksandra

    2008-01-01

    There has been an ample interest in delivery of therapeutic molecules using live cells. Oral delivery has been stipulated as best way to deliver live cells to humans for therapy. Colon, in particular, is a part of gastrointestinal (GI) tract that has been proposed to be an oral targeted site. The main objective of these oral therapy procedures is to deliver live cells not only to treat diseases like colorectal cancer, inflammatory bowel disease, and other GI tract diseases like intestinal obstruction and gastritis, but also to deliver therapeutic molecules for overall therapy in various diseases such as renal failure, coronary heart disease, hypertension, and others. This review provides a comprehensive summary of recent advancement in colon targeted live bacterial cell biotherapeutics. Current status of bacterial cell therapy, principles of artificial cells and its potentials in oral delivery of live bacterial cell biotherapeutics for clinical applications as well as biotherapeutic future perspectives are also discussed in our review. PMID:19707368

  3. PRIMARY MARROW DERIVED STROMAL CELLS: ISOLATION AND MANIPULATION

    PubMed Central

    Ramakrishnan, Aravind; Torok-Storb, Beverly; Pillai, Manoj M

    2013-01-01

    Marrow Stromal Cells (MSCs) are relatively rare cells difficult to visualize in marrow biopsies or detect in aspirated marrow. Under specific conditions MSC can be expanded in vitro and the population can give rise to several mesenchymal lineages. “MSC” also refers to mesenchymal stem cells which implies that all cells in the population are multipotent. It is generally agreed that while there may be a few multipotent stem cells in an MSC population the majority are not stem cells. In either case MSC do not produce hematopoietic cells. Although MSCs have been isolated and characterized from several tissues, bone marrow is their most common source for research and clinical use. Primary MSC populations can be derived from bone marrow mononuclear cells with relative ease, but it is important to recognize the cellular heterogeneity within a culture and how this may vary from donor to donor. In this chapter, we will describe methodology to derive primary MSCs from bone marrow screens, an otherwise discarded byproduct of bone marrow harvests used for clinical transplantation. We will also describe some useful techniques to characterize and manipulate MSCs – both primary and immortalized cell lines. PMID:23959984

  4. Primary marrow-derived stromal cells: isolation and manipulation.

    PubMed

    Ramakrishnan, Aravind; Torok-Storb, Beverly; Pillai, Manoj M

    2013-01-01

    Marrow stromal cells (MSCs) are relatively rare cells difficult to visualize in marrow biopsies or detect in aspirated marrow. Under specific conditions MSC can be expanded in vitro and the population can give rise to several mesenchymal lineages. "MSC" also refers to mesenchymal stem cells which implies that all cells in the population are multipotent. It is generally agreed that while there may be a few multipotent stem cells in an MSC population the majority are not stem cells. In either case MSCs do not produce hematopoietic cells. Although MSCs have been isolated and characterized from several tissues, bone marrow is their most common source for research and clinical use. Primary MSC populations can be derived from bone marrow mononuclear cells with relative ease, but it is important to recognize the cellular heterogeneity within a culture and how this may vary from donor to donor. In this chapter, we describe methodology to derive primary MSCs from bone marrow screens, an otherwise discarded by-product of bone marrow harvests used for clinical transplantation. We also describe some useful techniques to characterize and manipulate MSCs-both primary and immortalized cell lines.

  5. Primary cilia mechanics affects cell mechanosensation: A computational study.

    PubMed

    Khayyeri, Hanifeh; Barreto, Sara; Lacroix, Damien

    2015-08-21

    Primary cilia (PC) are mechanical cell structures linked to the cytoskeleton and are central to how cells sense biomechanical signals from their environment. However, it is unclear exactly how PC mechanics influences cell mechanosensation. In this study we investigate how the PC mechanical characteristics are involved in the mechanotransduction process whereby cilium deflection under fluid flow induces strains on the internal cell components that regulate the cell׳s mechanosensitive response. Our investigation employs a computational approach in which a finite element model of a cell consisting of a nucleus, cytoplasm, cortex, microtubules, actin bundles and a primary cilium was used together with a finite element representation of a flow chamber. Fluid-structure interaction analysis was performed by simulating perfusion flow of 1mm/s on the cell model. Simulations of cells with different PC mechanical characteristics, showed that the length and the stiffness of PC are responsible for the transmission of mechanical stimuli to the cytoskeleton. Fluid flow deflects the cilium, with the highest strains found at the base of the PC and in the cytoplasm. The PC deflection created further strains on the cell nucleus but did not influence microtubules and actin bundles significantly. Our results indicate that PC deflection under fluid flow stimulation transmits mechanical strain primarily to other essential organelles in the cytoplasm, such as the Golgi complex, that regulate cells' mechanoresponse. The simulations further suggest that cell mechanosensitivity can be altered by targeting PC length and rigidity.

  6. Filopodial morphology correlates to the capture efficiency of primary T-cells on nanohole arrays.

    PubMed

    Kim, Dong-Joo; Kim, Gil-Sung; Seol, Jin-Kyeong; Hyung, Jung-Hwan; Park, No-Won; Lee, Mi-Ri; Lee, Myung Kyu; Fan, Rong; Lee, Sang-Kwon

    2014-06-01

    Nanostructured surfaces emerge as a new class of material for capture and separation of cell populations including primary immune cells and disseminating rare tumor cells, but the underlying mechanism remains elusive. Although it has been speculated that nanoscale topological structures on cell surface are involved in the cell capture process, there are no studies that systematically analyze the relation between cell surface structures and the capture efficiency. Here we report on the first mechanistic study by quantifying the morphological parameters of cell surface nanoprotrusions, including filopodia, lamellipodia, and microvilli in the early stage of cell capture (< 20 min) in correlation to the efficiency of separating primary T lymphocytes. This was conducted by using a set of nanohole arrays (NHAs) with varying hole and pitch sizes. Our results showed that the formation of filopodia (e.g., width of filopodia and the average number of the filopodial filaments per cell) depends on the feature size of the nanostructures and the cell separation efficiency is strongly correlated to the number of filopodial fibers, suggesting a possible role of early stage mechanosensing and cell spreading in determining the efficiency of cell capture. In contrast, the length of filopodial filaments was less significantly correlated to the cell capture efficiency and the nanostructure dimensions of the NHAs. This is the first mechanistic study on nanostructure-based immune cell capture and provides new insights to not only the biology of cell-nanomaterial interaction but also the design of new rare cell capture technologies with improved efficiency and specificity.

  7. A randomized, controlled trial of disease management modules, including telepsychiatric care, for depression in rural primary care.

    PubMed

    Hilty, Donald M; Marks, Shayna; Wegelin, Jacob; Callahan, Edward J; Nesbitt, Thomas S

    2007-02-01

    Introduction. Disease management modules (DMM), including education, tracking, support, and medical care, have improved health for patients with asthma and diabetes. For rural patients, novel ways of delivery are needed to access clinical expertise from urban or academic specialists. Telemedicine (telephone and televideo) could be instrumental in this process, though no randomized, controlled trials have assessed their effectiveness.Methods. Self-report and structured psychiatric interviews were used to screen potential depressed subjects. Subjects were randomized to: 1) usual care with a DMM using telephone and self-report questionnaires; or 2) a DMM using telephone, questionnaires, and monthly televideo psychiatric consultation emphasizing primary care physician (PCP) skill development. Subjects' depressive symptoms, health status, and satisfaction with care were tabulated at three, six, and 12 months after study entry.Results. There was significant clinical improvement for depression in both groups, with a trend toward significance in the more intensive module. Satisfaction and retention was superior in the more intensive group. There was no overall change in health functioning in either group.Conclusions. Intensive modules using telepsychiatric educational interventions toward PCPs may be superior, but the most critical ingredient may be administrative tracking of patients, prompted intervention by PCPs, and (when necessary) new ideas by a specialist.

  8. A Randomized, Controlled Trial of Disease Management Modules, Including Telepsychiatric Care, for Depression in Rural Primary Care

    PubMed Central

    Marks, Shayna; Wegelin, Jacob; Callahan, Edward J.; Nesbitt, Thomas S.

    2007-01-01

    Introduction. Disease management modules (DMM), including education, tracking, support, and medical care, have improved health for patients with asthma and diabetes. For rural patients, novel ways of delivery are needed to access clinical expertise from urban or academic specialists. Telemedicine (telephone and televideo) could be instrumental in this process, though no randomized, controlled trials have assessed their effectiveness. Methods. Self-report and structured psychiatric interviews were used to screen potential depressed subjects. Subjects were randomized to: 1) usual care with a DMM using telephone and self-report questionnaires; or 2) a DMM using telephone, questionnaires, and monthly televideo psychiatric consultation emphasizing primary care physician (PCP) skill development. Subjects' depressive symptoms, health status, and satisfaction with care were tabulated at three, six, and 12 months after study entry. Results. There was significant clinical improvement for depression in both groups, with a trend toward significance in the more intensive module. Satisfaction and retention was superior in the more intensive group. There was no overall change in health functioning in either group. Conclusions. Intensive modules using telepsychiatric educational interventions toward PCPs may be superior, but the most critical ingredient may be administrative tracking of patients, prompted intervention by PCPs, and (when necessary) new ideas by a specialist. PMID:20805900

  9. Efficient Transformation of Primary Human Mesenchymal Stromal Cells by Adenovirus Early Region 1 Oncogenes

    PubMed Central

    Speiseder, Thomas; Hofmann-Sieber, Helga; Rodríguez, Estefanía; Schellenberg, Anna; Akyüz, Nuray; Dierlamm, Judith; Spruss, Thilo; Lange, Claudia

    2016-01-01

    ABSTRACT Previous observations that human amniotic fluid cells (AFC) can be transformed by human adenovirus type 5 (HAdV-5) E1A/E1B oncogenes prompted us to identify the target cells in the AFC population that are susceptible to transformation. Our results demonstrate that one cell type corresponding to mesenchymal stem/stroma cells (hMSCs) can be reproducibly transformed by HAdV-5 E1A/E1B oncogenes as efficiently as primary rodent cultures. HAdV-5 E1-transformed hMSCs exhibit all properties commonly associated with a high grade of oncogenic transformation, including enhanced cell proliferation, anchorage-independent growth, increased growth rate, and high telomerase activity as well as numerical and structural chromosomal aberrations. These data confirm previous work showing that HAdV preferentially transforms cells of mesenchymal origin in rodents. More importantly, they demonstrate for the first time that human cells with stem cell characteristics can be completely transformed by HAdV oncogenes in tissue culture with high efficiency. Our findings strongly support the hypothesis that undifferentiated progenitor cells or cells with stem cell-like properties are highly susceptible targets for HAdV-mediated cell transformation and suggest that virus-associated tumors in humans may originate, at least in part, from infections of these cell types. We expect that primary hMSCs will replace the primary rodent cultures in HAdV viral transformation studies and are confident that these investigations will continue to uncover general principles of viral oncogenesis that can be extended to human DNA tumor viruses as well. IMPORTANCE It is generally believed that transformation of primary human cells with HAdV-5 E1 oncogenes is very inefficient. However, a few cell lines have been successfully transformed with HAdV-5 E1A and E1B, indicating that there is a certain cell type which is susceptible to HAdV-mediated transformation. Interestingly, all those cell lines have been

  10. Generating induced pluripotent stem cells from common marmoset (Callithrix jacchus) fetal liver cells using defined factors, including Lin28.

    PubMed

    Tomioka, Ikuo; Maeda, Takuji; Shimada, Hiroko; Kawai, Kenji; Okada, Yohei; Igarashi, Hiroshi; Oiwa, Ryo; Iwasaki, Tsuyoshi; Aoki, Mikio; Kimura, Toru; Shiozawa, Seiji; Shinohara, Haruka; Suemizu, Hiroshi; Sasaki, Erika; Okano, Hideyuki

    2010-09-01

    Although embryonic stem (ES) cell-like induced pluripotent stem (iPS) cells have potential therapeutic applications in humans, they are also useful for creating genetically modified human disease models in nonhuman primates. In this study, we generated common marmoset iPS cells from fetal liver cells via the retrovirus-mediated introduction of six human transcription factors: Oct-3/4, Sox2, Klf4, c-Myc, Nanog, and Lin28. Four to five weeks after introduction, several colonies resembling marmoset ES cells were observed and picked for further expansion in ES cell medium. Eight cell lines were established, and validation analyses of the marmoset iPS cells followed. We detected the expression of ES cell-specific surface markers. Reverse transcription-PCR showed that these iPS cells expressed endogenous Oct-3/4, Sox2, Klf4, c-Myc, Nanog and Lin28 genes, whereas all of the transgenes were silenced. Karyotype analysis showed that two of three iPS cell lines retained a normal karyotype after a 2-month culture. Both embryoid body and teratoma formation showed that marmoset iPS cells had the developmental potential to give rise to differentiated derivatives of all three primary germ layers. In summary, we generated marmoset iPS cells via the transduction of six transcription factors; this provides a powerful preclinical model for studies in regenerative medicine.

  11. Primary cutaneous smoldering adult T-cell leukemia/ lymphoma.

    PubMed

    Gittler, Julia; Martires, Kathryn; Terushkin, Vitaly; Brinster, Nooshin; Ramsay, David

    2016-12-15

    HTLV-1 is a virus that is endemic in southwesternJapan and the Caribbean and has been implicatedin the development of ATLL. ATLL, which is anuncommon malignant condition of peripheralT-lymphocytes, is characterized by four clinicalsubtypes, which include acute, lymphomatous,chronic, and smoldering types, that are based onLDH levels, calcium levels, and extent of organinvolvement. We present a 52-year- old woman withpruritic patches with scale on the buttocks and withtender, hyperpigmented macules and papules oftwo-years duration. Histopathologic examinationwas suggestive of mycosis fungoides, laboratoryresults showed HTLV-I and II, and the patient wasdiagnosed with primary cutaneous ATLL. We reviewthe literature on HTLV-1 and ATLL and specifically theprognosis of cutaneous ATLL. The literature suggeststhat a diagnosis of ATLL should be considered amongpatients of Caribbean origin or other endemicareas with skin lesions that suggest a cutaneousT-cell lymphoma, with clinicopathologic features ofmycosis fungoides. Differentiation between ATLLand cutaneous T-cell lymphoma is imperative as theyhave different prognoses and treatment approaches.

  12. Primary cutaneous blastoid mantle cell lymphoma-case report.

    PubMed

    Estrozi, Bruna; Sanches, José A; Varela, Paulo C S; Bacchi, Carlos E

    2009-06-01

    Mantle cell lymphoma (MCL) commonly involves extranodal sites, usually as a manifestation of disseminated disease. In rare cases, MCLs may arise as a primary tumor in the skin. Blastoid mantle cell lymphoma (BV-MCL) is a rare variant and has a more aggressive clinical course. The phenotype of BV-MCL is characterized as CD20+, CD5+, cyclin D1+, CD23-, and CD10-. Interphase fluorescence in situ hybridization shows a characteristic t(11;14) fusion pattern. We report a case of a BV-MCL arising in skin as primary cutaneous MCL with the characteristic immunophenotype and translocation.

  13. Replication of human immunodeficiency virus type 1 in primary dendritic cell cultures.

    PubMed Central

    Langhoff, E; Terwilliger, E F; Bos, H J; Kalland, K H; Poznansky, M C; Bacon, O M; Haseltine, W A

    1991-01-01

    The ability of the human immunodeficiency virus type 1 (HIV-1) to replicate in primary blood dendritic cells was investigated. Dendritic cells compose less than 1% of the circulating leukocytes and are nondividing cells. Highly purified preparations of dendritic cells were obtained using recent advances in cell fractionation. The results of these experiments show that dendritic cells, in contrast to monocytes and T cells, support the active replication of all strains of HIV-1 tested, including T-cell tropic and monocyte/macrophage tropic isolates. The dendritic cell cultures supported much more virus production than did cultures of primary unseparated T cells, CD4+ T cells, and adherent as well as nonadherent monocytes. Replication of HIV-1 in dendritic cells produces no noticeable cytopathic effect nor does it decrease total cell number. The ability of the nonreplicating dendritic cells to support high levels of replication of HIV-1 suggests that this antigen-presenting cell population, which is also capable of supporting clonal T-cell growth, may play a central role in HIV pathogenesis, serving as a source of continued infection of CD4+ T cells and as a reservoir of virus infection. Images PMID:1910172

  14. The classification and diagnostic algorithm for primary lymphatic dysplasia: an update from 2010 to include molecular findings.

    PubMed

    Connell, F C; Gordon, K; Brice, G; Keeley, V; Jeffery, S; Mortimer, P S; Mansour, S; Ostergaard, P

    2013-10-01

    Historically, primary lymphoedema was classified into just three categories depending on the age of onset of swelling; congenital, praecox and tarda. Developments in clinical phenotyping and identification of the genetic cause of some of these conditions have demonstrated that primary lymphoedema is highly heterogenous. In 2010, we introduced a new classification and diagnostic pathway as a clinical and research tool. This algorithm has been used to delineate specific primary lymphoedema phenotypes, facilitating the discovery of new causative genes. This article reviews the latest molecular findings and provides an updated version of the classification and diagnostic pathway based on this new knowledge.

  15. Primary cell cultures of bovine colon epithelium: isolation and cell culture of colonocytes.

    PubMed

    Föllmann, W; Weber, S; Birkner, S

    2000-10-01

    Epithelial cells from bovine colon were isolated by mechanical preparation combined with an enzymatic digestion from colon specimens derived from freshly slaughtered animals. After digestion with collagenase I, the isolated tissue was centrifuged on a 2% D-sorbitol gradient to separate epithelial crypts which were seeded in collagen I-coated culture flasks. By using colon crypts and omitting the seeding of single cells a contamination by fibroblasts was prevented. The cells proliferated under the chosen culture conditions and formed monolayer cultures which were maintained for several weeks, including subcultivation steps. A population doubling time of about 21 hr was estimated in the log phase of the corresponding growth curve. During the culture period the cells were characterized morphologically and enzymatically. By using antibodies against cytokeratine 7 and 13 the isolated cells were identified as cells of epithelial origin. Antibodies against vimentin served as negative control. Morphological features such as microvilli, desmosomes and tight junctions, which demonstrated the ability of the cultured cells to restore an epithelial like monolayer, were shown by ultrastructural investigations. The preservation of the secretory function of the cultured cells was demonstrated by mucine cytochemistry with alcian blue staining. A stable expression of enzyme activities over a period of 6 days in culture occurred for gamma-glutamyltranspeptidase, acid phosphatase and NADH-dehydrogenase activity under the chosen culture conditions. Activity of alkaline phosphatase decreased to about 50% of basal value after 6 days in culture. Preliminary estimations of the metabolic competence of these cells revealed cytochrome P450 1A1-associated EROD activity in freshly isolated cells which was stable over 5 days in cultured cells. Then activity decreased completely. This culture system with primary epithelial cells from the colon will be used further as a model for the colon

  16. Primary cicatricial alopecia: Lymphocytic primary cicatricial alopecias, including chronic cutaneous lupus erythematosus, lichen planopilaris, frontal fibrosing alopecia, and Graham-Little syndrome.

    PubMed

    Bolduc, Chantal; Sperling, Leonard C; Shapiro, Jerry

    2016-12-01

    Both primary and secondary forms of cicatricial alopecia have been described. The hair follicles are the specific target of inflammation in primary cicatricial alopecias. Hair follicles are destroyed randomly with surrounding structures in secondary cicatricial alopecia. This 2-part continuing medical education article will review primary cicatricial alopecias according to the working classification suggested by the North American Hair Research Society. In this classification, the different entities are classified into 3 different groups according to their prominent inflammatory infiltrate (ie, lymphocytic, neutrophilic, and mixed). Part I discusses the following lymphocytic primary cicatricial alopecias: chronic cutaneous lupus erythematosus, lichen planopilaris, frontal fibrosing alopecia, and Graham-Little syndrome. Copyright © 2015 American Academy of Dermatology, Inc. Published by Elsevier Inc. All rights reserved.

  17. Surface topography regulates wnt signaling through control of primary cilia structure in mesenchymal stem cells

    NASA Astrophysics Data System (ADS)

    McMurray, R. J.; Wann, A. K. T.; Thompson, C. L.; Connelly, J. T.; Knight, M. M.

    2013-12-01

    The primary cilium regulates cellular signalling including influencing wnt sensitivity by sequestering β-catenin within the ciliary compartment. Topographic regulation of intracellular actin-myosin tension can control stem cell fate of which wnt is an important mediator. We hypothesized that topography influences mesenchymal stem cell (MSC) wnt signaling through the regulation of primary cilia structure and function. MSCs cultured on grooves expressed elongated primary cilia, through reduced actin organization. siRNA inhibition of anterograde intraflagellar transport (IFT88) reduced cilia length and increased active nuclear β-catenin. Conversely, increased primary cilia assembly in MSCs cultured on the grooves was associated with decreased levels of nuclear active β-catenin, axin-2 induction and proliferation, in response to wnt3a. This negative regulation, on grooved topography, was reversed by siRNA to IFT88. This indicates that subtle regulation of IFT and associated cilia structure, tunes the wnt response controlling stem cell differentiation.

  18. Surface topography regulates wnt signaling through control of primary cilia structure in mesenchymal stem cells.

    PubMed

    McMurray, R J; Wann, A K T; Thompson, C L; Connelly, J T; Knight, M M

    2013-12-18

    The primary cilium regulates cellular signalling including influencing wnt sensitivity by sequestering β-catenin within the ciliary compartment. Topographic regulation of intracellular actin-myosin tension can control stem cell fate of which wnt is an important mediator. We hypothesized that topography influences mesenchymal stem cell (MSC) wnt signaling through the regulation of primary cilia structure and function. MSCs cultured on grooves expressed elongated primary cilia, through reduced actin organization. siRNA inhibition of anterograde intraflagellar transport (IFT88) reduced cilia length and increased active nuclear β-catenin. Conversely, increased primary cilia assembly in MSCs cultured on the grooves was associated with decreased levels of nuclear active β-catenin, axin-2 induction and proliferation, in response to wnt3a. This negative regulation, on grooved topography, was reversed by siRNA to IFT88. This indicates that subtle regulation of IFT and associated cilia structure, tunes the wnt response controlling stem cell differentiation.

  19. Surface topography regulates wnt signaling through control of primary cilia structure in mesenchymal stem cells

    PubMed Central

    McMurray, R. J.; Wann, A. K. T.; Thompson, C. L.; Connelly, J. T.; Knight, M. M.

    2013-01-01

    The primary cilium regulates cellular signalling including influencing wnt sensitivity by sequestering β-catenin within the ciliary compartment. Topographic regulation of intracellular actin-myosin tension can control stem cell fate of which wnt is an important mediator. We hypothesized that topography influences mesenchymal stem cell (MSC) wnt signaling through the regulation of primary cilia structure and function. MSCs cultured on grooves expressed elongated primary cilia, through reduced actin organization. siRNA inhibition of anterograde intraflagellar transport (IFT88) reduced cilia length and increased active nuclear β-catenin. Conversely, increased primary cilia assembly in MSCs cultured on the grooves was associated with decreased levels of nuclear active β-catenin, axin-2 induction and proliferation, in response to wnt3a. This negative regulation, on grooved topography, was reversed by siRNA to IFT88. This indicates that subtle regulation of IFT and associated cilia structure, tunes the wnt response controlling stem cell differentiation. PMID:24346024

  20. Culturing primary rat inner medullary collecting duct cells.

    PubMed

    Faust, Dörte; Geelhaar, Andrea; Eisermann, Beate; Eichhorst, Jenny; Wiesner, Burkhard; Rosenthal, Walter; Klussmann, Enno; Klussman, Enno

    2013-06-21

    Arginine-vasopressin (AVP) facilitates water reabsorption by renal collecting duct principal cells and thereby fine-tunes body water homeostasis. AVP binds to vasopressin V2 receptors (V2R) on the surface of the cells and thereby induces synthesis of cAMP. This stimulates cellular signaling processes leading to changes in the phosphorylation of the water channel aquaporin-2 (AQP2). Protein kinase A phoshorylates AQP2 and thereby triggers the translocation of AQP2 from intracellular vesicles into the plasma membrane facilitating water reabsorption from primary urine. Aberrations of AVP release from the pituitary or AVP-activated signaling in principal cells can cause central or nephrogenic diabetes insipidus, respectively; an elevated blood plasma AVP level is associated with cardiovascular diseases such as chronic heart failure and the syndrome of inappropriate antidiuretic hormone secretion. Here, we present a protocol for cultivation of primary rat inner medullary collecting duct (IMCD) cells, which express V2R and AQP2 endogenously. The cells are suitable for elucidating molecular mechanisms underlying the control of AQP2 and thus to discover novel drug targets for the treatment of diseases associated with dysregulation of AVP-mediated water reabsorption. IMCD cells are obtained from rat renal inner medullae and are used for experiments six to eight days after seeding. IMCD cells can be cultured in regular cell culture dishes, flasks and micro-titer plates of different formats, the procedure only requires a few hours, and is appropriate for standard cell culture laboratories.

  1. Culturing Primary Rat Inner Medullary Collecting Duct Cells

    PubMed Central

    Faust, Dörte; Geelhaar, Andrea; Eisermann, Beate; Eichhorst, Jenny; Wiesner, Burkhard; Rosenthal, Walter; Klussman, Enno

    2013-01-01

    Arginine-vasopressin (AVP) facilitates water reabsorption by renal collecting duct principal cells and thereby fine-tunes body water homeostasis. AVP binds to vasopressin V2 receptors (V2R) on the surface of the cells and thereby induces synthesis of cAMP. This stimulates cellular signaling processes leading to changes in the phosphorylation of the water channel aquaporin-2 (AQP2). Protein kinase A phoshorylates AQP2 and thereby triggers the translocation of AQP2 from intracellular vesicles into the plasma membrane facilitating water reabsorption from primary urine. Aberrations of AVP release from the pituitary or AVP-activated signaling in principal cells can cause central or nephrogenic diabetes insipidus, respectively; an elevated blood plasma AVP level is associated with cardiovascular diseases such as chronic heart failure and the syndrome of inappropriate antidiuretic hormone secretion. Here, we present a protocol for cultivation of primary rat inner medullary collecting duct (IMCD) cells, which express V2R and AQP2 endogenously. The cells are suitable for elucidating molecular mechanisms underlying the control of AQP2 and thus to discover novel drug targets for the treatment of diseases associated with dysregulation of AVP-mediated water reabsorption. IMCD cells are obtained from rat renal inner medullae and are used for experiments six to eight days after seeding. IMCD cells can be cultured in regular cell culture dishes, flasks and micro-titer plates of different formats, the procedure only requires a few hours, and is appropriate for standard cell culture laboratories. PMID:23852264

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

    PubMed

    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.

  3. The Primary Cilium in Cell Signaling and Cancer

    SciTech Connect

    Michaud III, Edward J; Yoder, Bradley

    2006-01-01

    The primary cilium is a microtubule-based antenna-like structure that emanates from the surface of virtually all cells in the mammalian body. It is anchored to the cell by the basal body, which develops from the mother centriole of the centrosome in a manner that is coordinately regulated with the cell cycle. The primary cilium is a sensory organelle that receives both mechanical and chemical signals from other cells and the environment, and transmits these signals to the nucleus to elicit a cellular response. Recent studies revealed that multiple components of the Sonic hedgehog and plateletderived growth factor receptor-A signal transduction pathways localize to the primary cilium, and that loss of the cilium blocks ligand-induced signaling by both pathways. In light of the major role that these pathways play in numerous types of cancer, we anticipate that the emerging discoveries being made about the function of the primary cilium in signaling pathways that are critical for embryonic development and tissue homeostasis in adults will also provide novel insights into the molecular mechanisms of carcinogenesis. (Cancer Res 2006; 66 13): 6463-7)

  4. High rate of detection of primary aldosteronism, including surgically treatable forms, after 'non-selective' screening of hypertensive patients.

    PubMed

    Stowasser, Michael; Gordon, Richard D; Gunasekera, Thanuja G; Cowley, Diane C; Ward, Gregory; Archibald, Colin; Smithers, B Mark

    2003-11-01

    Wide testing of the aldosterone : renin ratio among hypertensive individuals has revealed primary aldosteronism to be common, with most patients normokalaemic. Some investigators, however, have reported aldosterone-producing adenoma to be rare among patients so detected. To test the hypothesis that differences among reported studies in the rate of detection of aldosterone-producing adenoma (as opposed to bilateral adrenal hyperplasia) reflect differences in the procedures used for diagnosis of primary aldosteronism, and the methods used to identify aldosterone-producing adenomas. In the newly established Princess Alexandra Hospital Hypertension Unit (PAHHU), we used procedures developed by Greenslopes Hospital Hypertension Unit (which reports that more than 30% of patients with primary aldosteronism have aldosterone-producing adenomas) to diagnose primary aldosteronism and determine the subtype. All patients with an increased aldosterone : renin ratio (measured after correction for hypokalaemia and while the patient was not receiving interfering medications) underwent fludrocortisone suppression testing to confirm or exclude primary aldosteronism; if they were positive, they underwent genetic testing to exclude glucocorticoid-remediable aldosteronism before adrenal venous sampling was used to differentiate lateralizing from bilateral primary aldosteronism. This approach allowed PAHHU to diagnose, within 2 years, 54 patients [only seven (13%) hypokalaemic] with primary aldosteronism. All tested negative for glucocorticoid-remediable aldosteronism. Aldosterone production was lateralized to one adrenal in 15 patients (31%; only six hypokalaemic) and was bilateral in 34 (69%; all normokalaemic) of 49 patients who underwent adrenal venous sampling. Among patients with lateralizing adrenal hyperplasia, computed tomography revealed an ipsilateral mass in only six and a contralateral lesion in one. Fourteen patients underwent unilateral adrenalectomy, which cured the

  5. Arsenic is Cytotoxic and Genotoxic to Primary Human Lung Cells

    PubMed Central

    Xie, Hong; Huang, ShouPing; Martin, Sarah; Wise, John P.

    2014-01-01

    Arsenic originates from both geochemical and numerous anthropogenic activities. Exposure of the general public to significant levels of arsenic is widespread. Arsenic is a well-documented human carcinogen. Long-term exposure to high levels of arsenic in drinking water have been linked to bladder, lung, kidney, liver, prostate, and skin cancer. Among them, lung cancer is of great public concern. However, little is known about how arsenic causes lung cancer and few studies have considered effects in normal human lung cells. The purpose of this study was to determine the cytotoxicity and genotoxicity of arsenic in human primary bronchial fibroblast and epithelial cells. Our data show that arsenic induces a concentration-dependent decrease in cell survival after short (24 h) or long (120 h) exposures. Arsenic induces concentration-dependent but not time-dependent increases in chromosome damage in fibroblasts. No chromosome damage is induced after either 24 h or 120 h arsenic exposure in epithelial cells. Using neutral comet assay and gamma-H2A.X foci forming assay, we found that 24 h or 120 h exposure to arsenic induces increases in DNA double strand breaks in both cell lines. These data indicate that arsenic is cytotoxic and genotoxic to human lung primary cells but lung fibroblasts are more sensitive to arsenic than epithelial cells. Further research is needed to understand the specific mechanisms involved in arsenic-induced genotoxicity in human lung cells. PMID:24291234

  6. Lysis of primary hepatic tumours by lymphokine activated killer cells.

    PubMed Central

    Hsieh, K H; Shu, S Y; Lee, C S; Chu, C T; Yang, C S; Chang, K J

    1987-01-01

    Lymphokine activated killer cell is a newly described lytic system against a variety of solid tumours and is distinct in several respects from the classic cytolytic T cell and the natural killer systems. This study was conducted to evaluate the lytic activity of lymphokine activated killer cells against fresh autologous and allogeneic, as well as cultured hepatocellular carcinoma cells. Lymphokine activated killer cell was generated by incubating peripheral blood mononuclear cells with various concentrations of recombinant IL-2 (rIL-2, Cetus, USA) for various periods of time. A four hour 51Cr release assay was used to measure cytotoxicity. The results show that fresh and cultured hepatocellular carcinoma cells were only slightly susceptible to natural killer cells. Normal hepatocytes were resistant to lymphokine activated killer-mediated lysis. Lymphokine activated killer cells could be generated from mononuclear cells of hepatocellular carcinoma patients and normal subjects with lytic activity against fresh autologous and allogeneic and cultured hepatocellular carcinoma cells, but lymphokine activated killer cells from the former was less efficient than that from the latter. It is concluded that the adoptive immunotherapy with combined rIL-2 and lymphokine activated killer may be worth trying in early cases of primary hepatocellular carcinoma. PMID:3030899

  7. Pulp tissue from primary teeth: new source of stem cells

    PubMed Central

    TELLES, Paloma Dias; MACHADO, Maria Aparecida de Andrade Moreira; SAKAI, Vivien Thiemy; NÖR, Jacques Eduardo

    2011-01-01

    SHED (stem cells from human exfoliated deciduous teeth) represent a population of postnatal stem cells capable of extensive proliferation and multipotential differentiation. Primary teeth may be an ideal source of postnatal stem cells to regenerate tooth structures and bone, and possibly to treat neural tissue injury or degenerative diseases. SHED are highly proliferative cells derived from an accessible tissue source, and therefore hold potential for providing enough cells for clinical applications. In this review, we describe the current knowledge about dental pulp stem cells and discuss tissue engineering approaches that use SHED to replace irreversibly inflamed or necrotic pulps with a healthy and functionally competent tissue that is capable of forming new dentin. PMID:21625731

  8. Extension to PV OPTICS to include front electrode design in solar cells

    NASA Astrophysics Data System (ADS)

    Guhabiswas, Debraj

    Proper optical designing of solar cells and modules is of paramount importance towards achieving high photovoltaic conversion efficiencies. Modeling softwares such as PV OPTICS, BIRANDY and SUNRAYS have been created to aid such optical designing of cells and modules; but none of these modeling packages take the front metal electrode architecture of a solar cell into account. A new model, has been developed to include the front metal electrode architecture to finished solar cells for optical calculations. This has been implemented in C++ in order to add a new module to PV OPTICS (NREL's photovoltaic modeling tool) to include front metallization patterns for optical design and simulation of solar cells. This new addition also calculates the contribution of light that diffuses out of the illuminated (non-metallized) regions to the solar cell current. It also determines the optical loss caused by the absorption in the front metal and separates metallic losses due to front and back contacts. This added capability also performs the following functions: • calculates the total current that can be generated in a solar cell due to optical absorption in each region, including the region beneath the front metal electrodes for the radiation spectrum of AM 1.5, • calculates various losses in the solar cell due to front electrode shading, metal absorption, and reflectance, • makes a plot of how light is absorbed in the metal as well as silicon under the shaded region in the solar cell. Although Finite Difference Time Domain (FDTD) is the numerical technique of choice to solve Maxwell's equations for a propagating electromagnetic wave, it is both time consuming and very demanding on the computer processors. Furthermore, for complicated geometric structures, FDTD poses various limitations. Hence, ray tracing has been chosen as the means of implementing this new model. This new software has been used to carry out a detailed investigation on the effect of various parameters of

  9. Time course of programmed cell death, which included autophagic features, in hybrid tobacco cells expressing hybrid lethality.

    PubMed

    Ueno, Naoya; Nihei, Saori; Miyakawa, Naoto; Hirasawa, Tadashi; Kanekatsu, Motoki; Marubashi, Wataru; van Doorn, Wouter G; Yamada, Tetsuya

    2016-12-01

    PCD with features of vacuolar cell death including autophagy-related features were detected in hybrid tobacco cells, and detailed time course of features of vacuolar cell death were established. A type of interspecific Nicotiana hybrid, Nicotiana suaveolens × N. tabacum exhibits temperature-sensitive lethality. This lethality results from programmed cell death (PCD) in hybrid seedlings, but this PCD occurs only in seedlings and suspension-cultured cells grown at 28 °C, not those grown at 36 °C. Plant PCD can be classified as vacuolar cell death or necrotic cell death. Induction of autophagy, vacuolar membrane collapse and actin disorganization are each known features of vacuolar cell death, but observed cases of PCD showing all these features simultaneously are rare. In this study, these features of vacuolar cell death were evident in hybrid tobacco cells expressing hybrid lethality. Ion leakage, plasma membrane disruption, increased activity of vacuolar processing enzyme, vacuolar membrane collapse, and formation of punctate F-actin foci were each evident in these cells. Transmission electron microscopy revealed that macroautophagic structures formed and tonoplasts ruptured in these cells. The number of cells that contained monodansylcadaverine (MDC)-stained structures and the abundance of nine autophagy-related gene transcripts increased just before cell death at 28 °C; these features were not evident at 36 °C. We assessed whether an autophagic inhibitor, wortmannin (WM), influenced lethality in hybrid cells. After the hybrid cell began to die, WM suppressed increases in ion leakage and cell deaths, and it decreased the number of cells containing MDC-stained structures. These results showed that several features indicative of autophagy and vacuolar cell death were evident in the hybrid tobacco cells subject to lethality. In addition, we documented a detailed time course of these vacuolar cell death features.

  10. Nonhematopoietic cells are the primary source of bone marrow-derived lung epithelial cells.

    PubMed

    Kassmer, Susannah H; Bruscia, Emanuela M; Zhang, Ping-Xia; Krause, Diane S

    2012-03-01

    Previous studies have demonstrated that bone marrow (BM)-derived cells differentiate into nonhematopoietic cells of multiple tissues. To date, it remains unknown which population(s) of BM cells are primarily responsible for this engraftment. To test the hypothesis that nonhematopoietic stem cells in the BM are the primary source of marrow-derived lung epithelial cells, either wild-type hematopoietic or nonhematopoietic BM cells were transplanted into irradiated surfactant-protein-C (SPC)-null mice. Donor-derived, SPC-positive type 2 pneumocytes were predominantly detected in the lungs of mice receiving purified nonhematopoietic cells and were absent from mice receiving purified hematopoietic stem and progenitor cells. We conclude that cells contained in the nonhematopoietic fraction of the BM are the primary source of marrow-derived lung epithelial cells. These nonhematopoietic cells may represent a primitive stem cell population residing in adult BM.

  11. Nonhematopoietic Cells are the Primary Source of Bone Marrow-Derived Lung Epithelial Cells

    PubMed Central

    Kassmer, Susannah H.; Bruscia, Emanuela M.; Zhang, Ping-Xia; Krause, Diane S.

    2013-01-01

    Previous studies have demonstrated that bone marrow (BM)-derived cells differentiate into nonhematopoietic cells of multiple tissues. To date, it remains unknown which population(s) of BM cells are primarily responsible for this engraftment. To test the hypothesis that nonhematopoietic stem cells in the BM are the primary source of marrow-derived lung epithelial cells, either wild-type hematopoietic or nonhematopoietic BM cells were transplanted into irradiated surfactant-protein-C (SPC)-null mice. Donor-derived, SPC-positive type 2 pneumocytes were predominantly detected in the lungs of mice receiving purified nonhematopoietic cells and were absent from mice receiving purified hematopoietic stem and progenitor cells. We conclude that cells contained in the nonhematopoietic fraction of the BM are the primary source of marrow-derived lung epithelial cells. These nonhematopoietic cells may represent a primitive stem cell population residing in adult BM. PMID:22162244

  12. Uncorrectable ptosis: primary cutaneous signet-ring cell carcinoma.

    PubMed

    Hansen, Mark S; Chi, Sulene L; Cummings, Thomas; Woodward, Julie A

    2013-09-14

    Primary cutaneous signet-ring cell carcinoma (PCSRCC) is a rare but aggressive tumor. Our case highlights a 60-year-old man who presented with eyelid ptosis, for which he underwent multiple surgical procedures over a 3-year period prior to referral to our clinic. These procedures were complicated by scarring, delayed healing, and poor cosmetic outcome. In addition, the patient was noted to develop progressive enophthalmos. These concerning signs led to a CT scan and subsequent eyelid biopsy, which revealed a diagnosis of PCSRCC. Further management has involved an MRI and orbitotomy with biopsy revealing widespread extension of the carcinoma. Exenteration was performed to reduce the likelihood of metastasis. There are few documented case reports of PCSRCC of the eyelid in the literature. Of the 33 published cases of PCSRCC, 27 cases involve the eyelids and the other 6 cases involve the axilla. The unique clinical features of this case will be discussed, in particular the presentation as ptosis, an otherwise commonplace complaint in the oculoplastics clinic. The surgical course and histopathologic findings will be presented. The literature regarding PCSRCC will be reviewed including demographics, management, and prognosis. Although rare, PCSRCC follows an aggressive course with characteristically delayed diagnosis. Early identification and treatment likely offer a better prognosis. Thus, description of the clinical presentation of this rare tumor may aid in recognition and earlier treatment.

  13. The cellular and molecular toxicity of lead in primary and clonal osteoblastic bone cells

    SciTech Connect

    Long, G.J.

    1989-01-01

    First, steady state kinetic models of lead metabolism and calcium homeostasis were developed in both primary and clonal osteoblastic bone cells. Secondly, the effect of lead on cellular calcium homeostasis was determined. Finally, the effect of lead on 1,25 (OH){sub 2}D{sub 3} induced production of osteocalcin, a protein synthesized and secreted by osteoblasts, was investigated. Lead metabolism in osteoblastic bone cells was characterized by three intracellular pools. The largest of these, S{sub 3}, included mitochondrial lead and accounted for 70 percent of total cell lead in primary osteoblastic bone cells and 85 percent of total lead in clonal osteoblastic bone cells. None of the kinetic pools were saturated at lead concentrations up to 100 {mu}M lead. Calcium homeostasis in osteoblastic bone cells was also described by a three compartment, intracellular kinetic model.

  14. The Challenges of Implementing Group Work in Primary School Classrooms and Including Pupils with Special Educational Needs

    ERIC Educational Resources Information Center

    Baines, Ed; Blatchford, Peter; Webster, Rob

    2015-01-01

    Findings from two studies are discussed in relation to the experiences and challenges faced by teachers trying to implement effective group work in schools and classrooms and to reflect on the lessons learnt about how to involve pupils with special educational needs (SEN). The first study reports on UK primary school teachers' experiences of…

  15. The Challenges of Implementing Group Work in Primary School Classrooms and Including Pupils with Special Educational Needs

    ERIC Educational Resources Information Center

    Baines, Ed; Blatchford, Peter; Webster, Rob

    2015-01-01

    Findings from two studies are discussed in relation to the experiences and challenges faced by teachers trying to implement effective group work in schools and classrooms and to reflect on the lessons learnt about how to involve pupils with special educational needs (SEN). The first study reports on UK primary school teachers' experiences of…

  16. Satellite Cell Functional Alterations Following Cutaneous Burn in rats Include an Increase in Their Osteogenic Potential

    DTIC Science & Technology

    2013-10-07

    have yet to be identified. Interestingly, a therapeutic strategy for the treatment of HO for both nerve injury and burn is radiation , a methodology...Satellite cell functional alterations following cutaneous burn in rats include an increase in their osteogenic potential Xiaowu Wu, MD,* and...Skeletal muscle Muscle precursor cell Thermal injury Atrophy Heterotopic ossification a b s t r a c t Background: Significant consequences of severe burn

  17. Primary culture of glial cells from mouse sympathetic cervical ganglion: a valuable tool for studying glial cell biology.

    PubMed

    de Almeida-Leite, Camila Megale; Arantes, Rosa Maria Esteves

    2010-12-15

    Central nervous system glial cells as astrocytes and microglia have been investigated in vitro and many intracellular pathways have been clarified upon various stimuli. Peripheral glial cells, however, are not as deeply investigated in vitro despite its importance role in inflammatory and neurodegenerative diseases. Based on our previous experience of culturing neuronal cells, our objective was to standardize and morphologically characterize a primary culture of mouse superior cervical ganglion glial cells in order to obtain a useful tool to study peripheral glial cell biology. Superior cervical ganglia from neonatal C57BL6 mice were enzymatically and mechanically dissociated and cells were plated on diluted Matrigel coated wells in a final concentration of 10,000cells/well. Five to 8 days post plating, glial cell cultures were fixed for morphological and immunocytochemical characterization. Glial cells showed a flat and irregular shape, two or three long cytoplasm processes, and round, oval or long shaped nuclei, with regular outline. Cell proliferation and mitosis were detected both qualitative and quantitatively. Glial cells were able to maintain their phenotype in our culture model including immunoreactivity against glial cell marker GFAP. This is the first description of immunocytochemical characterization of mouse sympathetic cervical ganglion glial cells in primary culture. This work discusses the uses and limitations of our model as a tool to study many aspects of peripheral glial cell biology.

  18. Fabrication of contacts for silicon solar cells including printing burn through layers

    DOEpatents

    Ginley, David S; Kaydanova, Tatiana; Miedaner, Alexander; Curtis, Calvin J; Van Hest, Marinus Franciscus Antonius Maria

    2014-06-24

    A method for fabricating a contact (240) for a solar cell (200). The method includes providing a solar cell substrate (210) with a surface that is covered or includes an antireflective coating (220). For example, the substrate (210) may be positioned adjacent or proximate to an outlet of an inkjet printer (712) or other deposition device. The method continues with forming a burn through layer (230) on the coating (220) by depositing a metal oxide precursor (e.g., using an inkjet or other non-contact printing method to print or apply a volume of liquid or solution containing the precursor). The method includes forming a contact layer (240) comprising silver over or on the burn through layer (230), and then annealing is performed to electrically connect the contact layer (240) to the surface of the solar cell substrate (210) through a portion of the burn through layer (230) and the coating (220).

  19. Primary non-hodgkin B cell lymphoma in a man.

    PubMed

    Alhabshi, Sh M I; Ismail, Z; Arasaratnam, Sh A

    2011-03-01

    Malignant breast lymphoma is a rare condition and primary breast lymphoma is extremely rare in the male population. We present a case of a 26-year-old man (transgender) who presented with a large palpable mass in the right breast. This mass was rapidly growing in size associated with right axillary lymphadenopathy. Ultrasound and MRI findings were consistent with BIRADS IV lesion which was suspicious of malignancy. Core biopsy was performed and histopathology confirmed the diagnosis of primary non Hodgkin B cell lymphoma of the breast.

  20. Combining a BCL2 inhibitor with the retinoid derivative fenretinide targets melanoma cells including melanoma initiating cells.

    PubMed

    Mukherjee, Nabanita; Reuland, Steven N; Lu, Yan; Luo, Yuchun; Lambert, Karoline; Fujita, Mayumi; Robinson, William A; Robinson, Steven E; Norris, David A; Shellman, Yiqun G

    2015-03-01

    Investigations from multiple laboratories support the existence of melanoma initiating cells (MICs) that potentially contribute to melanoma's drug resistance. ABT-737, a small molecule BCL-2/BCL-XL/BCL-W inhibitor, is promising in cancer treatments, but not very effective against melanoma, with the antiapoptotic protein MCL-1 as the main contributor to resistance. The synthetic retinoid fenretinide N-(4-hydroxyphenyl)retinamide (4-HPR) has shown promise for treating breast cancers. Here, we tested whether the combination of ABT-737 with 4-HPR is effective in killing both the bulk of melanoma cells and MICs. The combination synergistically decreased cell viability and caused cell death in multiple melanoma cells lines (carrying either BRAF or NRAS mutations) but not in normal melanocytes. The combination increased the NOXA expression and caspase-dependent MCL-1 degradation. Knocking down NOXA protected cells from combination-induced apoptosis, implicating the role of NOXA in the drug synergy. The combination treatment also disrupted primary spheres (a functional assay for MICs) and decreased the percentage of aldehyde dehydrogenase (high) cells (a marker of MICs) in melanoma cell lines. Moreover, the combination inhibited the self-renewal capacity of MICs, measured by secondary sphere-forming assays. In vivo, the combination inhibited tumor growth. Thus, this combination is a promising treatment strategy for melanoma, regardless of mutation status of BRAF or NRAS.

  1. Cytotoxicity and accumulation of ergot alkaloids in human primary cells.

    PubMed

    Mulac, Dennis; Humpf, Hans-Ulrich

    2011-04-11

    Ergot alkaloids are secondary metabolites produced by fungi of the species Claviceps. Toxic effects after consumption of contaminated grains are described since mediaeval times. Of the more than 40 known ergot alkaloids six are found predominantly. These are ergotamine, ergocornine, ergocryptine, ergocristine, ergosine and ergometrine, along with their corresponding isomeric forms (-inine-forms). Toxic effects are known to be induced by an interaction of the ergot alkaloids as neurotransmitters, like dopamine or serotonin. Nevertheless data concerning cytotoxic effects are missing and therefore a screening of the six main ergot alkaloids was performed in human primary cells in order to evaluate the toxic potential. As it is well known that ergot alkaloids isomerize easily the stability was tested in the cell medium. Based on these results factors were calculated to correct the used concentration values to the biologically active lysergic (-ine) form. These factors range from 1.4 for the most stable compound ergometrine to 5.0 for the most unstable ergot alkaloid ergocristine. With these factors, reflecting the instability, several controverse literature data concerning the toxicity could be explained. To evaluate the cytotoxic effects of ergot alkaloids, human cells in primary culture were used. These cells remain unchanged in contrast to cell lines and the data allow a better comparison to the in vivo situation than using immortalized cell lines. To characterize the effects on primary cells, renal proximal tubule epithelial cells (RPTEC) and normal human astrocytes (NHA) were used. The parameters necrosis (LDH-release) and apoptosis (caspase-3-activation, DNA condensation and fragmentation) were distinguished. The results show that depending on the individual structure of the peptide ergot alkaloids the toxic properties change. While ergometrine as a lysergic acid amide did not show any effect, the peptide ergot alkaloids revealed a different toxic potential. Of

  2. Second primary germ cell tumors in patients with seminoma of the testis.

    PubMed

    Cockburn, A G; Vugrin, D; Batata, M; Hajdu, S; Whitmore, W F

    1983-08-01

    In a review of our experience with seminoma 9 cases of bilateral primary testis germ cell tumors were encountered, including 2 simultaneous and 7 successive. Of the 9 cases 6 were bilateral seminomas and 7 were stage I, contributing to the good survival experience. Treatment policy is specified and discussed.

  3. The Histone Demethylase Jumonji Coordinates Cellular Senescence Including Secretion of Neural Stem Cell-attracting Cytokines

    PubMed Central

    Perrigue, Patrick M.; Silva, Michael E.; Warden, Charles D.; Feng, Nathan L.; Reid, Michael A.; Mota, Daniel J.; Joseph, Lauren P.; Tian, Yangzi Isabel; Glackin, Carlotta A.; Gutova, Margarita; Najbauer, Joseph; Aboody, Karen S.; Barish, Michael E.

    2016-01-01

    Jumonji domain-containing protein 3 (JMJD3/KDM6B) demethylates lysine 27 on histone H3 (H3K27me3), a repressive epigenetic mark controlling chromatin organization and cellular senescence. To better understand the functional consequences of JMJD3 its expression was investigated in brain tumor cells. Querying patient expression profile databases confirmed JMJD3 over-expression in high-grade glioma. Immunochemical staining of two glioma cell lines, U251 and U87, indicated intrinsic differences in JMJD3 expression levels that were reflected in changes in cell phenotype and variations associated with cellular senescence, including senescence-associated β-galactosidase (SA-β-gal) activity and the senescence associated secretory phenotype (SASP). Over-expressing wild type JMJD3 (JMJD3wt) activated SASP-associated genes, enhanced SA-βgal activity, and induced nuclear blebbing. Conversely, over-expression of a catalytically inactive dominant negative mutant JMJD3 (JMJD3mut) increased proliferation. In addition, a large number of transcripts were identified by RNA-seq as altered in JMJD3 over-expressing cells, including cancer- and inflammation-related transcripts as defined by IPA analysis. These results suggest that expression of the SASP in the context of cancer undermines normal tissue homeostasis and contributes to tumorigenesis and tumor progression. These studies are therapeutically relevant because inflammatory cytokines have been linked to homing of neural stem cells and other stem cells to tumor loci. PMID:25652587

  4. Primary antitumor immune response mediated by CD4+ T cells.

    PubMed

    Corthay, Alexandre; Skovseth, Dag K; Lundin, Katrin U; Røsjø, Egil; Omholt, Hilde; Hofgaard, Peter O; Haraldsen, Guttorm; Bogen, Bjarne

    2005-03-01

    Gene-targeted mice have recently revealed a role for lymphocytes and interferon-gamma (IFNgamma) in conferring protection against cancer, but the mechanisms remain unclear. Here, we have characterized a successful primary antitumor immune response initiated by naive CD4+ T cells. Major histocompatibility complex class II (MHC-II)-negative myeloma cells injected subcutaneously into syngeneic mice were surrounded within 3 days by macrophages that captured tumor antigens. Within 6 days, naive myeloma-specific CD4+ T cells became activated in draining lymph nodes and subsequently migrated to the incipient tumor site. Upon recognition of tumor-derived antigenic peptides presented on MHC-II by macrophages, the myeloma-specific CD4+ T cells were reactivated and started to secrete cytokines. T cell-derived IFNgamma activated macrophages in close proximity to the tumor cells. Tumor cell growth was completely inhibited by such locally activated macrophages. These data indicate a mechanism for immunosurveillance of MHC-II-negative cancer cells by tumor-specific CD4+ T cells through collaboration with macrophages.

  5. Universal cell frame for high-pressure water electrolyzer and electrolyzer including the same

    DOEpatents

    Schmitt, Edwin W.; Norman, Timothy J.

    2013-01-08

    Universal cell frame generic for use as an anode frame and as a cathode frame in a water electrolyzer. According to one embodiment, the universal cell frame includes a unitary annular member having a central opening. Four trios of transverse openings are provided in the annular member, each trio being spaced apart by about 90 degrees. A plurality of internal radial passageways fluidly interconnect the central opening and each of the transverse openings of two diametrically-opposed trios of openings, the other two trios of openings lacking corresponding radial passageways. Sealing ribs are provided on the top and bottom surfaces of the annular member. The present invention is also directed at a water electrolyzer that includes two such cell frames, one being used as the anode frame and the other being used as the cathode frame, the cathode frame being rotated 90 degrees relative to the anode frame.

  6. A&M. TAN607. Special service cubicle (hot cell). Details include Zpipe ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    A&M. TAN-607. Special service cubicle (hot cell). Details include Z-pipe and stepped plug penetrations through shielding wall. Ralph M. Parsons 902-3-ANP-607-A116. Date: December 1952. Approved by INEEL Classification Office for public release. INEEL index code no. 034-0607-693-106767 - Idaho National Engineering Laboratory, Test Area North, Scoville, Butte County, ID

  7. Primary Mediastinal Large B-cell Lymphoma Exhibiting Endobronchial Involvement

    PubMed Central

    Shimada, Midori; Fukuda, Minoru; Horio, Kensuke; Suyama, Takayuki; Kitazaki, Takeshi; Hashiguchi, Kohji; Fukuda, Masaaki; Shigematsu, Kazuto; Nakamura, Yoichi; Honda, Takuya; Ashizawa, Kazuto; Mukae, Hiroshi

    2016-01-01

    Primary mediastinal large B-cell lymphoma (PMLBCL) is one of the subtypes of diffuse large B-cell lymphoma. We experienced a rare case of PMLBCL that exhibited endobronchial involvement. A 33-year-old Japanese female with the chief complaints of epigastralgia, back pain, and nausea visited a primary care hospital. Computed tomography of the chest and abdomen demonstrated a bulky mass in the left anterior mediastinum, multiple pulmonary nodules, axillary lymph node swelling, and a pancreatic tumor. Fiberoptic bronchoscopy showed a white-tinged irregularly shaped endobronchial tumor accompanied by capillary vessel dilation in the left upper lobar bronchus. Taken together, these findings resulted in a diagnosis of PMLBCL. PMID:27803409

  8. Monocyte-derived factors including PLA2G7 induced by macrophage-nasopharyngeal carcinoma cell interaction promote tumor cell invasiveness

    PubMed Central

    Low, Heng Boon; Png, Chin Wen; Li, Chunwei; Wang, De Yun; Wong, Soon Boon Justin; Zhang, Yongliang

    2016-01-01

    The non-keratinizing undifferentiated subtype of nasopharyngeal carcinoma (NPC) is a malignancy characterized by an intimate relationship between neoplastic cells and a non-neoplastic lymphoid component. Tumor-associated macrophages (TAMs) foster tumor progression through production of soluble mediators that support proliferation, angiogenesis, survival and invasion of malignant cells. However, the role of macrophages in the progression of NPC remains poorly understood. This study aims to investigate the functional and phenotypic changes that occur to macrophages in macrophage-NPC cell co-culture systems, and how these changes influence tumor cells. We found that monocytes, including THP-1 cells and primary human monocytes, co-cultured with C666-1 NPC cells upregulate expression of pro-inflammatory cytokines at the early stages, followed by the induction of metastasis-related genes and interferon-stimulated genes at the later stage of coculture, indicating that TAMs are “educated” by NPC cells for cancer progression. Importantly, the induction of these factors from the TAMs was also found to enhance the migratory capabilities of the NPC cells. We have also identified one of these macrophage-derived factor, phospholipase A2 Group 7 (PLA2G7), to be important in regulating tumor cell migration and a novel tumor-promoting factor in NPC. Further studies to characterize the role of PLA2G7 in tumor metastasis may help determine its potential as a therapeutic target in NPC. PMID:27487154

  9. Bortezomib induces apoptosis in primitive chronic myeloid leukemia cells including LTC-IC and NOD/SCID repopulating cells

    PubMed Central

    Heaney, Nicholas B.; Pellicano, Francesca; Zhang, Bin; Crawford, Lisa; Chu, Su; Kazmi, Syed M. A.; Allan, Elaine K.; Jorgensen, Heather G.; Irvine, Alexandra E.; Bhatia, Ravi

    2010-01-01

    Chronic myeloid leukemia (CML) is treated effectively with tyrosine kinase inhibitors (TKIs); however, 2 key problems remain—the insensitivity of CML stem and progenitor cells to TKIs and the emergence of TKI-resistant BCR-ABL mutations. BCR-ABL activity is associated with increased proteasome activity and proteasome inhibitors (PIs) are cytotoxic against CML cell lines. We demonstrate that bortezomib is antiproliferative and induces apoptosis in chronic phase (CP) CD34+ CML cells at clinically achievable concentrations. We also show that bortezomib targets primitive CML cells, with effects on CD34+38−, long-term culture-initiating (LTC-IC) and nonobese diabetic/severe combined immunodeficient (NOD/SCID) repopulating cells. Bortezomib is not selective for CML cells and induces apoptosis in normal CD34+38− cells. The effects against CML cells are seen when bortezomib is used alone and in combination with dasatinib. Bortezomib causes proteasome but not BCR-ABL inhibition and is also effective in inhibiting proteasome activity and inducing apoptosis in cell lines expressing BCR-ABL mutations, including T315I. By targeting both TKI-insensitive stem and progenitor cells and TKI-resistant BCR-ABL mutations, we believe that bortezomib offers a potential therapeutic option in CML. Because of known toxicities, including myelosuppression, the likely initial clinical application of bortezomib in CML would be in resistant and advanced disease. PMID:20068223

  10. A flexible and qualitatively stable model for cell cycle dynamics including DNA damage effects.

    PubMed

    Jeffries, Clark D; Johnson, Charles R; Zhou, Tong; Simpson, Dennis A; Kaufmann, William K

    2012-01-01

    This paper includes a conceptual framework for cell cycle modeling into which the experimenter can map observed data and evaluate mechanisms of cell cycle control. The basic model exhibits qualitative stability, meaning that regardless of magnitudes of system parameters its instances are guaranteed to be stable in the sense that all feasible trajectories converge to a certain trajectory. Qualitative stability can also be described by the signs of real parts of eigenvalues of the system matrix. On the biological side, the resulting model can be tuned to approximate experimental data pertaining to human fibroblast cell lines treated with ionizing radiation, with or without disabled DNA damage checkpoints. Together these properties validate a fundamental, first order systems view of cell dynamics. Classification Codes: 15A68.

  11. The spindle-shaped cells in cutaneous Kaposi's sarcoma. Histologic simulators include factor XIIIa dermal dendrocytes.

    PubMed Central

    Nickoloff, B. J.; Griffiths, C. E.

    1989-01-01

    Kaposi's sarcoma is a neoplasm that develops as multifocal lesions, often involving the skin, characterized by a complex histologic picture including numerous vascular spaces, perivascular and interstitial spindle-shaped cells, and extravasated erythrocytes, lymphocytes, and plasma cells. Using an antibody against factor XIIIa, which identifies dermal dendrocytes, numerous factor XIIIa-positive dermal dendrocytes were detected among the spindle-shaped cells in 12 acquired immune deficiency syndrome (AIDS)-associated, and five non-AIDS-associated Kaposi's sarcoma lesions. The factor XIIIa-positive dermal dendrocytes were also increased in histologic simulators of Kaposi's sarcoma such as dermatofibroma, angiomatoid malignant fibrous histiocytoma, granuloma annulare, and early wound healing, but were absent in keloids. The increased number of dermal dendrocytes, which are often in an angiocentric configuration and which also express CD4, lymphocyte function associated antigen-1 (LFA-1), and Leu M3 in Kaposi's sarcoma, may be important to the angioproliferative response. The results suggested that the spindle-shaped cells that are present in a variety of cutaneous lesions are dermal dendrocytes and belong to the reticuloendothelial system, unlike other mesenchymal cell types such as the endothelial cell. Apparently a diverse array of stimuli, including human immunodeficiency virus type-1 (HIV-1) infection and trauma, can stimulate the accumulation of factor XIIIa expressing dermal dendrocytes in the skin. These cells can then participate in different stages of a variety of cutaneous alterations including Kaposi's sarcoma, dermatofibroma, granuloma annulare, and early wound healing. Thus, the factor XIIIa-positive dermal dendrocyte is a common cellular denominator among diverse clinical entities that share some histologic features. Images Figure 1 Figure 2 Figure 3 p797-a PMID:2573283

  12. Primary small-cell carcinoma of the breast

    PubMed Central

    Dao, Tuoc; Howard, Evan; Bredeweg, Arthur

    2017-01-01

    Early diagnosis of rare breast cancers is expected to occur more frequently as screening compliance improves and diagnostic modalities become more sensitive. Well-defined treatment algorithms exist for the management of ductal and lobular carcinomas; however, less information is available to guide the treatment of atypical breast cancers. This case report describes a 38-year-old African American woman with primary small cell carcinoma of the breast and her treatment.

  13. A Literature Revision in Primary Cutaneous B-cell Lymphoma

    PubMed Central

    Selva, R La; Violetti, S Alberti; Delfino, C; Grandi, V; Cicchelli, S; Tomasini, C; Fierro, M T; Berti, E; Pimpinelli, N; Quaglino, P

    2017-01-01

    The term “Primary Cutaneous B-Cell Lymphoma” (PCBCL) comprehends a variety of lymphoproliferative disorders characterized by a clonal proliferation of B-cells primarily involving the skin. The absence of evident extra-cutaneous disease must be confirmed after six-month follow-up in order to exclude a nodal non-Hodgkin's lymphoma (NHL) with secondary cutaneous involvement, which may have a completely different clinical behavior and prognosis. In this article, we have summarized the clinico-pathological features of main types of PCBCL and we outline the guidelines for management based on a review of the available literature. PMID:28400634

  14. Primary B-cell malignant lymphoma of the lung.

    PubMed

    Canver, C C

    1993-10-01

    A 52-year-old asymptomatic man was evaluated for two right lung lesions discovered on a chest roentgenogram during a routine physical examination. A computed tomographic scan revealed the absence of mediastinal nodal involvement. Guided-needle aspiration cytology was inconclusive. A subsequent right thoracotomy was necessary to perform biopsy of these masses, which proved to be B-cell malignant lymphomas of the lung. This case represents a rare example of a primary low-grade B-cell pulmonary lymphoma of mucosa-associated lymphoid tissue, with its distinct clinicopathologic features.

  15. Primary pure squamous cell carcinoma of the duodenum: a case report.

    PubMed

    Graur, Florin; Mois, Emil; Al Hajjar, Nadim

    2014-09-01

    Primary pure squamous cell carcinoma of the duodenum is a very rare type of duodenal neoplasm and is more likely to be presented as a metastatic tumor. The literature offers little information on this subject and includes very few articles and case reports. Laboratory tests, CT and ultrasound examinations, x-rays and immunohistochemical markers assisted us in making this rare diagnosis of primary squamous cell carcinoma of the duodenum in a 47 year old female patient, who presented with weight loss and melena. The 8 cm duodenal tumor with pancreas invasion was resected by a cephalic duodenopancreatectomy. The pathology examination revealed a primary duodenal squamous cell carcinoma moderately differentiated (G2), invasive in the head of the pancreas, with keratinization, stage II B (pT4N0MxL0V0R0). Positive outcome after surgery was highlighted, no recurrence being registered at the 6 month CT scan follow-up.

  16. Primary and secondary room temperature molten salt electrochemical cells

    NASA Astrophysics Data System (ADS)

    Reynolds, G. F.; Dymek, C. J., Jr.

    1985-07-01

    Three novel primary cells which use room temperature molten salt electrolytes are examined and found to have high open circuit potentials in the 1.75-2.19 V range, by comparison with the Al/AlCl3-MEICl concentration cell; their cathodes were of FeCl3-MEICl, WCl6-MEICl, and Br2/reticulated vitreous carbon together with Pt. Also, secondary electrochemical cell candidates were examined which combined the reversible Al/AlCl3-MEICl electrode with reversible zinc and cadmium molten salt electrodes to yield open circuit potentials of about 0.7 and 1.0 V, respectively. Room temperature molten salts' half-cell reduction potentials are given.

  17. T regulatory cells distinguish two types of primary hypophysitis.

    PubMed

    Mirocha, S; Elagin, R B; Salamat, S; Jaume, J C

    2009-03-01

    Numerous cases of primary hypophysitis have been described over the past 25 years with, however, little insight into the cause(s) of this disease. In order to guide treatment, a better understanding of the pathogenesis is needed. We studied the pathogenesis of primary hypophysitis by analysing systematically the immune response at the pituitary tissue level of consecutive cases of 'lymphocytic' hypophysitis who underwent pituitary biopsy. In order to investigate further the pathogenesis of their diseases we characterized two cases at clinical, cellular and molecular levels. We show here, for the first time, that lymphocytic hypophysitis probably encompasses at least two separate entities. One entity, in agreement with the classical description of lymphocytic hypophysitis, demonstrates an autoimmune process with T helper 17 cell dominance and lack of T regulatory cells. The other entity represents a process in which T regulatory cells seem to control the immune response, which may not be self- but foreign-targeted. Our data suggest that it may be necessary to biopsy suspected primary hypophysitis and to analyse pituitary tissue with immune markers to guide treatment. Based on our results, hypophysitis driven by an immune homeostatic process should not be treated with immunosuppression, while autoimmune-defined hypophysitis may benefit from it. We show here for the first time two different pathogenic processes classified under one disease type and how to distinguish them. Because of our findings, changes in current diagnostic and therapeutic approaches may need to be considered.

  18. Erythropoietin (EPO)-receptor signaling induces cell death of primary myeloma cells in vitro.

    PubMed

    Våtsveen, Thea Kristin; Sponaas, Anne-Marit; Tian, Erming; Zhang, Qing; Misund, Kristine; Sundan, Anders; Børset, Magne; Waage, Anders; Brede, Gaute

    2016-08-31

    Multiple myeloma is an incurable complex disease characterized by clonal proliferation of malignant plasma cells in a hypoxic bone marrow environment. Hypoxia-dependent erythropoietin (EPO)-receptor (EPOR) signaling is central in various cancers, but the relevance of EPOR signaling in multiple myeloma cells has not yet been thoroughly investigated. Myeloma cell lines and malignant plasma cells isolated from bone marrow of myeloma patients were used in this study. Transcript levels were analysed by quantitative PCR and cell surface levels of EPOR in primary cells by flow cytometry. Knockdown of EPOR by short interfering RNA was used to show specific EPOR signaling in the myeloma cell line INA-6. Flow cytometry was used to assess viability in primary cells treated with EPO in the presence and absence of neutralizing anti-EPOR antibodies. Gene expression data for total therapy 2 (TT2), total therapy 3A (TT3A) trials and APEX 039 and 040 were retrieved from NIH GEO omnibus and EBI ArrayExpress. We show that the EPOR is expressed in myeloma cell lines and in primary myeloma cells both at the mRNA and protein level. Exposure to recombinant human EPO (rhEPO) reduced viability of INA-6 myeloma cell line and of primary myeloma cells. This effect could be partially reversed by neutralizing antibodies against EPOR. In INA-6 cells and primary myeloma cells, janus kinase 2 (JAK-2) and extracellular signal regulated kinase 1 and 2 (ERK-1/2) were phosphorylated by rhEPO treatment. Knockdown of EPOR expression in INA-6 cells reduced rhEPO-induced phospo-JAK-2 and phospho-ERK-1/2. Co-cultures of primary myeloma cells with bone marrow-derived stroma cells did not protect the myeloma cells from rhEPO-induced cell death. In four different clinical trials, survival data linked to gene expression analysis indicated that high levels of EPOR mRNA were associated with better survival. Our results demonstrate for the first time active EPOR signaling in malignant plasma cells. EPO

  19. Electrolyte for use in high energy lithium based rechargeable electrochemical cell and rechargeable electrochemical cell including the electrolyte

    NASA Astrophysics Data System (ADS)

    Mammone, R. J.; Binder, M.

    1986-04-01

    The general object of this invention is to provide a lithium based rechargeable electrochemical cell having an improved capacity. A more specific object of the invention is to provide an electrolyte for such a cell. A still further object of the invention is to provide such a cell. A still further object of the invention is to provide such a rechargeable electrochemical cell that permits the oxidation of dithionite to occur without using chlorine as an intermediate oxidizing agent. It has now been found that the aforementioned objects can be attained by providing an electrolyte including bromine dissolved in the liquid complex Li(s02)3A1C14.

  20. Primary cilia regulate the osmotic stress response of renal epithelial cells through TRPM3.

    PubMed

    Siroky, Brian J; Kleene, Nancy K; Kleene, Steven J; Varnell, Charles D; Comer, Raven G; Liu, Jialiu; Lu, Lu; Pachciarz, Nolan W; Bissler, John J; Dixon, Bradley P

    2017-04-01

    Primary cilia sense environmental conditions, including osmolality, but whether cilia participate in the osmotic response in renal epithelial cells is not known. The transient receptor potential (TRP) channels TRPV4 and TRPM3 are osmoresponsive. TRPV4 localizes to cilia in certain cell types, while renal subcellular localization of TRPM3 is not known. We hypothesized that primary cilia are required for maximal activation of the osmotic response of renal epithelial cells and that ciliary TRPM3 and TRPV4 mediate that response. Ciliated [murine epithelial cells from the renal inner medullary collecting duct (mIMCD-3) and 176-5] and nonciliated (176-5Δ) renal cells expressed Trpv4 and Trpm3 Ciliary expression of TRPM3 was observed in mIMCD-3 and 176-5 cells and in wild-type mouse kidney tissue. TRPV4 was identified in cilia and apical membrane of mIMCD-3 cells by electrophysiology and in the cell body by immunofluorescence. Hyperosmolal stress at 500 mOsm/kg (via NaCl addition) induced the osmotic response genes betaine/GABA transporter (Bgt1) and aldose reductase (Akr1b3) in all ciliated cell lines. This induction was attenuated in nonciliated cells. A TRPV4 agonist abrogated Bgt1 and Akr1b3 induction in ciliated and nonciliated cells. A TRPM3 agonist attenuated Bgt1 and Akr1b3 induction in ciliated cells only. TRPM3 knockout attenuated Akr1b3 induction. Viability under osmotic stress was greater in ciliated than nonciliated cells. Akr1b3 induction was also less in nonciliated than ciliated cells when mannitol was used to induce hyperosmolal stress. These findings suggest that primary cilia are required for the maximal osmotic response in renal epithelial cells and that TRPM3 is involved in this mechanism. TRPV4 appears to modulate the osmotic response independent of cilia.

  1. Transport Mechanism of Nicotine in Primary Cultured Alveolar Epithelial Cells.

    PubMed

    Takano, Mikihisa; Nagahiro, Machi; Yumoto, Ryoko

    2016-02-01

    Nicotine is absorbed from the lungs into the systemic circulation during cigarette smoking. However, there is little information concerning the transport mechanism of nicotine in alveolar epithelial cells. In this study, we characterized the uptake of nicotine in rat primary cultured type II (TII) and transdifferentiated type I-like (TIL) epithelial cells. In both TIL and TII cells, [(3)H]nicotine uptake was time and temperature-dependent, and showed saturation kinetics. [(3)H]Nicotine uptake in these cells was not affected by Na(+), but was sensitive to extracellular and intracellular pH, suggesting the involvement of a nicotine/proton antiport system. The uptake of [(3)H]nicotine in these cells was potently inhibited by organic cations such as clonidine, diphenhydramine, and pyrilamine, but was not affected by substrates and/or inhibitors of known organic cation transporters such as carnitine, 1-methyl-4-phenylpyridinium, and tetraethylammonium. In addition, the uptake of [(3)H]nicotine in TIL cells was stimulated by preloading the cells with unlabeled nicotine, pyrilamine, and diphenhydramine, but not with tetraethylammonium. These results suggest that a novel proton-coupled antiporter is involved in the uptake of nicotine in alveolar epithelial cells and its absorption from the lungs into the systemic circulation.

  2. Control of hair cell excitability by vestibular primary sensory neurons

    PubMed Central

    Brugeaud, Aurore; Travo, Cécile; Demêmes, Danielle; Lenoir, Marc; Llorens, Jordi; Puel, Jean-Luc; Chabbert, Christian

    2007-01-01

    In the rat utricle, synaptic contacts between hair cells and the nerve fibers arising from the vestibular primary neurons form during the first week after birth. During that period, the sodium-based excitability that characterizes neonate utricle sensory cells is switched off. To investigate whether the establishment of synaptic contacts was responsible for the modulation of the hair cell excitability, we used an organotypic culture of rat utricle in which the setting of synapses was prevented. Under this condition, the voltage-gated sodium current and the underlying action potentials persisted in a large proportion of non-afferented hair cells. We then studied whether impairment of nerve terminals in utricle of adult rats may also affect hair cell excitability. We induced selective and transient damages of afferent terminals using glutamate excitotoxicity in vivo. The efficiency of the excitotoxic injury was attested by selective swellings of the terminals and underlying altered vestibular behavior. Under this condition, the sodium-based excitability transiently recovered in hair cells. These results indicate that the modulation of hair cells excitability depends on the state of the afferent terminals. In adult utricle hair cells this property may be essential to set the conditions required for restoration of the sensory network after damage. This is achieved via re-expression of a biological process that occurs during synaptogenesis. PMID:17392466

  3. Effects of cigarette smoke extract on primary activated T cells.

    PubMed

    Hernandez, Claudia P; Morrow, Kevin; Velasco, Cruz; Wyczechowska, Dorota D; Naura, Amarjit S; Rodriguez, Paulo C

    2013-03-01

    Tobacco smoking predisposes the development of diseases characterized by chronic inflammation and T cell dysfunction. In this study, we aimed to determine the direct effects of cigarette smoke on primary T cells and to identify the corresponding molecular mediators. Activated T cells cultured in the presence of cigarette smoke extract (CSE) displayed a dose-dependent decrease in cell proliferation, which associated with the induction of cellular apoptosis. T cell apoptosis by CSE was independent of caspases and mediated through reactive oxygen and nitrogen species endogenously contained within CSE. Additional results showed that exposure of T cells to CSE induced phosphorylation of the stress mediator eukaryotic-translation-initiation-factor 2 alpha (eIF2α). Inhibition of the phosphorylation of eIF2α in T cells prevented the cellular apoptosis induced by CSE. Altogether, the results show the direct effects of CSE on T cells, which advance in the understanding of how cigarette smoking promotes chronic inflammation and immune dysfunction. Copyright © 2013 Elsevier Inc. All rights reserved.

  4. Limited Effector Memory B-Cell Response to Envelope Glycoprotein B During Primary Human Cytomegalovirus Infection.

    PubMed

    Dauby, Nicolas; Sartori, Delphine; Kummert, Caroline; Lecomte, Sandra; Haelterman, Edwige; Delforge, Marie-Luce; Donner, Catherine; Mach, Michael; Marchant, Arnaud

    2016-05-15

    Following primary human cytomegalovirus (HCMV) infection, the production of antibodies against envelope glycoprotein B (gB) is delayed, compared with production of antibodies against tegument proteins, and this likely reduces the control of HCMV dissemination. The frequency and the phenotype of gB-specific and tegument protein-specific B cells were studied in a cohort of pregnant women with primary HCMV infection. Healthy adults who had chronic HCMV infection or were recently immunized with tetanus toxoid (TT) were included as controls. Primary HCMV infection was associated with high and similar frequencies of gB-specific and tegument protein-specific B cells following primary HCMV infection. During primary infection, tegument protein-specific B cells expressed an activated (CD21(low)) memory B-cell (MBC) phenotype. Activated MBCs were also induced by TT booster immunization, indicating that the expansion of this subset is part of the physiological B-cell response to protein antigens. In contrast, gB-specific B cells had a predominant classical (CD21(+)) MBC phenotype during both primary and chronic infections. The delayed production of gB-specific immunoglobulin G (IgG) during primary HCMV infection is associated with a limited induction of MBCs with effector potential. This novel mechanism by which HCMV may interfere with the production of neutralizing antibodies could represent a target for therapeutic immunization. © The Author 2015. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail journals.permissions@oup.com.

  5. Investigating the cell death mechanisms in primary prostate cancer cells using low-temperature plasma treatment

    NASA Astrophysics Data System (ADS)

    O'Connell, Deborah; Hirst, A. M.; Packer, J. R.; Simms, M. S.; Mann, V. M.; Frame, F. M.; Maitland, N. J.

    2016-09-01

    Atmospheric pressure plasmas have shown considerable promise as a potential cancer therapy. An atmospheric pressure plasma driven with kHz kV excitation, operated with helium and oxygen admixtures is used to investigate the interaction with prostate cancer cells. The cytopathic effect was verified first in two commonly used prostate cancer cell lines (BPH-1 and PC-3 cells) and further extended to examine the effects in paired normal and tumour prostate epithelial cells cultured directly from patient tissues. Through the formation of reactive species in cell culture media, and potentially other plasma components, we observed high levels of DNA damage, together with reduced cell viability and colony-forming ability. We observed differences in response between the prostate cell lines and primary cells, particularly in terms of the mechanism of cell death. The primary cells ultimately undergo necrotic cell death in both the normal and tumour samples, in the complete absence of apoptosis. In addition, we provide the first evidence of an autophagic response in primary cells. This work highlights the importance of studying primary cultures in order to gain a more realistic insight into patient efficacy. EPSRC EP/H003797/1 & EP/K018388/1, Yorkshire Cancer Research: YCR Y257PA.

  6. Primary thyroid spindle cell tumors: spindle cell variant of papillary thyroid carcinoma?

    PubMed

    Ma, Xiaomei; Xia, Chunyan; Liu, Huimin; Zhu, Weijian

    2015-01-01

    Primary thyroid spindle cell tumors or spindle cell component in the thyroid tumors are very rare. The spindle tumor cells were positive for thyroid papillary carcinoma markers. So these tumors were diagnosed as spindle cell variant of papillary thyroid carcinoma (PTC). To further delineate clinico-pathological features of primary thyroid spindle cell tumors and discuss differential diagnosis, we reported a 67-year-old man with a mass in the right thyroid without clinical symptom. Microscopy revealed that an encapsulated tumor with lot criss spindle cells arranged in bundles. Nuclear grooves were easy to see and rare displayed pseudoinclusions. Immunohistochemical studied showed that the spindle cells were all strong positive for TTF-1, Pax-8, thyroglobulin. Rare follicular were seen in the periphery of the tumor near the thyroid tissue. The cells formed follicular but the spindle tumor cells were positive for pan-keratins. The pathological diagnosis was primary thyroid spindle cell tumors, suspected spindle cell variant of PTC. Primary thyroid spindle cell tumors were presence and without the unified name. The further reports and more discussion were need about these tumors.

  7. Variation in pestivirus growth in testicle primary cell culture is more dependent on the individual cell donor than cattle breed.

    PubMed

    Weber, Matheus N; Bauermann, Fernando V; Gómez-Romero, Ninnet; Herring, Andy D; Canal, Cláudio W; Neill, John D; Ridpath, Julia F

    2017-03-01

    The causes of bovine respiratory disease complex (BRDC) are multifactorial and include infection with both viral and bacterial pathogens. Host factors are also involved as different breeds of cattle appear to have different susceptibilities to BRDC. Infection with bovine pestiviruses, including bovine viral diarrhea virus 1 (BVDV1), BVDV2 and 'HoBi'-like viruses, is linked to the development of BRDC. The aim of the present study was to compare the growth of different bovine pestiviruses in primary testicle cell cultures obtained from taurine, indicine and mixed taurine and indicine cattle breeds. Primary cells strains, derived from testicular tissue, were generated from three animals from each breed. Bovine pestivirus strains used were from BVDV-1a, BVDV-1b, BVDV-2a and 'HoBi'-like virus. Growth was compared by determining virus titers after one passage in primary cells. All tests were run in triplicate. Virus titers were determined by endpoint dilution and RT-qPCR. Statistical analysis was performed using one way analysis of variance (ANOVA) followed by the Tukey's Multiple Comparison Test (P˂0.05). Significant differences in virus growth did not correlate with cattle breed. However, significant differences were observed between cells derived from different individuals regardless of breed. Variation in the replication of virus in primary cell strains may reflect a genetic predisposition that favors virus replication.

  8. Distinctions Among Circulating Antibody Secreting Cell Populations, Including B-1 Cells, in Human Adult Peripheral Blood1

    PubMed Central

    Quách, Tâm D.; Rodríguez-Zhurbenko, Nely; Hopkins, Thomas J.; Guo, Xiaoti; Vázquez, Ana María Hernández; Li, Wentian; Rothstein, Thomas L.

    2015-01-01

    Human antibody secreting cell (ASC) populations in circulation are not well studied. In addition to B-1 (CD20+CD27+CD38lo/intCD43+) cell and the conventional plasmablast (CD20-CD27hiCD38hi) cell populations, here we identified a novel B cell population termed 20+38hi B cells (CD20+CD27hiCD38hi) that spontaneously secretes antibody. At steady state, 20+38hi B cells are distinct from plasmablasts on the basis of CD20 expression, amount of antibody production, frequency of mutation, and diversity of B cell receptor repertoire. However, cytokine treatment of 20+38hi B cells induces loss of CD20 and acquisition of CD138, suggesting that 20+38hi B cells are precursors to plasmablasts, or pre-plasmablasts. We then evaluated similarities and differences between CD20+CD27+CD38lo/intCD43+ B-1 cells, CD20+CD27hiCD38hi 20+38hi B cells, CD20-CD27hiCD38hi plasmablasts, and CD20+CD27+CD38lo/intCD43- memory B cells. We found that B-1 cells differ from 20+38hi B cells and plasmablasts in numbers of ways, including antigen expression, morphological appearance, transcriptional profiling, antibody skewing, antibody repertoire, and secretory response to stimulation. In terms of gene expression, B-1 cells align more closely with memory B cells than with 20+38hi B cells or plasmablasts, but differ in that memory B cells do not express antibody secretion related genes. We found that, B-1 cell antibodies utilize Vh4-34, which is often associated with autoreactivity, 3 to 6-fold more often than other B cell populations. Along with selective production of IgM anti-PC, this data suggests that human B-1 cells might be preferentially selected for autoreactivity/natural-specificity. In sum, our results indicate that human healthy adult peripheral blood at steady state consists of 3 distinct ASC populations. PMID:26740107

  9. Dye laser amplifier including a dye cell contained within a support vessel

    DOEpatents

    Davin, James

    1992-01-01

    A large (high flow rate) dye laser amplifier in which a continous replenished supply of dye is excited by a first light beam, specifically a copper vapor laser beam, in order to amplify the intensity of a second different light beam, specifically a dye beam, passing through the dye is disclosed herein. This amplifier includes a dye cell defining a dye chamber through which a continuous stream of dye is caused to pass at a flow rate of greater than 30 gallons/minute at a static pressure greater than 150 pounds/square inch and a specifically designed support vessel for containing the dye cell.

  10. Dye laser amplifier including a dye cell contained within a support vessel

    DOEpatents

    Davin, J.

    1992-12-01

    A large (high flow rate) dye laser amplifier in which a continuous replenished supply of dye is excited by a first light beam, specifically a copper vapor laser beam, in order to amplify the intensity of a second different light beam, specifically a dye beam, passing through the dye is disclosed herein. This amplifier includes a dye cell defining a dye chamber through which a continuous stream of dye is caused to pass at a flow rate of greater than 30 gallons/minute at a static pressure greater than 150 pounds/square inch and a specifically designed support vessel for containing the dye cell. 6 figs.

  11. Identification of drugs that restore primary cilium expression in cancer cells

    PubMed Central

    Khan, Niamat Ali; Willemarck, Nicolas; Talebi, Ali; Marchand, Arnaud; Binda, Maria Mercedes; Dehairs, Jonas; Rueda-Rincon, Natalia; Daniels, Veerle W.; Bagadi, Muralidhararao; Raj, Deepak Balaji Thimiri Govinda; Vanderhoydonc, Frank; Munck, Sebastian; Chaltin, Patrick; Swinnen, Johannes V.

    2016-01-01

    The development of cancer is often accompanied by a loss of the primary cilium, a microtubule-based cellular protrusion that functions as a cellular antenna and that puts a break on cell proliferation. Hence, restoration of the primary cilium in cancer cells may represent a novel promising approach to attenuate tumor growth. Using a high content analysis-based approach we screened a library of clinically evaluated compounds and marketed drugs for their ability to restore primary cilium expression in pancreatic ductal cancer cells. A diverse set of 118 compounds stimulating cilium expression was identified. These included glucocorticoids, fibrates and other nuclear receptor modulators, neurotransmitter regulators, ion channel modulators, tyrosine kinase inhibitors, DNA gyrase/topoisomerase inhibitors, antibacterial compounds, protein inhibitors, microtubule modulators, and COX inhibitors. Certain compounds also dramatically affected the length of the cilium. For a selection of compounds (Clofibrate, Gefitinib, Sirolimus, Imexon and Dexamethasone) their ability to restore ciliogenesis was confirmed in a panel of human cancer cell line models representing different cancer types (pancreas, lung, kidney, breast). Most compounds attenuated cell proliferation, at least in part through induction of the primary cilium, as demonstrated by cilium removal using chloral hydrate. These findings reveal that several commonly used drugs restore ciliogenesis in cancer cells, and warrant further investigation of their antineoplastic properties. PMID:26862738

  12. Potential of primary kidney cells for somatic cell nuclear transfer mediated transgenesis in pig

    PubMed Central

    2012-01-01

    Background Somatic cell nuclear transfer (SCNT) is currently the most efficient and precise method to generate genetically tailored pig models for biomedical research. However, the efficiency of this approach is crucially dependent on the source of nuclear donor cells. In this study, we evaluate the potential of primary porcine kidney cells (PKCs) as cell source for SCNT, including their proliferation capacity, transfection efficiency, and capacity to support full term development of SCNT embryos after additive gene transfer or homologous recombination. Results PKCs could be maintained in culture with stable karyotype for up to 71 passages, whereas porcine fetal fibroblasts (PFFs) and porcine ear fibroblasts (PEFs) could be hardly passaged more than 20 times. Compared with PFFs and PEFs, PKCs exhibited a higher proliferation rate and resulted in a 2-fold higher blastocyst rate after SCNT and in vitro cultivation. Among the four transfection methods tested with a GFP expression plasmid, best results were obtained with the NucleofectorTM technology, resulting in transfection efficiencies of 70% to 89% with high fluorescence intensity, low cytotoxicity, good cell proliferation, and almost no morphological signs of cell stress. Usage of genetically modified PKCs in SCNT resulted in approximately 150 piglets carrying at least one of 18 different transgenes. Several of those pigs originated from PKCs that underwent homologous recombination and antibiotic selection before SCNT. Conclusion The high proliferation capacity of PKCs facilitates the introduction of precise and complex genetic modifications in vitro. PKCs are thus a valuable cell source for the generation of porcine biomedical models by SCNT. PMID:23140586

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

    PubMed Central

    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. Evaluation of high-energy lithium thionyl chloride primary cells

    NASA Technical Reports Server (NTRS)

    Frank, H. A.

    1980-01-01

    An advanced commercial primary lithium cell (LiSoCl2) was evaluated in order to establish baseline data for improved lithium batteries for aerospace applications. The cell tested had nominal capacity of 6 Ah. Maximum energy density at low rates (less than C/30, where C is the cell capacity in amp-hrs and 30 corresponds to a 30 hr discharge time) was found to be near 300 Wh/kg. An equation which predicts the operating voltage of these cells as a function of current and state of charge is presented. Heat generation rates of these cells were determined as a function of current in a calorimeter. It was found that heat rates could be theoretically predicted with some degree of accuracy at currents less than 1 amp or the C/6 rate. No explosions were observed in the cells during the condition of overdischarge or reversal nor during high rate discharge. It was found, however, that the cells can vent when overdischarge currents are greater than C/30 and when discharge rates are greater than 1.5C.

  15. Production of avian influenza virus vaccine using primary cell cultures generated from host organs.

    PubMed

    Babar, Mustafeez Mujtaba; Riaz, Muhammad Suleman; Zaidi, Najam-us-Sahar Sadaf; Afzal, Farhan; Farooq, Muhammad Sabir

    2013-06-01

    The global availability of a therapeutically effective influenza virus vaccine during a pandemic remains a major challenge for the biopharmaceutical industry. Long production time, coupled with decreased supply of embryonated chicken eggs (ECE), significantly affects the conventional vaccine production. Transformed cell lines have attained regulatory approvals for vaccine production. Based on the fact that the avian influenza virus would infect the cells derived from its natural host, the viral growth characteristics were studied on chicken embryo-derived primary cell cultures. The viral propagation was determined on avian origin primary cell cultures, transformed mammalian cell lines, and in ECE. A comparison was made between these systems by utilizing various cell culture-based assays. In-vitro substrate susceptibility and viral infection characteristics were evaluated by performing hemagglutination assay (HA), 50 % tissue culture infectious dose (TCID₅₀) and monitoring of cytopathic effects (CPE) caused by the virus. The primary cell culture developed from chicken embryos showed stable growth characteristics with no contamination. HA, TCID₅₀, and CPE exhibited that these cell systems were permissive to viral infection, yielding 2-10 times higher viral titer as compared to mammalian cell lines. Though the viral output from the ECE was equivalent to the chicken cell culture, the time period for achieving it was decreased to half. Some of the prerequisites of inactivated influenza virus vaccine production include generation of higher vial titer, independence from exogenous sources, and decrease in the production time lines. Based on the tests, it can be concluded that chicken embryo primary cell culture addresses these issues and can serve as a potential alternative for influenza virus vaccine production.

  16. Primary cerebellar extramedullary myeloid cell tumor mimicking oligodendroglioma.

    PubMed

    Ho, D M; Wong, T T; Guo, W Y; Chang, K P; Yen, S H

    1997-10-01

    Extramedullary myeloid cell tumors (EMCTs) are tumors consisting of immature cells of the myeloid series that occur outside the bone marrow. Most of them are associated with acute myelogenous leukemia or other myeloproliferative disorders, and a small number occur as primary lesions, i.e., are not associated with hematological disorders. Occurrence inside the cranium is rare, and there has been only one case of primary EMCT involving the cerebellum reported in the literature. The case we report here is a blastic EMCT occurring in the cerebellum of a 3-year-old boy who had no signs of leukemia or any hematological disorder throughout the entire course. The cerebellar tumor was at first misdiagnosed as an "oligodendroglioma" because of the uniformity and "fried egg" artifact of the tumor cells. The tumor disappeared during chemotherapy consisting of 12 treatments. However, it recurred and metastasized to the cerebrospinal fluid (CSF) shortly after the therapy was completed. A diagnosis of EMCT was suspected because of the presence of immature myeloid cells in the CSF, and was confirmed by anti-myeloperoxidase and anti-lysozyme immunoreactivity of the cerebellar tumor. The patient succumbed 1 year and 3 months after the first presentation of the disease.

  17. CXCR4 heterogeneity in primary cells: possible role of ubiquitination.

    PubMed

    Lapham, Cheryl K; Romantseva, Tatiana; Petricoin, Emmanuel; King, Lisa R; Manischewitz, Jody; Zaitseva, Marina B; Golding, Hana

    2002-12-01

    The chemokine receptor CXCR4 is a primary coreceptor for the HIV-1 virus. The predicted molecular weight (MW) of glycosylated CXCR4 is 45-47 kDa. However, immunoblots of whole cell lysates from human lymphocytes, monocytes, macrophages, and the Jurkat T-lymphocyte line revealed multiple MW isoforms of CXCR4. Three of the bands could be precipitated by anti-CXCR4 monoclonal antibodies (101 and 47 kDa) or coprecipitated with CD4 (62 kDa). Expression of these isoforms was enhanced by infection with a recombinant vaccinia virus encoding CXCR4. In immunoblots of two-dimensional gels, antiubiquitin antibodies reacted with the 62-kDa CXCR4 species from monocytes subsequent to coprecipitation with anti-CD4 antibodies. Culturing of monocytes and lymphocytes with lactacystin enhanced the amount of the 101-kDa CXCR4 isoform in immunoblots by three- to sevenfold. In lymphocytes, lactacystin also increased cell-surface expression of CXCR4, which correlated with enhanced fusion with HIV-1 envelope-expressing cells. Similar increases in the intensity of the 101-kDa isoform were seen after treatment with the lysosomal inhibitors monensin and ammonium chloride. Antiubiquitin antibodies reacted with multiple proteins above 62 kDa, which were precipitated with anti-CXCR4 antibodies. Our data indicate that ubiquitination may contribute to CXCR4 heterogeneity and suggest roles for proteasomes and lysosomes in the constitutive turnover of CXCR4 in primary human cells.

  18. Brucella pinnipedialis in hooded seal (Cystophora cristata) primary epithelial cells.

    PubMed

    Larsen, Anett Kristin; Godfroid, Jacques; Nymo, Ingebjørg Helena

    2016-01-25

    Marine Brucella spp. have been isolated from numerous pinniped and cetacean species, but pathological findings in association with infection with Brucella pinnipedialis in pinnipeds have been sparse. The capacity of brucellae to survive and replicate within host macrophages underlies their important ability to produce chronic infections, but previous work has shown that B. pinnipedialis spp. are rapidly eliminated from hooded seal (Cystophora cristata) alveolar macrophages. To investigate if multiplication could take place in other hooded seal cell types, primary epithelial cells were isolated, verified to express the epithelial marker cytokeratin and challenged with three different strains of B. pinnipedialis; B. pinnipedialis sp. nov., B. pinnipedialis hooded seal strain B17, and B. pinnipedialis hooded seal strain 22F1. All strains were steadily eliminated and the amounts of intracellular bacteria were reduced to less than one-third by 48 h post infection. Intracellular presence was verified using immunocytochemistry. So far, intracellular multiplication in seal cells has not been documented for B. pinnipedialis. The lack of intracellular survival in macrophages, as well as in epithelial cells, together with the fact that pathological changes due to B. pinnipedialis infection is not yet identified in seals, suggests that the bacteria may only cause a mild, acute and transient infection. These findings also contribute to substantiate the hypothesis that seals may not be the primary host of B. pinnipedialis and that the transmission to seals are caused by other species in the marine environment.

  19. Low-temperature plasma treatment induces DNA damage leading to necrotic cell death in primary prostate epithelial cells

    PubMed Central

    Hirst, A M; Simms, M S; Mann, V M; Maitland, N J; O'Connell, D; Frame, F M

    2015-01-01

    Background: In recent years, the rapidly advancing field of low-temperature atmospheric pressure plasmas has shown considerable promise for future translational biomedical applications, including cancer therapy, through the generation of reactive oxygen and nitrogen species. Method: The cytopathic effect of low-temperature plasma was first verified in two commonly used prostate cell lines: BPH-1 and PC-3 cells. The study was then extended to analyse the effects in paired normal and tumour (Gleason grade 7) prostate epithelial cells cultured directly from patient tissue. Hydrogen peroxide (H2O2) and staurosporine were used as controls throughout. Results: Low-temperature plasma (LTP) exposure resulted in high levels of DNA damage, a reduction in cell viability, and colony-forming ability. H2O2 formed in the culture medium was a likely facilitator of these effects. Necrosis and autophagy were recorded in primary cells, whereas cell lines exhibited apoptosis and necrosis. Conclusions: This study demonstrates that LTP treatment causes cytotoxic insult in primary prostate cells, leading to rapid necrotic cell death. It also highlights the need to study primary cultures in order to gain more realistic insight into patient response. PMID:25839988

  20. Gene expression profiles of circulating tumor cells versus primary tumors in metastatic breast cancer.

    PubMed

    Onstenk, Wendy; Sieuwerts, Anieta M; Weekhout, Marleen; Mostert, Bianca; Reijm, Esther A; van Deurzen, Carolien H M; Bolt-de Vries, Joan B; Peeters, Dieter J; Hamberg, Paul; Seynaeve, Caroline; Jager, Agnes; de Jongh, Felix E; Smid, Marcel; Dirix, Luc Y; Kehrer, Diederik F S; van Galen, Anne; Ramirez-Moreno, Raquel; Kraan, Jaco; Van, Mai; Gratama, Jan W; Martens, John W M; Foekens, John A; Sleijfer, Stefan

    2015-06-28

    Before using circulating tumor cells (CTCs) as liquid biopsy, insight into molecular discrepancies between CTCs and primary tumors is essential. We characterized CellSearch-enriched CTCs from 62 metastatic breast cancer (MBC) patients with ≥5 CTCs starting first-line systemic treatment. Expression levels of 35 tumor-associated, CTC-specific genes, including ESR1, coding for the estrogen receptor (ER), were measured by reverse transcription quantitative polymerase chain reaction and correlated to corresponding primary tumors. In 30 patients (48%), gene expression profiles of 35 genes were discrepant between CTCs and the primary tumor, but this had no prognostic consequences. In 15 patients (24%), the expression of ER was discrepant. Patients with ER-negative primary tumors and ER-positive CTCs had a longer median TTS compared to those with concordantly ER-negative CTCs (8.5 versus 2.1 months, P = 0.05). From seven patients, an axillary lymph node metastasis was available. In two patients, the CTC profiles better resembled the lymph node metastasis than the primary tumor. Our findings suggest that molecular discordances between CTCs and primary tumors frequently occur, but that this bears no prognostic consequences. Alterations in ER-status between primary tumors and CTCs might have prognostic implications.

  1. Characterization of the interaction between human respiratory syncytial virus and the cell cycle in continuous cell culture and primary human airway epithelial cells.

    PubMed

    Wu, Weining; Munday, Diane C; Howell, Gareth; Platt, Gareth; Barr, John N; Hiscox, Julian A

    2011-10-01

    Viruses can modify conditions inside cells to make them more favorable for replication and progeny virus production. One way of doing this is through manipulation of the cell cycle, a process that describes the ordered growth and division of cells. Analysis of model cell lines, such as A549 cells and primary airway epithelial cells, infected with human respiratory syncytial virus (HRSV) has shown alteration of the cell cycle during infection, although the signaling events were not clearly understood. In this study, targeted transcriptomic analysis of HRSV-infected primary airway epithelial cells revealed alterations in the abundances of many mRNAs encoding cell cycle-regulatory molecules, including decreases in the D-type cyclins and corresponding cyclin-dependent kinases (CDK4 and CDK6 [CDK4/6]). These alterations were reflected in changes in protein abundance and/or relocalization in HRSV-infected cells; taken together, they were predicted to result in G(0)/G(1) phase arrest. In contrast, there was no change in the abundances of D-type cyclins in A549 cells infected with HRSV. However, the abundance of the G(1)/S phase progression inhibitor p21(WAF1/CIP1) was increased over that in mock-treated cells, and this, again, was predicted to result in G(0)/G(1) phase arrest. The G(0)/G(1) phase arrest in both HRSV-infected primary cells and A549 cells was confirmed using dual-label flow cytometry that accurately measured the different stages of the cell cycle. Comparison of progeny virus production in primary and A549 cells enriched in G(0)/G(1) using a specific CDK4/6 kinase inhibitor with asynchronously replicating cells indicated that this phase of the cell cycle was more efficient for virus production.

  2. Docosahexaenoic Acid Induces Apoptosis in Primary Chronic Lymphocytic Leukemia Cells

    PubMed Central

    Gyan, Emmanuel; Tournilhac, Olivier; Halty, Christelle; Veyrat-Masson, Richard; Akil, Saïda; Berger, Marc; Hérault, Olivier; Callanan, Mary; Bay, Jacques-Olivier

    2015-01-01

    Chronic lymphocytic leukemia is an indolent disorder with an increased infectious risk remaining one of the main causes of death. Development of therapies with higher safety profile is thus a challenging issue. Docosahexaenoic acid (DHA, 22:6) is an omega-3 fatty acid, a natural compound of normal cells, and has been shown to display antitumor potency in cancer. We evaluated the potential in vitro effect of DHA in primary CLL cells. DHA induces high level of in vitro apoptosis compared to oleic acid in a dose-dependent and time-dependent manner. Estimation of IC50 was only of 4.813 µM, which appears lower than those reported in solid cancers. DHA is highly active on CLL cells in vitro. This observation provides a rationale for further studies aiming to understand its mechanisms of action and its potent in vivo activity. PMID:26734128

  3. Second Unrelated Donor Hematopoietic Cell Transplantation for Primary Graft Failure

    PubMed Central

    Schriber, Jeffrey; Agovi, Manza-A.; Ho, Vincent; Ballen, Karen K.; Bacigalupo, Andrea; Lazarus, Hillard M.; Bredeson, Christopher N.; Gupta, Vikas; Maziarz, Richard T.; Hale, Gregory A.; Litzow, Mark R.; Logan, Brent; Bornhauser, Martin; Giller, Roger H.; Isola, Luis; Marks, David I.; Rizzo, J. Douglas; Pasquini, Marcelo C.

    2010-01-01

    Failure to engraft donor cells is a devastating complication after allogeneic hematopoietic cell transplantation (HCT). We describe the results of 122 patients reported to the National Marrow Donor Program between 1990 and 2005, who received a second unrelated donor HCT after failing to achieve an absolute neutrophil count of ≥ 500/ μL without recurrent disease. Patients were transplanted for leukemia (n=83), myelodysplastic disorders (n=16), severe aplastic anemia (n=20) and other diseases (n=3). The median age was 29 years. Twenty-four patients received second grafts from a different unrelated donor. Among 98 patients who received a second graft from the same donor, 28 received products that were previously collected and cryopreserved for the first transplantation. One-year overall survival after second transplant was 11% with 10 patients alive at last follow up. We observed no differences between patients who received grafts from the same or different donors, or in those who received fresh or cryopreserved product. The outcomes after a second allogeneic HCT for primary graft failure are dismal. Identifying risk factors for primary graft failure can decrease the incidence of this complication. Further studies are needed to test whether early recognition and hastened procurement of alternative grafts can improve transplant outcomes for primary graft failure. PMID:20172038

  4. Primary retroperitoneal Merkel cell carcinoma: Case report and literature review

    PubMed Central

    Quiroz-Sandoval, Osvaldo A.; Cuellar-Hubbe, Mario; Lino-Silva, Leonardo S.; Salcedo-Hernández, Rosa A.; López-Basave, Horacio N.; Padilla-Rosciano, Alejandro E.; León-Takahashi, Alberto M.; Herrera-Gómez, Ángel

    2015-01-01

    Background Merkel cell carcinoma (MCC) is an aggressive cutaneous neuroendocrine carcinoma that affects elderly patients and typically arises in sun-exposed skin. The disease is very rare and only few cases present with no apparent skin lesion. In the retroperitoneum there are only two cases reported in the literature. Case presentation We report a case of a 54-year-old Mexican male with MCC, which presented as a large retroperitoneal mass. Pathological and immunohistochemical analysis of the transabdominal CT-guided biopsy specimen revealed a MCC. The patient underwent preoperative chemotherapy followed by a laparotomy and the mass was successfully excised. Discussion There are two possible explanations for what occurred in our patient. The most plausible theory is the retroperitoneal mass could be a massively enlarged lymph node where precursor cells became neoplastic. This would be consistent with a presumptive diagnosis of primary nodal disease. Moreover, metastasis to the retroperitoneal lymph nodes has been reported as relatively common when compared to other sites such as liver, bone, brain and skin. The less probable theory is the non-described “regression” phenomena of a cutaneous MCC, but we are not found a primary skin lesion. Conclusion Preoperative chemotherapy and excision of the primary tumor is the surgical treatment of choice for retroperitoneal MCC. We propose that further studies are needed to elucidate the true efficacy of chemotherapy in conventional and unconventional patients with MCC. PMID:26708276

  5. Immunohistochemical Expression of Cathepsin D in Primary and Recurrent Squamous Cell Carcinoma.

    PubMed

    Satelur, Krishnanand P; Kumar, G S

    2017-09-01

    The aim of this study is to analyze and compare the immunohistochemical expression of cathepsin B in primary oral squamous cell carcinoma (OSCC) and recurrent OSCC. A total of 50 cases were studied immunohistochemically for rabbit polyclonal antihuman cathepsin D expression. A total of 10 cases of breast carcinoma were taken as positive controls. Immunohistochemical staining was performed using labeled streptavidin-biotin technique. All the 45 cases of OSCC, both primary and recurrent cases included, showed varying grades of cathepsin D immu-noreactivity. Statistical significance at 5% level was observed in cathepsin D expression between the different grades of well, moderate, and poorly differentiated primary squamous cell carcinomas. In the comparison of cathepsin D staining intensity among primary squamous cell carcinomas with and without recurrence, a statistical significance between the groups was observed when the p-value was at 10%, but the same comparison was not significant when the p-value was at 5%. Cathepsin D expression in primary squamous cell carcinomas with recurrences was very variable as compared with primary squamous cell carcinomas without recurrences. Comparison of cathepsin D expression in primary with their recurrent counterparts showed mostly similar intensity of expression in recurrent carcinomas, thus suggesting its limited usefulness in predicting recurrence. Although cathepsin D might have shown limited usefulness in predicting cancer recurrence, it, however, is a proven valuable tool to detect the aggressiveness of various other tumors, and if corroborated with a larger sample may hold the key to early, more effective, and more specific treatment modalities for cases of oral cancer also.

  6. The small GTPase Cdc42 is necessary for primary ciliogenesis in renal tubular epithelial cells.

    PubMed

    Zuo, Xiaofeng; Fogelgren, Ben; Lipschutz, Joshua H

    2011-06-24

    Primary cilia are found on many epithelial cell types, including renal tubular epithelial cells, where they participate in flow sensing. Disruption of cilia function has been linked to the pathogenesis of polycystic kidney disease. We demonstrated previously that the exocyst, a highly conserved eight-protein membrane trafficking complex, localizes to primary cilia of renal tubular epithelial cells, is required for ciliogenesis, biochemically and genetically interacts with polycystin-2 (the protein product of the polycystic kidney disease 2 gene), and, when disrupted, results in MAPK pathway activation both in vitro and in vivo. The small GTPase Cdc42 is a candidate for regulation of the exocyst at the primary cilium. Here, we demonstrate that Cdc42 biochemically interacts with Sec10, a crucial component of the exocyst complex, and that Cdc42 colocalizes with Sec10 at the primary cilium. Expression of dominant negative Cdc42 and shRNA-mediated knockdown of both Cdc42 and Tuba, a Cdc42 guanine nucleotide exchange factor, inhibit ciliogenesis in Madin-Darby canine kidney cells. Furthermore, exocyst Sec8 and polycystin-2 no longer localize to primary cilia or the ciliary region following Cdc42 and Tuba knockdown. We also show that Sec10 directly interacts with Par6, a member of the Par complex that itself directly interacts with Cdc42. Finally, we show that Cdc42 knockdown results in activation of the MAPK pathway, something observed in cells with dysfunctional primary cilia. These data support a model in which Cdc42 localizes the exocyst to the primary cilium, whereupon the exocyst then targets and docks vesicles carrying proteins necessary for ciliogenesis.

  7. The Small GTPase Cdc42 Is Necessary for Primary Ciliogenesis in Renal Tubular Epithelial Cells*

    PubMed Central

    Zuo, Xiaofeng; Fogelgren, Ben; Lipschutz, Joshua H.

    2011-01-01

    Primary cilia are found on many epithelial cell types, including renal tubular epithelial cells, where they participate in flow sensing. Disruption of cilia function has been linked to the pathogenesis of polycystic kidney disease. We demonstrated previously that the exocyst, a highly conserved eight-protein membrane trafficking complex, localizes to primary cilia of renal tubular epithelial cells, is required for ciliogenesis, biochemically and genetically interacts with polycystin-2 (the protein product of the polycystic kidney disease 2 gene), and, when disrupted, results in MAPK pathway activation both in vitro and in vivo. The small GTPase Cdc42 is a candidate for regulation of the exocyst at the primary cilium. Here, we demonstrate that Cdc42 biochemically interacts with Sec10, a crucial component of the exocyst complex, and that Cdc42 colocalizes with Sec10 at the primary cilium. Expression of dominant negative Cdc42 and shRNA-mediated knockdown of both Cdc42 and Tuba, a Cdc42 guanine nucleotide exchange factor, inhibit ciliogenesis in Madin-Darby canine kidney cells. Furthermore, exocyst Sec8 and polycystin-2 no longer localize to primary cilia or the ciliary region following Cdc42 and Tuba knockdown. We also show that Sec10 directly interacts with Par6, a member of the Par complex that itself directly interacts with Cdc42. Finally, we show that Cdc42 knockdown results in activation of the MAPK pathway, something observed in cells with dysfunctional primary cilia. These data support a model in which Cdc42 localizes the exocyst to the primary cilium, whereupon the exocyst then targets and docks vesicles carrying proteins necessary for ciliogenesis. PMID:21543338

  8. Inefficiency in macromolecular transport of SCS-based microcapsules affects viability of primary human mesenchymal stem cells but not of immortalized cells.

    PubMed

    Sanz-Nogués, Clara; Horan, Jason; Thompson, Kerry; Howard, Linda; Ryan, Gerard; Kassem, Moustapha; O'Brien, Timothy

    2015-11-01

    Microcapsules made of sodium cellulose sulphate (SCS) and poly-diallyl-dimethyl-ammonium chloride (pDADMAC) have been employed to encapsulate a wide range of established cell lines for several applications. However, little is known about the encapsulation of primary cells including human mesenchymal stem cells (hMSCs). Human MSCs are of interest in regenerative medicine applications due to pro-angiogenic, anti-inflammatory and immunomodulatory properties, which result from paracrine effects of this cell type. In the present work we have encapsulated primary hMSCs and hMSC-TERT immortalized cells and compared their behavior and in vitro angiogenic potential. We found that, although both cell types were able to secret angiogenic factors such as VEGF, there was a marked reduction of primary hMSC viability compared to hMSC-TERT cells when cultured in these microcapsules. Moreover, this applied to other primary cell cultures such as primary human fibroblasts but not to other cell lines such as human embryonic kidney 293 (HEK293) cells. We found that the microcapsule membrane had a molecular weight cut-off below a critical size, which caused impairment in the diffusion of essential nutrients and had a more detrimental effect on the viability of primary cell cultures compared to cell lines and immortalized cells.

  9. Oncogenic NRAS Primes Primary Acute Myeloid Leukemia Cells for Differentiation

    PubMed Central

    Millahn, Axel; Stiewe, Thorsten; Krause, Michael; Stabla, Kathleen; Ross, Petra; Huynh, Minh; Illmer, Thomas; Mernberger, Marco; Barckhausen, Christina; Neubauer, Andreas

    2015-01-01

    RAS mutations are frequently found among acute myeloid leukemia patients (AML), generating a constitutively active signaling protein changing cellular proliferation, differentiation and apoptosis. We have previously shown that treatment of AML patients with high-dose cytarabine is preferentially beneficial for those harboring oncogenic RAS. On the basis of a murine AML cell culture model, we ascribed this effect to a RAS-driven, p53-dependent induction of differentiation. Hence, in this study we sought to confirm the correlation between RAS status and differentiation of primary blasts obtained from AML patients. The gene expression signature of AML blasts with oncogenic NRAS indeed corresponded to a more mature profile compared to blasts with wildtype RAS, as demonstrated by gene set enrichment analysis (GSEA) and real-time PCR analysis of myeloid ecotropic viral integration site 1 homolog (MEIS1) in a unique cohort of AML patients. In addition, in vitro cell culture experiments with established cell lines and a second set of primary AML cells showed that oncogenic NRAS mutations predisposed cells to cytarabine (AraC) driven differentiation. Taken together, our findings show that AML with inv(16) and NRAS mutation have a differentiation gene signature, supporting the notion that NRAS mutation may predispose leukemic cells to AraC induced differentiation. We therefore suggest that promotion of differentiation pathways by specific genetic alterations could explain the superior treatment outcome after therapy in some AML patient subgroups. Whether a differentiation gene expression status may generally predict for a superior treatment outcome in AML needs to be addressed in future studies. PMID:25901794

  10. Mesenchymal Stem Cells Support Survival and Proliferation of Primary Human Acute Myeloid Leukemia Cells through Heterogeneous Molecular Mechanisms

    PubMed Central

    Brenner, Annette K.; Nepstad, Ina; Bruserud, Øystein

    2017-01-01

    Acute myeloid leukemia (AML) is a bone marrow malignancy, and various bone marrow stromal cells seem to support leukemogenesis, including osteoblasts and endothelial cells. We have investigated how normal bone marrow mesenchymal stem cells (MSCs) support the in vitro proliferation of primary human AML cells. Both MSCs and primary AML cells show constitutive release of several soluble mediators, and the mediator repertoires of the two cell types are partly overlapping. The two cell populations were cocultured on transwell plates, and MSC effects on AML cells mediated through the local cytokine/soluble mediator network could thus be evaluated. The presence of normal MSCs had an antiapoptotic and growth-enhancing effect on primary human AML cells when investigating a group of 51 unselected AML patients; this was associated with increased phosphorylation of mTOR and its downstream targets, and the effect was independent of cytogenetic or molecular-genetic abnormalities. The MSCs also supported the long-term proliferation of the AML cells. A subset of the patients also showed an altered cytokine network with supra-additive levels for several cytokines. The presence of cytokine-neutralizing antibodies or receptor inhibitors demonstrated that AML cells derived from different patients were heterogeneous with regard to effects of various cytokines on AML cell proliferation or regulation of apoptosis. We conclude that even though the effects of single cytokines derived from bone marrow MSCs on human AML cells differ among patients, the final cytokine-mediated effects of the MSCs during coculture is growth enhancement and inhibition of apoptosis. PMID:28232835

  11. Serglycin in Quiescent and Proliferating Primary Endothelial Cells

    PubMed Central

    Reine, Trine M.; Vuong, Tram T.; Rutkovskiy, Arkady; Meen, Astri J.; Vaage, Jarle; Jenssen, Trond G.; Kolset, Svein O.

    2015-01-01

    Proteoglycans are fundamental components of the endothelial barrier, but the functions of the proteoglycan serglycin in endothelium are less described. Our aim was to describe the roles of serglycin in processes relevant for endothelial dysfunction. Primary human umbilical vein endothelial cells (HUVEC) were cultured in vitro and the expression of proteoglycans was investigated. Dense cell cultures representing the quiescent endothelium coating the vasculature was compared to sparse activated cell cultures, relevant for diabetes, cancer and cardiovascular disease. Secretion of 35S- proteoglycans increased in sparse cultures, and we showed that serglycin is a major component of the cell-density sensitive proteoglycan population. In contrast to the other proteoglycans, serglycin expression and secretion was higher in proliferating compared to quiescent HUVEC. RNAi silencing of serglycin inhibited proliferation and wound healing, and serglycin expression and secretion was augmented by hypoxia, mechanical strain and IL-1β induced inflammation. Notably, the secretion of the angiogenic chemokine CCL2 resulting from IL-1β activation, was increased in serglycin knockdown cells, while angiopoietin was not affected. Both serglycin and CCL2 were secreted predominantly to the apical side of polarized HUVEC, and serglycin and CCL2 co-localized both in perinuclear areas and in vesicles. These results suggest functions for serglycin in endothelial cells trough interactions with partner molecules, in biological processes with relevance for diabetic complications, cardiovascular disease and cancer development. PMID:26694746

  12. Primary retroperitoneal myxoid/round cell liposarcoma is a nonexisting disease: an immunohistochemical and molecular biological analysis.

    PubMed

    de Vreeze, Ronald S A; de Jong, Daphne; Tielen, Ivon H G; Ruijter, Henrique J; Nederlof, Petra M; Haas, Rick L; van Coevorden, Frits

    2009-02-01

    Almost all primary retroperitoneal liposarcomas can be classified as well-/dedifferentiated liposarcoma. Rarely, however, primary retroperitoneal liposarcoma is classified as myxoid/round cell liposarcoma, based on the presence of myxoid areas and vascular crow's feet pattern, which has resulted in a debate on the classification of liposarcoma in the retroperitoneum. Genetically, myxoid/round cell liposarcoma and well-/dedifferentiated liposarcoma are different diseases. Myxoid/round cell liposarcoma is characterized by a translocation causing FUS-CHOP or EWSR1-CHOP fusion, whereas well-/dedifferentiated liposarcoma is characterized by an amplification of the 12q13-15 region, including MDM2 and CDK4 genes. As myxoid/round cell liposarcoma is highly radio- and chemosensitive, differentiation between subtypes is important to optimize treatment. We studied whether primary retroperitoneal liposarcomas diagnosed as myxoid/round cell liposarcoma represent molecularly true myxoid/round cell liposarcoma or are histopathological mimics and represent well-/dedifferentiated liposarcoma. Primary retroperitoneal myxoid/round cell liposarcoma (n=16) were compared to primary extremity myxoid/round cell liposarcoma (n=20). Histopathological and immunohistochemical features were studied. Amplification status of the 12q13-15 region was studied using a multiplex ligation-dependent probe amplification analysis, and FUS-CHOP or EWS-CHOP translocations were studied using RT-PCR. In primary retroperitoneal myxoid/round cell liposarcoma, MDM2 and CDK4 staining was both positive in 12 of 15 cases. In primary extremity myxoid/round cell liposarcoma, MDM2 was negative in 18/20 and CDK4 was negative in all cases. Multiplex ligation-dependent probe amplification showed the amplification of 12q13-15 region in 16/16 primary retroperitoneal myxoid/round cell liposarcomas and in 1/20 primary extremity myxoid/round cell liposarcomas. Translocation was present in all (18/18) primary extremity myxoid

  13. Basal cells are the progenitors of primary tracheal epithelial cell cultures

    SciTech Connect

    Ford, J.R.; Terzaghi-Howe, M. Oak Ridge National Lab., TN )

    1992-01-01

    The goal of this study was to identify the cells from the rat tracheal epithelium which attach and proliferate in primary culture. When cells isolated from tracheas by enzymatic digestion were held in suspension at 37C for several hours most of the differentiated cells dies. The kinetics of this selective cell death were not dependent on the constituents of the holding medium. With time in suspension, the colony forming efficiency of the surviving cells increased two- to threefold. Comparison of the growth curves of cells held or plated directly showed no difference in the number of cells in the proliferating populations. Using two lectins, it was possible to monitor the loss of specific populations in suspension. BS1-B4 is a marker for basal cells and UEA-1 is a secretory cell marker. Only those cells that were BS1-B4 positive survived in suspension. Further, the colonies that formed in primary culture were positive for this marker. Single cell suspensions of cells were sorted by flow cytometry and a fivefold increase in the colony forming efficiency of BS1-B4 positive cells compared to that of the negative cells was observed. These findings suggest that the cells that survived in suspension and proliferated in culture originated from the basal cells of the trachea.

  14. Cell longevity and sustained primary growth in palm stems.

    PubMed

    Tomlinson, P Barry; Huggett, Brett A

    2012-12-01

    Longevity, or organismal life span, is determined largely by the period over which constituent cells can function metabolically. Plants, with modular organization (the ability continually to develop new organs and tissues) differ from animals, with unitary organization (a fixed body plan), and this difference is reflected in their respective life spans, potentially much longer in plants than animals. We draw attention to the observation that palm trees, as a group of monocotyledons without secondary growth comparable to that of lignophytes (plants with secondary growth from a bifacial cambium), retain by means of sustained primary growth living cells in their trunks throughout their organismal life span. Does this make palms the longest-lived trees because they can grow as individuals for several centuries? No conventional lignophyte retains living metabolically active differentiated cell types in its trunk for this length of time, even though the tree as a whole can exist for millennia. Does this contrast also imply that the long-lived cells in a palm trunk have exceptional properties, which allows this seeming immortality? We document the long-life of many tall palm species and their inherent long-lived stem cell properties, comparing such plants to conventional trees. We provide a summary of aspects of cell age and life span in animals and plants. Cell replacement is a feature of animal function, whereas conventional trees rely on active growth centers (meristems) to sustain organismal development. However, the long persistence of living cells in palm trunks is seen not as evidence for unique metabolic processes that sustain longevity, but is a consequence of unique constructional features. This conclusion suggests that the life span of plant cells is not necessarily genetically determined.

  15. Attachment of human primary osteoblast cells to modified polyethylene surfaces.

    PubMed

    Poulsson, Alexandra H C; Mitchell, Stephen A; Davidson, Marcus R; Johnstone, Alan J; Emmison, Neil; Bradley, Robert H

    2009-04-09

    Ultra-high-molecular-weight polyethylene (UHMWPE) has a long history of use in medical devices, primarily for articulating surfaces due to its inherent low surface energy which limits tissue integration. To widen the applications of UHMWPE, the surface energy can be increased. The increase in surface energy would improve the adsorption of proteins and attachment of cells to allow tissue integration, thereby allowing UHMWPE to potentially be used for a wider range of implants. The attachment and function of human primary osteoblast-like (HOB) cells to surfaces of UHMWPE with various levels of incorporated surface oxygen have been investigated. The surface modification of the UHMWPE was produced by exposure to a UV/ozone treatment. The resulting surface chemistry was studied using X-ray photoelectron spectroscopy (XPS), and the topography and surface structure were probed by atomic force microscopy (AFM) and scanning electron microscopy (SEM), which showed an increase in surface oxygen from 11 to 26 atom % with no significant change to the surface topography. The absolute root mean square roughness of both untreated and UV/ozone-treated surfaces was within 350-450 nm, and the water contact angles decreased with increasing oxygen incorporation, i.e., showing an increase in surface hydrophilicity. Cell attachment and functionality were assessed over a 21 day period for each cell-surface combination studied; these were performed using SEM and the alamarBlue assay to study cell attachment and proliferation and energy-dispersive X-ray (EDX) analysis to confirm extracellular mineral deposits, and total protein assay to examine the intra- and extracellular protein expressed by the cells. HOB cells cultured for 21 days on the modified UHMWPE surfaces with 19 and 26 atom % oxygen incorporated showed significantly higher cell densities compared to cells cultured on tissue culture polystyrene (TCPS) from day 3 onward. This indicated that the cells attached and proliferated more

  16. Primary Signet Ring Cell Carcinoma of the Prostate

    PubMed Central

    Warner, Jonathan N.; Nakamura, Leah Y.; Pacelli, Anna; Humphreys, Mitchell R.; Castle, Erik P.

    2010-01-01

    Nine patients treated with primary signet ring cell carcinoma of the prostate were identified among 29,783 cases of prostate cancer evaluated at Mayo Clinic from January 15, 1970, until January 2, 2009. A PubMed search of the English-language literature published from January 1, 1980, to January 1, 2010, was then performed using the key words signet ring cell and prostate, identifying 42 cases. This study reviews those cases, along with the additional 9 reported herein, and evaluates clinical characteristics, histologic diagnoses, treatment modalities, and outcomes. Mean age at diagnosis was 68 years (range, 50-85 years), and mean prostate-specific antigen level was 95.3 ng/mL (range, 1.9-536.0 ng/mL; to convert to μg/L, multiply by 1). Most patients (66%) had non–stage IV carcinoma, the most common Gleason sum was 8 (33%), and mean survival was 29 months. The presence of a primary signet ring cell carcinoma of the prostate was best confirmed by negative findings on gastrointestinal work-up, a positive stain for prostate-specific acid phosphatase, and negative carcinoembryonic antigen test results. PMID:21123640

  17. Primary cilia in the basal cells of equine epididymis: a serendipitous finding.

    PubMed

    Arrighi, Silvana

    2013-04-01

    Occurrence of a solitary cilium was an unexpected discovery while studying the ultrastructure of epididymal epithelium in equidae. Primary cilia were detected in epididymal basal cells of all individuals of the equines studied - horses, donkey and mules - independently from age and tract of the duct, emerging from the basal cell surface and insinuating into the intercellular spaces. More rarely solitary cilia occurred also at the luminal surface of the principal cells. The ciliary apparatus was constituted by a structurally typical basal body continuous with the finger-like ciliary shaft extending from the cell surface, and an adjacent centriole oriented at right angles to the basal body. The cilium was structured as the typical primary, non-motile cilia found in many mammalian cells, having a 9+0 microtubular pattern. The basal diplosome was randomly associated with other cellular organelles including the Golgi complex, the endoplasmic reticulum, the microfilament network, the plasma membrane, vesicles and pits. Primary ciliogenesis is a new and unexpected finding in the epididymal epithelium. A monitoring role of luminal factors and extracellular liquids might be attributed to this organelle, likely acting as chemical receptor of the luminal environment, thus modulating the epithelial function by a cell-to-cell crosstalk involving the entire epithelium.

  18. Arsenic trioxide promotes mitochondrial DNA mutation and cell apoptosis in primary APL cells and NB4 cell line.

    PubMed

    Meng, Ran; Zhou, Jin; Sui, Meng; Li, ZhiYong; Feng, GuoSheng; Yang, BaoFeng

    2010-01-01

    This study aimed to investigate the effects of arsenic trioxide (As(2)O(3)) on the mitochondrial DNA (mtDNA) of acute promyelocytic leukemia (APL) cells. The NB4 cell line was treated with 2.0 micromol/L As(2)O(3) in vitro, and the primary APL cells were treated with 2.0 micromol/L As(2)O(3) in vitro and 0.16 mg kg(-1) d(-1) As(2)O(3) in vivo. The mitochondrial DNA of all the cells above was amplified by PCR, directly sequenced and analyzed by Sequence Navigatore and Factura software. The apoptosis rates were assayed by flow cytometry. Mitochondrial DNA mutation in the D-loop region was found in NB4 and APL cells before As(2)O(3) use, but the mutation spots were remarkably increased after As(2)O(3) treatment, which was positively correlated to the rates of cellular apoptosis, the correlation coefficient: r (NB4-As2O3)=0.973818, and r (APL-As2O3)=0.934703. The mutation types include transition, transversion, codon insertion or deletion, and the mutation spots in all samples were not constant and regular. It is revealed that As(2)O(3) aggravates mtDNA mutation in the D-loop region of acute promyelocytic leukemia cells both in vitro and in vivo. Mitochondrial DNA might be one of the targets of As(2)O(3) in APL treatment.

  19. Poxvirus tropism for primary human leukocytes and hematopoietic cells.

    PubMed

    Yu, Qigui; Hu, Ningjie; Ostrowski, Mario

    2009-01-01

    Poxviruses including canarypox (ALVAC) and vaccinia viruses have, in recent years, received considerable attention as live vectors for the development of vaccines against infectious diseases such as AIDS, malaria, and tuberculosis. However, the cellular targets for viral infection within the human immune system and the consequences of infection for cells involved in the generation of immune responses have not been clearly delineated. Using recombinant enhanced green fluorescence protein (EGFP)-expressing ALVAC and vaccinia viruses, we have focused here on a side-by-side comparison of ALVAC and vaccinia virus tropism for cells from human peripheral blood and bone marrow. Both ALVAC and vaccinia viruses showed a strong bias toward monocyte infection. ALVAC minimally infected CD19(+) B cells and was unable to infect ex vivo NK cells and T lymphocytes, whereas vaccinia virus could infect B lymphocytes and NK cell populations. Vaccinia virus was also able to infect T lymphocytes at low but detectable levels which could be enhanced upon their activation. Both ALVAC and vaccinia viruses could infect immature monocyte-derived dendritic cells (MDDCs), but only ALVAC infection induced their subsequent maturation. Infection in human bone marrow cells showed that ALVAC infection was restricted to a myelomonocytoid cell-specific CD33(+) cell population, while vaccinia virus showed a strong, but not exclusive, preference for these cells.

  20. Cancer Stem Cells in Primary Liver Cancers: Pathological Concepts and Imaging Findings

    PubMed Central

    Joo, Ijin; Kim, Haeryoung

    2015-01-01

    There is accumulating evidence that cancer stem cells (CSCs) play an integral role in the initiation of hepatocarcinogenesis and the maintaining of tumor growth. Liver CSCs derived from hepatic stem/progenitor cells have the potential to differentiate into either hepatocytes or cholangiocytes. Primary liver cancers originating from CSCs constitute a heterogeneous histopathologic spectrum, including hepatocellular carcinoma, combined hepatocellular-cholangiocarcinoma, and intrahepatic cholangiocarcinoma with various radiologic manifestations. In this article, we reviewed the recent concepts of CSCs in the development of primary liver cancers, focusing on their pathological and radiological findings. Awareness of the pathological concepts and imaging findings of primary liver cancers with features of CSCs is critical for accurate diagnosis, prediction of outcome, and appropriate treatment options for patients. PMID:25598674

  1. Primary signet ring cell carcinoma of the colon and rectum.

    PubMed

    Arifi, Samia; Elmesbahi, Omar; Amarti Riffi, Afaf

    2015-10-01

    Colorectal primary signet ring cell carcinoma (SRCC) is a rare entity accounting for nearly 1% of all colorectal carcinomas. It is an independent prognostic factor associated with less favorable outcome. This aggressiveness is mainly due to the intrinsic biology of these tumors. Here is an overview of the literature related to clinicopathological features, molecular biology, and management of SRCC of the colon and the rectum. Copyright © 2015 Société Française du Cancer. Published by Elsevier Masson SAS. All rights reserved.

  2. Exome-level comparison of primary well-differentiated neuroendocrine tumors and their cell lines.

    PubMed

    Boora, Ganesh K; Kanwar, Rahul; Kulkarni, Amit A; Pleticha, Josef; Ames, Matthew; Schroth, Gary; Beutler, Andreas S; Banck, Michaela S

    2015-01-01

    Neuroendocrine cancer cell lines are used to investigate therapeutic targets in neuroendocrine tumors (NET) and have been instrumental in the design of clinical trials targeting the PI3K/AKT/mTOR pathways, VEGF inhibitors, and somatostatin analogues. It remains unknown, however, whether the genomic makeup of NET cell lines reflect that of primary NET since comprehensive unbiased genome sequencing has not been performed on the cell lines. Four bronchopulmonary NET (BP-NET)-NCI-H720, NCI-H727, NCI-H835, and UMC11-and two pancreatic neuroendocrine tumors (panNET)-BON-1 and QGP1-were cultured. DNA was isolated, and exome sequencing was done. GATK and EXCAVATOR were used for bioinformatic analysis. We detected a total of 1,764 nonsynonymous single nucleotide variants at a rate of 8 per Mb in BP-NET and 4.3 per Mb in panNET cell lines, including 52 mutated COSMIC cancer genes in these cell lines, such as TP53, BRCA1, RB1, TSC2, NOTCH1, EP300, GNAS, KDR, STK11, and APC but not ATRX, DAXX, nor MEN1. Our data suggest that mutation rate, the pattern of copy number variations, and the mutational spectra in the BP-NET cell lines are more similar to the changes observed in small cell lung cancer than those found in primary BP-NET. Likewise, mutation rate and pattern including the absence of mutations in ATRX/DAXX, MEN1, and YY1 in the panNET cell lines BON1 and QGP1 suggest that these cell lines do not have the genetic signatures of a primary panNET. These results suggest that results from experiments with BP-NET and panNET cell lines need to be interpreted with caution.

  3. Defining an EPOR- Regulated Transcriptome for Primary Progenitors, including Tnfr-sf13c as a Novel Mediator of EPO- Dependent Erythroblast Formation

    PubMed Central

    Singh, Seema; Dev, Arvind; Verma, Rakesh; Pradeep, Anamika; Sathyanarayana, Pradeep; Green, Jennifer M.; Narayanan, Aishwarya; Wojchowski, Don M.

    2012-01-01

    Certain concepts concerning EPO/EPOR action modes have been challenged by in vivo studies: Bcl-x levels are elevated in maturing erythroblasts, but not in their progenitors; truncated EPOR alleles that lack a major p85/PI3K recruitment site nonetheless promote polycythemia; and Erk1 disruption unexpectedly bolsters erythropoiesis. To discover novel EPO/EPOR action routes, global transcriptome analyses presently are applied to interrogate EPO/EPOR effects on primary bone marrow-derived CFUe-like progenitors. Overall, 160 EPO/EPOR target transcripts were significantly modulated 2-to 21.8-fold. A unique set of EPO-regulated survival factors included Lyl1, Gas5, Pim3, Pim1, Bim, Trib3 and Serpina 3g. EPO/EPOR-modulated cell cycle mediators included Cdc25a, Btg3, Cyclin-d2, p27-kip1, Cyclin-g2 and CyclinB1-IP-1. EPO regulation of signal transduction factors was also interestingly complex. For example, not only Socs3 plus Socs2 but also Spred2, Spred1 and Eaf1 were EPO-induced as negative-feedback components. Socs2, plus five additional targets, further proved to comprise new EPOR/Jak2/Stat5 response genes (which are important for erythropoiesis during anemia). Among receptors, an atypical TNF-receptor Tnfr-sf13c was up-modulated >5-fold by EPO. Functionally, Tnfr-sf13c ligation proved to both promote proerythroblast survival, and substantially enhance erythroblast formation. The EPOR therefore engages a sophisticated set of transcriptome response circuits, with Tnfr-sf13c deployed as one novel positive regulator of proerythroblast formation. PMID:22808010

  4. Defining an EPOR- regulated transcriptome for primary progenitors, including Tnfr-sf13c as a novel mediator of EPO- dependent erythroblast formation.

    PubMed

    Singh, Seema; Dev, Arvind; Verma, Rakesh; Pradeep, Anamika; Sathyanarayana, Pradeep; Green, Jennifer M; Narayanan, Aishwarya; Wojchowski, Don M

    2012-01-01

    Certain concepts concerning EPO/EPOR action modes have been challenged by in vivo studies: Bcl-x levels are elevated in maturing erythroblasts, but not in their progenitors; truncated EPOR alleles that lack a major p85/PI3K recruitment site nonetheless promote polycythemia; and Erk1 disruption unexpectedly bolsters erythropoiesis. To discover novel EPO/EPOR action routes, global transcriptome analyses presently are applied to interrogate EPO/EPOR effects on primary bone marrow-derived CFUe-like progenitors. Overall, 160 EPO/EPOR target transcripts were significantly modulated 2-to 21.8-fold. A unique set of EPO-regulated survival factors included Lyl1, Gas5, Pim3, Pim1, Bim, Trib3 and Serpina 3g. EPO/EPOR-modulated cell cycle mediators included Cdc25a, Btg3, Cyclin-d2, p27-kip1, Cyclin-g2 and CyclinB1-IP-1. EPO regulation of signal transduction factors was also interestingly complex. For example, not only Socs3 plus Socs2 but also Spred2, Spred1 and Eaf1 were EPO-induced as negative-feedback components. Socs2, plus five additional targets, further proved to comprise new EPOR/Jak2/Stat5 response genes (which are important for erythropoiesis during anemia). Among receptors, an atypical TNF-receptor Tnfr-sf13c was up-modulated >5-fold by EPO. Functionally, Tnfr-sf13c ligation proved to both promote proerythroblast survival, and substantially enhance erythroblast formation. The EPOR therefore engages a sophisticated set of transcriptome response circuits, with Tnfr-sf13c deployed as one novel positive regulator of proerythroblast formation.

  5. Use of Composite Protein Database including Search Result Sequences for Mass Spectrometric Analysis of Cell Secretome

    PubMed Central

    Shin, Jihye; Kim, Gamin; Kabir, Mohammad Humayun; Park, Seong Jun; Lee, Seoung Taek; Lee, Cheolju

    2015-01-01

    Mass spectrometric (MS) data of human cell secretomes are usually run through the conventional human database for identification. However, the search may result in false identifications due to contamination of the secretome with fetal bovine serum (FBS) proteins. To overcome this challenge, here we provide a composite protein database including human as well as 199 FBS protein sequences for MS data search of human cell secretomes. Searching against the human-FBS database returned more reliable results with fewer false-positive and false-negative identifications compared to using either a human only database or a human-bovine database. Furthermore, the improved results validated our strategy without complex experiments like SILAC. We expect our strategy to improve the accuracy of human secreted protein identification and to also add value for general use. PMID:25822838

  6. Efficient Transformation of Primary Human Mesenchymal Stromal Cells by Adenovirus Early Region 1 Oncogenes.

    PubMed

    Speiseder, Thomas; Hofmann-Sieber, Helga; Rodríguez, Estefanía; Schellenberg, Anna; Akyüz, Nuray; Dierlamm, Judith; Spruss, Thilo; Lange, Claudia; Dobner, Thomas

    2017-01-01

    Previous observations that human amniotic fluid cells (AFC) can be transformed by human adenovirus type 5 (HAdV-5) E1A/E1B oncogenes prompted us to identify the target cells in the AFC population that are susceptible to transformation. Our results demonstrate that one cell type corresponding to mesenchymal stem/stroma cells (hMSCs) can be reproducibly transformed by HAdV-5 E1A/E1B oncogenes as efficiently as primary rodent cultures. HAdV-5 E1-transformed hMSCs exhibit all properties commonly associated with a high grade of oncogenic transformation, including enhanced cell proliferation, anchorage-independent growth, increased growth rate, and high telomerase activity as well as numerical and structural chromosomal aberrations. These data confirm previous work showing that HAdV preferentially transforms cells of mesenchymal origin in rodents. More importantly, they demonstrate for the first time that human cells with stem cell characteristics can be completely transformed by HAdV oncogenes in tissue culture with high efficiency. Our findings strongly support the hypothesis that undifferentiated progenitor cells or cells with stem cell-like properties are highly susceptible targets for HAdV-mediated cell transformation and suggest that virus-associated tumors in humans may originate, at least in part, from infections of these cell types. We expect that primary hMSCs will replace the primary rodent cultures in HAdV viral transformation studies and are confident that these investigations will continue to uncover general principles of viral oncogenesis that can be extended to human DNA tumor viruses as well. It is generally believed that transformation of primary human cells with HAdV-5 E1 oncogenes is very inefficient. However, a few cell lines have been successfully transformed with HAdV-5 E1A and E1B, indicating that there is a certain cell type which is susceptible to HAdV-mediated transformation. Interestingly, all those cell lines have been derived from human

  7. Cost-Effectiveness of Including a Nurse Specialist in the Treatment of Urinary Incontinence in Primary Care in the Netherlands

    PubMed Central

    Holtzer-Goor, K. M.; Gaultney, J. G.; van Houten, P.; Wagg, A. S.; Huygens, S. A.; Nielen, M. M. J.; Albers-Heitner, C. P.; Redekop, W. K.; Rutten-van Mölken, M. P.; Al, M. J.

    2015-01-01

    Objective Incontinence is an important health problem. Effectively treating incontinence could lead to important health gains in patients and caregivers. Management of incontinence is currently suboptimal, especially in elderly patients. To optimise the provision of incontinence care a global optimum continence service specification (OCSS) was developed. The current study evaluates the costs and effects of implementing this OCSS for community-dwelling patients older than 65 years with four or more chronic diseases in the Netherlands. Method A decision analytic model was developed comparing the current care pathway for urinary incontinence in the Netherlands with the pathway as described in the OCSS. The new care strategy was operationalised as the appointment of a continence nurse specialist (NS) located with the general practitioner (GP). This was assumed to increase case detection and to include initial assessment and treatment by the NS. The analysis used a societal perspective, including medical costs, containment products (out-of-pocket and paid by insurer), home care, informal care, and implementation costs. Results With the new care strategy a QALY gain of 0.005 per patient is achieved while saving €402 per patient over a 3 year period from a societal perspective. In interpreting these findings it is important to realise that many patients are undetected, even in the new care situation (36%), or receive care for containment only. In both of these groups no health gains were achieved. Conclusion Implementing the OCSS in the Netherlands by locating a NS in the GP practice is likely to reduce incontinence, improve quality of life, and reduce costs. Furthermore, the study also highlighted that various areas of the continence care process lack data, which would be valuable to collect through the introduction of the NS in a study setting. PMID:26426124

  8. Breast Implant Informed Consent Should Include the Risk of Anaplastic Large Cell Lymphoma.

    PubMed

    Clemens, Mark W; Miranda, Roberto N; Butler, Charles E

    2016-04-01

    Breast implant-associated anaplastic large cell lymphoma (ALCL) is a rare T-cell lymphoma arising around breast implants. Public awareness has increased following a safety communication warning of the association of breast implant-associated ALCL by the U.S. Food and Drug Administration in 2011. Difficulty with determining an accurate assessment of risk, including diagnosis, or standardized treatment regimen has led surgeons to commonly omit preoperative discussion of this rare and frequently misunderstood cancer. Risk disclosure is a form of respect for patient autonomy, and informed consent has positive practical and moral consequences for the practice of plastic surgery. A model of breast implant-associated ALCL informed consent implementation and health care provider education are reviewed with 1-year process follow-up at a tertiary cancer center. Breast implant-associated ALCL should be included during preoperative counseling on the risks of breast implantation when obtaining informed consent. Pertinent aspects of decision-making include disease awareness, presenting symptoms, and resources for concerned patients. Education of health care professionals and provision of patient-focused materials ensures effectiveness of the informed consent process.

  9. Reduction of CD147 surface expression on primary T cells leads to enhanced cell proliferation.

    PubMed

    Biegler, Brian; Kasinrerk, Watchara

    2012-12-01

    CD147 is a ubiquitously expressed membrane glycoprotein that has numerous functional associations in health and disease. However, the molecular mechanisms by which CD147 participates in these processes are unclear. Establishing physiologically relevant silencing of CD147 in primary T cells could provide clues essential for elucidating some aspects of CD147 biology. To date, achieving the knockdown of CD147 in primary T cells has remained elusive. Utilizing RNA interference and the Nucleofector transfection system, we were able to reduce the expression of CD147 in primary T cells. Comparison of basic functions, such as proliferation and CD25 expression, were then made between control populations and populations with reduced expression. Up-regulation of CD147 was found upon T-cell activation, indicating a role in T-cell responses. To better understand the possible importance of this up-regulation, we knocked down the expression of CD147 using RNA interference. When compared to control populations the CD147 knockdown populations exhibited increased proliferation. This alteration of cell proliferation, however, was not linked to a change in CD25 expression. We achieved reduction of CD147 surface expression in primary T cells by siRNA-mediated gene silencing. Our results point to CD147 having a possible negative regulatory role in T cell-mediated immune responses.

  10. 9 CFR 113.51 - Requirements for primary cells used for production of biologics.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Requirements for primary cells used... VECTORS STANDARD REQUIREMENTS Ingredient Requirements § 113.51 Requirements for primary cells used for production of biologics. Primary cells used to prepare biological products shall be derived from...

  11. 9 CFR 113.51 - Requirements for primary cells used for production of biologics.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Requirements for primary cells used... VECTORS STANDARD REQUIREMENTS Ingredient Requirements § 113.51 Requirements for primary cells used for production of biologics. Primary cells used to prepare biological products shall be derived from...

  12. Exposure to zidovudine adversely affects mitochondrial turnover in primary T cells.

    PubMed

    Wallace, Zoë R; Sanderson, Sharon; Simon, Anna Katarina; Dorrell, Lucy

    2016-09-01

    Zidovudine (ZDV) is a widely used component of antiretroviral therapy (ART) in resource-limited settings, despite its known adverse effects, which include mitochondrial toxicity in muscle, liver and adipose tissue. It has also been associated with impaired immunological recovery. We hypothesised that ZDV might impair mitochondrial health and survival of primary T cells. We performed a cross-sectional analysis of mitochondrial function, mitophagy and susceptibility to apoptosis in healthy donor primary T cells after exposure to ZDV in vitro, together with T cells from patients who were virologically suppressed on ZDV-containing ART regimens for ≥1 year and age-matched subjects receiving non-ZDV ART regimens. The proportion of T cells expressing mitochondrial reactive oxygen species (mtROS) was significantly higher after in vitro (CD4(+) T cells and CD8(+) T cells) and in vivo (CD4(+) T cells) exposure to ZDV than other antiretroviral agents. We did not detect any effect of ZDV on mitophagy, as indicated by change in autophagic flux. However, spontaneous apoptosis, indicated by upregulation of caspase-3 was greater in ZDV-exposed T cells. In conclusion, ZDV exposure was associated with impaired mitochondrial turnover and increased susceptibility to apoptosis in T cells. These mechanisms could contribute to sub-optimal immune reconstitution.

  13. Cyclooxygenase-2 expression in primary and metastatic Merkel cell carcinoma.

    PubMed

    Joachims, Zohar; Feinmesser, Raphael; Purim, Ofer; Halpern, Marisa; Brenner, Baruch; Fenig, Eyal; Roizman, Pepi; Sulkes, Jaqueline; Feinmesser, Meora

    2008-10-01

    Cyclooxygenase-2 (COX-2) is involved in the development and progression of many tumors, and its inhibition has been shown to block tumor growth. This study examined COX-2 expression in primary and metastatic Merkel cell carcinoma (MCC). Formalin-fixed paraffin-embedded tissues from 26 primary MCCs and 7 lymph node metastases were stained immunohistochemically with a monoclonal antibody directed against COX-2, and the percentage and intensity of staining were analyzed semiquantitatively. Immunopositivity for COX-2 was found in 20 primary tumors (77%), and was diffuse in 16 of them (80%). Staining intensity was strong in 5 tumors (19%), moderate in 6 (23%), and weak in 9 (35%). Five metastases (71%) showed similar staining. Prominent mitotic activity was associated with more diffuse COX-2 immunopositivity. No association was found between COX-2 expression and outcome. This study confirms that most MCCs express COX-2 and shows that COX-2 expression is related to one parameter of aggressive behavior--a high mitotic rate--but not to any others. The possibility of treating MCC with COX-2 inhibitors should be considered.

  14. Expression of tight junction proteins in epithelium including Ck20-positive M-like cells of human adenoids in vivo and in vitro.

    PubMed

    Takano, Ken-ichi; Kojima, Takashi; Ogasawara, Noriko; Go, Mitsuru; Kikuchi, Shin; Ninomiya, Takafumi; Kurose, Makoto; Koizumi, Jun-ichi; Kamekura, Ryuta; Murata, Masaki; Tanaka, Satoshi; Chiba, Hideki; Himi, Tetsuo; Sawada, Norimasa

    2008-06-01

    The human adenoid epithelium forms a continuous barrier against a wide variety of exogenous antigens. In this study, to elucidate the structures of the epithelial barrier in the human adenoid, including M-cells, we identified M-cells using an anti-cytokeratin 20 (Ck20) antibody and investigated expression of tight junction proteins in human adenoid epithelium in vivo and in vitro. In human adenoid epithelium and primary cultures, mRNAs of occludin, junctional adhesion molecule-A, ZO-1, and claudin-1, -4, -7, and -8 were detected by reverse transcription-polymerase chain reaction, whereas claudin-2 and -9 were expressed in vitro. In the epithelium in vivo, some Ck20-positive cells were randomly observed and indicated pocket-like structures, whereas Ck7 was positive in almost cells. Transmission electron microscopy revealed that Ck20-associated gold particles could be identified in M-like cells which had short microvilli and harboured the lymphocyte in the pocket-like structure. In primary cultures in vitro, Ck20-positive cells were also detected and had a function to take up fluorescent microparticles. In Ck20-positive cells in vivo and in vitro, expression of occludin, ZO-1, claudin-1 and -7 were observed at cell borders. These results indicate that the epithelial barrier of the human adenoid is stably maintained by expression of tight junction proteins in the epithelium including Ck20-positive M-like cells.

  15. Selective isolation and characterization of primary cells from normal breast and tumors reveal plasticity of adipose derived stem cells.

    PubMed

    Weigand, Annika; Boos, Anja M; Tasbihi, Kereshmeh; Beier, Justus P; Dalton, Paul D; Schrauder, Michael; Horch, Raymund E; Beckmann, Matthias W; Strissel, Pamela L; Strick, Reiner

    2016-03-12

    There is a need to establish more cell lines from breast tumors in contrast to immortalized cell lines from metastatic effusions in order to represent the primary tumor and not principally metastatic biology of breast cancer. This investigation describes the simultaneous isolation, characterization, growth and function of primary mammary epithelial cells (MEC), mesenchymal cells (MES) and adipose derived stem cells (ADSC) from four normal breasts, one inflammatory and one triple-negative ductal breast tumors. A total of 17 cell lines were established and gene expression was analyzed for MEC and MES (n = 42) and ADSC (n = 48) and MUC1, pan-KRT, CD90 and GATA-3 by immunofluorescence. DNA fingerprinting to track cell line identity was performed between original primary tissues and isolates. Functional studies included ADSC differentiation, tumor MES and MEC invasion co-cultured with ADSC-conditioned media (CM) and MES adhesion and growth on 3D-printed scaffolds. Comparative analysis showed higher gene expression of EPCAM, CD49f, CDH1 and KRTs for normal MEC lines; MES lines e.g. Vimentin, CD10, ACTA2 and MMP9; and ADSC lines e.g. CD105, CD90, CDH2 and CDH11. Compared to the mean of all four normal breast cell lines, both breast tumor cell lines demonstrated significantly lower ADSC marker gene expression, but higher expression of mesenchymal and invasion gene markers like SNAI1 and MMP2. When compared with four normal ADSC differentiated lineages, both tumor ADSC showed impaired osteogenic and chondrogenic but enhanced adipogenic differentiation and endothelial-like structures, possibly due to high PDGFRB and CD34. Addressing a functional role for overproduction of adipocytes, we initiated 3D-invasion studies including different cell types from the same patient. CM from ADSC differentiating into adipocytes induced tumor MEC 3D-invasion via EMT and amoeboid phenotypes. Normal MES breast cells adhered and proliferated on 3D-printed scaffolds containing 20 fibers

  16. Inhibition of glutaminase selectively suppresses the growth of primary acute myeloid leukemia cells with IDH mutations.

    PubMed

    Emadi, Ashkan; Jun, Sung Ah; Tsukamoto, Takashi; Fathi, Amir T; Minden, Mark D; Dang, Chi V

    2014-04-01

    The incidence of mutations in isocitrate dehydrogenase 1 and 2 (IDH1/2) in de novo acute myeloid leukemia (AML) is approximately 20%. These mutations result in distinct metabolic characteristics including dependency of cancer cells on glutamine as the main source for α-ketoglutarate, which is consumed by leukemia cells to produce a cancer-derived metabolite, 2-hydroxyglutarate. We sought to exploit this glutamine addiction therapeutically in mutant IDH primary AML cells from patients by measuring cell growth after exposure to a small molecule glutaminase inhibitor, BPTES. We found that BPTES only suppressed the growth of AML cells expressing mutant IDH compared with those expressing wild type IDH. This study lays the groundwork for strategies to target a specific subtype of AML metabolically with IDH mutations with a unique reprogramming of intermediary metabolism that culminates in glutamine dependency of cancer cells for survival.

  17. Sheep primary cells as in vitro models to investigate Mycoplasma agalactiae host cell interactions.

    PubMed

    Hegde, Shrilakshmi; Gabriel, Cordula; Kragl, Martin; Chopra-Dewasthaly, Rohini

    2015-10-01

    Appropriate infection models are imperative for the understanding of pathogens like mycoplasmas that are known for their strict host and tissue specificity, and lack of suitable cell and small animal models has hindered pathogenicity studies. This is particularly true for the economically important group of ruminant mycoplasmas whose virulence factors need to be elucidated for designing effective intervention strategies. Mycoplasma agalactiae serves as a useful role model especially because it is phylogenetically very close to M. bovis and causes similar symptoms by as yet unknown mechanisms. Here, we successfully prepared and characterized four different primary sheep cell lines, namely the epithelial and stromal cells from the mammary gland and uterus, respectively. Using immunohistochemistry, we identified vimentin and cytokeratin as specific markers to confirm the typical cell phenotypes of these primary cells. Furthermore, M. agalactiae's consistent adhesion and invasion into these primary cells proves the reliability of these cell models. Mimicking natural infections, mammary epithelial and stromal cells showed higher invasion and adhesion rates compared to the uterine cells as also seen via double immunofluorescence staining. Altogether, we have generated promising in vitro cell models to study host-pathogen interactions of M. agalactiae and related ruminant pathogens in a more authentic manner.

  18. Primary Cell Wall Composition of Bryophytes and Charophytes

    PubMed Central

    POPPER, ZOË A.; FRY, STEPHEN C.

    2003-01-01

    Major differences in primary cell wall (PCW) components between non‐vascular plant taxa are reported. (1) Xyloglucan: driselase digestion yielded isoprimeverose (the diagnostic repeat unit of xyloglucan) from PCW‐rich material of Anthoceros (a hornwort), mosses and both leafy and thalloid liverworts, as well as numerous vascular plants, showing xyloglucan to be a PCW component in all land plants tested. In contrast, charophycean green algae (Klebsormidium flaccidium, Coleochaete scutata and Chara corallina), thought to be closely related to land plants, did not contain xyloglucan. They did not yield isoprimeverose; additionally, charophyte material was not digestible with xyloglucan‐specific endoglucanase or cellulase to give xyloglucan‐derived oligosaccharides. (2) Uronic acids: acid hydrolysis of PCW‐rich material from the charophytes, the hornwort, thalloid and leafy liverworts and a basal moss yielded higher concentrations of glucuronic acid than that from the remaining land plants including the less basal mosses and all vascular plants tested. Polysaccharides of the hornwort Anthoceros contained an unusual repeat‐unit, glucuronic acid‐α(1→3)‐galactose, not found in appreciable amounts in any other plants tested. Galacturonic acid was consistently the most abundant PCW uronic acid, but was present in higher concentrations in acid hydrolysates of bryophytes and charophytes than in those of any of the vascular plants. Mannuronic acid was not detected in any of the species surveyed. (3) Mannose: acid hydrolysis of charophyte and bryophyte PCW‐rich material also yielded appreciably higher concentrations of mannose than are found in vascular plant PCWs. (4) Mixed‐linkage glucan (MLG) was absent from all algae and bryophytes tested; however, upon digestion with licheninase, PCW‐rich material from the alga Ulva lactuca and the leafy liverwort Lophocolea bidentata yielded penta‐ to decasaccharides, indicating the presence of MLG

  19. Primary cell wall composition of bryophytes and charophytes.

    PubMed

    Popper, Zoë A; Fry, Stephen C

    2003-01-01

    Major differences in primary cell wall (PCW) components between non-vascular plant taxa are reported. (1) Xyloglucan: driselase digestion yielded isoprimeverose (the diagnostic repeat unit of xyloglucan) from PCW-rich material of Anthoceros (a hornwort), mosses and both leafy and thalloid liverworts, as well as numerous vascular plants, showing xyloglucan to be a PCW component in all land plants tested. In contrast, charophycean green algae (Klebsormidium flaccidium, Coleochaete scutata and Chara corallina), thought to be closely related to land plants, did not contain xyloglucan. They did not yield isoprimeverose; additionally, charophyte material was not digestible with xyloglucan-specific endoglucanase or cellulase to give xyloglucan-derived oligosaccharides. (2) Uronic acids: acid hydrolysis of PCW-rich material from the charophytes, the hornwort, thalloid and leafy liverworts and a basal moss yielded higher concentrations of glucuronic acid than that from the remaining land plants including the less basal mosses and all vascular plants tested. Polysaccharides of the hornwort Anthoceros contained an unusual repeat-unit, glucuronic acid-alpha(1-->3)-galactose, not found in appreciable amounts in any other plants tested. Galacturonic acid was consistently the most abundant PCW uronic acid, but was present in higher concentrations in acid hydrolysates of bryophytes and charophytes than in those of any of the vascular plants. Mannuronic acid was not detected in any of the species surveyed. (3) Mannose: acid hydrolysis of charophyte and bryophyte PCW-rich material also yielded appreciably higher concentrations of mannose than are found in vascular plant PCWs. (4) Mixed-linkage glucan (MLG) was absent from all algae and bryophytes tested; however, upon digestion with licheninase, PCW-rich material from the alga Ulva lactuca and the leafy liverwort Lophocolea bidentata yielded penta- to decasaccharides, indicating the presence of MLG-related polysaccharides. Our

  20. A Multiplex Immunoassay Using the Guthrie Specimen to Detect T-Cell Deficiencies Including Severe Combined Immunodeficiency Disease

    PubMed Central

    Janik, David K.; Lindau-Shepard, Barbara; Comeau, Anne Marie; Pass, Kenneth A.

    2011-01-01

    BACKGROUND Severe combined immunodeficiency (SCID) fulfills many of the requirements for addition to a newborn screening panel. Two newborn screening SCID pilot studies are now underway using the T-cell receptor excision circle (TREC) assay, a molecular technique. Here we describe an immunoassay with CD3 as a marker for T cells and CD45 as a marker for total leukocytes that can be used with the Guthrie specimen. METHODS The multiplexing capabilities of the Luminex platform were used. Antibody pairs were used to capture and detect CD3 and CD45 from a single 3-mm punch of the Guthrie specimen. The assay for each bio-marker was developed separately in identical buffers and then combined to create a multiplex assay. RESULTS Using calibrators made from known amounts of leukocytes, a detection limit of 0.25 × 106 cells/mL for CD3 and 0.125 × 106 cells/mL for CD45 was obtained. Affinity tests showed no cross-reactivity between the antibodies to CD3 and CD45. The multiplex assay was validated against 8 coded specimens of known clinical status and linked to results from the TREC assay that had identified them. All were correctly identified by the CD345 assay. CONCLUSIONS The performance parameters of the CD345 assay met the performance characteristics generally accepted for immunoassays. Our assay classifications of positive specimens concur with previous TREC results. This CD345 assay warrants evaluation as a viable alternative or complement to the TREC assay as a primary screening tool for detecting T-cell immunodeficiencies, including SCID, in Guthrie specimens. PMID:20660143

  1. Primary large cell neuroendocrine carcinoma of the urinary bladder.

    PubMed

    Evans, Andrew J; Al-Maghrabi, Jaudah; Tsihlias, John; Lajoie, Ginette; Sweet, Joan M; Chapman, William B

    2002-10-01

    Reports of primary large cell neuroendocrine carcinomas of the urinary bladder are few; we identified only 2 cases in the literature. Both of these cases involved male patients with rapid progression of disease culminating in death with widespread metastases. We report a case of primary large cell neuroendocrine carcinoma of the bladder, with an admixed minor element of adenocarcinoma, in an 82-year-old man. This solitary lesion arose in a bladder diverticulum lateral to the left ureteric orifice. Two attempts at transurethral resection were unsuccessful at achieving local control. The patient underwent a partial cystectomy with left-sided pelvic lymphadenectomy following preoperative staging investigations that found no metastatic disease. Pathologically, the tumor invaded into the deep aspect of the muscularis propria, without extension into perivesical fat. The lateral resection margin was microscopically positive for tumor, but no malignancy was found in the pelvic lymph nodes. The adenocarcinoma comprised less than 5% of total tumor volume, and areas of transition between the neuroendocrine and adenocarcinoma components were apparent. The patient developed a local recurrence 8 months postoperatively, which was managed by a combination of transurethral resection and radiation therapy. Currently, the patient has no evidence of local or metastatic disease 2 years after initial diagnosis.

  2. (18)F-FDG PET/CT in bilateral primary adrenal T-cell lymphoma.

    PubMed

    Santhosh, Sampath; Mittal, Bhagwant Rai; Shankar, Praveen; Kashyap, Raghava; Bhattacharya, Anish; Singh, Baljinder; Das, Ashim; Bhansali, Anil

    2011-01-01

    Primary adrenal lymphoma is extremely rare. We report a young patient who presented with non- specific symptoms of fever and abdominal pain. Conventional imaging modalities demonstrated bilateral bulky adrenal masses, and whole-body fluorine-18-fluorodesoxyglucose ((18)F-FDG) positron emission tomography/computed tomography showed intense (18)F-FDG-avid bilateral adrenal masses with no evidence of extra-adrenal spread. A pathological diagnosis of non-Hodgkin lymphoma of peripheral T-cell type was made. The present case indicates that primary adrenal lymphoma should be included in the differential diagnosis of bilateral adrenal masses.

  3. T Helper 17 Cells in Primary Sjögren’s Syndrome

    PubMed Central

    Matsui, Kiyoshi; Sano, Hajime

    2017-01-01

    Primary Sjögren’s syndrome is an autoimmune disease characterized by diffuse infiltration of lymphocytes into exocrine glands and other tissues. The infiltrating lymphocytes have been identified as subsets of B cells and T cells, including T helper 17 cells, T regulatory cells and follicular helper T cells. The role of these cells in the development of the syndrome is now known, as is their impact on the production of proinflammatory cytokines such as IL-6, IL-17, IL-22 and IL-23. In particular, experimental animal models and patients suggest that a shift in Th17/Treg balance toward the proinflammatory Th17 axis exacerbates primary Sjögren’s syndrome and other autoimmune disorders. Nevertheless, the pathogenesis of the disorder is not yet fully elucidated. This review summarizes the recent advances in therapeutic control of the Treg/Th17 balance, as well as the efficacy of candidate therapeutics against primary Sjögren’s syndrome. PMID:28678161

  4. Conditions for initiating Lake Victoria haplochromine (Oreochromis esculentus) primary cell cultures from caudal fin biopsies.

    PubMed

    Filice, Melissa; Lee, C; Mastromonaco, Gabriela F

    2014-10-01

    The global decline of freshwater fishes has created a need to cryopreserve biological materials from endangered species in an effort to conserve the biodiversity within this taxon. Since maternal gametes and embryos from fish are difficult to cryopreserve, somatic cells obtained from caudal fins have become an increasingly popular resource as they contain both maternal and paternal DNA ensuring valuable traits are not lost from the population. Somatic cells stored in cryobanks can be used to supplement endangered populations with genetically valuable offspring with the use of assisted reproductive technologies. However, initiating primary cell cultures from caudal fin biopsies of endangered species can be challenging as standardized protocols have not yet been developed. The objective of this study was to identify culture conditions, including antibiotic supplementation, biopsy size, and culture temperature, suitable for establishing primary cell cultures of ngege (Oreochromis esculentus), a critically endangered African cichlid. Six-millimeter caudal fin biopsies provided sufficient material to develop a primary cell culture when incubated at 25°C using standard fish cell culture medium containing 1× Primocin. Further investigation and application of these culture conditions for other endangered freshwater fishes is necessary.

  5. Increased power generation from primary sludge in microbial fuel cells coupled with prefermentation.

    PubMed

    Choi, Jeongdong; Ahn, Youngho

    2014-12-01

    Raw primary sludge and the prefermentation liquor (PL) of primary sludge were used to generate electricity in single-chambered air-cathode microbial fuel cells (MFCs). The MFCs treating the primary sludge produced 0.53 V and 370 mW/m(2) for the maximum potential and power density, respectively. In the primary sludge-fed MFCs, only 5 % of the total energy production was produced from direct electricity generation, whereas 95 % of that resulted from the conversion of methane to electricity. MFCs treating the PL generated the maximum potential of 0.58 V and maximum power density of 885 mW/m(2), respectively. In the energy production analysis, direct electricity production (1,921 Wh/kg TCODrem) in the MFCs treating the PL was much higher than that of the primary sludge-fed MFC (138 Wh/kg TCODrem). Volatile suspended solids during 10 days were reduced to 18.3 and 38 % in the primary sludge-fed MFCs and prefermentation reactor, respectively. These findings suggest that a two-stage process including prefermentation and MFCs is of great benefit on sludge reduction and higher electricity generation from primary sludge.

  6. Primary Clear Cell Microcystic Adenoma of the Sinonasal Cavity: Pathological or Fortuitous Association?

    PubMed Central

    Markham, Hannah; Theaker, Jeffery; Bateman, Adrian; Sommerlad, Matthew; Crawford, Gillian; Eccles, Diana

    2017-01-01

    Primary clear cell microcystic adenoma of the sinonasal cavity is rare. It has previously been described only as a VHL-associated tumour. Von Hippel-Lindau (VHL) syndrome is an inherited cancer syndrome characterised by an elevated risk of neoplasia including clear cell renal cell carcinoma (ccRCC), haemangioblastoma, and phaeochromocytoma. We describe the second reported case of a primary clear cell microcystic adenoma of the sinonasal cavity. The 39-year-old patient with VHL syndrome had previously undergone resection and ablation of ccRCC. He presented with epistaxis. Imaging demonstrated a mass in the ethmoid sinus. Initial clinical suspicion was of metastatic ccRCC. However, tumour morphology and immunoprofile were distinct from the previous ccRCC and supported a diagnosis of primary microcystic adenoma. Analysis of DNA extracted from sinonasal tumour tissue did not show loss of the wild-type allele at the VHL locus. Although this did not support tumour association with VHL disease, it was not possible to look for a loss-of-function mutation. The association of primary microcystic adenoma of the sinonasal cavity with VHL disease remains speculative. These lesions are benign but are likely to require regular surveillance. Such tumours may require repeated surgical excision. PMID:28261513

  7. Cathode catalysts for primary phosphoric acid fuel cells

    NASA Technical Reports Server (NTRS)

    1981-01-01

    Alkylation or carbon Vulcan XC-72, the support carbon, was shown to provide the most stable bond type for linking cobalt dehydrodibenzo tetraazannulene (CoTAA) to the surface of the carbon; this result is based on data obtained by cyclic voltammetry, pulse voltammetry and by release of 14C from bonded CoTAA. Half-cell tests at 100 C in 85% phosphoric acid showed that CoTAA bonded to the surface of carbon (Vulcan XC-72) via an alkylation procedure is a more active catalyst than is platinum based on a factor of two improvement in Tafel slope; dimeric CoTAA had catalytic activity equal to platinum. Half-cell tests also showed that bonded CoTAA catalysts do not suffer a loss in potential when air is used as a fuel rather than oxygen. Commercially available polytetrafluroethylene (PTFE) was shown to be unstable in the fuel cell environment with degradation occurring in 2000 hours or less. The PTFE was stressed at 200 C in concentrated phosphoric acid as well as electrochemically stressed in 150 C concentrated phosphoric acid; the surface chemistry of PTFE was observed to change significantly. Radiolabeled PTFE was prepared and used to verify that such chemical changes also occur in the primary fuel cell environment.

  8. Activity of resveratrol triesters against primary acute lymphoblastic leukemia cells.

    PubMed

    Urbaniak, Alicja; Delgado, Magdalena; Kacprzak, Karol; Chambers, Timothy C

    2017-06-15

    Resveratrol is a common polyphenol of plant origin known for its cancer prevention and other properties. Its wider application is limited due to poor water solubility, low stability, and weak bioavailability. To overcome these limitations, a series of 13 novel resveratrol triesters were synthesized previously. In this paper, we describe the synthesis of 3 additional derivatives and the activity of all 16 against primary acute lymphoblastic leukemia cells. Of these, 3 compounds were more potent than resveratrol (IC50=10.5µM) namely: resveratryl triacetate (IC50=3.4µM), resveratryl triisobutyrate (IC50=5.1µM), and resveratryl triisovalerate (IC50=4.9µM); all other derivatives had IC50 values of >10µM. Further studies indicated that the active compounds caused G1 phase arrest, increased expression of p53, and induced characteristics of apoptotic cell death. Moreover, the compounds were only effective in cycling cells, with cells arrested in G1 phase being refractory. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Kinetic modeling of rhamnolipid production by Pseudomonas aeruginosa PAO1 including cell density-dependent regulation.

    PubMed

    Henkel, Marius; Schmidberger, Anke; Vogelbacher, Markus; Kühnert, Christian; Beuker, Janina; Bernard, Thomas; Schwartz, Thomas; Syldatk, Christoph; Hausmann, Rudolf

    2014-08-01

    The production of rhamnolipid biosurfactants by Pseudomonas aeruginosa is under complex control of a quorum sensing-dependent regulatory network. Due to a lack of understanding of the kinetics applicable to the process and relevant interrelations of variables, current processes for rhamnolipid production are based on heuristic approaches. To systematically establish a knowledge-based process for rhamnolipid production, a deeper understanding of the time-course and coupling of process variables is required. By combining reaction kinetics, stoichiometry, and experimental data, a process model for rhamnolipid production with P. aeruginosa PAO1 on sunflower oil was developed as a system of coupled ordinary differential equations (ODEs). In addition, cell density-based quorum sensing dynamics were included in the model. The model comprises a total of 36 parameters, 14 of which are yield coefficients and 7 of which are substrate affinity and inhibition constants. Of all 36 parameters, 30 were derived from dedicated experimental results, literature, and databases and 6 of them were used as fitting parameters. The model is able to describe data on biomass growth, substrates, and products obtained from a reference batch process and other validation scenarios. The model presented describes the time-course and interrelation of biomass, relevant substrates, and products on a process level while including a kinetic representation of cell density-dependent regulatory mechanisms.

  10. Clinical Analysis of Primary Colorectal Signet-Ring Cell Carcinoma.

    PubMed

    Liang, ZhengZi; Yan, DengGuo; Li, GuoSheng; Cheng, HaiYu

    2017-07-08

    The objective of the study was to investigate the clinicopathological features of primary colorectal signet-ring cell carcinoma. We retrospectively analyzed the clinical and survival data of 37 patients with primary colorectal signet-ring cell carcinoma. The mean survival time of patients in stage II, III, and IV were estimated using Student t test and the cumulative survival rates were estimated according to the method of Kaplan-Meier. The significance of the differences in survival rates were calculated using the log rank test. The incidence of primary colorectal signet-ring cell carcinoma was 1.40%, the median age of 37 patients was 50 years, the male to female ratio was 1.47:1, and 21 patients (56.8%) received a radical resection. Most patients 33 (89.2%) had an advanced tumor stage at the time of diagnosis (17 patients 45.9% stage III and 16 patients 43.2% stage IV), 34 (94.5%) patients showed a tumor depth of >T3, lymph node involvement occurred in 26 patients (70.3%), patients had a high incidence of peritoneal metastasis (16 patients 43.2% at presentation, 30 patients 81.1% at presentation and recurrence) and a low incidence of liver metastases (1 patients 2.7% at presentation, 5 patients 13.5% at presentation and recurrence). The 5-year survival rate after the initial surgery was 10.8%, the mean survival time of 37 patients was 27.1 ± 3.3 months, the mean survival time of patients in stage II, III, and IV were 47.0 ± 12.8 months, 37.1 ± 3.9 months, and 10.5 ± 1.4 months, respectively (P < .000). Colorectal signet-ring cell carcinoma is a rare neoplasm with a predominance in men. Its characteristic features were the advanced stage at the time of diagnosis, a high incidence of peritoneal metastases, a low incidence of liver metastasis, and a poor prognosis. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Reference gene for primary culture of prostate cancer cells.

    PubMed

    Souza, Aline Francielle Damo; Brum, Ilma Simoni; Neto, Brasil Silva; Berger, Milton; Branchini, Gisele

    2013-04-01

    Selection of reference genes to normalize mRNA levels between samples is critical for gene expression studies because their expression can vary depending on the tissues or cells used and the experimental conditions. We performed ten cell cultures from samples of prostate cancer. Cells were divided into three groups: control (with no transfection protocol), cells transfected with siRNA specific to knockdown the androgen receptor and cells transfected with inespecific siRNAs. After 24 h, mRNA was extracted and gene expression was analyzed by Real-time qPCR. Nine candidates to reference genes for gene expression studies in this model were analyzed (aminolevulinate, delta-, synthase 1 (ALAS1); beta-actin (ACTB); beta-2-microglobulin (B2M); glyceraldehyde-3-phosphate dehydrogenase (GAPDH); hypoxanthine phosphoribosyltransferase 1 (HPRT1); succinate dehydrogenase complex, subunit A, flavoprotein (Fp) (SDHA); TATA box binding protein (TBP); ubiquitin C (UBC); tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein, zeta polypeptide (YWHAZ)). Expression stability was calculated NormFinder algorithm to find the most stable genes. NormFinder calculated SDHA as the most stable gene and the gene with the lowest intergroup and intragroup variation, and indicated GAPDH and SDHA as the best combination of two genes for the purpose of normalization. Androgen receptor mRNA expression was evaluated after normalization by each candidate gene and showed statistical difference in the transfected group compared to control group only when normalized by combination of GAPDH and SDHA. Based on the algorithm analysis, the combination of SDHA and GAPDH should be used to normalize target genes mRNA levels in primary culture of prostate cancer cells submitted to transfection with siRNAs.

  12. Tetherin Can Restrict Cell-Free and Cell-Cell Transmission of HIV from Primary Macrophages to T Cells

    PubMed Central

    Giese, Sebastian; Marsh, Mark

    2014-01-01

    Bst-2/Tetherin inhibits the release of HIV by tethering newly formed virus particles to the plasma membrane of infected cells. Although the mechanisms of Tetherin-mediated restriction are increasingly well understood, the biological relevance of this restriction in the natural target cells of HIV is unclear. Moreover, whether Tetherin exerts any restriction on the direct cell-cell spread of HIV across intercellular contacts remains controversial. Here we analyse the restriction endogenous Tetherin imposes on HIV transmission from primary human macrophages, one of the main targets of HIV in vivo. We find that the mRNA and protein levels of Tetherin in macrophages are comparable to those in T cells from the same donors, and are highly upregulated by type I interferons. Improved immunocytochemistry protocols enable us to demonstrate that Tetherin localises to the cell surface, the trans-Golgi network, and the macrophage HIV assembly compartments. Tetherin retains budded virions in the assembly compartments, thereby impeding the release and cell-free spread of HIV, but it is not required for the maintenance of these compartments per se. Notably, using a novel assay to quantify cell-cell spread, we show that Tetherin promotes the transfer of virus clusters from macrophages to T cells and thereby restricts the direct transmission of a dual-tropic HIV-1. Kinetic analyses provide support for the notion that this direct macrophage-T cell spread is mediated, at least in part, by so-called virological synapses. Finally, we demonstrate that the viral Vpu protein efficiently downregulates the cell surface and overall levels of Tetherin, and thereby abrogates this HIV restriction in macrophages. Together, our study shows that Tetherin, one of the most potent HIV restriction factors identified to date, can inhibit virus spread from primary macrophages, regardless of the mode of transmission. PMID:24991932

  13. Tumor necrosis factor (cachetin) decreases adipose cell differentiation in primary cell culture

    SciTech Connect

    Martin, R.J.; Jones, D.D.; Jewell, D.E.; Hausman, G.J.

    1986-03-05

    Cachetin has been shown to effect gene product expression in the established adipose cell line 3T3-L1. Expression of messenger RNA for lipoprotein lipase is suppressed in cultured adipocytes. The purpose of this study was to determine the effect of Cachetin on adipose cell differentiation in primary cell culture. Stromalvascular cells obtained from the inguinal fat pad of 4-5 week old Sprague-Dawley rats were grown in culture for two weeks. During the proliferative growth phase all cells were grown on the same medium and labelled with /sup 3/H-thymidine. Cachetin treatment (10/sup -6/ to 10/sup -10/ M) was initiated on day 5, the initial phase of preadipocyte differentiation. Adipocytes and stromal cells were separated using density gradient, and /sup 3/H-thymidine was determined for both cell types. Thymidine incorporation into adipose cells was decreased maximally (approx. 50%) at 10/sup -10/ M. Stromalvascular cells were not influenced at any of the doses tested. Adipose cell lipid content as indicated by oil red-O staining was decreased by Cachetin. Esterase staining by adipose cells treated with Cachetin was increased indicating an increase in intracellular lipase. These studies show that Cachetin has specific effects on primary adipose cell differentiation.

  14. Stem cell therapy for the treatment of Leydig cell dysfunction in primary hypogonadism

    PubMed Central

    Peak, Taylor C; Haney, Nora M; Wang, William; DeLay, Kenneth J; Hellstrom, Wayne J

    2016-01-01

    The production of testosterone occurs within the Leydig cells of the testes. When production fails at this level from either congenital, acquired, or systemic disorders, the result is primary hypogonadism. While numerous testosterone formulations have been developed, none are yet fully capable of replicating the physiological patterns of testosterone secretion. Multiple stem cell therapies to restore androgenic function of the testes are under investigation. Leydig cells derived from bone marrow, adipose tissue, umbilical cord, and the testes have shown promise for future therapy for primary hypogonadism. In particular, the discovery and utilization of a group of progenitor stem cells within the testes, known as stem Leydig cells (SLCs), has led not only to a better understanding of testicular development, but of treatment as well. When combining this with an understanding of the mechanisms that lead to Leydig cell dysfunction, researchers and physicians will be able to develop stem cell therapies that target the specific step in the steroidogenic process that is deficient. The current preclinical studies highlight the complex nature of regenerating this steroidogenic process and the problems remain unresolved. In summary, there appears to be two current directions for stem cell therapy in male primary hypogonadism. The first method involves differentiating adult Leydig cells from stem cells of various origins from bone marrow, adipose, or embryonic sources. The second method involves isolating, identifying, and transplanting stem Leydig cells into testicular tissue. Theoretically, in-vivo re-activation of SLCs in men with primary hypogonadism due to age would be another alternative method to treat hypogonadism while eliminating the need for transplantation. PMID:27822338

  15. Effects of Simulated Microgravity on Primary Human NK Cells

    PubMed Central

    Li, Qi; Mei, Qibing; Huyan, Ting; Xie, Li; Che, Su; Yang, Hui; Zhang, Mingjie

    2013-01-01

    Abstract The deleterious effects of microgravity on lymphocytes have been demonstrated in previous studies. However, research on the effects of microgravity on human natural killer (NK) cells remains exceedingly limited. In this study, we demonstrated that NK cell cytotoxicity was significantly decreased under simulated microgravity (SMG) conditions (p<0.05). Several processes, including apoptosis, receptor expression, and cytokine secretion, were investigated in human NK cells under SMG. We observed decreased cytotoxicity, concurrent with increased apoptosis and necrosis, in NK cells after exposure to SMG (p<0.05). Additionally, interferon (IFN)-γ and perforin expression decreased significantly, and the expression of granzyme-B was only slightly reduced. Meanwhile, SMG selectively inhibited the expression of certain surface receptors on NK cells. Specifically, the expression of NKG2A and NKG2D were significantly downregulated under SMG, but the expression of NKp30 and NKp44 was not affected. We also found that interleukin (IL)–15 alone or in combination with IL-12 could counteract the inhibition of NK cell cytotoxicity under SMG. Our findings indicate that human NK cells were sensitive to SMG, as reflected by their decreased cytotoxicity. Factors such as increased early apoptosis and late apoptosis/necrosis and the decreased expression of INF-γ, cytolytic proteins, and cell surface receptors may be responsible for the loss of cytotoxicity in human NK cells under SMG. A combination of IL-12 and IL-15 may be useful as a therapeutic strategy for overcoming the effects of microgravity on human NK cells during long space missions. Key Words: Simulated microgravity (SMG)—Natural killer (NK) cells—Cytotoxicity. Astrobiology 13, 703–714. PMID:23919749

  16. Individuals with Primary Osteoarthritis Have Different Phenotypes Depending on the Affected Joint - A Case Control Study from Southern Sweden Including 514 Participants

    PubMed Central

    Karlsson, Magnus K; Karlsson, Caroline; Magnusson, Håkan; Cöster, Maria; von Schewelov, Tord; Nilsson, Jan Åke; Brudin, Lars; Rosengren, Björn E

    2014-01-01

    Objective: The aim of this study was to evaluate whether primary osteoarthritis (OA), independent of affected joint, is associated with a phenotype that is different from the phenotype in a normative cohort. Material and Methods: We included 274 patients with primary OA, 30 women and 32 men (mean age 66 years, range 42-84) with primary hip OA, 38 women and 74 men (mean age 61 years; range 34-85) with primary knee OA, 42 women and 19 men (men age 64 years, range 42-87) with primary ankle or foot OA and 20 women and 19 men (mean age 66 years, range 47-88) with primary hand or finger OA. Of all patients included with OA, 23% had hip OA, 41% knee OA, 22% ankle or foot OA and 14% hand or finger OA. Serving as references were 122 women and 118 men of the same ages who were population-based, included as a control cohort. We measured total body BMD (g/cm2) and proportion of fat and lean mass (%) with dual energy X-ray absorptiometry. Height, weight and BMI (kg/m2) were also assessed. We then calculated Z-scores (number of standard deviations difference from the mean value of the control cohort) in the OA patients and compared these between the groups. Results: Individuals with hand OA and controls had similar phenotype. Individuals with lower extremity OA, irrespective of the affected joint, had similar weight, BMI and BMD, but higher than in individuals with hand OA and controls (all p<0.05). Individuals with lower extremity OA had higher fat and lower lean mass than individuals with hand OA and controls (all p<0.001). Conclusion: Individuals with primary OA in the lower extremity have a phenotype with higher BMD, higher BMI, proportionally higher fat content and lower lean body mass content. The different skeletal phenotypes in our patients with OA in the lower extremity and patients with hand OA indicate that separate pathophysiologic pathways may be responsible for primary OA in different joints PMID:25614774

  17. Reconstituting development of pancreatic intraepithelial neoplasia from primary human pancreas duct cells

    PubMed Central

    Lee, Jonghyeob; Snyder, Emily R.; Liu, Yinghua; Gu, Xueying; Wang, Jing; Flowers, Brittany M.; Kim, Yoo Jung; Park, Sangbin; Szot, Gregory L.; Hruban, Ralph H.; Longacre, Teri A.; Kim, Seung K.

    2017-01-01

    Development of systems that reconstitute hallmark features of human pancreatic intraepithelial neoplasia (PanINs), the precursor to pancreatic ductal adenocarcinoma, could generate new strategies for early diagnosis and intervention. However, human cell-based PanIN models with defined mutations are unavailable. Here, we report that genetic modification of primary human pancreatic cells leads to development of lesions resembling native human PanINs. Primary human pancreas duct cells harbouring oncogenic KRAS and induced mutations in CDKN2A, SMAD4 and TP53 expand in vitro as epithelial spheres. After pancreatic transplantation, mutant clones form lesions histologically similar to native PanINs, including prominent stromal responses. Gene expression profiling reveals molecular similarities of mutant clones with native PanINs, and identifies potential PanIN biomarker candidates including Neuromedin U, a circulating peptide hormone. Prospective reconstitution of human PanIN development from primary cells provides experimental opportunities to investigate pancreas cancer development, progression and early-stage detection. PMID:28272465

  18. Quantitative proteomics of extracellular vesicles derived from human primary and metastatic colorectal cancer cells.

    PubMed

    Choi, Dong-Sic; Choi, Do-Young; Hong, Bok Sil; Jang, Su Chul; Kim, Dae-Kyum; Lee, Jaewook; Kim, Yoon-Keun; Kim, Kwang Pyo; Gho, Yong Song

    2012-01-01

    Cancer cells actively release extracellular vesicles (EVs), including exosomes and microvesicles, into surrounding tissues. These EVs play pleiotropic roles in cancer progression and metastasis, including invasion, angiogenesis, and immune modulation. However, the proteomic differences between primary and metastatic cancer cell-derived EVs remain unclear. Here, we conducted comparative proteomic analysis between EVs derived from human primary colorectal cancer cells (SW480) and their metastatic derivatives (SW620). Using label-free quantitation, we identified 803 and 787 proteins in SW480 EVs and SW620 EVs, respectively. Based on comparison between the estimated abundance of EV proteins, we identified 368 SW480 EV-enriched and 359 SW620 EV-enriched proteins. SW480 EV-enriched proteins played a role in cell adhesion, but SW620 EV-enriched proteins were associated with cancer progression and functioned as diagnostic indicators of metastatic cancer; they were overexpressed in metastatic colorectal cancer and played roles in multidrug resistance. As the first proteomic analysis comparing primary and metastatic cancer-derived EVs, this study increases our understanding of the pathological function of EVs in the metastatic process and provides useful biomarkers for cancer metastasis.

  19. Collision tumor of the thyroid gland: primary squamous cell and papillary thyroid carcinoma.

    PubMed

    Warman, Meir; Lipschitz, Noga; Ikher, Sergey; Halperin, Doron

    2011-01-01

    Introduction. Collision tumor of the thyroid gland is defined when independent and histologically distinct tumors coexist within the gland. The presence of both papillary and squamous cell carcinoma in the thyroid gland is unusual. Suggested etiologies include embryonic remanents of squamous epithelium, chronic inflammation, or thyroid malignancies promoting squamous metaplasia. Case Presentation. An elderly patient presented with a rapid enlargement of a long-standing right thyroid nodule. The tumor was locally invasive and unresectable. Pathology revealed the diagnosis of papillary and squamous cell carcinoma of the thyroid gland. Possible primary sites for squamous cell carcinoma in upper aerodigestive tract were excluded. The patient outcome was fatal although palliative chemoradiotherapy. Discussion. Collision tumor of papillary and squamous cell carcinoma of the thyroid gland is a rare entity that may imply bad prognosis, as to the presence of the squamous portion. The best treatment includes resection of the tumor; unfortunately it is not possible in most cases.

  20. Modern management of primary T-cell immunodeficiencies.

    PubMed

    Pachlopnik Schmid, Jana; Güngör, Tayfun; Seger, Reinhard

    2014-06-01

    The study of human T-cell PIDs with Mendelian inheritance has enabled the molecular characterization of important key functions and pathways in T-cell biology. In most cases, T-cell PIDs become apparent as combined T- and B-cell deficiencies. Severe combined immunodeficiencies (SCIDs) are characterized by a complete lack of T-cell development and, in some cases, a developmental block in other lymphoid lineages and manifest within the first year of life. Combined immunodeficiency syndromes (CIDs) result from hypomorphic mutations in typical SCID associated genes or from partial defects of T-cell development and manifest later in childhood by increased susceptibility to infection often combined with disturbances in immune homeostasis, e.g., autoimmunity and increased incidence in lymphoproliferation. The discovery of mutations and characterization of the cellular changes that underlie lymphocyte defects and immune dysregulation have led to novel, specific, successful therapies for severe diseases which are often fatal if left untreated. Over the last few years, impressive progress has been made in understanding the disease mechanisms of T-cell immunodeficiencies and in improving the long-term outcomes of potentially curative treatments, including gene therapy.

  1. Differential Cytotoxic Activity of a Novel Palladium-Based Compound on Prostate Cell Lines, Primary Prostate Epithelial Cells and Prostate Stem Cells

    PubMed Central

    Cevatemre, Buse; Pellacani, Davide; Walker, Hannah; Mann, Vincent M.; Simms, Matthew S.; Stower, Michael J.; Yilmaz, Veysel T.; Maitland, Norman J.

    2013-01-01

    The outcome for patients with advanced metastatic and recurrent prostate cancer is still poor. Therefore, new chemotherapeutics are required, especially for killing cancer stem cells that are thought to be responsible for disease recurrence. In this study, we screened the effect of a novel palladium-based anticancer agent (Pd complex) against six different prostate cancer cell lines, and primary cultures from seven Gleason 6/7 prostate cancer, three Gleason 8/9 prostate cancer and four benign prostate hyperplasia patient samples, as well as cancer stem cells selected from primary cultures. MTT and ATP viability assays were used to assess cell growth and flow cytometry to assess cell cycle status. In addition, immunofluorescence was used to detect γH2AX nuclear foci, indicative of DNA damage, and Western blotting to assess the induction of apoptosis and autophagy. The Pd complex showed a powerful growth-inhibitory effect against both cell lines and primary cultures. More importantly, it successfully reduced the viability of cancer stem cells as first reported in this study. The Pd complex induced DNA damage and differentially induced evidence of cell death, as well as autophagy. In conclusion, this novel agent may be promising for use against the bulk of the tumour cell population as well as the prostate cancer stem cells, which are thought to be responsible for the resistance of metastatic prostate cancer to chemotherapy. This study also indicates that the combined use of the Pd complex with an autophagy modulator may be a more promising approach to treat prostate cancer. In addition, the differential effects observed between cell lines and primary cells emphasise the importance of the model used to test novel drugs including its genetic background, and indeed the necessity of using cells cultured from patient samples. PMID:23675532

  2. Differential cytotoxic activity of a novel palladium-based compound on prostate cell lines, primary prostate epithelial cells and prostate stem cells.

    PubMed

    Ulukaya, Engin; Frame, Fiona M; Cevatemre, Buse; Pellacani, Davide; Walker, Hannah; Mann, Vincent M; Simms, Matthew S; Stower, Michael J; Yilmaz, Veysel T; Maitland, Norman J

    2013-01-01

    The outcome for patients with advanced metastatic and recurrent prostate cancer is still poor. Therefore, new chemotherapeutics are required, especially for killing cancer stem cells that are thought to be responsible for disease recurrence. In this study, we screened the effect of a novel palladium-based anticancer agent (Pd complex) against six different prostate cancer cell lines, and primary cultures from seven Gleason 6/7 prostate cancer, three Gleason 8/9 prostate cancer and four benign prostate hyperplasia patient samples, as well as cancer stem cells selected from primary cultures. MTT and ATP viability assays were used to assess cell growth and flow cytometry to assess cell cycle status. In addition, immunofluorescence was used to detect γH2AX nuclear foci, indicative of DNA damage, and Western blotting to assess the induction of apoptosis and autophagy. The Pd complex showed a powerful growth-inhibitory effect against both cell lines and primary cultures. More importantly, it successfully reduced the viability of cancer stem cells as first reported in this study. The Pd complex induced DNA damage and differentially induced evidence of cell death, as well as autophagy. In conclusion, this novel agent may be promising for use against the bulk of the tumour cell population as well as the prostate cancer stem cells, which are thought to be responsible for the resistance of metastatic prostate cancer to chemotherapy. This study also indicates that the combined use of the Pd complex with an autophagy modulator may be a more promising approach to treat prostate cancer. In addition, the differential effects observed between cell lines and primary cells emphasise the importance of the model used to test novel drugs including its genetic background, and indeed the necessity of using cells cultured from patient samples.

  3. Primary cutaneous diffuse large B-cell lymphoma (PCDLBCL), leg-type and other: an update on morphology and treatment.

    PubMed

    Paulli, M; Lucioni, M; Maffi, A; Croci, G A; Nicola, M; Berti, E

    2012-12-01

    Primary cutaneous B-cell lymphoma (PCBCL) is an heterogeneous group of lymphoproliferative disorders, which account for 25-30% of all primary cutaneous lymphoma and include three main histotypes: 1) primary cutaneous marginal zone B-cell lymphoma (PCMZL); 2) primary cutaneous follicular center cell lymphoma (PCFCL); 3) primary cutaneous diffuse large B-cell lymphoma (DLBCL), leg type (PCDLBCL-LT). PCMZL and PCFCL are indolent lymphomas, with an excellent prognosis despite an high rate of cutaneous recurrences; in contrast, PCDLBCL-LT is clinically more aggressive and usually requires to be treated with multi-agent chemotherapy and anti-CD20 monoclonal antibodies. PCDLBCL-LT histologically consists of large round cells (centroblasts and immunoblasts), is characterized by strong bcl-2 expression, in the absence of t(14;18) translocation, and resembles the activated B-cell type of nodal DLBCL. Recently, the term primary cutaneous DLBCL-other (PCDLBCL-O) has been proposed to include diffuse lymphomas composed of large transformed B-cells that lack the typical features of PCDLBCL-LT and do not conform to the definition of PCFCL. Some clinical studies suggested that such cases have an indolent clinical course and may be treated in a conservative manner; however, data regarding the actual prognosis and clinical behaviour of these peculiar cases are still too limited. The spectrum of primary cutaneous DLBCL also encompasses some rare morphological variants, such as anaplastic or plasmablastic subtypes and T-cell rich B-cell lymphoma, and some recently described, exceedingly rare DLBCL subtypes, such as intravascular large B-cell lymphoma and EBV-associated large B-cell lymphoma of the elderly, which often present in the skin.

  4. Antioxidant enzymes in malignant prostate cell lines and in primary cultured prostatic cells.

    PubMed

    Jung, K; Seidel, B; Rudolph, B; Lein, M; Cronauer, M V; Henke, W; Hampel, G; Schnorr, D; Loening, S A

    1997-01-01

    The antioxidant enzymes catalase, glutathione reductase (GR), glutathione S-transferase (GST), glutathione peroxidase (GPx), and superoxide dismutase (SOD) were determined in the androgen-response LNCaP and androgen-nonresponsive PC-3 and DU 145 cells as well as in prostatic epithelial cell cultures of benign and malignant human prostatic tissue. There were no differences between the enzyme activities of the human primary cell cultures from cancerous tissue and their normal counterparts. The enzyme activities of the three permanent cell lines were either higher (SOD, catalase, GR) or lower (GST, GPx) than in the primary cell cultures. In LNCaP cells catalase and GR were significantly higher, GST, in contrast, was significantly lower than in PC-3 and DU 145 cells. GST in PC-3 and DU 145 cells, and SOD in all the three cell lines showed no significant differences. Catalase, GPx and GR values were significantly different in the three permanent cell lines. The different enzymatic equipment of the prostate cancer cell lines provides the basis for experimental testing of new concepts of cancer treatment with the help of systematic modulations of the antioxidant defence systems in prostate cancer.

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

    PubMed

    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. Sickle cell children traveling abroad: primary risk is infection.

    PubMed

    Runel-Belliard, Camille; Lesprit, Emmanuelle; Quinet, Béatrice; Grimprel, Emmanuel

    2009-01-01

    Pediatricians taking care of sickle cell children in France are concerned about giving travel advice. Very few articles are published and no study has been done about it. A lot of pediatricians are using their own experience to decide if sickle cell children can travel abroad. Studying the consequences of such travel for sickle cell children is important to discuss common recommendations. We conducted a prospective study from June 2006 to December 2007 on desires to travel expressed during our consultations with sickle cell children. We studied notable events that occurred during travel and at least 2 months after return. Of 52 desires to travel, 10 were cancelled. All of the 42 trips were to Africa. Median duration of travel was 1.29 months (0.5-3). Median age at travel was 7.6 years (0.2-17.7). Events during travel were two hospitalizations (4.8%), a transfusion (2.4%), and four paramedical or medical examinations (9.6%). After return, four events occurred: two SS children had Plasmodium falciparum malaria (4.8%) and two had digestive bacteremia (4.8%) in SC and Sbeta+ children. No event occurred during plane travel. None of our patients died. The primary risk for sickle cell children traveling to Africa is infection: malaria first and digestive septicemia second. These risks are increased by long travel and poor sanitary conditions. Each travel should be prepared a long time before departure, and each pediatrician should insist on malaria prophylaxis and sanitary conditions, especially for young children. Trips should be shorter than 1 month when possible. A longer prospective study will be done to confirm these results.

  7. Expression and functional activity of bitter taste receptors in primary renal tubular epithelial cells and M-1 cells.

    PubMed

    Liang, Jie; Chen, Fuxue; Gu, Fu; Liu, Xin; Li, Feng; Du, Dongshu

    2017-04-01

    The kidney is essential in the maintenance of in vivo homeostasis by body fluid and electrolyte conservation and metabolic waste removal. Previously, we reported the expression of a novel G protein family (Tas2rs), which includes bitter taste receptors, in the kidney tubule system, including the nephrons and the collecting duct system. Bitter taste receptors could affect kidney function via Ca(2+) intake. Alkaloids such as phenylthiocarbamide stimulate these receptors and cause an increase in Ca(2+) intake. In this study, we determined the expression of bitter taste receptors in the immature kidney and small intestine and in primary renal epithelial cells and M-1 (collecting tubule cell line) cells, by using QPCR and immunostaining. We found no expression of bitter taste receptors in the immature kidney and small intestine several days after birth; the relative abundance of Tas2rs transcripts varied depending on the developmental stage. Tas2rs were expressed in primary renal epithelial cells and M-1 cells. The traditional Chinese medicinal plant extracts phellodendrine and coptisine caused a rapid rise in intracellular Ca(2+) concentration, which was inhibited by the phospholipase C (PLC) inhibitor U-73122. Thus, phellodendrine and coptisine could change the physiological status of renal cells in vitro by mediation of bitter taste receptors in a PLC-dependent manner. Our results provide new insights on the expression and role of bitter taste receptors in renal development and function.

  8. Isolation and characterization of cancer stem cells from primary head and neck squamous cell carcinoma tumors

    PubMed Central

    Kim, Hong S.; Pearson, Alexander T.; Nör, Jacques E.

    2017-01-01

    Summary Drug resistance remains a significant problem in the treatment of patients with head and neck squamous cell carcinoma (HNSCC). Recent reports showed that a sub-population of highly tumorigenic cells (cancer stem cells) is uniquely resistant to chemotherapy, and suggesting that these cells play an important role in the relapse of HNSCC. The development of methods for the isolation and culture of cancer stem cells is a key step to enable studies exploring the mechanisms underlying the role of these cells in chemoresistance. Here, we describe a method to isolate cancer stem cells from primary head and neck tumors and for the generation of orospheres, i.e. the culture of these cells in suspension in ultra-low attachment plates. PMID:26910078

  9. Are primary renal cell carcinoma and metastases of renal cell carcinoma the same cancer?

    PubMed

    Semeniuk-Wojtaś, Aleksandra; Stec, Rafał; Szczylik, Cezary

    2016-05-01

    Metastasis is a process consisting of cells spreading from the primary site of the cancer to distant parts of the body. Our understanding of this spread is limited and molecular mechanisms causing particular characteristics of metastasis are still unknown. There is some evidence that primary renal cell carcinoma (RCC) and metastases of RCC exhibit molecular differences that may effect on the biological characteristics of the tumor. Some authors have detected differences in clear cell and nonclear cell component between these 2 groups of tumors. Investigators have also determined that primary RCC and metastases of RCC diverge in their range of renal-specific markers and other protein expression, gene expression pattern, and microRNA expression. There are also certain proteins that are variously expressed in primary RCCs and their metastases and have effect on clinical outcome, e.g., endothelin receptor type B, phos-S6, and CD44. However, further studies are needed on large cohorts of patients to identify differences representing promising targets for prognostic purposes predicting disease-free survival and the metastatic burden of a patient as well as their suitability as potential therapeutic targets. To sum up, in this review we have attempted to summarize studies connected with differences between primary RCC and its metastases and their influence on the biological characteristics of renal cancer.

  10. A mathematical model of a lithium/thionyl chloride primary cell

    NASA Astrophysics Data System (ADS)

    Evans, T. I.; Nguyen, T. V.; White, R. E.

    1987-08-01

    A 1-D mathematical model for the lithium/thionyl chloride primary cell was developed to investigate methods of improving its performance and safety. The model includes many of the components of a typical lithium/thionyl chloride cell such as the porous lithium chloride film which forms on the lithium anode surface. The governing equations are formulated from fundamental conservation laws using porous electrode theory and concentrated solution theory. The model is used to predict 1-D, time dependent profiles of concentration, porosity, current, and potential as well as cell temperature and voltage. When a certain discharge rate is required, the model can be used to determine the design criteria and operating variables which yield high cell capacities. Model predictions can be used to establish operational and design limits within which the thermal runaway problem, inherent in these cells, can be avoided.

  11. A mathematical model of a lithium/thionyl chloride primary cell

    NASA Technical Reports Server (NTRS)

    Evans, T. I.; Nguyen, T. V.; White, R. E.

    1987-01-01

    A 1-D mathematical model for the lithium/thionyl chloride primary cell was developed to investigate methods of improving its performance and safety. The model includes many of the components of a typical lithium/thionyl chloride cell such as the porous lithium chloride film which forms on the lithium anode surface. The governing equations are formulated from fundamental conservation laws using porous electrode theory and concentrated solution theory. The model is used to predict 1-D, time dependent profiles of concentration, porosity, current, and potential as well as cell temperature and voltage. When a certain discharge rate is required, the model can be used to determine the design criteria and operating variables which yield high cell capacities. Model predictions can be used to establish operational and design limits within which the thermal runaway problem, inherent in these cells, can be avoided.

  12. Evaluation of cytokine gene expression after avian influenza virus infection in avian cell lines and primary cell cultures

    USDA-ARS?s Scientific Manuscript database

    The innate immune responses elicited by avian influenza virus (AIV) infection has been studied by measuring cytokine gene expression by relative real time PCR (rRT-PCR) in vitro, using both cell lines and primary cell cultures. Continuous cell lines offer advantages over the use of primary cell cult...

  13. FAM65B controls the proliferation of transformed and primary T cells

    PubMed Central

    Froehlich, Jeanne; Versapuech, Margaux; Megrelis, Laura; Largeteau, Quitterie; Meunier, Sylvain; Tanchot, Corinne; Bismuth, Georges

    2016-01-01

    Cell quiescence is controlled by regulated genome-encoded programs that actively express genes which are often down-regulated or inactivated in transformed cells. Among them is FoxO1, a transcription factor that imposes quiescence in several cell types, including T lymphocytes. In these cells, the FAM65B encoding gene is a major target of FOXO1. Here, we show that forced expression of FAM65B in transformed cells blocks their mitosis because of a defect of the mitotic spindle, leading to G2 cell cycle arrest and apoptosis. Upon cell proliferation arrest, FAM65B is engaged in a complex containing two proteins well known to be involved in cell proliferation i.e. the HDAC6 deacetylase and the 14.3.3 scaffolding protein. In primary T cells, FAM65B is down-regulated upon T cell receptor engagement, and maintaining its expression blocks their proliferation, establishing that the decrease of FAM65B expression is required for proliferation. Conversely, inhibiting FAM65B expression in naive T lymphocytes decreases their activation threshold. These results identify FAM65B as a potential new target for controlling proliferation of both transformed and normal cells. PMID:27556504

  14. Multi-cellular 3D human primary liver cell culture elevates metabolic activity under fluidic flow.

    PubMed

    Esch, Mandy B; Prot, Jean-Matthieu; Wang, Ying I; Miller, Paula; Llamas-Vidales, Jose Ricardo; Naughton, Brian A; Applegate, Dawn R; Shuler, Michael L

    2015-05-21

    We have developed a low-cost liver cell culture device that creates fluidic flow over a 3D primary liver cell culture that consists of multiple liver cell types, including hepatocytes and non-parenchymal cells (fibroblasts, stellate cells, and Kupffer cells). We tested the performance of the cell culture under fluidic flow for 14 days, finding that hepatocytes produced albumin and urea at elevated levels compared to static cultures. Hepatocytes also responded with induction of P450 (CYP1A1 and CYP3A4) enzyme activity when challenged with P450 inducers, although we did not find significant differences between static and fluidic cultures. Non-parenchymal cells were similarly responsive, producing interleukin 8 (IL-8) when challenged with 10 μM bacterial lipoprotein (LPS). To create the fluidic flow in an inexpensive manner, we used a rocking platform that tilts the cell culture devices at angles between ±12°, resulting in a periodically changing hydrostatic pressure drop between reservoirs and the accompanying periodically changing fluidic flow (average flow rate of 650 μL min(-1), and a maximum shear stress of 0.64 dyne cm(-2)). The increase in metabolic activity is consistent with the hypothesis that, similar to unidirectional fluidic flow, primary liver cell cultures increase their metabolic activity in response to fluidic flow periodically changes direction. Since fluidic flow that changes direction periodically drastically changes the behavior of other cells types that are shear sensitive, our findings support the theory that the increase in hepatic metabolic activity associated with fluidic flow is either activated by mechanisms other than shear sensing (for example increased opportunities for gas and metabolite exchange), or that it follows a shear sensing mechanism that does not depend on the direction of shear. Our mode of device operation allows us to evaluate drugs under fluidic cell culture conditions and at low device manufacturing and operation

  15. Management of primary germ cell tumors of the mediastinum.

    PubMed

    Economou, J S; Trump, D L; Holmes, E C; Eggleston, J E

    1982-05-01

    Twenty-eight patients with primary malignant germ cell tumors (GCT) of the mediastinum were treated at the University of California at Los Angeles and The Johns Hopkins Hospital in the past 30 years. Of 11 patients with pure seminomas, nine (82%) are free of disease from 6 months to 15 years following therapy. The primary treatment modality in these patients was mediastinal radiation; one patient with metastatic disease had a complete remission and prolonged survival following combination chemotherapy. Seventeen patients had GCT with nonseminomatous elements. Only three (18%) are alive and free of disease. One patient treated only surgically is alive at 15 years and two patients treated with combination chemotherapy and operation are alive and free of disease at 6 months and 3 years. When analyzed by a Kaplan-Meier actuarial survival estimate, patients with nonseminomatous GCT who were treated with cisplatin-bleomycin-based chemotherapy had a median survival of 14.0 months whereas those treated with chemotherapy regimens not employing these agents had a median survival of 4.0 months (generalized Wilcoxon test, p = 0.0495). Patients with pure seminomas are effectively treated with radiation therapy. Patients with nonseminomatous tumors have a much poorer prognosis and deserve aggressive multimodality therapy with cisplatin-bleomycin-based chemotherapy.

  16. Primary large-cell lymphoma of the central nervous system

    SciTech Connect

    Amendola, B.E.; McClatchey, K.D.; Amendola, M.A.; Gebarski, S.S.

    1986-06-01

    Primary non-Hodgkin's lymphoma of the central nervous system (CNS) is a rare disease. Seven patients were seen and treated at the University of Michigan Medical Center between January 1969 and December 1983. All patients had histologically proven diagnoses of large cell lymphoma with clinical and radiologic evidence of involvement limited to the CNS. Five of seven patients received postoperative radiation therapy, two of whom have had apparent local control at 1- and 2-year follow-up. The two patients without postoperative radiation died of local recurrence 2 and 3 months following subtotal resection. These poor results suggest that adjuvant therapy may be required for improved control of this type of extranodal lymphoma.

  17. Hematopoietic stem cell transplantation for primary immunodeficiency diseases.

    PubMed

    Slatter, Mary A; Cant, Andrew J

    2011-11-01

    Hematopoietic stem cell transplantation (HSCT) is now highly successfully curing a widening range of primary immunodeficiencies (PIDs). Better tissue typing, matching of donors, less toxic chemotherapy, better virus detection and treatment, improved supportive care, and graft-versus-host disease prophylaxis mean up to a 90% cure for severe combined immunodeficiency patients and a 70-80% cure for other PIDs given a matched unrelated donor, and rising to 95% for young patients with specific PIDs, such as Wiskott-Aldrich syndrome. Precise molecular diagnosis, detailed data on prognosis, and careful pre-HSCT assessment of infective lung and liver damage will ensure an informed benefit analysis of HSCT and the best outcome. It is now recognized that the best treatment option for chronic granulomatous disease is HSCT, which can also be curative for CD40 ligand deficiency and complex immune dysregulation disorders.

  18. Reactivity of alveolar epithelial cells in primary culture with type I cell monoclonal antibodies.

    PubMed

    Danto, S I; Zabski, S M; Crandall, E D

    1992-03-01

    An understanding of the process of alveolar epithelial cell growth and differentiation requires the ability to trace and analyze the phenotypic transitions that the cells undergo. This analysis demands specific phenotypic probes to type II and, especially, type I pneumocytes. To this end, monoclonal antibodies have been generated to type I alveolar epithelial cells using an approach designed to enhance production of lung-specific clones from a crude lung membrane preparation. The monoclonal antibodies were screened by a combination of enzyme-linked immunosorbent assay and immunohistochemical techniques, with the determination of type I cell specificity resting primarily on immunoelectron microscopic localization. Two of these new markers of the type I pneumocyte phenotype (II F1 and VIII B2) were used to analyze primary cultures of type II cells growing on standard tissue culture plastic and on a variety of substrata reported to affect the morphology of these cells in culture. On tissue culture plastic, the antibodies fail to react with early (days 1 to 3) type II cell cultures. The cells become progressively more reactive with time in culture to a plateau of approximately 6 times background by day 8, with a maximum rate of increase between days 3 and 5. This finding is consistent with the hypothesis that type II cells in primary culture undergo at least partial differentiation into type I cells. Type II cells grown on laminin, which reportedly delays the loss of type II cell appearance, and on fibronectin, which has been reported to facilitate cell spreading and loss of type II cell features, develop the type I cell markers during cultivation in vitro with kinetics similar to those on uncoated tissue culture plastic. Cells on type I collagen and on tissue culture-treated Nuclepore filters, which have been reported to support monolayers with type I cell-like morphology, also increase their expression of the II F1 and VIII B2 epitopes around days 3 to 5. Taken

  19. Cytotoxicity testing of silver-containing burn treatments using primary and immortal skin cells.

    PubMed

    Boonkaew, Benjawan; Kempf, Margit; Kimble, Roy; Cuttle, Leila

    2014-12-01

    A novel burn wound hydrogel dressing has been previously developed which is composed of 2-acrylamido-2-methylpropane sulfonic acid sodium salt with silver nanoparticles (silver AMPS). This study compared the cytotoxicity of this dressing to the commercially available silver products; Acticoat™, PolyMem Silver(®) and Flamazine™ cream. Human keratinocytes (HaCaT and primary HEK) and normal human fibroblasts (NHF) were exposed to dressings incubated on Nunc™ polycarbonate inserts for 24, 48 and 72h. Four different cytotoxicity assays were performed including; Trypan Blue cell count, MTT, Celltiter-Blue™ and Toluidine Blue surface area assays. The results were expressed as relative cell viability compared to an untreated control. The cytotoxic effects of Acticoat™ and Flamazine™ cream were dependent on exposure time and cell type. After 24h exposure, Acticoat™ and Flamazine™ cream were toxic to all tested cell lines. Surprisingly, HaCaTs treated with Acticoat™ and Flamazine™ had an improved ability to survive at 48 and 72h while HEKs and NHFs had no improvement in survival with any treatment. The novel silver hydrogel and PolyMem Silver(®) showed low cytotoxicity to all tested cell lines at every time interval and these results support the possibility of using the novel silver hydrogel as a burn wound dressing. Researchers who rely on HaCaT cells as an accurate keratinocyte model should be aware that they can respond differently to primary skin cells.

  20. Isolation of Highly Pure Primary Mouse Alveolar Epithelial Type II Cells by Flow Cytometric Cell Sorting

    PubMed Central

    Lowell, Clifford A.

    2017-01-01

    In this protocol, we describe the method for isolating highly pure primary alveolar epithelial type II (ATII) cells from lungs of naïve mice. The method combines negative selection for a variety of lineage markers along with positive selection for EpCAM, a pan-epithelial cell marker. This method yields 2-3 × 106 ATII cells per mouse lung. The cell preps are highly pure and viable and can be used for genomic or proteomic analyses or cultured ex vivo to understand their roles in various biological processes. PMID:28180137

  1. Isolation of primary murine brain microvascular endothelial cells.

    PubMed

    Ruck, Tobias; Bittner, Stefan; Epping, Lisa; Herrmann, Alexander M; Meuth, Sven G

    2014-11-14

    The blood-brain-barrier is ultrastructurally assembled by a monolayer of brain microvascular endothelial cells (BMEC) interconnected by a junctional complex of tight and adherens junctions. Together with other cell-types such as astrocytes or pericytes, they form the neurovascular unit (NVU), which specifically regulates the interchange of fluids, molecules and cells between the peripheral blood and the CNS. Through this complex and dynamic system BMECs are involved in various processes maintaining the homeostasis of the CNS. A dysfunction of the BBB is observed as an essential step in the pathogenesis of many severe CNS diseases. However, specific and targeted therapies are very limited, as the underlying mechanisms are still far from being understood. Animal and in vitro models have been extensively used to gain in-depth understanding of complex physiological and pathophysiological processes. By reduction and simplification it is possible to focus the investigation on the subject of interest and to exclude a variety of confounding factors. However, comparability and transferability are also reduced in model systems, which have to be taken into account for evaluation. The most common animal models are based on mice, among other reasons, mainly due to the constantly increasing possibilities of methodology. In vitro studies of isolated murine BMECs might enable an in-depth analysis of their properties and of the blood-brain-barrier under physiological and pathophysiological conditions. Further insights into the complex mechanisms at the BBB potentially provide the basis for new therapeutic strategies. This protocol describes a method to isolate primary murine microvascular endothelial cells by a sequence of physical and chemical purification steps. Special considerations for purity and cultivation of MBMECs as well as quality control, potential applications and limitations are discussed.

  2. Transforming primary healthcare by including the stakeholders involved in delivering care to people living in poverty: EQUIhealThY study protocol.

    PubMed

    Loignon, Christine; Hudon, Catherine; Boudreault-Fournier, Alexandrine; Dupéré, Sophie; Macaulay, Ann C; Pluye, Pierre; Gaboury, Isabelle; Haggerty, Jeannie L; Fortin, Martin; Goulet, Émilie; Lambert, Mireille; Pelissier-Simard, Luce; Boyer, Sophie; de Laat, Marianne; Lemire, Francine; Champagne, Louise; Lemieux, Martin

    2013-03-11

    Ensuring access to timely and appropriate primary healthcare for people living in poverty is an issue facing all countries, even those with universal healthcare systems. The transformation of healthcare practices and organization could be improved by involving key stakeholders from the community and the healthcare system in the development of research interventions. The aim of this project is to stimulate changes in healthcare organizations and practices by encouraging collaboration between care teams and people living in poverty. Our objectives are twofold: 1) to identify actions required to promote the adoption of professional practices oriented toward social competence in primary care teams; and 2) to examine factors that would encourage the inclusion of people living in poverty in the process of developing social competence in healthcare organizations. This study will use a participatory action research design applied in healthcare organizations. Participatory research is an increasingly recognized approach that is helpful for involving the people for whom the research results are intended. Our research team consists of 19 non-academic researchers, 11 academic researchers and six partners. A steering committee composed of academic researchers and stakeholders will have a decision-making role at each step, including knowledge dissemination and recommendations for new interventions. In this project we will adopt a multiphase approach and will use a variety of methods, including photovoice, group discussions and interviews. The proposed study will be one of only a few using participatory research in primary care to foster changes aimed at enhancing quality and access to care for people living in poverty. To our knowledge this will be the first study to use photovoice in healthcare organizations to promote new interventions. Our project includes partners who are targeted for practice changes and improvements in delivering primary care to persons living in poverty

  3. Lovastatin Induces Multiple Stress Pathways Including LKB1/AMPK Activation That Regulate Its Cytotoxic Effects in Squamous Cell Carcinoma Cells

    PubMed Central

    Ma, Laurie; Niknejad, Nima; Gorn-Hondermann, Ivan; Dayekh, Khalil; Dimitroulakos, Jim

    2012-01-01

    Background Cellular stress responses trigger signaling cascades that inhibit proliferation and protein translation to help alleviate the stress or if the stress cannot be overcome induce apoptosis. In recent studies, we demonstrated the ability of lovastatin, an inhibitor of mevalonate synthesis, to induce the Integrated Stress Response as well as inhibiting epidermal growth factor receptor (EGFR) activation. Methodology/Principal Findings In this study, we evaluated the effects of lovastatin on the activity of the LKB1/AMPK pathway that is activated upon cellular energy shortage and can interact with the above pathways. In the squamous cell carcinoma (SCC) cell lines SCC9 and SCC25, lovastatin treatment (1–25 µM, 24 hrs) induced LKB1 and AMPK activation similar to metformin (1–10 mM, 24 hrs), a known inducer of this pathway. Lovastatin treatment impaired mitochondrial function and also decreased cellular ADP/ATP ratios, common triggers of LKB1/AMPK activation. The cytotoxic effects of lovastatin were attenuated in LKB1 null MEFs indicating a role for this pathway in regulating lovastatin-induced cytotoxicity. Of clinical relevance, lovastatin induces synergistic cytotoxicity in combination with the EGFR inhibitor gefitinib. In LKB1 deficient (A549, HeLa) and expressing (SCC9, SCC25) cell lines, metformin enhanced gefitinib cytotoxicity only in LKB1 expressing cell lines while both groups showed synergistic cytotoxic effects with lovastatin treatments. Furthermore, the combination of lovastatin with gefitinib induced a potent apoptotic response without significant induction of autophagy that is often induced during metabolic stress inhibiting cell death. Conclusion/Significance Thus, targeting multiple metabolic stress pathways including the LKB1/AMPK pathway enhances lovastatin’s ability to synergize with gefitinib in SCC cells. PMID:23029387

  4. Differentiation "in vitro" of primary and immortalized porcine mesenchymal stem cells into cardiomyocytes for cell transplantation.

    PubMed

    Moscoso, I; Centeno, A; López, E; Rodriguez-Barbosa, J I; Santamarina, I; Filgueira, P; Sánchez, M J; Domínguez-Perles, R; Peñuelas-Rivas, G; Domenech, N

    2005-01-01

    Cell transplantation to regenerate injured tissues is a promising new treatment for patients suffering several diseases. Bone marrow contains a population of progenitor cells known as mesenchymal stem cells (MSCs), which have the capability to colonize different tissues, replicate, and differentiate into multilineage cells. Our goal was the isolation, characterization, and immortalization of porcine MSCs (pMSCs) to study their potential differentiation "in vitro" into cardiomyocytes. pMSCs were obtained from the aspirated bone marrow of Large-White pigs. After 4 weeks in culture, adherent cells were phenotypically characterized by flow cytometry and immunochemistry by using monoclonal antibodies. Primary pMSCs were transfected with the plasmid pRNS-1 to obtain continuous growing cloned cell lines. Fresh pMSCs and immortalized cells were treated with 5-azacytidine to differentiate them into cardiomyocytes. Flow cytometry analysis of isolated pMSCs demonstrated the following phenotype, CD90(pos), CD29(pos), CD44(pos), SLA-I(pos), CD106(pos), CD46(pos) and CD45(neg), CD14(neg), CD31(neg), and CD11b(neg), similar to that described for human MSC. We derived several stable immortalized MSC cell lines. One of these, called pBMC-2, was chosen for further characterization. After "in vitro" stimulation of both primary or immortalized cells with 5-azacytidine, we obtained different percentages (30%-50%) of cells with cardiomyocyte characteristics, namely, positive for alpha-Actin and T-Troponin. Thus, primary or immortalized pMSCs derived from bone marrow and cultured were able to differentiate "ex vivo" into cardiac-like muscle cells. These elements may be potentials tools to improve cardiac function in a swine myocardial infarct model.

  5. Robotic surgery for primary head and neck squamous cell carcinoma of unknown site.

    PubMed

    Patel, Sapna A; Magnuson, J Scott; Holsinger, F Christopher; Karni, Ron J; Richmon, Jeremy D; Gross, Neil D; Bhrany, Amit D; Ferrell, Jay K; Ford, Samuel E; Kennedy, Aimee A; Méndez, Eduardo

    2013-11-01

    Identification of the primary site in head and neck squamous cell carcinoma (HNSCC) is crucial because it improves the patient's prognosis and minimizes morbidity from treatment. To determine the efficacy of transoral robotic surgery (TORS) in identifying unknown primary sites of head and neck squamous cell carcinoma. Retrospective, multi-institutional case series from January 1, 2010, to February 28, 2013, in which data were pooled from the following 6 institutions: University of Washington Medical Center, The University of Texas MD Anderson Cancer Center, University of Alabama-Birmingham Hospital, The University of Texas Medical School at Houston, Johns Hopkins Hospital, and Oregon Health Sciences University. All patients diagnosed as having HNSCC of an unknown primary site who underwent TORS to identify the primary site were included in the study. We excluded those with recurrent disease, a history of radiation therapy to the head and neck, or evidence of a primary tumor site based on previous biopsy results. Identification of the primary tumor site. Forty-seven patients were eligible for the study. The tumor site was identified by TORS in 34 of 47 patients (72.3%). The primary site was located in the base of tongue for 20 patients (58.8%) and the palatine tonsil for 13 patients (38.2%), with 1 patient having a primary site in both the base of tongue and the palatine tonsil. Suspicious physical examination findings were present in 23 of 47 patients (48.9%), with positive and negative predictive values of 56.5% and 25.0%, respectively. Of those who underwent any imaging, 16 patients had suspicious findings, with positive and negative predictive values of 50.0% and 16.7%, respectively. In 18 of 47 patients (38.3%), both preoperative radiographic and physical examination failed to suggest a primary site. Of these 18 patients, 13 (72.2%) were identified after undergoing TORS. We demonstrate that TORS is a useful approach to identify and treat the primary site in

  6. Electrolytic/fuel cell bundles and systems including a current collector in communication with an electrode thereof

    DOEpatents

    Hawkes, Grant L.; Herring, James S.; Stoots, Carl M.; O& #x27; Brien, James E.

    2013-03-05

    Electrolytic/fuel cell bundles and systems including such bundles include an electrically conductive current collector in communication with an anode or a cathode of each of a plurality of cells. A cross-sectional area of the current collector may vary in a direction generally parallel to a general direction of current flow through the current collector. The current collector may include a porous monolithic structure. At least one cell of the plurality of cells may include a current collector that surrounds an outer electrode of the cell and has at least six substantially planar exterior surfaces. The planar surfaces may extend along a length of the cell, and may abut against a substantially planar surface of a current collector of an adjacent cell. Methods for generating electricity and for performing electrolysis include flowing current through a conductive current collector having a varying cross-sectional area.

  7. Expression and regulation of Schlafen (SLFN) family members in primary human monocytes, monocyte-derived dendritic cells and T cells

    PubMed Central

    Puck, Alexander; Aigner, Regina; Modak, Madhura; Cejka, Petra; Blaas, Dieter; Stöckl, Johannes

    2015-01-01

    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. PMID:26623250

  8. Expression and regulation of Schlafen (SLFN) family members in primary human monocytes, monocyte-derived dendritic cells and T cells.

    PubMed

    Puck, Alexander; Aigner, Regina; Modak, Madhura; Cejka, Petra; Blaas, Dieter; Stöckl, Johannes

    2015-01-01

    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.

  9. Murine Stem Cell-Based Retrovirus Production for Marking Primary Mouse Mammary Cells for Metastasis Studies.

    PubMed

    Beverly, Levi J; Podsypanina, Katrina

    2016-02-01

    Since the introduction of retroviral vector technology, permanent genetic marking of cells has considerably contributed to the understanding of different physiological and disease processes in vivo. Recent marking strategies aim to elucidate the contribution of cells on the clonal level, and the advent of fluorescent proteins has opened new avenues for the in vivo analysis of gene-marked cells. Gene-modified cells are easily identifiable (e.g., via the introduced fluorescent protein) within whole organ structures, allowing one to measure the contribution of transduced cells to malignant outgrowth. In our laboratory, we use the tetracycline-inducible system to study oncogene cooperation in metastatic progression. We use bicistronic retroviruses expressing the tetracycline transactivator (tTA) and the candidate gene (MIT-gene) or the tTA alone (MIT-Rx) to infect primary mammary cells from mice harboring tetracycline-inducible transgenes. This allows for constitutive expression of the candidate gene and tTA-dependent expression of the inducible oncogene. We also use MIG-based vectors, which allow for constitutive expression of the candidate gene and a green fluorescent protein. Here we describe how to produce retroviral particles carrying both MIT- and MIG-based vectors. Because of the fragility of the retroviral envelope, we do not attempt to concentrate the virus, and we directly use packaging cell media to infect primary epithelial cells (either normal or tumor). Infected cells can be transplanted into recipient mice to investigate metastatic colonization. © 2016 Cold Spring Harbor Laboratory Press.

  10. Decellularized extracellular matrices produced from immortal cell lines derived from different parts of the placenta support primary mesenchymal stem cell expansion

    PubMed Central

    Kusuma, Gina D.; Brennecke, Shaun P.; O’Connor, Andrea J.; Kalionis, Bill

    2017-01-01

    Mesenchymal stem/stromal cells (MSCs) exhibit undesired phenotypic changes during ex vivo expansion, limiting production of the large quantities of high quality primary MSCs needed for both basic research and cell therapies. Primary MSCs retain many desired MSC properties including proliferative capacity and differentiation potential when expanded on decellularized extracellular matrix (dECM) prepared from primary MSCs. However, the need to use low passage number primary MSCs (passage 3 or lower) to produce the dECM drastically limits the utility and impact of this technology. Here, we report that primary MSCs expanded on dECM prepared from high passage number (passage 25) human telomerase reverse transcriptase (hTERT) transduced immortal MSC cell lines also exhibit increased proliferation and osteogenic differentiation. Two hTERT-transduced placenta-derived MSC cell lines, CMSC29 and DMSC23 [derived from placental chorionic villi (CMSCs) and decidua basalis (DMSCs), respectively], were used to prepare dECM-coated substrates. These dECM substrates showed structural and biochemical differences. Primary DMSCs cultured on dECM-DMSC23 showed a three-fold increase in cell number after 14 days expansion in culture and increased osteogenic differentiation compared with controls. Primary CMSCs cultured on the dECM-DMSC23 exhibited a two-fold increase in cell number and increased osteogenic differentiation. We conclude that immortal MSC cell lines derived from different parts of the placenta produce dECM with varying abilities for supporting increased primary MSC expansion while maintaining important primary MSC properties. Additionally, this is the first demonstration of using high passage number cells to produce dECM that can promote primary MSC expansion, and this advancement greatly increases the feasibility and applicability of dECM-based technologies. PMID:28152107

  11. Loss of Proliferation and Antigen Presentation Activity following Internalization of Polydispersed Carbon Nanotubes by Primary Lung Epithelial Cells

    PubMed Central

    Kumari, Mandavi; Sachar, Sumedha; Saxena, Rajiv K.

    2012-01-01

    Interactions between poly-dispersed acid functionalized single walled carbon nanotubes (AF-SWCNTs) and primary lung epithelial (PLE) cells were studied. Peritoneal macrophages (PMs, known phagocytic cells) were used as positive controls in this study. Recovery of live cells from cultures of PLE cells and PMs was significantly reduced in the presence of AF-SWCNTs, in a time and dose dependent manner. Both PLE cells as well as PMs could take up fluorescence tagged AF-SWCNTs in a time dependent manner and this uptake was significantly blocked by cytochalasin D, an agent that blocks the activity of acto-myosin fibers and therefore the phagocytic activity of cells. Confocal microscopic studies confirmed that AF-SWCNTs were internalized by both PLE cells and PMs. Intra-trachially instilled AF-SWCNTs could also be taken up by lung epithelial cells as well as alveolar macrophages. Freshly isolated PLE cells had significant cell division activity and cell cycling studies indicated that treatment with AF-SWCNTs resulted in a marked reduction in S-phase of the cell cycle. In a previously standardized system to study BCG antigen presentation by PLE cells and PMs to sensitized T helper cells, AF-SWCNTs could significantly lower the antigen presentation ability of both cell types. These results show that mouse primary lung epithelial cells can efficiently internalize AF-SWCNTs and the uptake of nanotubes interfered with biological functions of PLE cells including their ability to present BCG antigens to sensitized T helper cells. PMID:22384094

  12. Loss of proliferation and antigen presentation activity following internalization of polydispersed carbon nanotubes by primary lung epithelial cells.

    PubMed

    Kumari, Mandavi; Sachar, Sumedha; Saxena, Rajiv K

    2012-01-01

    Interactions between poly-dispersed acid functionalized single walled carbon nanotubes (AF-SWCNTs) and primary lung epithelial (PLE) cells were studied. Peritoneal macrophages (PMs, known phagocytic cells) were used as positive controls in this study. Recovery of live cells from cultures of PLE cells and PMs was significantly reduced in the presence of AF-SWCNTs, in a time and dose dependent manner. Both PLE cells as well as PMs could take up fluorescence tagged AF-SWCNTs in a time dependent manner and this uptake was significantly blocked by cytochalasin D, an agent that blocks the activity of acto-myosin fibers and therefore the phagocytic activity of cells. Confocal microscopic studies confirmed that AF-SWCNTs were internalized by both PLE cells and PMs. Intra-trachially instilled AF-SWCNTs could also be taken up by lung epithelial cells as well as alveolar macrophages. Freshly isolated PLE cells had significant cell division activity and cell cycling studies indicated that treatment with AF-SWCNTs resulted in a marked reduction in S-phase of the cell cycle. In a previously standardized system to study BCG antigen presentation by PLE cells and PMs to sensitized T helper cells, AF-SWCNTs could significantly lower the antigen presentation ability of both cell types. These results show that mouse primary lung epithelial cells can efficiently internalize AF-SWCNTs and the uptake of nanotubes interfered with biological functions of PLE cells including their ability to present BCG antigens to sensitized T helper cells.

  13. Plasma cells and the chronic nonsuppurative destructive cholangitis of primary biliary cirrhosis.

    PubMed

    Takahashi, Toru; Miura, Tomofumi; Nakamura, Junichiro; Yamada, Satoshi; Miura, Tsutomu; Yanagi, Masahiko; Matsuda, Yasunobu; Usuda, Hiroyuki; Emura, Iwao; Tsuneyama, Koichi; He, Xiao-Song; Gershwin, M Eric

    2012-03-01

    There has been increased interest in the role of B cells in the pathogenesis of primary biliary cirrhosis (PBC). Although the vast majority of patients with this disease have anti-mitochondrial antibodies, there is no correlation of anti-mitochondrial antibody titer and/or presence with disease severity. Furthermore, in murine models of PBC, it has been suggested that depletion of B cells may exacerbate biliary pathology. To address this issue, we focused on a detailed phenotypic characterization of mononuclear cell infiltrates surrounding the intrahepatic bile ducts of patients with PBC, primary sclerosing cholangitis, autoimmune hepatitis, chronic hepatitis C, and graft-versus-host disease, including CD3, CD4, CD8, CD20, CD38, and immunoglobulin classes, as well as double immunohistochemical staining for CD38 and IgM. Interestingly, CD20 B lymphocytes, which are a precursor of plasma cells, were found in scattered locations or occasionally forming follicle-like aggregations but were not noted at the proximal location of chronic nonsuppurative destructive cholangitis. In contrast, there was a unique and distinct coronal arrangement of CD38 cells around the intrahepatic ducts in PBC but not controls; the majority of such cells were considered plasma cells based on their expression of intracellular immunoglobulins, including IgM and IgG, but not IgA. Patients with PBC who manifest this unique coronal arrangement were those with significantly higher titers of anti-mitochondrial antibodies. These data collectively suggest a role for plasma cells in the specific destruction of intrahepatic bile ducts in PBC and confirm the increasing interest in plasma cells and autoimmunity. Copyright © 2011 American Association for the Study of Liver Diseases.

  14. MYCN induces neuroblastoma in primary neural crest cells.

    PubMed

    Olsen, R R; Otero, J H; García-López, J; Wallace, K; Finkelstein, D; Rehg, J E; Yin, Z; Wang, Y-D; Freeman, K W

    2017-08-31

    Neuroblastoma (NBL) is an embryonal cancer of the sympathetic nervous system (SNS), which causes 15% of pediatric cancer deaths. High-risk NBL is characterized by N-Myc amplification and segmental chromosomal gains and losses. Owing to limited disease models, the etiology of NBL is largely unknown, including both the cell of origin and the majority of oncogenic drivers. We have established a novel system for studying NBL based on the transformation of neural crest cells (NCCs), the progenitor cells of the SNS, isolated from mouse embryonic day 9.5 trunk neural tube explants. Based on pathology and gene expression analysis, we report the first successful transformation of wild-type NCCs into NBL by enforced expression of N-Myc, to generate phenotypically and molecularly accurate tumors that closely model human MYCN-amplified NBL. Using comparative genomic hybridization, we found that NCC-derived NBL tumors acquired copy number gains and losses that are syntenic to those observed in human MYCN-amplified NBL including 17q gain, 2p gain and loss of 1p36. When p53-compromised NCCs were transformed with N-Myc, we generated primitive neuroectodermal tumors with divergent differentiation including osteosarcoma. These subcutaneous tumors were metastatic to regional lymph nodes, liver and lung. Our novel experimental approach accurately models human NBL and establishes a new system with potential to study early stages of NBL oncogenesis, to functionally assess NBL oncogenic drivers and to characterize NBL metastasis.

  15. Primary outgrowth cultures are a reliable source of human pancreatic stellate cells.

    PubMed

    Han, Song; Delitto, Daniel; Zhang, Dongyu; Sorenson, Heather L; Sarosi, George A; Thomas, Ryan M; Behrns, Kevin E; Wallet, Shannon M; Trevino, Jose G; Hughes, Steven J

    2015-11-01

    Recent advances demonstrate a critical yet poorly understood role for the pancreatic stellate cell (PSC) in the pathogenesis of chronic pancreatitis (CP) and pancreatic cancer (PC). Progress in this area has been hampered by the availability, fidelity, and/or reliability of in vitro models of PSCs. We examined whether outgrowth cultures from human surgical specimens exhibited reproducible phenotypic and functional characteristics of PSCs. PSCs were cultured from surgical specimens of healthy pancreas, CP and PC. Growth dynamics, phenotypic characteristics, soluble mediator secretion profiles and co-culture with PC cells both in vitro and in vivo were assessed. Forty-seven primary cultures were established from 52 attempts, demonstrating universal α-smooth muscle actin and glial fibrillary acidic protein but negligible epithelial surface antigen expression. Modification of culture conditions consistently led to cytoplasmic lipid accumulation, suggesting induction of a quiescent phenotype. Secretion of growth factors, chemokines and cytokines did not significantly differ between donor pathologies, but did evolve over time in culture. Co-culture of PSCs with established PC cell lines resulted in significant changes in levels of multiple secreted mediators. Primary PSCs co-inoculated with PC cells in a xenograft model led to augmented tumor growth and metastasis. Therefore, regardless of donor pathology, outgrowth cultures produce PSCs that demonstrate consistent growth and protein secretion properties. Primary cultures from pancreatic surgical specimens, including malignancies, may represent a reliable source of human PSCs.

  16. Investigation of primary Li-Si/FeS2 cells

    NASA Astrophysics Data System (ADS)

    Redey, L.; Smaga, J. A.; Battles, J. E.; Guidotti, R.

    1987-04-01

    The factors that limit the performance of thermally activated Li-Si/FeS2 batteries were defined through the use of electrochemical characterization tests and post-test examinations. For the characterization tests, 82 individual cells were instrumented with multiple voltage sensors and discharged under isothermal and isobaric conditions. The voltage data for the sensors were recorded to determine the ohmic and electrochemical impedances of each cell component at different levels of discharge. The data analysis completed to date has demonstrated that this approach can successfully differentiate the influence of various operating parameters (e.g., temperature, current density), electrode structures (e.g., FeS2 particle size), and additives on cell capacity, specific energy, and power capability. Thirty cells selected from these tests and additional tests at SNL were examined using optical and scanning electron microscopy, energy dispersive spectroscopy, and X-ray diffraction. These analyses documented microstructural and compositional changes in the active materials and electrolyte. In general, the electrochemical impedance of the FeS electrode limited cell performance. Several methods (including use of fine FeS2 particle size, graphite additions, and higher operating temperatures) produced measurable reductions in this impedance and yielded significant improvements in specific energy and power.

  17. Prospects for primary stroke prevention in children with sickle cell anaemia.

    PubMed

    Jordan, Lori C; Casella, James F; DeBaun, Michael R

    2012-04-01

    This review will focus on the strengths and limitations associated with the current standard of care for primary prevention of ischaemic strokes in children with sickle cell anaemia (SCA) - transcranial Doppler ultrasound (TCD) screening followed by regular blood transfusion therapy when TCD measurement is above a threshold defined by a randomized clinical trial (RCT). The theoretical basis for potential alternative strategies for primary prevention of neurological injury in SCA is also discussed. These strategies will include, but will not be limited to: immunizations to prevent bacterial infections, particularly in low income countries; management of elevated blood pressure; and targeted strategies to increase baseline haemoglobin levels with therapies such as hyroxycarbamide or potentially definitive haematopoietic stem cell transplant.

  18. Mechanisms of Ionizing Radiation-Induced Cell Death in Primary Lung Cells

    DTIC Science & Technology

    2013-03-05

    Zhai M, et al. 2007. Protein oxidation implicated as the primary determinant of bacterial radioresistance. PLoS Biol 5:e92 43. Demidenko ZN...oxidative stress induces senescence in retinal pigment epithelial cells via TGF-beta release. Investigative ophthalmology & visual science 50:926- 35

  19. Primary metastatic Ewing's family tumors: results of the Italian Sarcoma Group and Scandinavian Sarcoma Group ISG/SSG IV Study including myeloablative chemotherapy and total-lung irradiation.

    PubMed

    Luksch, R; Tienghi, A; Hall, K Sundby; Fagioli, F; Picci, P; Barbieri, E; Gandola, L; Eriksson, M; Ruggieri, P; Daolio, P; Lindholm, P; Prete, A; Bisogno, G; Tamburini, A; Grignani, G; Abate, M E; Podda, M; Smeland, S; Ferrari, S

    2012-11-01

    The Italian Sarcoma Group and the Scandinavian Sarcoma Group designed a joint study to improve the prognosis for patients with Ewing's family tumors and synchronous metastatic disease limited to the lungs, or the pleura, or a single bone. The study was opened in 1999 and closed to the enrollment in 2008. The program consisted of intensive five-drug combination chemotherapy, surgery and/or radiotherapy as local treatment, and consolidation treatment with high-dose busulfan/melphalan plus autologous stem cell rescue and total-lung irradiation. During the study period, 102 consecutive patients were enrolled. The median follow-up was 62 months (range 24-124). The 5-year event-free survival probability was 0.43 [standard deviation (SD) = 0.05] and the 5-year overall survival probability was 0.52 (SD = 0.052). Unfavorable prognostic factors emerging on multivariate analysis were a poor histological/radiological response at the site of the primary tumor [relative risk (RR) = 3.4], and incomplete radiological remission of lung metastases after primary chemotherapy (RR = 2.6). One toxic death and one secondary leukemia were recorded. This intensive approach is feasible and long-term survival is achievable in ∼50% of patients. New treatment approaches are warranted for patients responding poorly to primary chemotherapy.

  20. Stem cells decreased neuronal cell death after hypoxic stress in primary fetal rat neurons in vitro.

    PubMed

    Sakai, Tetsuro; Xu, Yan

    2012-01-01

    To explore stem cell-mediated neuronal protection through extracellular signaling pathways by transplanted stem cells, we sought to identify potential candidate molecules responsible for neuronal protection using an in vitro coculture system. Primary fetal rat hippocampal neurons underwent hypoxia (≤1% oxygen) for 96 h nad then were returned to a normoxic condition. The study group then received rat umbilical cord matrix-derived stem cells, while the control group received fresh media only. The experimental group showed decreased neuronal apoptosis compared to the control group [44.5 ± 1.6% vs. 71.0 ± 4.2% (mean ± SD, p = 0.0005) on day 5] and higher neuronal survival (4.9 ± 1.2 cells/100× field vs. 2.2 ± 0.3, p = 0.02 on day 5). Among 90 proteins evaluated using a protein array, stem cell coculture media showed increased protein secretion of TIMP-1 (5.61-fold), TIMP-2 (4.88), CNTF-Rα (3.42), activin A (2.20), fractalkine (2.04), CCR4 (2.02), and decreased secretion in MIP-2 (0.30-fold), AMPK α1 (0.43), TROY (0.48), and TIMP-3 (0.50). This study demonstrated that coculturing stem cells with primary neurons in vitro decreased neuronal cell death after hypoxia with significantly altered protein secretion. The results suggest that stem cells may offer neuronal protection through extracellular signaling.

  1. Structural characterization and primary in vitro cell culture of locust male germline stem cells and their niche.

    PubMed

    Dorn, David C; Dorn, August

    2011-03-01

    The establishment of in vitro culture systems to expand stem cells and to elucidate the niche/stem cell interaction is among the most sought-after culture systems of our time. To further investigate niche/stem cell interactions, we evaluated in vitro cultures of isolated intact male germline-niche complexes (i.e., apical complexes), complexes with empty niche spaces, and completely empty niches (i.e., isolated apical cells) from the testes of Locusta migratoria and the interaction of these complexes with isolated germline stem cells, spermatogonia (of transit-amplifying stages), cyst progenitor cells, cyst progenitor cell-like cells, cyst cells, and follicle envelope cells. The structural characteristics of these cell types allow the identification of the different cell types in primary cultures, which we studied in detail by light and electron microscopy. In intact testes germline stem cells strongly adhere to their niche (the apical cell), but emigrate from their niche and form filopodia if the apical complex is put into culture with "standard media." The lively movements of the long filopodia of isolated germline stem cells and spermatogonia may be indicative of their search for specific signals to home to their niche. All other incubated cell types (except for follicle envelope cells) expressed rhizopodia and lobopodia. Nevertheless isolated germline stem cells in culture do not migrate to empty niche spaces of nearby apical cells. This could indicate that apical cells lose their germline stem cell attracting ability in vitro, although apical cells devoid of germline stem cells either by emigration of germline stem cells or by mechanical removal of germline stem cells are capable of surviving in vitro up to 56 days, forming many small lobopodia and performing amoeboid movements. We hypothesize that the breakdown of the apical complex in vitro with standard media interrupts the signaling between the germline stem cells and the niche (and conceivably the cyst

  2. Primary stroke in a woman with sickle cell anemia responsive to hydroxyurea therapy.

    PubMed

    Ballas, Samir K; Martinez, Ubaldo; Savage, Michael

    2014-01-01

    The most common cause of stroke in children with sickle cell anemia is infarction due to ischemia. In adults, however, stroke is most commonly hemorrhagic in nature. Other causes of stroke in patients with sickle cell disease are very rare. In this short communication, we describe a woman with sickle cell anemia responsive to hydroxyurea (HU) therapy who had primary stroke due to paradoxical embolization caused by a large atrial septal defect. Successful management of the stroke included surgical closure of the defect with trans-esophageal echocardiographic guidance. To the best of our knowledge, this is the first patient with sickle cell anemia and stroke due to congenital heart disease who did not require open heart surgery for successful management.

  3. Development method of Hybrid Energy Storage System, including PEM fuel cell and a battery

    NASA Astrophysics Data System (ADS)

    Ustinov, A.; Khayrullina, A.; Borzenko, V.; Khmelik, M.; Sveshnikova, A.

    2016-09-01

    Development of fuel cell (FC) and hydrogen metal-hydride storage (MH) technologies continuously demonstrate higher efficiency rates and higher safety, as hydrogen is stored at low pressures of about 2 bar in a bounded state. A combination of a FC/MH system with an electrolyser, powered with a renewable source, allows creation of an almost fully autonomous power system, which could potentially replace a diesel-generator as a back-up power supply. However, the system must be extended with an electro-chemical battery to start-up the FC and compensate the electric load when FC fails to deliver the necessary power. Present paper delivers the results of experimental and theoretical investigation of a hybrid energy system, including a proton exchange membrane (PEM) FC, MH- accumulator and an electro-chemical battery, development methodology for such systems and the modelling of different battery types, using hardware-in-the-loop approach. The economic efficiency of the proposed solution is discussed using an example of power supply of a real town of Batamai in Russia.

  4. Time-lapse imaging of primary preneoplastic mammary epithelial cells derived from genetically engineered mouse models of breast cancer.

    PubMed

    Nakles, Rebecca E; Millman, Sarah L; Cabrera, M Carla; Johnson, Peter; Mueller, Susette; Hoppe, Philipp S; Schroeder, Timm; Furth, Priscilla A

    2013-02-08

    Time-lapse imaging can be used to compare behavior of cultured primary preneoplastic mammary epithelial cells derived from different genetically engineered mouse models of breast cancer. For example, time between cell divisions (cell lifetimes), apoptotic cell numbers, evolution of morphological changes, and mechanism of colony formation can be quantified and compared in cells carrying specific genetic lesions. Primary mammary epithelial cell cultures are generated from mammary glands without palpable tumor. Glands are carefully resected with clear separation from adjacent muscle, lymph nodes are removed, and single-cell suspensions of enriched mammary epithelial cells are generated by mincing mammary tissue followed by enzymatic dissociation and filtration. Single-cell suspensions are plated and placed directly under a microscope within an incubator chamber for live-cell imaging. Sixteen 650 μm x 700 μm fields in a 4x4 configuration from each well of a 6-well plate are imaged every 15 min for 5 days. Time-lapse images are examined directly to measure cellular behaviors that can include mechanism and frequency of cell colony formation within the first 24 hr of plating the cells (aggregation versus cell proliferation), incidence of apoptosis, and phasing of morphological changes. Single-cell tracking is used to generate cell fate maps for measurement of individual cell lifetimes and investigation of cell division patterns. Quantitative data are statistically analyzed to assess for significant differences in behavior correlated with specific genetic lesions.

  5. Mouse newborn cells allow highly productive mouse cytomegalovirus replication, constituting a novel convenient primary cell culture system.

    PubMed

    Le-Trilling, Vu Thuy Khanh; Trilling, Mirko

    2017-01-01

    Mammalian cell culture is indispensable for most aspects of current biomedical research. Immortalized cell lines are very convenient, but transforming principles (e.g. oncogenic viruses or their oncogenes) can heavily influence the experimental outcome. Primary cells do not share this apparent disadvantage but are more laborious to generate. Certain viruses (e.g. mouse cytomegalovirus) do not replicate efficiently in most transformed cell lines. In the past, such viruses have been routinely propagated on primary mouse embryonic fibroblasts (MEF) established around day 17 (d17) of gestation. According to new regulations of the European Union, experiments using gravid mammals and/or their embryos in the last trimester (>d14 in the case of mice) of gestation do require explicit permission of the local authorities responsible for animal care and use. Applying for such permission is time-consuming and often inflexible. Embryonic fibroblasts could also be produced at earlier time points of pregnancy from younger and smaller embryos. Obviously, this approach consumes more pregnant mice and embryos. Newborn mice are larger thus yielding more cells per sacrificed animal and the new Directive (2010/63/EU) excludes the killing of animals solely for the use of their organs or tissues. We established a convenient protocol to generate adherent mouse newborn cells (MNC). A direct comparison of MNC with MEF revealed that MNC fully recapitulate all tested aspects of a broad panel of virological parameters (plaque size, final titers, viral replication kinetics, viral gene expression, drug and interferon susceptibility as well as species specificity). The herein described approach allows researchers the legal use of primary cells and contributes to the 3R (replace, reduce, refine) guiding principles-especially the 'reduce' aspect-for the use of animals in scientific research. Additionally, it offers the option to directly compare in vitro and in vivo experiments when MNC are

  6. Mouse newborn cells allow highly productive mouse cytomegalovirus replication, constituting a novel convenient primary cell culture system

    PubMed Central

    Le-Trilling, Vu Thuy Khanh

    2017-01-01

    Mammalian cell culture is indispensable for most aspects of current biomedical research. Immortalized cell lines are very convenient, but transforming principles (e.g. oncogenic viruses or their oncogenes) can heavily influence the experimental outcome. Primary cells do not share this apparent disadvantage but are more laborious to generate. Certain viruses (e.g. mouse cytomegalovirus) do not replicate efficiently in most transformed cell lines. In the past, such viruses have been routinely propagated on primary mouse embryonic fibroblasts (MEF) established around day 17 (d17) of gestation. According to new regulations of the European Union, experiments using gravid mammals and/or their embryos in the last trimester (>d14 in the case of mice) of gestation do require explicit permission of the local authorities responsible for animal care and use. Applying for such permission is time-consuming and often inflexible. Embryonic fibroblasts could also be produced at earlier time points of pregnancy from younger and smaller embryos. Obviously, this approach consumes more pregnant mice and embryos. Newborn mice are larger thus yielding more cells per sacrificed animal and the new Directive (2010/63/EU) excludes the killing of animals solely for the use of their organs or tissues. We established a convenient protocol to generate adherent mouse newborn cells (MNC). A direct comparison of MNC with MEF revealed that MNC fully recapitulate all tested aspects of a broad panel of virological parameters (plaque size, final titers, viral replication kinetics, viral gene expression, drug and interferon susceptibility as well as species specificity). The herein described approach allows researchers the legal use of primary cells and contributes to the 3R (replace, reduce, refine) guiding principles—especially the ‘reduce’ aspect—for the use of animals in scientific research. Additionally, it offers the option to directly compare in vitro and in vivo experiments when MNC are

  7. Integrated management of atrial fibrillation including tailoring of anticoagulation in primary care: study design of the ALL-IN cluster randomised trial.

    PubMed

    van den Dries, Carline J; Oudega, Ruud; Elvan, Arif; Rutten, Frans H; van de Leur, Sjef J C M; Bilo, Henk J G; Hoes, Arno W; Moons, Karel G M; Geersing, Geert-Jan

    2017-09-18

    In our ageing society, we are at the merge of an expected epidemic of atrial fibrillation (AF). AF management requires an integrated approach, including rate or rhythm control, stroke prevention with anticoagulation and treatment of comorbidities such as heart failure or type 2 diabetes. As such, primary care seems to be the logical healthcare setting for the chronic management of patients with AF. However, primary care has not yet played a dominant role in AF management, which has been in fact more fragmented between different healthcare providers. This fragmentation might have contributed to high healthcare costs. To demonstrate the feasibility of managing AF in primary care, studies are needed that evaluate the safety and (cost-)effectiveness of integrated AF management in primary care. The ALL-IN trial is a multicentre, pragmatic, cluster randomised, non-inferiority trial performed in primary care practices in a suburban region in the Netherlands. We aim to include a minimum of 1000 patients with AF aged 65 years or more from around 18 to 30 practices. Duration of the study is 2 years. Practices will be randomised to either the intervention arm (providing integrated AF management, involving a trained practice nurse and collaboration with secondary care) or the control arm (care as usual). The primary endpoint is all-cause mortality. Secondary endpoints are cardiovascular mortality, (non)-cardiovascular hospitalisation, major adverse cardiac events, stroke, major bleeding, clinically relevant non-major bleeding, quality of life and cost-effectiveness. The protocol was approved by the Medical Ethical Committee of the Isala Hospital Zwolle, the Netherlands. Patients in the intervention arm will be asked informed consent for participating in the intervention. Results are expected in 2019 and will be disseminated through both national and international journals and conferences. This trial is registered at the Netherlands Trial Register (NTR5532). © Article author

  8. Interactions of silver nanoparticles with primary mouse fibroblasts and liver cells

    SciTech Connect

    Arora, S.; Jain, J.; Rajwade, J.M.; Paknikar, K.M.

    2009-05-01

    Primary cells are ideal for in vitro toxicity studies since they closely resemble tissue environment. Here, we report a detailed study on the in vitro interactions of 7-20 nm spherical silver nanoparticles (SNP) with primary fibroblasts and primary liver cells isolated from Swiss albino mice. The intended use of silver nanoparticles is in the form of a topical antimicrobial gel formulation for the treatment of burns and wounds. Upon exposure to SNP for 24 h, morphology of primary fibroblasts and primary liver cells remained unaltered up to 25 {mu}g/mL and 100 {mu}g/mL SNP, respectively, although with minor decrease in confluence. IC{sub 50} values for primary fibroblasts and primary liver cells as revealed by XTT assay were 61 {mu}g/mL and 449 {mu}g/mL, respectively. Ultra-thin sections of primary cells exposed to 1/2 IC{sub 50} SNP for 24 h, visualized under Transmission electron microscope showed the presence of dark, electron dense, spherical aggregates inside the mitochondria, and cytoplasm, probably representing the intracellular SNP. When the cells were challenged with {approx} 1/2 IC{sub 50} concentration of SNP (i.e. 30 {mu}g/mL and 225 {mu}g/mL for primary fibroblasts and primary liver cells, respectively), enhancement of GSH ({approx} 1.2 fold) and depletion of lipid peroxidation ({approx} 1.4 fold) were seen in primary fibroblasts which probably protect the cells from functional damage. In case of primary liver cells; increased levels of SOD ({approx} 1.4 fold) and GSH ({approx} 1.1 fold) as compared to unexposed cells were observed. Caspase-3 activity assay indicated that the SNP concentrations required for the onset of apoptosis were found to be much lower (3.12 {mu}g/mL in primary fibroblasts, 12.5 {mu}g/mL in primary liver cells) than the necrotic concentration (100 {mu}g/mL in primary fibroblasts, 500 {mu}g/mL in primary liver cells). These observations were confirmed by CLSM studies by exposure of cells to 1/2 IC{sub 50} SNP (resulting in apoptosis

  9. Comparative Gene Expression Profiling of Primary and Metastatic Renal Cell Carcinoma Stem Cell-Like Cancer Cells.

    PubMed

    Khan, Mohammed I; Czarnecka, Anna M; Lewicki, Sławomir; Helbrecht, Igor; Brodaczewska, Klaudia; Koch, Irena; Zdanowski, Robert; Król, Magdalena; Szczylik, Cezary

    2016-01-01

    Recent advancement in cancer research has shown that tumors are highly heterogeneous, and multiple phenotypically different cell populations are found in a single tumor. Cancer development and tumor growth are driven by specific types of cells-stem cell-like cancer cells (SCLCCs)-which are also responsible for metastatic spread and drug resistance. This research was designed to verify the presence of SCLCCs in renal cell cancer cell lines. Subsequently, we aimed to characterize phenotype and cell biology of CD105+ cells, defined previously as renal cell carcinoma tumor-initiating cells. The main goal of the project was to describe the gene-expression profile of stem cell-like cancer cells of primary tumor and metastatic origin. Real-time PCR analysis of stemness genes (Oct-4, Nanog and Ncam) and soft agar colony formation assay were conducted to check the stemness properties of renal cell carcinoma (RCC) cell lines. FACS analysis of CD105+ and CD133+ cells was performed on RCC cells. Isolated CD105+ cells were verified for expression of mesenchymal markers-CD24, CD146, CD90, CD73, CD44, CD11b, CD19, CD34, CD45, HLA-DR and alkaline phosphatase. Hanging drop assay was used to investigate CD105+ cell-cell cohesion. Analysis of free-floating 3D spheres formed by isolated CD105+ was verified, as spheres have been hypothesized to contain undifferentiated multipotent progenitor cells. Finally, CD105+ cells were sorted from primary (Caki-2) and metastatic (ACHN) renal cell cancer cell lines. Gene-expression profiling of sorted CD105+ cells was performed with Agilent's human GE 4x44K v2 microarrays. Differentially expressed genes were further categorized into canonical pathways. Network analysis and downstream analysis were performed with Ingenuity Pathway Analysis. Metastatic RCC cell lines (ACHN and Caki-1) demonstrated higher colony-forming ability in comparison to primary RCC cell lines. Metastatic RCC cell lines harbor numerous CD105+ cell subpopulations and have

  10. Performance characteristics of lithium primary cells after controlled storage. [on-orbit for energy power supply

    NASA Technical Reports Server (NTRS)

    Deligiannis, F.; Shen, D. H.; Halpert, G.; Ang, V.; Donley, S.

    1991-01-01

    A program was initiated to investigate the effects of storage on the performance of lithium primary cells. Two types of liquid cathode cells were chosen to investigate these effects. The cell types included Li-SOCl2/BCX cells, Li-SO2 cells from two different manufacturers, and a small sample size of 8-year-old Li-SO2 cells. The following measurements are performed at each test interval: open circuit voltage, resistance and weight, microcalorimetry, ac impedance, capacity, and voltage delay. The authors examine the performance characteristics of these cells after one year of controlled storage at two temperatures (10 and 30 C). The Li-SO2 cells experienced little to no voltage and capacity degradation after one year storage. The Li-SOCl2/BCX cells exhibited significant voltage and capacity degradation after 30 C storage. Predischarging shortly prior to use appears to be an effective method of reducing the initial voltage drop. Studies are in progress to correlate ac impedance and microcalorimetry measurements with capacity losses and voltage delay.

  11. Large cell variant of small cell carcinoma, hypercalcemic type, of primary peritoneal origin.

    PubMed

    Popiolek, Dorota A; Kumar, Asok R; Mittal, Khush

    2005-01-01

    Large cell variant of small cell carcinoma hypercalcemic type (SCC-HT) is extremely rare. All reported cases involved an ovary, and one with primary peritoneal origin has not been described. Also, convincing neuroendocrine granules have not been illustrated. A 35-year-old woman underwent an exploratory laparotomy for leiomyomas. Intraoperative impression of peritoneal carcinomatosis was confirmed on frozen section. TAH/BSO, debulking/omentectomy followed. The tumor was present on the pelvic/abdominal peritoneum. The normal-sized ovaries were free of tumor grossly. The tumor had features of large cell variant of SCC-HT, described in the ovary. Furthermore, unequivocal neuroendocrine granules were present. The patient received standard chemotherapy for SCC. At 22 months she is NED. SCC-HT should be considered in the differential diagnosis of primary neoplasms of the peritoneum.

  12. Pericardial, pleural and peritoneal involvement in a patient with primary gastric mantle cell lymphoma.

    PubMed

    Keklik, Muzaffer; Yildirim, Afra; Keklik, Ertugrul; Ertan, Sirac; Deniz, Kemal; Ozturk, Fahir; Ileri, Ibrahim; Cerci, Ilkcan; Camlica, Demet; Cetin, Mustafa; Eser, Bulent

    2015-05-01

    Primary gastric mantle cell lymphoma is a rare form of gastointestinal tumour. Although peritoneal carcinomatosis accompanied by malignant ascites is relatively common, mantle cell lymphoma presenting with ascites is rare. Also, effusions involving pericardial and pleural cavities are uncommon during the course of lymphomas. We report the first case in which pericardial, pleural and peritoneal effusion of a primary gastric mantle cell lymphoma.

  13. Comparative Gene Expression Profiling of Primary and Metastatic Renal Cell Carcinoma Stem Cell-Like Cancer Cells

    PubMed Central

    Czarnecka, Anna M.; Lewicki, Sławomir; Helbrecht, Igor; Brodaczewska, Klaudia; Koch, Irena; Zdanowski, Robert; Król, Magdalena; Szczylik, Cezary

    2016-01-01

    Background Recent advancement in cancer research has shown that tumors are highly heterogeneous, and multiple phenotypically different cell populations are found in a single tumor. Cancer development and tumor growth are driven by specific types of cells—stem cell-like cancer cells (SCLCCs)—which are also responsible for metastatic spread and drug resistance. This research was designed to verify the presence of SCLCCs in renal cell cancer cell lines. Subsequently, we aimed to characterize phenotype and cell biology of CD105+ cells, defined previously as renal cell carcinoma tumor-initiating cells. The main goal of the project was to describe the gene-expression profile of stem cell-like cancer cells of primary tumor and metastatic origin. Materials and Methods Real-time PCR analysis of stemness genes (Oct-4, Nanog and Ncam) and soft agar colony formation assay were conducted to check the stemness properties of renal cell carcinoma (RCC) cell lines. FACS analysis of CD105+ and CD133+ cells was performed on RCC cells. Isolated CD105+ cells were verified for expression of mesenchymal markers—CD24, CD146, CD90, CD73, CD44, CD11b, CD19, CD34, CD45, HLA-DR and alkaline phosphatase. Hanging drop assay was used to investigate CD105+ cell-cell cohesion. Analysis of free-floating 3D spheres formed by isolated CD105+ was verified, as spheres have been hypothesized to contain undifferentiated multipotent progenitor cells. Finally, CD105+ cells were sorted from primary (Caki-2) and metastatic (ACHN) renal cell cancer cell lines. Gene-expression profiling of sorted CD105+ cells was performed with Agilent’s human GE 4x44K v2 microarrays. Differentially expressed genes were further categorized into canonical pathways. Network analysis and downstream analysis were performed with Ingenuity Pathway Analysis. Results Metastatic RCC cell lines (ACHN and Caki-1) demonstrated higher colony-forming ability in comparison to primary RCC cell lines. Metastatic RCC cell lines harbor

  14. Resveratrol induces cell death and inhibits human herpesvirus 8 replication in primary effusion lymphoma cells.

    PubMed

    Tang, Feng-Yi; Chen, Chang-Yu; Shyu, Huey-Wen; Hong, Shin; Chen, Hung-Ming; Chiou, Yee-Hsuan; Lin, Kuan-Hua; Chou, Miao-Chen; Wang, Lin-Yu; Wang, Yi-Fen

    2015-12-05

    Resveratrol (3,4',5-trihydroxy-trans-stilbene) has been reported to inhibit proliferation of various cancer cells. However, the effects of resveratrol on the human herpesvirus 8 (HHV8) harboring primary effusion lymphoma (PEL) cells remains unclear. The anti-proliferation effects and possible mechanisms of resveratrol in the HHV8 harboring PEL cells were examined in this study. Results showed that resveratrol induced caspase-3 activation and the formation of acidic vacuoles in the HHV8 harboring PEL cells, indicating resveratrol treatment could cause apoptosis and autophagy in PEL cells. In addition, resveratrol treatment increased ROS generation but did not lead to HHV8 reactivation. ROS scavenger (N-acetyl cysteine, NAC) could attenuate both the resveratrol induced caspase-3 activity and the formation of acidic vacuoles, but failed to attenuate resveratrol induced PEL cell death. Caspase inhibitor, autophagy inhibitors and necroptosis inhibitor could not block resveratrol induced PEL cell death. Moreover, resveratrol disrupted HHV8 latent infection, inhibited HHV8 lytic gene expression and decreased virus progeny production. Overexpression of HHV8-encoded viral FLICE inhibitory protein (vFLIP) could partially block resveratrol induced cell death in PEL cells. These data suggest that resveratrol-induced cell death in PEL cells may be mediated by disruption of HHV8 replication. Resveratrol may be a potential anti-HHV8 drug and an effective treatment for HHV8-related tumors.

  15. Metabolic detoxication pathways for sterigmatocystin in primary tracheal epithelial cells.

    PubMed

    Cabaret, Odile; Puel, Olivier; Botterel, Françoise; Pean, Michel; Khoufache, Khaled; Costa, Jean-Marc; Delaforge, Marcel; Bretagne, Stéphane

    2010-11-15

    Human health effects of inhaled mycotoxins remain poorly documented, despite the large amounts present in bioaerosols. Among these mycotoxins, sterigmatocystin is one of the most prevalent. Our aim was to study the metabolism and cellular consequences of sterigmatocystin once it is in contact with the airway epithelium. Metabolites were analyzed first in vitro, using recombinant P450 1A1, 1A2, 2A6, 2A13, and 3A4 enzymes, and subsequently in porcine tracheal epithelial cell (PTEC) primary cultures at an air-liquid interface. Expressed enzymes and PTECs were exposed to sterigmatocystin, uniformly enriched with (13)C to confirm the relationship between sterigmatocystin and metabolites. Induction of the expression of xenobiotic-metabolizing enzymes upon sterigmatocystin exposure was examined by real-time quantitative real-time polymerase chain reaction. Incubation of 50 μM sterigmatocystin with recombinant P450 1A1 led to the formation of three metabolites: monohydroxy-sterigmatocystin (M1), dihydroxy-sterigmatocystin (M2), and one glutathione adduct (M3), the latter after the formation of a transient epoxide. Recombinant P450 1A2 also led to M1 and M3. P450 3A4 led to only M3. In PTEC, 1 μM sterigmatocystin metabolism resulted in a glucuro conjugate (M4) mainly excreted at the basal side of cells. If PTEC were treated with β-naphthoflavone prior to sterigmatocystin incubation, two other products were detected, i.e., a sulfo conjugate (M5) and a glucoro conjugate (M6) of hydroxy-sterigmatocystin. Exposure of PTEC for 24 h to 1 μM sterigmatocystin induced an 18-fold increase in the mRNA levels of P450 1A1, without significantly induced 7-ethoxyresorufin O-deethylation activity. These data suggest that sterigmatocystin is mainly detoxified and is unable to produce significant amounts of reactive epoxide metabolites in respiratory cells. However, sterigmatocystin increases the P450 1A1 mRNA levels with unknown long-term consequences. These in vitro results obtained in

  16. Multiple defects, including premature apoptosis, prevent Kaposi's sarcoma-associated herpesvirus replication in murine cells.

    PubMed

    Austgen, Kathryn; Oakes, Scott A; Ganem, Don

    2012-02-01

    The development of a mouse model for Kaposi's sarcoma-associated herpesvirus (KSHV) infection has been impeded by the limited host range of the virus. Here, we have examined the molecular basis of this host range restriction. KSHV efficiently enters murine cells and establishes latency. However, ectopic expression of the lytic switch protein RTA (replication and transcription activator) in these cells induces little viral gene expression and no virus production. Upon treatment with histone deacetylase inhibitors, KSHV-infected murine cells display more extensive but aberrant viral transcription and do not support either viral DNA synthesis or the production of infectious virions. These aberrantly infected cells also display markedly enhanced apoptosis. Genetic ablation of the mitochondrial apoptotic pathway in these cells prolongs their survival and permits viral DNA replication but does not rescue the generation of virions. We conclude that multiple defects, both prior to and following DNA synthesis, restrict lytic KSHV infection in murine cells.

  17. Expansion of cytotoxic CD8+ CD28- T cells in healthy ageing people, including centenarians.

    PubMed Central

    Fagnoni, F F; Vescovini, R; Mazzola, M; Bologna, G; Nigro, E; Lavagetto, G; Franceschi, C; Passeri, M; Sansoni, P

    1996-01-01

    Ageing is associated with complex remodelling in the phenotypic and functional profiles of T lymphocytes. We investigated whether expression of CD28 antigen on T cells is conserved throughout adulthood and ageing in humans. For this purpose we analysed T cells obtained from peripheral blood of 102 healthy people of ages ranging from 20 to 105 years. We found an age-related increase of CD28- T cells in percentage and absolute number, predominantly among CD8+ T cells. CD28- T cells from aged donors analysed by flow cytometry appeared as resting cells (not expressing CD25, CD38, CD69, CD71, DR), bearing markers of cytotoxic activity (CD 11b and CD 57) and with a phenotype compatible with 'memory' cells (up-regulated CD2 and CD11a; CD62L absent). At the functional level, freshly isolated purified CD28- CD8+ T cells showed high anti-CD3 redirected cytotoxic activity against Fc-bearing P815 cells. The same activity tested on freshly isolated bulk T lymphocytes was significantly augmented with age. We found a positive correlation between age, number of CD8+ CD28- T cells and anti-CD3 redirected cytotoxicity by freshly isolated T cells. These data suggest that an activation of unknown nature within the cytotoxic arm of the immune system occurs with age. We speculate that these cytotoxic T lymphocytes (CTL) in vivo may constitute armed effector cells for immediate killing of targets bearing peptides from pathogens of intracellular origin. PMID:8881749

  18. Human embryonic stem cells in culture possess primary cilia with hedgehog signaling machinery

    PubMed Central

    Kiprilov, Enko N.; Awan, Aashir; Desprat, Romain; Velho, Michelle; Clement, Christian A.; Byskov, Anne Grete; Andersen, Claus Y.; Satir, Peter; Bouhassira, Eric E.; Christensen, Søren T.; Hirsch, Rhoda Elison

    2008-01-01

    Human embryonic stem cells (hESCs) are potential therapeutic tools and models of human development. With a growing interest in primary cilia in signal transduction pathways that are crucial for embryological development and tissue differentiation and interest in mechanisms regulating human hESC differentiation, demonstrating the existence of primary cilia and the localization of signaling components in undifferentiated hESCs establishes a mechanistic basis for the regulation of hESC differentiation. Using electron microscopy (EM), immunofluorescence, and confocal microscopies, we show that primary cilia 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. PMID:18332216

  19. Human embryonic stem cells in culture possess primary cilia with hedgehog signaling machinery.

    PubMed

    Kiprilov, Enko N; Awan, Aashir; Desprat, Romain; Velho, Michelle; Clement, Christian A; Byskov, Anne Grete; Andersen, Claus Y; Satir, Peter; Bouhassira, Eric E; Christensen, Søren T; Hirsch, Rhoda Elison

    2008-03-10

    Human embryonic stem cells (hESCs) are potential therapeutic tools and models of human development. With a growing interest in primary cilia in signal transduction pathways that are crucial for embryological development and tissue differentiation and interest in mechanisms regulating human hESC differentiation, demonstrating the existence of primary cilia and the localization of signaling components in undifferentiated hESCs establishes a mechanistic basis for the regulation of hESC differentiation. Using electron microscopy (EM), immunofluorescence, and confocal microscopies, we show that primary cilia 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.

  20. Early and sustained expression of latent and host modulating genes in coordinated transcriptional program of KSHV productive primary infection of human primary endothelial cells

    PubMed Central

    Yoo, Seung Min; Zhou, Fu-Chun; Ye, Feng-Chun; Pan, Hong-Yi; Gao, Shou-Jiang

    2009-01-01

    Coordinated expression of viral genes in primary infection is essential for successful infection of host cells. We examined the expression profiles of Kaposi’s sarcoma-associated herpesvirus (KSHV) transcripts in productive primary infection of primary human umbilical vein endothelial cells by whole-genome reverse-transcription real-time quantitative PCR. The latent transcripts were expressed early and sustained at high levels throughout the infection while the lytic transcripts were expressed in the order of immediate early, early, and lytic transcripts, all of which culminated before the production of infectious virions. Significantly, transcripts encoding genes with host modulating functions, including mitogenic and cell cycle-regulatory, immune-modulating, and anti-apoptotic genes, were expressed before those encoding viral structure and replication genes, and sustained at high levels throughout the infection, suggesting KSHV manipulation of host environment to facilitate infection. The KSHV transcriptional program in a primary infection defined in this study should provide a basis for further investigation of virus–cell interactions. PMID:16154170

  1. Complete androgen insensitivity syndrome: factors influencing gonadal histology including germ cell pathology.

    PubMed

    Kaprova-Pleskacova, Jana; Stoop, Hans; Brüggenwirth, Hennie; Cools, Martine; Wolffenbuttel, Katja P; Drop, Stenvert L S; Snajderova, Marta; Lebl, Jan; Oosterhuis, J Wolter; Looijenga, Leendert H J

    2014-05-01

    Patients with complete androgen insensitivity syndrome are at an increased risk for the development of gonadal germ cell cancer. Residual androgen receptor (AR) activity and abnormal gonadal location may influence the survival of atypical germ cells and the development of other histopathological features. To assess this, we evaluated 37 gonads from 19 patients with complete androgen insensitivity (ranging in age from 3 months to 18 years). Histological abnormalities were examined using hematoxylin and eosin-stained sections and sections stained for POU5F1 and KITLG, markers of early changes in germ cells at risk for malignant transformation. Hamartomatous nodules (HNs), Leydig cell hyperplasia (LCH), decreased germ cells, tubular atrophy and stromal fibrosis were more pronounced as age increased (P<0.001). Expected residual AR activity acted as a positive predictor only for non-malignant germ cell survival in (post)pubertal patients (P<0.05). Immunohistochemical studies indicated that delayed maturation of germ cells was present in three patients, whereas intermediate changes that occurred between delayed maturation and intratubular germ cell neoplasia, designated pre-intratubular germ cell neoplasia, were identified in four cases. Intratubular germ cell neoplasia was observed in one patient. Neither POU5F1 nor KITLG expression was dependent on expected residual AR activity. An independent effect of inguinal versus abdominal position of the gonads was difficult to assess because inguinal gonads were present primarily in the youngest individuals. In conclusion, many histological changes occur increasingly with age. Expected residual AR activity contributes to better survival of the general germ cell population in (post)pubertal age; however, it did not seem to have an important role in the survival of the germ cells at risk for malignant transformation (defined by POU5F1 positivity and KITLG overexpression) in complete androgen insensitivity. Comparison of the high

  2. Primary mesenchyme cell migration requires a chondroitin sulfate/dermatan sulfate proteoglycan.

    PubMed

    Lane, M C; Solursh, M

    1991-02-01

    Primary mesenchyme cell migration in the sea urchin embryo is inhibited by sulfate deprivation and exposure to exogenous beta-D-xylosides, two treatments known to disrupt proteoglycan synthesis. We show that in the developing sea urchin, exogenous xyloside affects the synthesis by the primary mesenchyme cells of a very large, cell surface chondroitin sulfate/dermatan sulfate proteoglycan. This proteoglycan is present in a partially purified fraction that restores migratory ability to defective cells in vitro. The integrity of this chondroitin sulfate/dermatan sulfate proteoglycan appears essential for primary mesenchyme cell migration since treatment of actively migrating cells with chondroitinase ABC reversibly inhibited their migration in vitro.

  3. Regulation of human renin expression in chorion cell primary cultures

    SciTech Connect

    Duncan, K.G.; Haidar, M.A.; Baxter, J.D.; Reudelhuber, T.L. )

    1990-10-01

    The human renin gene is expressed in the kidney, placenta, and several other sites. The release of renin or its precursor, prorenin, can be affected by several regulatory agents. In this study, primary cultures of human placental cells were used to examine the regulation of prorenin release and renin mRNA levels and of the transfected human renin promoter linked to chloramphenicol acetyltransferase reporter sequences. Treatment of the cultures with a calcium ionophore alone, calcium ionophore plus forskolin (that activates adenylate cyclase), or forskolin plus a phorbol ester increased prorenin release and renin mRNA levels 1.3{endash} to 6{endash}fold, but several classes of steroids did not affect prorenin secretion or renin RNA levels. These results suggest that (i) the first 584 base pairs of the renin gene 5'{endash}flanking DNA do not contain functional glucocorticoid or estrogen response elements, (ii) placental prorenin release and renin mRNA are regulated by calcium ion and by the combinations of cAMP with either C kinase or calcium ion, and (iii) the first 100 base pairs of the human renin 5'{endash}flanking DNA direct accurate initiation of transcription and can be regulated by cAMP. Thus, some control of renin release in the placenta (and by inference in other tissues) occurs via transcriptional influences on its promoter.

  4. [Primary squamous cell carcinoma of the breast: rare form carcinoma].

    PubMed

    Maksimović, Sinisa

    2009-01-01

    Primary squamous cell carcinoma (SCC) is a rare form of breast carcinoma. Incidence is reported to be 0.1-3.6%. We report a case of a young woman, 37-year-old, with history of a lump in the upper outer quadrant of the left breast with ulceration of the skin surface. Menarche occurred at age of 12. The patient was married, had two deliveries and had her first child at age of 26. She did not use contraceptive pills. Diagnosis of the tumour of the breast was made at the Department of surgery in General Hospital in Bijeljina in September 2007. Clinical examination, mammography and ultrasonography were performed. Physical examination revealed a circumscribed and firm mass measuring 60 x 60 x 80 mm. Mammogram showed a round, high-density mass with almost regular but partially irregular margin. Ultrasonogram of the left breast tumor identified an irregularly shaped hypoechoic lesion. After clinical staging of the disease, we performed incision biopsy of the skin and tumour of the left breast with histopathology examination (standard hematoxylin and eosin). Patient had estrogen and progesteron receptors negative and was HER2/neu negative. After histopathology, patient's case was presented to the working group for breast tumors which decided to start with the neoadjuvant chemotherapy using platinum. After six cycles of neoadjuvant chemotherapy, regression of breast tumor was confirmed. Working group decided that radical mastectomy of left breast should be performed.

  5. Porcine Endogenous Retrovirus Infects but Does Not Replicate in Nonhuman Primate Primary Cells and Cell Lines

    PubMed Central

    Ritzhaupt, Armin; van der Laan, Luc J. W.; Salomon, Daniel R.; Wilson, Carolyn A.

    2002-01-01

    Porcine endogenous retroviruses (PERV) can infect human cell lines in vitro; hence, there is a presumed risk of viral exposure to a recipient when pig cells are transplanted into humans (xenotransplantation). Nonhuman primates (NHP) are considered a potential permissive animal model to study the risk of in vivo infection of PERV after xenotransplantation. We set out to determine whether PERV can infect and replicate in NHP primary cells or established cell lines from African green monkey, rhesus macaque, and baboon. We confirm that the NHP cell lines under investigation were infected with PERV as measured by detection of viral DNA and RNA by PCR and reverse transcription (RT)-PCR, respectively, indicating that a functional receptor must be present on the cell surface. However, the load of detectable viral DNA in infected NHP cells declined over time, and the cells never had detectable reverse transcriptase activity. Utilizing quantitative real-time TaqMan PCR we found detectable levels of unintegrated DNA intermediates, but the levels were approximately 100-fold lower compared to HEK 293 cells infected with PERV. Virions released from infected NHP cells could productively infect naïve human cell lines, HEK 293 and HeLa, as shown by RT-PCR and RT assay. However, naïve NHP cells remained negative in RT-PCR and RT assay after exposure to virions from infected NHP cells. Together our data demonstrate that NHP cells are not permissive to productive replication by PERV, presumably due to inefficient cell entry and replication. In light of these observations, the appropriateness of NHP as suitable animal models to study PERV infection in vivo needs to be reevaluated. PMID:12388691

  6. Global target profile of the kinase inhibitor bosutinib in primary chronic myeloid leukemia cells.

    PubMed

    Remsing Rix, L L; Rix, U; Colinge, J; Hantschel, O; Bennett, K L; Stranzl, T; Müller, A; Baumgartner, C; Valent, P; Augustin, M; Till, J H; Superti-Furga, G

    2009-03-01

    The detailed molecular mechanism of action of second-generation BCR-ABL tyrosine kinase inhibitors, including perturbed targets and pathways, should contribute to rationalized therapy in chronic myeloid leukemia (CML) or in other affected diseases. Here, we characterized the target profile of the dual SRC/ABL inhibitor bosutinib employing a two-tiered approach using chemical proteomics to identify natural binders in whole cell lysates of primary CML and K562 cells in parallel to in vitro kinase assays against a large recombinant kinase panel. The combined strategy resulted in a global survey of bosutinib targets comprised of over 45 novel tyrosine and serine/threonine kinases. We have found clear differences in the target patterns of bosutinib in primary CML cells versus the K562 cell line. A comparison of bosutinib with dasatinib across the whole kinase panel revealed overlapping, but distinct, inhibition profiles. Common among those were the SRC, ABL and TEC family kinases. Bosutinib did not inhibit KIT or platelet-derived growth factor receptor, but prominently targeted the apoptosis-linked STE20 kinases. Although in vivo bosutinib is inactive against ABL T315I, we found this clinically important mutant to be enzymatically inhibited in the mid-nanomolar range. Finally, bosutinib is the first kinase inhibitor shown to target CAMK2G, recently implicated in myeloid leukemia cell proliferation.

  7. Primary Cilia on Horizontal Basal Cells Regulate Regeneration of the Olfactory Epithelium.

    PubMed

    Joiner, Ariell M; Green, Warren W; McIntyre, Jeremy C; Allen, Benjamin L; Schwob, James E; Martens, Jeffrey R

    2015-10-07

    The olfactory epithelium (OE) is one of the few tissues to undergo constitutive neurogenesis throughout the mammalian lifespan. It is composed of multiple cell types including olfactory sensory neurons (OSNs) that are readily replaced by two populations of basal stem cells, frequently dividing globose basal cells and quiescent horizontal basal cells (HBCs). However, the precise mechanisms by which these cells mediate OE regeneration are unclear. Here, we show for the first time that the HBC subpopulation of basal stem cells uniquely possesses primary cilia that are aligned in an apical orientation in direct apposition to sustentacular cell end feet. The positioning of these cilia suggests that they function in the detection of growth signals and/or differentiation cues. To test this idea, we generated an inducible, cell type-specific Ift88 knock-out mouse line (K5rtTA;tetOCre;Ift88(fl/fl)) to disrupt cilia formation and maintenance specifically in HBCs. Surprisingly, the loss of HBC cilia did not affect the maintenance of the adult OE but dramatically impaired the regeneration of OSNs following lesion. Furthermore, the loss of cilia during development resulted in a region-specific decrease in neurogenesis, implicating HBCs in the establishment of the OE. Together, these results suggest a novel role for primary cilia in HBC activation, proliferation, and differentiation. We show for the first time the presence of primary cilia on a quiescent population of basal stem cells, the horizontal basal cells (HBCs), in the olfactory epithelium (OE). Importantly, our data demonstrate that cilia on HBCs are necessary for regeneration of the OE following injury. Moreover, the disruption of HBC cilia alters neurogenesis during the development of the OE, providing evidence that HBCs participate in the establishment of this tissue. These data suggest that the mechanisms of penetrance for ciliopathies in the OE extend beyond that of defects in olfactory sensory neurons and may

  8. Transforming primary healthcare by including the stakeholders involved in delivering care to people living in poverty: EQUIhealThY study protocol

    PubMed Central

    2013-01-01

    Background Ensuring access to timely and appropriate primary healthcare for people living in poverty is an issue facing all countries, even those with universal healthcare systems. The transformation of healthcare practices and organization could be improved by involving key stakeholders from the community and the healthcare system in the development of research interventions. The aim of this project is to stimulate changes in healthcare organizations and practices by encouraging collaboration between care teams and people living in poverty. Our objectives are twofold: 1) to identify actions required to promote the adoption of professional practices oriented toward social competence in primary care teams; and 2) to examine factors that would encourage the inclusion of people living in poverty in the process of developing social competence in healthcare organizations. Methods/design This study will use a participatory action research design applied in healthcare organizations. Participatory research is an increasingly recognized approach that is helpful for involving the people for whom the research results are intended. Our research team consists of 19 non-academic researchers, 11 academic researchers and six partners. A steering committee composed of academic researchers and stakeholders will have a decision-making role at each step, including knowledge dissemination and recommendations for new interventions. In this project we will adopt a multiphase approach and will use a variety of methods, including photovoice, group discussions and interviews. Discussion The proposed study will be one of only a few using participatory research in primary care to foster changes aimed at enhancing quality and access to care for people living in poverty. To our knowledge this will be the first study to use photovoice in healthcare organizations to promote new interventions. Our project includes partners who are targeted for practice changes and improvements in delivering

  9. Circulating follicular helper T cells presented distinctively different responses toward bacterial antigens in primary biliary cholangitis.

    PubMed

    Zhou, Zun-Qiang; Tong, Da-Nian; Guan, Jiao; Li, Mei-Fang; Feng, Qi-Ming; Zhou, Min-Jie; Zhang, Zheng-Yun

    2017-10-01

    Primary biliary cholangitis (PBC) is a chronic and progressive cholestatic liver disease with unknown causes. The initiation of PBC is associated with bacterial infections and abnormal immune correlates, such as the presence of self-reactive anti-mitochondrial antibodies and shifted balance of T cell subsets. In particular, the CD4(+)CXCR5(+) follicular helper T (Tfh) cells are highly activated in PBC patients and are significantly associated with PBC severity, but the underlying reasons are unknown. In this study, we found that the circulating CD4(+)CXCR5(+) T cells were enriched with the interferon (IFN)-γ-secreting Th1-subtype and the interleukin (IL)-17-secreting Th17-subtype, but not the IL-4-secreting Th2 subtype. We further demonstrated that a host of microbial motifs, including Pam3CSK4, poly(I:C), LPS, imiquimod, and CpG, could significantly stimulate IFN-γ, IL-17, and/or IL-21 from circulating CD4(+)CXCR5(+) T cells in PBC patients, especially in the presence of monocytes and B cells. Whole bacterial cells of Escherichia coli, Novosphingobium aromaticivorans, and Mycobacterium gordonae, could also potently stimulate IFN-γ, IL-17, and/or IL-21 production from circulating CD4(+)CXCR5(+) T cells. But interestingly, while the whole cell could potently stimulate circulating CD4(+)CXCR5(+) T cells from both healthy controls and PBC patients, the cell protein lysate could only potently stimulate circulating CD4(+)CXCR5(+) T cells from PBC patients, but not those from healthy controls, suggesting that circulating CD4(+)CXCR5(+) T cells in PBC patients had distinctive antigen-specificity from those in healthy individuals. Together, these data demonstrated that bacterial antigen stimulation is a potential source of aberrant Tfh cell activation in PBC patients. Copyright © 2017. Published by Elsevier B.V.

  10. Fabrication method for cores of structural sandwich materials including star shaped core cells

    DOEpatents

    Christensen, Richard M.

    1997-01-01

    A method for fabricating structural sandwich materials having a core pattern which utilizes star and non-star shaped cells. The sheets of material are bonded together or a single folded sheet is used, and bonded or welded at specific locations, into a flat configuration, and are then mechanically pulled or expanded normal to the plane of the sheets which expand to form the cells. This method can be utilized to fabricate other geometric cell arrangements than the star/non-star shaped cells. Four sheets of material (either a pair of bonded sheets or a single folded sheet) are bonded so as to define an area therebetween, which forms the star shaped cell when expanded.

  11. Fabrication method for cores of structural sandwich materials including star shaped core cells

    DOEpatents

    Christensen, R.M.

    1997-07-15

    A method for fabricating structural sandwich materials having a core pattern which utilizes star and non-star shaped cells is disclosed. The sheets of material are bonded together or a single folded sheet is used, and bonded or welded at specific locations, into a flat configuration, and are then mechanically pulled or expanded normal to the plane of the sheets which expand to form the cells. This method can be utilized to fabricate other geometric cell arrangements than the star/non-star shaped cells. Four sheets of material (either a pair of bonded sheets or a single folded sheet) are bonded so as to define an area therebetween, which forms the star shaped cell when expanded. 3 figs.

  12. Treatment of prostate cancer cell lines and primary cells using low temperature plasma

    NASA Astrophysics Data System (ADS)

    O'Connell, Deborah; Hirst, Adam; Frame, Fiona F.; Maitland, Norman J.

    2014-10-01

    The mechanisms of cell death after plasma treatment of both benign and cancerous prostate epithelial cells are investigated. Prostate cancer tissue was obtained with patient consent from targeted needle core biopsies following radical prostatectomy. Primary cells were cultured from cancer tissue and plated onto a chamber slide at a density of 10,000 cells per well in 200 microliter of stem cell media (SCM). The treated sample was previously identified as Gleason grade 7 cancer through tissue histo-pathology. A dielectric barrier discharge (DBD) jet configuration, with helium as a carrier gas, and 0.3% O2 admixture was used for treating the cells. Reactive oxygen and nitrogen species (RONS) produced by the plasma are believed to be the main mediators of the plasma-cell interaction and response. We found the concentration of reactive oxygen species (ROS) induced inside the cells increased with plasma exposure. Exposure to the plasma for >3 minutes showed high levels of DNA damage compared to untreated and hydrogen peroxide controls. Cell viability and cellular recovery are also investigated and will be presented. All findings were common to both cell lines, suggesting the potential of LTP therapy for both benign and malignant disease.

  13. Two Different Cell Populations Is an Important Clue for Diagnosis of Primary Cutaneous Adenoid Cystic Carcinoma: Immunohistochemical Study

    PubMed Central

    Alkan, Banu Ince; Karadeniz, Müjde; Bozdoğan, Nazan

    2017-01-01

    Primary cutaneous adenoid cystic carcinoma (PCACC) is a very rare malignancy. The differential diagnosis of PCACCs in pathology practice can be difficult and a group of primary and metastatic lesions, including adenoid basal cell carcinoma of the skin, should be considered in the differential diagnosis. Besides histomorphological clues, immunohistochemistry studies are very helpful in the differential diagnosis of PCACC. We report herein a case of PCACC with extensive immunohistochemical studies and review the literature from an immunohistochemistry perspective. PMID:28243477

  14. Melleolides induce rapid cell death in human primary monocytes and cancer cells.

    PubMed

    Bohnert, Markus; Scherer, Olga; Wiechmann, Katja; König, Stefanie; Dahse, Hans-Martin; Hoffmeister, Dirk; Werz, Oliver

    2014-08-01

    The melleolides are structurally unique and bioactive natural products of the basidiomycete genus Armillaria. Here, we report on cytotoxic effects of melleolides from Armillaria mellea towards non-transformed human primary monocytes and human cancer cell lines, respectively. In contrast to staurosporine or pretubulysin that are less cytotoxic for monocytes, the cytotoxic potency of the active melleolides in primary monocytes is comparable to that in cancer cells. The onset of the cytotoxic effects of melleolides was rapid (within <1 h), as compared to the apoptosis inducer staurosporine, the protein biosynthesis inhibitor cycloheximide, and the DNA transcription inhibitor actinomycin D (>5 h, each). Side-by-side comparison with the detergent triton X-100 and staurosporine in microscopic and flow cytometric analysis studies as well as analysis of the viability of mitochondria exclude cell lysis and apoptosis as relevant or primary mechanisms. Our results rather point to necrotic features of cell death mediated by an as yet elusive but rapid mechanism. Copyright © 2014 Elsevier Ltd. All rights reserved.

  15. Characterization of the Interaction between Human Respiratory Syncytial Virus and the Cell Cycle in Continuous Cell Culture and Primary Human Airway Epithelial Cells▿

    PubMed Central

    Wu, Weining; Munday, Diane C.; Howell, Gareth; Platt, Gareth; Barr, John N.; Hiscox, Julian A.

    2011-01-01

    Viruses can modify conditions inside cells to make them more favorable for replication and progeny virus production. One way of doing this is through manipulation of the cell cycle, a process that describes the ordered growth and division of cells. Analysis of model cell lines, such as A549 cells and primary airway epithelial cells, infected with human respiratory syncytial virus (HRSV) has shown alteration of the cell cycle during infection, although the signaling events were not clearly understood. In this study, targeted transcriptomic analysis of HRSV-infected primary airway epithelial cells revealed alterations in the abundances of many mRNAs encoding cell cycle-regulatory molecules, including decreases in the D-type cyclins and corresponding cyclin-dependent kinases (CDK4 and CDK6 [CDK4/6]). These alterations were reflected in changes in protein abundance and/or relocalization in HRSV-infected cells; taken together, they were predicted to result in G0/G1 phase arrest. In contrast, there was no change in the abundances of D-type cyclins in A549 cells infected with HRSV. However, the abundance of the G1/S phase progression inhibitor p21WAF1/CIP1 was increased over that in mock-treated cells, and this, again, was predicted to result in G0/G1 phase arrest. The G0/G1 phase arrest in both HRSV-infected primary cells and A549 cells was confirmed using dual-label flow cytometry that accurately measured the different stages of the cell cycle. Comparison of progeny virus production in primary and A549 cells enriched in G0/G1 using a specific CDK4/6 kinase inhibitor with asynchronously replicating cells indicated that this phase of the cell cycle was more efficient for virus production. PMID:21795354

  16. Imbalances of T cell subpopulations in primary immunodeficiencies and systemic lupus erythematosus.

    PubMed

    Pandolfi, F; Quinti, I; Sirianni, M C; Fiorilli, M; Zolla, S; Aiuti, F

    1981-01-01

    In the present report we describe a recently proposed technique for the enumeration of T lymphocyte subpopulations. The study was performed on peripheral blood lymphocytes (PBL) of 21 patients with primary immunodeficiencies and 10 with systemic lupus erythematosus (SLE). With this methodology, sheep rosettes are evaluated before and after preincubation with theophylline. Previous reports have shown that cells rosetting after the preincubation (T-res) contain most of the percentages of the helper activity, whereas sensitive cells (T-sens) exert suppression. T-res and T-sens cells were compared to several clinical and immunological parameters, including the detection of lymphocytes with Fc IgM and IgG receptors. We demonstrated a significant positive correlation between T-res and and EAox IgM cells and between T-sens and EAox IgG cells. In addition, patients with Ig defects demonstrated heterogeneity regarding alterations in the proportions of T-res and T-sens cells. On the contrary, in 8 patients with SLE, there was a markedly reduced proportion of T-sens cells. Variations in the balance of subpopulations were observed after treatment with thymic hormones and levamisole. We may conclude that theophylline rosettes represent an easy technique that can be profitably used in the evaluation of T cell subpopulations in immunological diseases.

  17. Organelle Transport in Cultured Drosophila Cells: S2 Cell Line and Primary Neurons.

    PubMed Central

    Gelfand, Vladimir I.

    2013-01-01

    Drosophila S2 cells plated on a coverslip in the presence of any actin-depolymerizing drug form long unbranched processes filled with uniformly polarized microtubules. Organelles move along these processes by microtubule motors. Easy maintenance, high sensitivity to RNAi-mediated protein knock-down and efficient procedure for creating stable cell lines make Drosophila S2 cells an ideal model system to study cargo transport by live imaging. The results obtained with S2 cells can be further applied to a more physiologically relevant system: axonal transport in primary neurons cultured from dissociated Drosophila embryos. Cultured neurons grow long neurites filled with bundled microtubules, very similar to S2 processes. Like in S2 cells, organelles in cultured neurons can be visualized by either organelle-specific fluorescent dyes or by using fluorescent organelle markers encoded by DNA injected into early embryos or expressed in transgenic flies. Therefore, organelle transport can be easily recorded in neurons cultured on glass coverslips using living imaging. Here we describe procedures for culturing and visualizing cargo transport in Drosophila S2 cells and primary neurons. We believe that these protocols make both systems accessible for labs studying cargo transport. PMID:24300413

  18. Identification of T cell-signaling pathways that stimulate latent HIV in primary cells

    PubMed Central

    Brooks, David G.; Arlen, Philip A.; Gao, Lianying; Kitchen, Christina M. R.; Zack, Jerome A.

    2003-01-01

    Eradication of HIV infection depends on the elimination of a small, but stable population of latently infected T cells. After the discontinuation of therapy, activation of latent virus can rekindle infection. To purge this reservoir, it is necessary to define cellular signaling pathways that lead to activation of latent HIV. We used the SCID-hu (Thy/Liv) mouse model of HIV latency to analyze a broad array of T cell-signaling pathways and show in primary, quiescent cells that viral induction depends on the activation of two primary intracellular signaling pathways, protein kinase C or nuclear factor of activated T cells (NF-AT). In contrast, inhibition or activation of other important T cell stimulatory pathways (such as mitogen-activated protein kinase, calcium flux, or histone deacetylation) do not significantly induce virus expression. We found that the activation of NF-κB is critical to viral reactivation; however, all pathways that stimulate NF-κBdonot reactivate latent virus. Our studies further show that inhibition of NF-κB does not prevent activation of HIV by NF-AT, indicating that these pathways can function independently to activate the HIV LTR. Thus, we define several molecular pathways that trigger HIV reactivation from latency and provide evidence that latent HIV infection is maintained by the functional lack of particular transcription factors in quiescent cells. PMID:14569007

  19. Organelle transport in cultured Drosophila cells: S2 cell line and primary neurons.

    PubMed

    Lu, Wen; Del Castillo, Urko; Gelfand, Vladimir I

    2013-11-20

    Drosophila S2 cells plated on a coverslip in the presence of any actin-depolymerizing drug form long unbranched processes filled with uniformly polarized microtubules. Organelles move along these processes by microtubule motors. Easy maintenance, high sensitivity to RNAi-mediated protein knock-down and efficient procedure for creating stable cell lines make Drosophila S2 cells an ideal model system to study cargo transport by live imaging. The results obtained with S2 cells can be further applied to a more physiologically relevant system: axonal transport in primary neurons cultured from dissociated Drosophila embryos. Cultured neurons grow long neurites filled with bundled microtubules, very similar to S2 processes. Like in S2 cells, organelles in cultured neurons can be visualized by either organelle-specific fluorescent dyes or by using fluorescent organelle markers encoded by DNA injected into early embryos or expressed in transgenic flies. Therefore, organelle transport can be easily recorded in neurons cultured on glass coverslips using living imaging. Here we describe procedures for culturing and visualizing cargo transport in Drosophila S2 cells and primary neurons. We believe that these protocols make both systems accessible for labs studying cargo transport.

  20. Methods of improving the efficiency of photovoltaic cells. [including X ray analysis

    NASA Technical Reports Server (NTRS)

    Loferski, J. J.; Roessler, B.; Crisman, E. E.; Chen, L. Y.; Kaul, R.

    1974-01-01

    Work on aluminum-alloyed silicon grating cells is continued. Optimization of the geometry (grating line width and spacing) confirms the analysis of such cells. A 1 sq cm grating cell was fabricated and its i-V characteristic was measured under an AMO solar simulator. It is found that the efficiency of this cell would be about 7.9%, if it were covered by the usual antireflection coating. The surface of the cell is not covered by a diffused junction. The response is blue shifted; the current is somewhat higher than that produced by a commercial Si cell. However, the open circuit voltage is low, and attempts to optimize the open circuit voltage of the aluminum-alloy junctions are described. A preliminary X-ray topographic examination of GaAs specimens of the type commonly used to make solar cells is studied. The X-ray study shows that the wafers are filled with regions having strain gradients, possibly caused by precipitates. It is possible that a correlation exists between the presence of low mechanical perfection and minority carrier diffusion lengths of GaAs crystals.

  1. GFR prediction from cystatin C and creatinine in children: effect of including body cell mass.

    PubMed

    Andersen, Trine Borup; Jødal, Lars; Boegsted, Martin; Erlandsen, Erland J; Morsing, Anni; Frøkiær, Jørgen; Brøchner-Mortensen, Jens

    2012-01-01

    Aiming to develop a more accurate cystatin C-based model for estimation of glomerular filtration rate (GFR) in children, we hypothesized that inclusion of body cell mass (BCM) would increase the accuracy of the GFR estimate in comparison to a well-established GFR reference method. Diagnostic test accuracy study. 119 children (mean age, 8.8; range, 2.3-14.9 years) referred for GFR measurement by chromium 51 ethylenediaminetetraacetic acid ((51)Cr-EDTA) clearance (mean GFR, 98; range, 13.7-147.4 mL/min/1.73 m(2)). GFR estimations by the 2 prediction models resulting from theoretical considerations corroborated by forward stepwise variable selection: GFR (mL/min) = 0.542 × (BCM/SCysC)(0.40) × (height × BSA/SCr)(0.65) and GFR (mL/min) = 0.426 × (weight/SCysC)(0.39) × (height × BSA/SCr)(0.64), where SCysC is serum cystatin C level, BSA is body surface area, and SCr is serum creatinine level. The accuracy and precision of these models were compared with 7 previously published prediction models using random subsampling cross-validation. Local constants and coefficients were calculated for all models. Root mean square error, R(2), and percentage of predictions within ±10% and ±30% of the reference GFR were calculated for all models. Based on 1,000 runs of the cross-validation procedure, median values and 2.5th and 97.5th quantiles of the validation parameters were calculated. GFR measurement by (51)Cr-EDTA clearance. The BCM model predicted 98% within ±30% of reference GFR and 66% within ±10%, which was higher than for any other model. The weight model predicted 97.5% within ±30% of reference GFR and 62% within ±10%. The BCM model had the highest R(2) and the smallest root mean square error. Included only 9 children with GFR <60 mL/min/1.73 m(2). Lack of independent validation cohort. The novel BCM model predicts GFR with higher accuracy than previously published models. The weight model is almost as accurate as the BCM model and allows for GFR estimation

  2. Molecular characterization of chordoma xenografts generated from a novel primary chordoma cell source and two chordoma cell lines.

    PubMed

    Karikari, Isaac O; Gilchrist, Christopher L; Jing, Liufang; Alcorta, David A; Chen, Jun; Richardson, William J; Gabr, Mostafa A; Bell, Richard D; Kelley, Michael J; Bagley, Carlos A; Setton, Lori A

    2014-09-01

    Chordoma cells can generate solid-like tumors in xenograft models that express some molecular characteristics of the parent tumor, including positivity for brachyury and cytokeratins. However, there is a dearth of molecular markers that relate to chordoma tumor growth, as well as the cell lines needed to advance treatment. The objective in this study was to isolate a novel primary chordoma cell source and analyze the characteristics of tumor growth in a mouse xenograft model for comparison with the established U-CH1 and U-CH2b cell lines. Primary cells from a sacral chordoma, called "DVC-4," were cultured alongside U-CH1 and U-CH2b cells for more than 20 passages and characterized for expression of CD24 and brachyury. While brachyury is believed essential for driving tumor formation, CD24 is associated with healthy nucleus pulposus cells. Each cell type was subcutaneously implanted in NOD/SCID/IL2Rγ(null) mice. The percentage of solid tumors formed, time to maximum tumor size, and immunostaining scores for CD24 and brachyury (intensity scores of 0-3, heterogeneity scores of 0-1) were reported and evaluated to test differences across groups. The DVC-4 cells retained chordoma-like morphology in culture and exhibited CD24 and brachyury expression profiles in vitro that were similar to those for U-CH1 and U-CH2b. Both U-CH1 and DVC-4 cells grew tumors at rates that were faster than those for U-CH2b cells. Gross tumor developed at nearly every site (95%) injected with U-CH1 and at most sites (75%) injected with DVC-4. In contrast, U-CH2b cells produced grossly visible tumors in less than 50% of injected sites. Brachyury staining was similar among tumors derived from all 3 cell types and was intensely positive (scores of 2-3) in a majority of tissue sections. In contrast, differences in the pattern and intensity of staining for CD24 were noted among the 3 types of cell-derived tumors (p < 0.05, chi-square test), with evidence of intense and uniform staining in a

  3. Rapid Selection of Mesenchymal Stem and Progenitor Cells in Primary Prostate Stromal Cultures

    PubMed Central

    Brennen, W. Nathaniel; Kisteman, L. Nelleke; Isaacs, John T.

    2016-01-01

    BACKGROUND Carcinoma-associated fibroblasts (CAFs) are a dominant component of the tumor microenvironment with pro-tumorigenic properties. Despite this knowledge, their physiologic origins remain poorly understood. Mesenchymal stem cells (MSCs) can be recruited from the bone marrow to areas of tissue damage and inflammation, including prostate cancer. MSCs can generate and have many overlapping properties with CAFs in preclinical models. METHODS Multiparameter flow cytometry and multipotent differentiation assays used to define MSCs in primary prostate stromal cultures derived from young (>25 yrs) organ donors and prostate cancer patients compared with bone marrow-derived stromal cultures. Population doubling times, population doublings, cell size, and differentiation potential determined under multiple culture conditions, including normoxia, hypoxia, and a variety of media. TGF-β measured by ELISA. RESULTS MSCs and stromal progenitors are not only present in normal and malignant prostate tissue, but are quickly selected for in primary stromal cultures derived from these tissues; becoming the dominant population within just a few passages. Growth potential inversely associated with TGF-β concentrations. All conditions generated populations with an average cell diameter >15 μm. All cultures tested had the ability to undergo osteogenic and chondrogenic differentiation, but unlike bone marrow-derived MSCs, primary stromal cultures derived from normal prostate tissue lack adipogenic differentiation potential. In contrast, a subset of stromal cultures derived from prostate cancer patients retain the ability to differentiate into adipocytes; a property that is significantly suppressed under hypoxic conditions in both bone marrow- and prostate-derived MSCs. CONCLUSIONS Primary prostate stromal cultures are highly enriched in cells with an MSC or stromal progenitor phenotype. The use of primary cultures such as these to study CAFs raises interesting implications when

  4. Remote System Technologies for Deactivating Hanford Hot Cells (for WM'03 - abstract included)

    SciTech Connect

    BERLIN, G.T.

    2003-01-28

    Remote system technologies are being deployed by Fluor Hanford to help accelerate the deactivation of highly-radioactive hot cell facilities. This paper highlights the application of several remotely deployed technologies enabling the deactivation tasks.

  5. PAX-8 expression in primary and metastatic Merkel cell carcinoma: an immunohistochemical analysis.

    PubMed

    Sangoi, Ankur R; Cassarino, David S

    2013-06-01

    PAX-8, a nephric cell lineage transcription factor initially characterized in renal cell carcinomas, is also well recognized as a marker of Müllerian tract and thyroid tumors. From a previous tissue microarray study of nonrenal neoplasms including a variety of skin tumors, we identified PAX-8 staining in a small set of Merkel cell carcinomas, a finding not previously described. Herein, we explore PAX-8 immunoreactivity in 34 whole-section Merkel cell carcinomas (24 primary, 10 metastatic) using polyclonal PAX-8 (prediluted) and 2 varieties of monoclonal PAX-8 (prediluted clone MRQ-50 and 1:100 dilution clone BC12). Nuclear staining intensity and extent was semiquantitatively analyzed with a comparison of staining thresholds required for a "positive" result (≥ 2+ vs. 1+). Thirty-three of 34 (97%) cases showed positive Cell Marque polyclonal PAX-8 staining, whereas 31 of 34 (91%) cases showed positive Cell Marque monoclonal PAX-8 staining. There was no significant difference in staining between primary versus metastatic tumors. The Cell Marque polyclonal PAX-8 was superior to their monoclonal PAX-8, maintaining strong sensitivity using a ≥ 2+ versus 1+ staining cut point for positive results (79% vs. 18%, respectively), which may be important in cases with scant tissue or background staining. The Biocare monoclonal PAX-8 was negative in all cases. PAX-8 staining in Merkel cell carcinoma expands the spectrum of tumors showing immunoreactivity and may prove to be a useful addition to a diagnostic panel. Awareness of this immunoreactivity and recognition of the antibody source and clone are important to preclude diagnostic pitfalls with tumors in the differential diagnosis.

  6. Changes in Zn homeostasis during long term culture of primary endothelial cells and effects of Zn on endothelial cell senescence.

    PubMed

    Malavolta, Marco; Costarelli, Laura; Giacconi, Robertina; Basso, Andrea; Piacenza, Francesco; Pierpaoli, Elisa; Provinciali, Mauro; Ogo, Ogo A; Ford, Dianne

    2017-09-14

    Endothelial cell senescence and Zn nutritional status influence cardiovascular disease. The influence of Zn appears dichotomous, hence it is imperative to understand the relationship with cellular senescence to improve knowledge about the molecular and cellular basis of the disease. Here we aimed to determine: 1) the impact of chronic exposure to a moderately high dose of Zn on senescence of endothelial cells; 2) the changes in Zn homeostasis during the lifespan of primary cultured endothelial cells; and 3) the susceptibility of proliferating and senescent endothelial cells to cell death after short term exposure to increasing doses of Zn and of the Zn chelator TPEN. Chronic exposure to Zn accelerated senescence and untreated cells at later passages, where doubling time had increased, displayed relocation of labile Zn and altered expression of genes involved in the response to Zn toxicity, including SLC30A1, SLC39A6, SLC30A5, SLC30A10 and metallothioneins, indicating that senescent cells have altered zinc homeostasis. Most Zn-dependent genes that were expressed differently between early and late passages were correlated with changes in the expression of anti-apoptotic genes. Short-term treatment with a high dose of Zn leads to cell death, but only in the population of cells at both earlier and later passages that had already entered senescence. In contrast, Zn depletion led to death of cells at earlier but not later passages, which suggests that there are sub-populations of senescent cells that are resistant to Zn depletion. This resistant senescent cell population may accumulate under conditions of Zn deficiency and contribute to vascular pathology. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. Comparative In Vitro Immune Stimulation Analysis of Primary Human B Cells and B Cell Lines

    PubMed Central

    Van Belle, Kristien; Herman, Jean; Boon, Louis; Waer, Mark

    2016-01-01

    B cell specific immunomodulatory drugs still remain an unmet medical need. Utilisation of validated simplified in vitro models would allow readily obtaining new insights in the complexity of B cell regulation. For this purpose we investigated which human B lymphocyte stimulation assays may be ideally suited to investigate new B lymphocyte immunosuppressants. Primary polyclonal human B cells underwent in vitro stimulation and their proliferation, production of immunoglobulins (Igs) and of cytokines, and expression of cell surface molecules were analysed using various stimuli. ODN2006, a toll-like receptor 9 (TLR9) agonist, was the most potent general B cell stimulus. Subsequently, we investigated on which human B cell lines ODN2006 evoked the broadest immunostimulatory effects. The Namalwa cell line proved to be the most responsive upon TLR9 stimulation and hence may serve as a relevant, homogeneous, and stable B cell model in an in vitro phenotypic assay for the discovery of new targets and inhibitors of the B cell activation processes. As for the read-out for such screening assay, it is proposed that the expression of activation and costimulatory surface markers reliably reflects B lymphocyte activation. PMID:28116319

  8. Ectopic lignification in primary cellulose-deficient cell walls of maize cell suspension cultures.

    PubMed

    Mélida, Hugo; Largo-Gosens, Asier; Novo-Uzal, Esther; Santiago, Rogelio; Pomar, Federico; García, Pedro; García-Angulo, Penélope; Acebes, José Luis; Álvarez, Jesús; Encina, Antonio

    2015-04-01

    Maize (Zea mays L.) suspension-cultured cells with up to 70% less cellulose were obtained by stepwise habituation to dichlobenil (DCB), a cellulose biosynthesis inhibitor. Cellulose deficiency was accompanied by marked changes in cell wall matrix polysaccharides and phenolics as revealed by Fourier transform infrared (FTIR) spectroscopy. Cell wall compositional analysis indicated that the cellulose-deficient cell walls showed an enhancement of highly branched and cross-linked arabinoxylans, as well as an increased content in ferulic acid, diferulates and p-coumaric acid, and the presence of a polymer that stained positive for phloroglucinol. In accordance with this, cellulose-deficient cell walls showed a fivefold increase in Klason-type lignin. Thioacidolysis/GC-MS analysis of cellulose-deficient cell walls indicated the presence of a lignin-like polymer with a Syringyl/Guaiacyl ratio of 1.45, which differed from the sensu stricto stress-related lignin that arose in response to short-term DCB-treatments. Gene expression analysis of these cells indicated an overexpression of genes specific for the biosynthesis of monolignol units of lignin. A study of stress signaling pathways revealed an overexpression of some of the jasmonate signaling pathway genes, which might trigger ectopic lignification in response to cell wall integrity disruptions. In summary, the structural plasticity of primary cell walls is proven, since a lignification process is possible in response to cellulose impoverishment. © 2015 Institute of Botany, Chinese Academy of Sciences.

  9. Modeling the biological response of normal human cells, including repair processes, to fractionated carbon beam irradiation

    PubMed Central

    Wada, Mami; Suzuki, Masao; Liu, Cuihua; Kaneko, Yumiko; Fukuda, Shigekazu; Ando, Koichi; Matsufuji, Naruhiro

    2013-01-01

    To understand the biological response of normal cells to fractionated carbon beam irradiation, the effects of potentially lethal damage repair (PLDR) and sublethal damage repair (SLDR) were both taken into account in a linear-quadratic (LQ) model. The model was verified by the results of a fractionated cell survival experiment with normal human fibroblast cells. Cells were irradiated with 200-kV X-rays and monoenergetic carbon ion beams (290 MeV/u) at two irradiation depths, corresponding to linear energy transfers (LETs) of approximately 13 keV/μm and 75 keV/μm, respectively, at the Heavy Ion Medical Accelerator in Chiba of the National Institute of Radiological Sciences. When we only took into account the repair factor of PLDR, γ, which was derived from the delayed assay, the cell survival response to fractionated carbon ion irradiation was not fully explained in some cases. When both the effects of SLDR and PLDR were taken into account in the LQ model, the cell survival response was well reproduced. The model analysis suggested that PLDR occurs in any type of radiation. The γ factors ranged from 0.36–0.93. In addition, SLD was perfectly repaired during the fraction interval for the lower LET irradiations but remained at about 30% for the high-LET irradiation. PMID:23449640

  10. Renal cell carcinoma in kidney allografts: histologic types, including biphasic papillary carcinoma.

    PubMed

    Troxell, Megan L; Higgins, John P

    2016-11-01

    Kidney transplant recipients are at increased risk for malignancy, with about 5% incidence of cancer in native end-stage kidneys. Carcinoma in the renal allograft is far less common. Prior studies have demonstrated a propensity for renal cell carcinomas (RCCs) of papillary subtypes in end-stage kidneys, and perhaps in allograft kidneys, but most allograft studies lack detailed pathologic review and predate the current classification system. We reviewed our experience with renal carcinoma in kidney allografts at 2 academic centers applying the International Society of Urological Pathology classification, informed by immunohistochemistry. The incidence of renal allograft carcinoma was about 0.26% in our population. Of 12 allograft carcinomas, 6 were papillary (50%), 4 were clear cell (33%), 1 was clear cell (tubulo)papillary, and 1 chromophobe. Two of the papillary carcinomas had distinctive biphasic glomeruloid architecture matching the newly named "biphasic squamoid alveolar" pattern and were difficult to classify on core biopsies. The 2 cell types had different immunophenotypes in our hands (eosinophilic cells: RCC-/CK34betaE12+ weight keratin +/cyclin D1+; clear cells: RCC+/cytokeratin high molecular weight negative to weak/cyclin D1-). None of the patients experienced cancer recurrences or metastasis. Our study confirms the predilection for papillary RCCs in kidney allografts and highlights the occurrence of rare morphologic variants. Larger studies are needed with careful pathologic review, which has been lacking in the literature.

  11. Japan Society of Gynecologic Oncology guidelines 2015 for the treatment of ovarian cancer including primary peritoneal cancer and fallopian tube cancer.

    PubMed

    Komiyama, Shinichi; Katabuchi, Hidetaka; Mikami, Mikio; Nagase, Satoru; Okamoto, Aikou; Ito, Kiyoshi; Morishige, Kenichiro; Suzuki, Nao; Kaneuchi, Masanori; Yaegashi, Nobuo; Udagawa, Yasuhiro; Yoshikawa, Hiroyuki

    2016-06-01

    The fourth edition of the Japan Society of Gynecologic Oncology guidelines for the treatment of ovarian cancer including primary peritoneal cancer and fallopian tube cancer was published in 2015. The guidelines contain seven chapters and six flow charts. The major changes in this new edition are as follows-(1) the format has been changed from reviews to clinical questions (CQ), and the guidelines for optimal clinical practice in Japan are now shown as 41 CQs and answers; (2) the 'flow charts' have been improved and placed near the beginning of the guidelines; (3) the 'basic points', including tumor staging, histological classification, surgical procedures, chemotherapy, and palliative care, are described before the chapter; (4) the FIGO surgical staging of ovarian cancer, fallopian tube cancer, and primary peritoneal cancer was revised in 2014 and the guideline has been revised accordingly to take the updated version of this classification into account; (5) the procedures for examination and management of hereditary breast and ovarian cancer are described; (6) information on molecular targeting therapy has been added; (7) guidelines for the treatment of recurrent cancer based on tumor markers alone are described, as well as guidelines for providing hormone replacement therapy after treatment.

  12. Phenotypical and functional alterations of CD8 regulatory T cells in primary biliary cirrhosis

    PubMed Central

    Bernuzzi, Francesca; Fenoglio, Daniela; Battaglia, Florinda; Fravega, Marco; Gershwin, M. Eric; Indiveri, Francesco; Ansari, Aftab A.; Podda, Mauro; Invernizzi, Pietro; Filaci, Gilberto

    2011-01-01

    The mechanisms that lead to loss of tolerance in autoimmune disease have remained both elusive and diverse, including both genetic predisposition and generic dysregulation of critical mononuclear cell subsets. In primary biliary cirrhosis (PBC), patients exhibit a multilineage response to the E2 component of pyruvate dehydrogenase involving antibody as well as autoreactive CD4 and CD8 responses. Recent data from murine models of PBC have suggested that a critical mechanism of biliary destruction is mediated by liver-infiltrating CD8 cells. Further, the number of autoreactive liver-infiltrating CD4 and CD8 cells is significantly higher in liver than blood in patients with PBC. Based on this data, we have studied the frequencies and phenotypic characterization of both CD4 and CD8 regulatory T cell components in both patients with PBC and age–sex matched controls. Our data is striking and indicate that CD8 Treg populations from PBC patients, but not controls, have significant phenotypic alterations, including increased expression of CD127 and reduced CD39. Furthermore, in vitro induction of CD8 Tregs by incubation with IL10 is significantly reduced in PBC patients. Importantly, the frequencies of circulating CD4+CD25+ and CD8+ and CD28− T cell subpopulations are not significantly different between patients and controls. In conclusion, these data identify the CD8 Treg subset as a regulatory T cell subpopulation altered in patients with PBC. PMID:20638239

  13. Brain tumor - primary - adults

    MedlinePlus

    ... Vestibular schwannoma (acoustic neuroma) - adults; Meningioma - adults; Cancer - brain tumor (adults) ... Primary brain tumors include any tumor that starts in the brain. Primary brain tumors can start from brain cells, ...

  14. T cells bearing anti-CD19 and/or anti-CD38 chimeric antigen receptors effectively abrogate primary double-hit lymphoma cells.

    PubMed

    Mihara, Keichiro; Yoshida, Tetsumi; Takei, Yoshifumi; Sasaki, Naomi; Takihara, Yoshihiro; Kuroda, Junya; Ichinohe, Tatsuo

    2017-06-08

    Patients with B cell lymphomas bearing MYC translocation combined with translocation involving other genes, such as BCL2, BCL3, or BCL6, defined as double-hit lymphoma (DHL), have a poor prognosis. Recent studies expanded the concept to include double-expressing lymphoma (DEL) that co-overexpresses MYC protein with either of those proteins. Accordingly, we defined cytogenetic DHL and DEL as primary DHL. An adoptive T cell immunotherapy with a chimeric antigen receptor (CAR) has been clinically shown to exhibit cytotoxicity in refractory neoplasias. We revealed the marked cytotoxicity of anti-CD19- and/or anti-CD38-CAR T cells against primary DHL cells from patients. CD19- and/or CD38-specific T cells were co-cultured with cytogenetic DHL (n = 3) or DEL (n = 2) cells from five patients for 3 days. We examined whether T cells retrovirally transduced with each vector showed cytotoxicity against DHL cells. Anti-CD19- and/or anti-CD38-CAR T cells were co-cultured with primary DHL cells at an E:T ratio of 1:2 for 3 days. Anti-CD19- and anti-CD38-CAR T cells completely abrogated these DHL cells, respectively. Anti-CD19-CAR T cells synergistically exerted collaborative cytotoxicity against these primary DHL cells with anti-CD38-CAR T cells. Therefore, refractory DHL cells can be efficiently abrogated by the clinical use of T cells with anti-CD19- and/or anti-CD38-CAR.

  15. Phenotypic and Functional Alterations in Circulating Memory CD8 T Cells with Time after Primary Infection

    PubMed Central

    Martin, Matthew D.; Kim, Marie T.; Shan, Qiang; Sompallae, Ramakrishna; Xue, Hai-Hui; Harty, John T.; Badovinac, Vladimir P.

    2015-01-01

    Memory CD8 T cells confer increased protection to immune hosts upon secondary viral, bacterial, and parasitic infections. The level of protection provided depends on the numbers, quality (functional ability), and location of memory CD8 T cells present at the time of infection. While primary memory CD8 T cells can be maintained for the life of the host, the full extent of phenotypic and functional changes that occur over time after initial antigen encounter remains poorly characterized. Here we show that critical properties of circulating primary memory CD8 T cells, including location, phenotype, cytokine production, maintenance, secondary proliferation, secondary memory generation potential, and mitochondrial function change with time after infection. Interestingly, phenotypic and functional alterations in the memory population are not due solely to shifts in the ratio of effector (CD62Llo) and central memory (CD62Lhi) cells, but also occur within defined CD62Lhi memory CD8 T cell subsets. CD62Lhi memory cells retain the ability to efficiently produce cytokines with time after infection. However, while it is was not formally tested whether changes in CD62Lhi memory CD8 T cells over time occur in a cell intrinsic manner or are due to selective death and/or survival, the gene expression profiles of CD62Lhi memory CD8 T cells change, phenotypic heterogeneity decreases, and mitochondrial function and proliferative capacity in either a lymphopenic environment or in response to antigen re-encounter increase with time. Importantly, and in accordance with their enhanced proliferative and metabolic capabilities, protection provided against chronic LCMV clone-13 infection increases over time for both circulating memory CD8 T cell populations and for CD62Lhi memory cells. Taken together, the data in this study reveal that memory CD8 T cells continue to change with time after infection and suggest that the outcome of vaccination strategies designed to elicit protective memory

  16. Primary reference cell calibrations at SERI - History and methods. [solar cells

    NASA Technical Reports Server (NTRS)

    Osterwald, C. R.; Emery, K. A.; Myers, D. R.; Hart, R. E.

    1990-01-01

    The tabular calibration method used at the Solar Energy Research Institute (SERI) for primary reference cells is derived and described in detail. An uncertainty analysis shows that the tabular method should have a total uncertainty of + or - 1.0 percent; data from five years of calibrations are then shown to support this analysis. Results from SERI's tabular method are compared with terrestrial calibrations performed by the NASA Lewis Research Center in the late 1970s.

  17. Isolation and Proteomic Analysis of Microvesicles and Exosomes from HT22 Cells and Primary Neurons.

    PubMed

    Witas, Richard; Chaput, Dale; Khan, Hirah; Stevens, Stanley M; Kang, David

    2017-01-01

    Exosomes and microvesicles are extracellular vesicles (EVs) released by most cell types. The role of EVs as a method of intercellular communication has led to these vesicles becoming a major area of interest in a variety of scientific fields including neuroscience. Emerging evidence is now demonstrating that the biomolecular composition of EVs, especially exosomes, can play a role in the progression of disease including various neurodegenerative diseases and cancer. In addition to the miRNA profiles of EVs, these vesicles also show interesting changes in protein expression profiles under different physiological and pathological conditions. Characterization of these profiles could prove valuable for both understanding disease pathogenesis and for the discovery of new biomarkers of disease. In this chapter, we describe a protocol for isolation of exosomes and microvesicles from immortalized HT22 cells and primary cortical neurons with sufficient yield and low serum contamination required for downstream analysis and label-free relative quantitation by mass spectrometry.

  18. Specification of Region-Specific Neurons Including Forebrain Glutamatergic Neurons from Human Induced Pluripotent Stem Cells

    PubMed Central

    Martins-Taylor, Kristen; Wang, Xiaofang; Zhang, Zheng; Park, Jung Woo; Zhan, Shuning; Kronenberg, Mark S.; Lichtler, Alexander; Liu, Hui-Xia; Chen, Fang-Ping; Yue, Lixia; Li, Xue-Jun; Xu, Ren-He

    2010-01-01

    Background Directed differentiation of human induced pluripotent stem cells (hiPSC) into functional, region-specific neural cells is a key step to realizing their therapeutic promise to treat various neural disorders, which awaits detailed elucidation. Methodology/Principal Findings We analyzed neural differentiation from various hiPSC lines generated by others and ourselves. Although heterogeneity in efficiency of neuroepithelial (NE) cell differentiation was observed among different hiPSC lines, the NE differentiation process resembles that from human embryonic stem cells (hESC) in morphology, timing, transcriptional profile, and requirement for FGF signaling. NE cells differentiated from hiPSC, like those from hESC, can also form rostral phenotypes by default, and form the midbrain or spinal progenitors upon caudalization by morphogens. The rostrocaudal neural progenitors can further mature to develop forebrain glutamatergic projection neurons, midbrain dopaminergic neurons, and spinal motor neurons, respectively. Typical ion channels and action potentials were recorded in the hiPSC-derived neurons. Conclusions/Significance Our results demonstrate that hiPSC, regardless of how they were derived, can differentiate into a spectrum of rostrocaudal neurons with functionality, which supports the considerable value of hiPSC for study and treatment of patient-specific neural disorders. PMID:20686615

  19. Blastoid Variant Mantle Cell Lymphoma with Complex Karyotype Including 11q Duplication

    PubMed Central

    Özer, Özge; Toprak, Selami K.; Öte, Enver; Yılmaz, Zerrin; Şahin, Feride İffet

    2014-01-01

    We describe a case of blastoid mantle cell lymphoma with a complex karyotype. The blastoid variant is a rare type of non-Hodgkin lymphoma exhibiting an aggressive clinical course. Mantle cell lymphoma is a distinct entity of mature B-cell neoplasms genetically characterized by the presence of t(11;14). In the present case, conventional analysis revealed structural abnormalities of chromosomes 2, 4, 6, 10, 13, and 19, along with 3 additional marker chromosomes. The derivative 1 chromosome determined in the case was a result of t(1p;11q). Our interesting finding was the presence of a different translocation between 11q and chromosome 1 in addition to t(11;14). Thus, the resulting 11q duplication was believed to additionally increase the enhanced expression of cyclin D1 gene, which is responsible in the pathogenesis of the disease. Fluorescence in situ hybridization method by the t(11;14) probe revealed clonal numerical abnormalities of chromosomes 11 and 14 in some cells. The detection of multiple abnormalities explains the bad prognosis in the present case. On the basis of our findings, we can easily conclude that results of cytogenetic analyses of similar mantle cell lymphoma patients would provide clues about new responsible gene regions and disease prognosis. In conclusion, it has been suggested that the presence of multiple chromosomal aberrations in addition to the specific t(11;14) may have a negative impact on clinical course and survival rate. PMID:25330523

  20. Blastoid variant mantle cell lymphoma with complex karyotype including 11q duplication.

    PubMed

    Ozer, Ozge; Toprak, Selami Koçak; Ote, Enver; Yılmaz, Zerrin; Sahin, Feride Iffet

    2014-09-05

    We describe a case of blastoid mantle cell lymphoma with a complex karyotype. The blastoid variant is a rare type of non-Hodgkin lymphoma exhibiting an aggressive clinical course. Mantle cell lymphoma is a distinct entity of mature B-cell neoplasms genetically characterized by the presence of t(11;14). In the present case, conventional analysis revealed structural abnormalities of chromosomes 2, 4, 6, 10, 13, and 19, along with 3 additional marker chromosomes. The derivative 1 chromosome determined in the case was a result of t(1p;11q). Our interesting finding was the presence of a different translocation between 11q and chromosome 1 in addition to t(11;14). Thus, the resulting 11q duplication was believed to additionally increase the enhanced expression of cyclin D1 gene, which is responsible in the pathogenesis of the disease. Fluorescence in situ hybridization method by the t(11;14) probe revealed clonal numerical abnormalities of chromosomes 11 and 14 in some cells. The detection of multiple abnormalities explains the bad prognosis in the present case. On the basis of our findings, we can easily conclude that results of cytogenetic analyses of similar mantle cell lymphoma patients would provide clues about new responsible gene regions and disease prognosis. In conclusion, it has been suggested that the presence of multiple chromosomal aberrations in addition to the specific t(11;14) may have a negative impact on clinical course and survival rate.

  1. IMMORTALIZATION OF HUMAN AND RHESUS MACAQUE PRIMARY ANTIGEN-SPECIFIC T CELLS BY RETROVIRALLY TRANSDUCED TELOMERASE REVERSE TRANSCRIPTASE

    PubMed Central

    Barsov, Eugene V.

    2011-01-01

    Human and rhesus macaque primary antigen-specific T cells derived from infected or immunized individuals or animals are a valuable material with which to study cellular immune responses against pathogens and tumors. Antigen-specific T cells can be expanded in vitro but have a finite proliferative life span. After a limited period in culture, primary T cells undergo replicative senescence and stop dividing. This restricts their applicability to short term experiments and complicates their use in adoptive immunotherapy. The proliferative life span of primary human and rhesus macaque T cells can be considerably extended by ectopically expressed human telomerase reverse transcriptase (TERT). Antigen-specific T cells transduced with TERT-expressing retroviral vectors can proliferate and expand in culture for long periods of time while maintaining their primary T cell characteristics including antigen-specific responses. Thus, TERT-immortalized T cells are an important and valuable resource for studying T cell immune responses and, potentially, for adoptive immunotherapy. PMID:22048804

  2. Cell survival curve for primary hepatic carcinoma cells and relationship between SF2 of hepatic carcinoma cells and radiosensitivity

    PubMed Central

    Liu, Zhi-Zhong; Huang, Wen-Ying; Lin, Ju-Sheng; Li, Xiao-Sheng; Lan, Xiao; Cai, Xiao-Kun; Liang, Kuo-Huan; Zhou, Hai-Jun

    2005-01-01

    AIM: To establish the cell survival curve for primary hepatic carcinoma cells and to study the relationship between SF2 of primary hepatic carcinoma cells and radiosensitivity. METHODS: Hepatic carcinoma cells were cultured in vitro using 39 samples of hepatic carcinoma at stages II-IV. Twenty-nine samples were cultured successfully in the fifth generation cells. After these cells were radiated with different dosages, the cell survival ratio and SF2 were calculated by clonogenic assay and SF2 model respectively. The relationship between SF2 and the clinical pathological feature was analyzed. RESULTS: Twenty-nine of thirty-nine samples were successfully cultured. After X-ray radiation of the fifth generation cells with 0, 2, 4, 6, 8 Gy, the cell survival rate was 41%, 36.5%, 31.0%, 26.8%, and 19%, respectively. There was a negative correlation between cell survival and irradiation dosage (r = -0.973, P<0.05). SF2 ranged 0.28-0.78 and correlated with the clinical stage and pathological grade of hepatic carcinoma (P<0.05). There was a positive correlation between SF2 and D0.5 (r = 0.773, P<0.05). CONCLUSION: SF2 correlates with the clinical stage and pathological grade of hepatic carcinoma and is a marker for predicting the radiosensitivity of hepatic carcinomas. PMID:16437614

  3. Establishment, characterization, and toxicological application of loggerhead sea turtle (Caretta caretta) primary skin fibroblast cell cultures.

    PubMed

    Webb, Sarah J; Zychowski, Gregory V; Bauman, Sandy W; Higgins, Benjamin M; Raudsepp, Terje; Gollahon, Lauren S; Wooten, Kimberly J; Cole, Jennifer M; Godard-Codding, Céline

    2014-12-16

    Pollution is a well-known threat to sea turtles but its impact is poorly understood. In vitro toxicity testing presents a promising avenue to assess and monitor the effects of environmental pollutants in these animals within the legal constraints of their endangered status. Reptilian cell cultures are rare and, in sea turtles, largely derived from animals affected by tumors. Here we describe the full characterization of primary skin fibroblast cell cultures derived from biopsies of multiple healthy loggerhead sea turtles (Caretta caretta), and the subsequent optimization of traditional in vitro toxicity assays to reptilian cells. Characterization included validating fibroblast cells by morphology and immunocytochemistry, and optimizing culture conditions by use of growth curve assays with a fractional factorial experimental design. Two cell viability assays, MTT and lactate dehydrogenase (LDH), and an assay measuring cytochrome P4501A (CYP1A) expression by quantitative PCR were optimized in the characterized cells. MTT and LDH assays confirmed cytotoxicity of perfluorooctanoic acid at 500 μM following 72 and 96 h exposures while CYP1A5 induction was detected after 72 h exposure to 0.1-10 μM benzo[a]pyrene. This research demonstrates the validity of in vitro toxicity testing in sea turtles and highlights the need to optimize mammalian assays to reptilian cells.

  4. Standardized 3D Bioprinting of Soft Tissue Models with Human Primary Cells.

    PubMed

    Rimann, Markus; Bono, Epifania; Annaheim, Helene; Bleisch, Matthias; Graf-Hausner, Ursula

    2016-08-01

    Cells grown in 3D are more physiologically relevant than cells cultured in 2D. To use 3D models in substance testing and regenerative medicine, reproducibility and standardization are important. Bioprinting offers not only automated standardizable processes but also the production of complex tissue-like structures in an additive manner. We developed an all-in-one bioprinting solution to produce soft tissue models. The holistic approach included (1) a bioprinter in a sterile environment, (2) a light-induced bioink polymerization unit, (3) a user-friendly software, (4) the capability to print in standard labware for high-throughput screening, (5) cell-compatible inkjet-based printheads, (6) a cell-compatible ready-to-use BioInk, and (7) standard operating procedures. In a proof-of-concept study, skin as a reference soft tissue model was printed. To produce dermal equivalents, primary human dermal fibroblasts were printed in alternating layers with BioInk and cultured for up to 7 weeks. During long-term cultures, the models were remodeled and fully populated with viable and spreaded fibroblasts. Primary human dermal keratinocytes were seeded on top of dermal equivalents, and epidermis-like structures were formed as verified with hematoxylin and eosin staining and immunostaining. However, a fully stratified epidermis was not achieved. Nevertheless, this is one of the first reports of an integrative bioprinting strategy for industrial routine application.

  5. PLX4032 Mediated Melanoma Associated Antigen Potentiation in Patient Derived Primary Melanoma Cells

    PubMed Central

    George, Andrea L.; Suriano, Robert; Rajoria, Shilpi; Osso, Maria C.; Tuli, Neha; Hanly, Elyse; Geliebter, Jan; Arnold, Angelo N.; Wallack, Marc; Tiwari, Raj K.

    2015-01-01

    Over expression of various immunogenic melanoma associated antigens (MAAs) has been exploited in the development of immunotherapeutic melanoma vaccines. Expression of MAAs such as MART-1 and gp100 is modulated by the MAPK signaling pathway, which is often deregulated in melanoma. The protein BRAF, a member of the MAPK pathway, is mutated in over 60% of melanomas providing an opportunity for the identification and approval by the FDA of a small molecule MAPK signaling inhibitor PLX4032 that functions to inactivate mutant BRAFV600E. To this end, we characterized five patient derived primary melanoma cell lines with respect to treatment with PLX4032. Cells were treated with 5μM PLX4032 and harvested. Western blotting analysis, RT-PCR and in vitro transwell migration and invasion assays were utilized to determine treatment effects. PLX4032 treatment modulated phosphorylation of signaling proteins belonging to the MAPK pathway including BRAF, MEK, and ERK and abrogated cell phenotypic characteristics such as migration and invasion. Most significantly, PLX4032 led to an up regulation of many MAA proteins in three of the four BRAF mutated cell lines, as determined at the protein and RNA level. Interestingly, MAGE-A1 protein and mRNA levels were reduced upon PLX4032 treatment in two of the primary lines. Taken together, our findings suggest that the BRAFV600E inhibitor PLX4032 has therapeutic potential over and above its known target and in combination with specific melanoma targeting vaccine strategies may have further clinical utility. PMID:26640592

  6. Extension of Lithium Ion Cell Model to Include Transient and Low-Temperature Behaviour

    NASA Astrophysics Data System (ADS)

    Dudley, G.

    2014-08-01

    Current-interruption resistance measurements have been analysed in detail allowing the ESTEC lithium ion cell electrical/thermal model to be extended to allow modelling of cell voltage in response to imposed current changes at low temperatures and short time scales where activation polarisation becomes important. Whilst an unnecessary complication in most cases, this extension is needed under certain circumstances such as the simulation of Mars rover batteries forced to operate at low temperature and possible effects of battery voltage transients on battery-bus power subsystems. Comparison with test data show that the model is capable of giving a good fit in these circumstances.

  7. Development of primary cell cultures using hemocytes and phagocytic tissue cells of Locusta migratoria: an application for locust immunity studies.

    PubMed

    Duressa, Tewodros Firdissa; Huybrechts, Roger

    2016-01-01

    Insect cell cultures played central roles in unraveling many insect physiological and immunological processes. Regardless, despite imminent needs, insect cell lines were developed primarily from Dipteran and Lepidopteran orders, leaving many important insects such as Orthopteran locusts under-represented. Besides the lack of cell lines, the slow progress in development of in vitro techniques is attributed to poor communications between different laboratories regarding optimized primary cell cultures. Therefore, we report here about methods developed for primary cell culture of Locusta migratoria hemocyte and phagocytic tissue cells by which we could maintain viable hemocytes in vitro for over 5 d and phagocytic tissue cells for over 12 d. 2-Mercaptoethanol and phenyl-thiourea supplements in Grace's medium together with addition of fetal bovine serum 30 min after cell seeding resulted in a successful setup of the primary cell cultures and a week-long survival of the hemocytes and phagocytic tissue cells in vitro.

  8. Primary cultures of human colon cancer as a model to study cancer stem cells.

    PubMed

    Koshkin, Sergey; Danilova, Anna; Raskin, Grigory; Petrov, Nikolai; Bajenova, Olga; O'Brien, Stephen J; Tomilin, Alexey; Tolkunova, Elena

    2016-09-01

    The principal cause of death in cancer involves tumor progression and metastasis. Since only a small proportion of the primary tumor cells, cancer stem cells (CSCs), which are the most aggressive, have the capacity to metastasize and display properties of stem cells, it is imperative to characterize the gene expression of diagnostic markers and to evaluate the drug sensitivity in the CSCs themselves. Here, we have examined the key genes that are involved in the progression of colorectal cancer and are expressed in cancer stem cells. Primary cultures of colorectal cancer cells from a patient's tumors were studied using the flow cytometry and cytological methods. We have evaluated the clinical and stem cell marker expression in these cells, their resistance to 5-fluorouracil and irinotecan, and the ability of cells to form tumors in mice. The data shows the role of stem cell marker Oct4 in the resistance of primary colorectal cancer tumor cells to 5-fluorouracil.

  9. Primary Cultures of Glomerular Parietal Epithelial Cells or Podocytes with Proven Origin

    PubMed Central

    Schulte, Kevin; Sechi, Antonio; Sauer-Lehnen, Sibille; Tag, Carmen; Boor, Peter; Kuppe, Christoph; Warsow, Gregor; Schordan, Sandra; Mostertz, Jörg; Chilukoti, Ravi Kumar; Homuth, Georg; Endlich, Nicole; Tacke, Frank; Weiskirchen, Ralf; Fuellen, Georg; Endlich, Karlhans; Floege, Jürgen; Smeets, Bart; Moeller, Marcus J.

    2012-01-01

    Parietal epithelial cells (PECs) are crucially involved in the pathogenesis of rapidly progressive glomerulonephritis (RPGN) as well as in focal and segmental glomerulosclerosis (FSGS). In this study, transgenic mouse lines were used to isolate pure, genetically tagged primary cultures of PECs or podocytes using FACsorting. By this approach, the morphology of primary glomerular epithelial cells in culture could be resolved: Primary podocytes formed either large cells with intracytoplasmatic extensions or smaller spindle shaped cells, depending on specific culture conditions. Primary PECs were small and exhibited a spindle-shaped or polygonal morphology. In the very early phases of primary culture, rapid changes in gene expression (e.g. of WT-1 and Pax-2) were observed. However, after prolonged culture primary PECs and podocytes still segregated clearly in a transcriptome analysis - demonstrating that the origin of primary cell cultures is important. Of the classical markers, synaptopodin and podoplanin expression were differentially regulated the most in primary PEC and podocyte cultures. However, no expression of any endogenous gene allowed to differentiate between the two cell types in culture. Finally, we show that the transcription factor WT1 is also expressed by PECs. In summary, genetic tagging of PECs and podocytes is a novel and necessary tool to derive pure primary cultures with proven origin. These cultures will be a powerful tool for the emerging field of parietal epithelial cell biology. PMID:22529955

  10. Estimation of Cell-Type Composition Including T and B Cell Subtypes for Whole Blood Methylation Microarray Data

    PubMed Central

    Waite, Lindsay L.; Weaver, Benjamin; Day, Kenneth; Li, Xinrui; Roberts, Kevin; Gibson, Andrew W.; Edberg, Jeffrey C.; Kimberly, Robert P.; Absher, Devin M.; Tiwari, Hemant K.

    2016-01-01

    DNA methylation levels vary markedly by cell-type makeup of a sample. Understanding these differences and estimating the cell-type makeup of a sample is an important aspect of studying DNA methylation. DNA from leukocytes in whole blood is simple to obtain and pervasive in research. However, leukocytes contain many distinct cell types and subtypes. We propose a two-stage model that estimates the proportions of six main cell types in whole blood (CD4+ T cells, CD8+ T cells, monocytes, B cells, granulocytes, and natural killer cells) as well as subtypes of T and B cells. Unlike previous methods that only estimate overall proportions of CD4+ T cell, CD8+ T cells, and B cells, our model is able to estimate proportions of naïve, memory, and regulatory CD4+ T cells as well as naïve and memory CD8+ T cells and naïve and memory B cells. Using real and simulated data, we are able to demonstrate that our model is able to reliably estimate proportions of these cell types and subtypes. In studies with DNA methylation data from Illumina's HumanMethylation450k arrays, our estimates will be useful both for testing for associations of cell type and subtype composition with phenotypes of interest as well as for adjustment purposes to prevent confounding in epigenetic association studies. Additionally, our method can be easily adapted for use with whole genome bisulfite sequencing (WGBS) data or any other genome-wide methylation data platform. PMID:26925097

  11. Estimation of Cell-Type Composition Including T and B Cell Subtypes for Whole Blood Methylation Microarray Data.

    PubMed

    Waite, Lindsay L; Weaver, Benjamin; Day, Kenneth; Li, Xinrui; Roberts, Kevin; Gibson, Andrew W; Edberg, Jeffrey C; Kimberly, Robert P; Absher, Devin M; Tiwari, Hemant K

    2016-01-01

    DNA methylation levels vary markedly by cell-type makeup of a sample. Understanding these differences and estimating the cell-type makeup of a sample is an important aspect of studying DNA methylation. DNA from leukocytes in whole blood is simple to obtain and pervasive in research. However, leukocytes contain many distinct cell types and subtypes. We propose a two-stage model that estimates the proportions of six main cell types in whole blood (CD4+ T cells, CD8+ T cells, monocytes, B cells, granulocytes, and natural killer cells) as well as subtypes of T and B cells. Unlike previous methods that only estimate overall proportions of CD4+ T cell, CD8+ T cells, and B cells, our model is able to estimate proportions of naïve, memory, and regulatory CD4+ T cells as well as naïve and memory CD8+ T cells and naïve and memory B cells. Using real and simulated data, we are able to demonstrate that our model is able to reliably estimate proportions of these cell types and subtypes. In studies with DNA methylation data from Illumina's HumanMethylation450k arrays, our estimates will be useful both for testing for associations of cell type and subtype composition with phenotypes of interest as well as for adjustment purposes to prevent confounding in epigenetic association studies. Additionally, our method can be easily adapted for use with whole genome bisulfite sequencing (WGBS) data or any other genome-wide methylation data platform.

  12. Background and future considerations for human cord blood hematopoietic cell transplantation, including economic concerns.

    PubMed

    Broxmeyer, Hal E; Farag, Sherif

    2013-12-01

    Cord blood (CB) has been used since 1988 as a source of hematopoietic stem cells (HSCs) and progenitor cells for hematopoietic cell transplantation (HCT) to treat patients with malignant and nonmalignant disorders. CB has both advantages and disadvantages when compared with other tissue sources of HSCs such as bone marrow and mobilized peripheral blood, which are also being used in the setting of HCT. This short review focuses on some historical information, as well as current efforts that are being assessed to enhance the efficacy of CB HCT. Also of importance are the costs of CB, and the feasibility and economics of using such to be identified, and newly confirmed improvements worldwide for the greatest number of patients. In this context, simple methods that would not necessarily entail the need for selected cell-processing facilities to ex vivo expand or improve the CB graft's functional activity may be of interest, with one such possibility being the use of an orally active inhibitor of the enzyme dipeptidylpeptidase 4, alone or in combination with other new and innovative approaches for improving HSC engraftment and in vivo repopulating capability of CB.

  13. Proteasome inhibitors, including curcumin, improve pancreatic β-cell function and insulin sensitivity in diabetic mice

    PubMed Central

    Weisberg, S; Leibel, R; Tortoriello, D V

    2016-01-01

    Background: Type 2 diabetes stems from obesity-associated insulin resistance, and in the genetically susceptible, concomitant pancreatic β-cell failure can occur, which further exacerbates hyperglycemia. Recent work by our group and others has shown that the natural polyphenol curcumin attenuates the development of insulin resistance and hyperglycemia in mouse models of hyperinsulinemic or compensated type 2 diabetes. Although several potential downstream molecular targets of curcumin exist, it is now recognized to be a direct inhibitor of proteasome activity. We now show that curcumin also prevents β-cell failure in a mouse model of uncompensated obesity-related insulin resistance (Leprdb/db on the Kaliss background). Results: In this instance, dietary supplementation with curcumin prevented hyperglycemia, increased insulin production and lean body mass, and prolonged lifespan. In addition, we show that short-term in vivo treatment with low dosages of two molecularly distinct proteasome inhibitors celastrol and epoxomicin reverse hyperglycemia in mice with β-cell failure by increasing insulin production and insulin sensitivity. Conclusions: These studies suggest that proteasome inhibitors may prove useful for patients with diabetes by improving both β-cell function and relieving insulin resistance. PMID:27110686

  14. Ten-year treatment outcomes including blood cell count disturbances in patients with simple renal cysts.

    PubMed

    Bryniarski, Piotr; Kaletka, Zbigniew; Życzkowski, Marcin; Prokopowicz, Grzegorz; Muskała, Bartosz; Paradysz, Andrzej

    2013-07-01

    The simple renal cyst is the most common benign kidney disease. It may cause pain and hypertension, especially if significantly enlarged. As in polycystic kidney disease, blood cell count disturbances are frequently observed in simple renal cysts. The aim of our study was to assess such disturbances, changes in blood pressure, and complication rate in our patients undergoing surgery due to simple renal cyst in the last 10 years. 210 patients with simple renal cysts were underwent surgery between 2002 and 2012. Two different kinds of operation were conducted: aspiration of cyst fluid with injection of sclerosing agent, and laparoscopic/retroperitoneoscopic decortications of the cyst wall. A control group comprised 134 patients with benign prostate hyperplasia. The following data were obtained: cyst burden, hematocrit, hemoglobin, red blood cells, thrombocytes, occurrence of pain, and blood pressure before and after the operation. Complications were collected and presented in Clavien score. Hematocrit, hemoglobin, and red blood cells were significantly increased in the experimental group. A positive correlation was observed between cyst burden and the parameters mentioned above. Of 91 patients with hypertension, 56 (61.7%) had blood pressure reduction after the operation. Treatment relieved the loin pain in 132 (88%) patients. Complications occurred in 15 (7.4%) patients. Patients with simple renal cysts have high values of red blood cells, hematocrit, and hemoglobin. Treatment decreases blood pressure in patients with hypertension. Complications after treatment are rare and mild.

  15. Organ Preference of Cancer Metastasis and Metastasis-Related Cell Adhesion Molecules Including Carbohydrates.

    PubMed

    Kawaguchi, Takanori

    2016-01-01

    This review starts on one of our special interests, the organ preference of metastasis. We examined data on 1,117 autopsy cases and found that the organ distribution of metastasis of cancers of the lung, pancreas, stomach, colon, rectum, uterine cervix, liver, bile duct, and esophagus involved the lung, liver, adrenal gland, bone/bone marrow, lymph node, and pleura/peritoneum. Cancers of the kidney, thyroid, ovary, choriocarcinoma, and breast, however, manifested different metastatic patterns. The distribution of leukemia and lymphoma metastases was quite different from that of epithelial cancers. On the basis of experimental studies, we believe that the anatomical-mechanical hypothesis should be replaced by the microinjury hypothesis, which suggests that tissue microinjury induced by temporal tumor cell embolization is crucial for successful metastasis. This hypothesis may actually reflect the so-called inflammatory oncotaxis concept. To clarify the mechanisms underlying metastasis, we developed an experimental model system of a rat hepatoma AH7974 that embraced substrate adhesiveness. This model did not prove a relationship between substrate-adhesion potential and metastatic lung-colonizing potential of tumor cells, but metastatic potential was correlated with the expression of the laminin carbohydrate that was recognized by Griffonia (Bandeiraea) simplicifolia isolectin G4. Therefore, we investigated the relationship between carbohydrate expression profiles and metastasis and prognosis. We indeed found an intimate relationship between the carbohydrate expression of cancer cells and the progression of malignant tumors, organ preference of metastasis, metastatic potential of tumor cells, and prognosis of patients.

  16. HPV Status and second primary tumours in Oropharyngeal Squamous Cell Carcinoma

    PubMed Central

    2013-01-01

    Introductions The incidence of human papillomavirus (HPV)-related oropharyngeal squamous cell carcinoma (OPSCCs) is rising in developed nations. Studies have shown that these virally mediated tumours are epidemiologically, clinically, and biologically different than other head and neck squamous cell carcinomas and traditional concepts of field cancerization may not apply to HPV-related oropharyngeal cancer. Objective The purpose of this study was to evaluate the rate of second primary tumors and the diagnostic yield of field cancerization work up in the upper aerodigestive tract in patients with HPV-related and HPV-unrelated oropharyngeal squamous cell carcinoma. Design Retrospective review. Setting Tertiary cancer care centers in Alberta. Methods Retrospective review of 406 patients diagnosed with OPSCC in Alberta between 2005 and 2009. HPV-status of tumours was determined by tissue microarray using immunohistochemistry staining for p16. Main outcome measures Primary outcome: incidence of upper aerodigestive tract second primary tumours in p16-positive versus p16-negative OPSCC. Secondary outcomes: diagnostic yield of traditional field cancerization work-up in p16-positive versus negative patients. Results The overall rate of SPTs was 7.4% (30/406). The incidence rate of SPTs was significantly lower in p16-positive patients (0.7 per 100 patient-yrs vs. 8.5 in p16-negative, p < 0.0001). Field cancerization work-up for synchronous lesions in the upper aerodigestive tract, including panendoscopy and whole-body PET-CT, had decreased diagnostic yield in p16-positive patients (2.8% vs. 10.2% in HPV-negative patients, p=0.02). Conclusions Patients with HPV-related OPSCC, who are non-smokers have decreased risk of developing second primary tumours in the upper aerodigestive tract and have low yield on field cancerization work-up. This study provides further evidence that virally mediated OPSCC are distinct and may benefit from alternate diagnostic pathways. PMID:23718873

  17. 9 CFR 113.51 - Requirements for primary cells used for production of biologics.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Requirements for primary cells used for production of biologics. 113.51 Section 113.51 Animals and Animal Products ANIMAL AND PLANT HEALTH... VECTORS STANDARD REQUIREMENTS Ingredient Requirements § 113.51 Requirements for primary cells used...

  18. 9 CFR 113.51 - Requirements for primary cells used for production of biologics.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Requirements for primary cells used for production of biologics. 113.51 Section 113.51 Animals and Animal Products ANIMAL AND PLANT HEALTH... VECTORS STANDARD REQUIREMENTS Ingredient Requirements § 113.51 Requirements for primary cells used...

  19. 9 CFR 113.51 - Requirements for primary cells used for production of biologics.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Requirements for primary cells used for production of biologics. 113.51 Section 113.51 Animals and Animal Products ANIMAL AND PLANT HEALTH... VECTORS STANDARD REQUIREMENTS Ingredient Requirements § 113.51 Requirements for primary cells used...

  20. Cryopreservation and lentiviral-mediated genetic modification of human primary cultured corneal endothelial cells.

    PubMed

    Suh, Leejee H; Zhang, Cheng; Chuck, Roy S; Stark, Walter J; Naylor, Stuart; Binley, Katie; Chakravarti, Shukti; Jun, Albert S

    2007-07-01

    To determine the viability and potential usefulness of cryopreserved human primary cultured corneal endothelial cells by characterizing their morphology, gene expression, and ability for genetic modification by the lentiviral vector equine infectious anemia virus (EIAV). Primary cultured endothelial cells were dissociated from human corneas and grown in organ culture medium. Corneal endothelial cell origin was confirmed by morphology and immunostaining with polyclonal anti-collagen VIII antibodies. Cells of different passages were cryopreserved in medium containing dimethyl sulfoxide and were assessed after thawing for morphology, proliferative capacity, gene expression, and ability to form cell-cell junctions. EIAV encoding enhanced green fluorescent protein (eGFP) was used to transduce cryopreserved human corneal endothelial cells. Transduced cells were then sorted by fluorescence-activated cell sorting (FACS) and imaged with fluorescence microscopy. Cryopreserved, primary, cultured human corneal endothelial cells are viable and retain their ability to proliferate, produce collagen VIII, and express ZO-1, a tight-junction protein. EIAV-based gene transfer of eGFP is highly efficient and nontoxic to cryopreserved human primary cultured corneal endothelial cells. These genetically modified cells can be selected to nearly pure populations with FACS sorting. Human primary cultured corneal endothelial cells retain their phenotypic properties after cryopreservation. The ability to store, genetically modify, and sort these cells through FACS to pure populations has the potential to greatly expand their future therapeutic application to treat corneal endothelial disorders.

  1. Cell communication in a coculture system consisting of outgrowth endothelial cells and primary osteoblasts.

    PubMed

    Herzog, David Paul Eric; Dohle, Eva; Bischoff, Iris; Kirkpatrick, Charles James

    2014-01-01

    Bone tissue is a highly vascularized and dynamic system with a complex construction. In order to develop a construct for implant purposes in bone tissue engineering, a proper understanding of the complex dependencies between different cells and cell types would provide further insight into the highly regulated processes during bone repair, namely, angiogenesis and osteogenesis, and might result in sufficiently equipped constructs to be beneficial to patients and thereby accomplish their task. This study is based on an in vitro coculture model consisting of outgrowth endothelial cells and primary osteoblasts and is currently being used in different studies of bone repair processes with special regard to angiogenesis and osteogenesis. Coculture systems of OECs and pOBs positively influence the angiogenic potential of endothelial cells by inducing the formation of angiogenic structures in long-term cultures. Although many studies have focused on cell communication, there are still numerous aspects which remain poorly understood. Therefore, the aim of this study is to investigate certain growth factors and cell communication molecules that are important during bone repair processes. Selected growth factors like VEGF, angiopoietins, BMPs, and IGFs were investigated during angiogenesis and osteogenesis and their expression in the cultures was observed and compared after one and four weeks of cultivation. In addition, to gain a better understanding on the origin of different growth factors, both direct and indirect coculture strategies were employed. Another important focus of this study was to investigate the role of "gap junctions," small protein pores which connect adjacent cells. With these bridges cells are able to exchange signal molecules, growth factors, and other important mediators. It could be shown that connexins, the gap junction proteins, were located around cell nuclei, where they await their transport to the cell membrane. In addition, areas in which two

  2. Primary tumor induces sentinel lymph node lymphangiogenesis in oral squamous cell carcinoma.

    PubMed

    Ishii, Hiroki; Chikamatsu, Kazuaki; Sakakura, Koichi; Miyata, Masanori; Furuya, Nobuhiko; Masuyama, Keisuke

    2010-05-01

    The main factor that affects the prognosis of patients with oral squamous cell carcinoma (OSCC) is regional lymph node metastases, which usually spreads first to the sentinel lymph nodes (SLNs). Recent studies have demonstrated that tumor cells in several malignancies can induce lymphangiogenesis in SLNs before metastasizing. To elucidate the mechanisms of tumor dissemination of OSCC, we investigated whether primary tumors induce lymphangiogenesis within SLNs in patients with OSCC. The mRNA expression of lymphatic-specific markers, including VEGFR-3, Prox-1, and LYVE-1 in 23 metastasis-negative SLNs obtained from 10 patients with OSCC, was investigated using a quantitative real-time RT-PCR assay, and compared with control lymph nodes from patients with non-cancerous diseases. In addition, VEGF-C and VEGF-D expressions of the primary tumor were examined by immunohistochemistry. In SLNs, there were highly significant correlations between the three lymphatic markers examined. Interestingly, the level of LYVE-1 expression in SLNs, despite the absence of metastasis, was significantly higher than in control lymph nodes. Moreover, SLNs from patients with VEGF-C-positive tumor showed a significantly higher expression of VEGFR-3 than those from patients with VEGF-C-negative tumor. Our findings suggest that in OSCC, the primary tumor actively induces lymphangiogenesis in SLNs prior to the onset of metastases, and where tumor-derived VEGF-C plays an important role.

  3. Thermodynamic Modeling and Dispatch of Distributed Energy Technologies including Fuel Cell -- Gas Turbine Hybrids

    NASA Astrophysics Data System (ADS)

    McLarty, Dustin Fogle

    Distributed energy systems are a promising means by which to reduce both emissions and costs. Continuous generators must be responsive and highly efficiency to support building dynamics and intermittent on-site renewable power. Fuel cell -- gas turbine hybrids (FC/GT) are fuel-flexible generators capable of ultra-high efficiency, ultra-low emissions, and rapid power response. This work undertakes a detailed study of the electrochemistry, chemistry and mechanical dynamics governing the complex interaction between the individual systems in such a highly coupled hybrid arrangement. The mechanisms leading to the compressor stall/surge phenomena are studied for the increased risk posed to particular hybrid configurations. A novel fuel cell modeling method introduced captures various spatial resolutions, flow geometries, stack configurations and novel heat transfer pathways. Several promising hybrid configurations are analyzed throughout the work and a sensitivity analysis of seven design parameters is conducted. A simple estimating method is introduced for the combined system efficiency of a fuel cell and a turbine using component performance specifications. Existing solid oxide fuel cell technology is capable of hybrid efficiencies greater than 75% (LHV) operating on natural gas, and existing molten carbonate systems greater than 70% (LHV). A dynamic model is calibrated to accurately capture the physical coupling of a FC/GT demonstrator tested at UC Irvine. The 2900 hour experiment highlighted the sensitivity to small perturbations and a need for additional control development. Further sensitivity studies outlined the responsiveness and limits of different control approaches. The capability for substantial turn-down and load following through speed control and flow bypass with minimal impact on internal fuel cell thermal distribution is particularly promising to meet local demands or provide dispatchable support for renewable power. Advanced control and dispatch

  4. T-cell number and subtype influence the disease course of primary chronic lymphocytic leukaemia xenografts in alymphoid mice

    PubMed Central

    Oldreive, Ceri E.; Skowronska, Anna; Davies, Nicholas J.; Parry, Helen; Agathanggelou, Angelo; Krysov, Sergey; Packham, Graham; Rudzki, Zbigniew; Cronin, Laura; Vrzalikova, Katerina; Murray, Paul; Odintsova, Elena; Pratt, Guy; Taylor, A. Malcolm R.; Moss, Paul; Stankovic, Tatjana

    2015-01-01

    ABSTRACT Chronic lymphocytic leukaemia (CLL) cells require microenvironmental support for their proliferation. This can be recapitulated in highly immunocompromised hosts in the presence of T cells and other supporting cells. Current primary CLL xenograft models suffer from limited duration of tumour cell engraftment coupled with gradual T-cell outgrowth. Thus, a greater understanding of the interaction between CLL and T cells could improve their utility. In this study, using two distinct mouse xenograft models, we investigated whether xenografts recapitulate CLL biology, including natural environmental interactions with B-cell receptors and T cells, and whether manipulation of autologous T cells can expand the duration of CLL engraftment. We observed that primary CLL xenografts recapitulated both the tumour phenotype and T-cell repertoire observed in patients and that engraftment was significantly shorter for progressive tumours. A reduction in the number of patient T cells that were injected into the mice to 2-5% of the initial number or specific depletion of CD8+ cells extended the limited xenograft duration of progressive cases to that characteristic of indolent disease. We conclude that manipulation of T cells can enhance current CLL xenograft models and thus expand their utility for investigation of tumour biology and pre-clinical drug assessment. PMID:26398941

  5. Unusual Presentation of Primary Squamous Cell Carcinoma of Mandible

    PubMed Central

    Shanmugasundaram, Karpagavalli; Subramanian, Sathasiva; Kumar, Vimal

    2016-01-01

    Carcinoma arising primarily from the jaw is a locally aggressive lesion with poor prognosis. Primary intraosseous carcinoma (PIOC) lesion develops either de novo remnants of odontogenic epithelium, odontogenic cyst/tumor, epithelium remnants, or/and salivary gland residues. We describe very interesting case of primary intraosseous carcinoma of mandible. This extensive lesion was sent for oncological opinion and further management. Due to the uncertainty of diagnostic criteria of PIOC, only few cases of this lesion with a typical presentation have been reported. This article presents a case of primary intraosseous carcinoma with a unique appearance and detailed review stating its clinicopathological correlation. PMID:28078158

  6. Chemical proteomic map of dimethyl fumarate–sensitive cysteines in primary human T cells

    PubMed Central

    Blewett, Megan M.; Xie, Jiji; Zaro, Balyn W.; Backus, Keriann M.; Altman, Amnon; Teijaro, John R.; Cravatt, Benjamin F.

    2016-01-01

    Dimethyl fumarate (DMF) is an electrophilic drug that is used to treat autoimmune conditions, including multiple sclerosis and psoriasis. The mechanism of action of DMF is unclear, but may involve the covalent modification of proteins or DMF serving as a pro-drug that is converted to monomethyl fumarate (MMF). Here, we found that DMF, but not MMF, blocked the activation of primary human and mouse T cells. Using a quantitative, site-specific chemical proteomic platform, we determined the DMF-sensitivity of > 2400 cysteine residues in human T cells. Cysteines sensitive to DMF, but not MMF, were identified in several proteins with established biochemical or genetic links to T cell function, including protein kinase C θ (PKCθ). Furthermore, DMF blocked the association of PKCθ with the costimulatory receptor CD28 by perturbing a CXXC motif in the C2 domain of this kinase. Mutation of these DMF-sensitive cysteines also impaired PKCθ-CD28 interactions and T cell activation, designating the C2 domain of PKCθ as a key functional, electrophile-sensing module important for T cell biology. PMID:27625306

  7. Propionibacterium acnes inhibits FOXM1 and induces cell cycle alterations in human primary prostate cells.

    PubMed

    Sayanjali, Behnam; Christensen, Gitte J M; Al-Zeer, Munir A; Mollenkopf, Hans-Joachim; Meyer, Thomas F; Brüggemann, Holger

    2016-11-01

    Propionibacterium acnes has been detected in diseased human prostate tissue, and cell culture experiments suggest that the bacterium can establish a low-grade inflammation. Here, we investigated its impact on human primary prostate epithelial cells. Microarray analysis confirmed the inflammation-inducing capability of P. acnes but also showed deregulation of genes involved in the cell cycle. qPCR experiments showed that viable P. acnes downregulates a master regulator of cell cycle progression, FOXM1. Flow cytometry experiments revealed that P. acnes increases the number of cells in S-phase. We tested the hypothesis that a P. acnes-produced berninamycin-like thiopeptide is responsible for this effect, since it is related to the FOXM1 inhibitor siomycin. The thiopeptide biosynthesis gene cluster was strongly expressed; it is present in subtype IB of P. acnes, but absent from type IA, which is most abundant on human skin. A knock-out mutant lacking the gene encoding the berninamycin-like peptide precursor was unable to downregulate FOXM1 and to halt the cell cycle. Our study reveals a novel host cell-interacting activity of P. acnes.

  8. Cancer risks related to low-level RF/MW exposures, including cell phones.

    PubMed

    Szmigielski, Stanislaw

    2013-09-01

    For years, radiofrequency (RF) and microwave (MW) radiations have been applied in the modern world. The rapidly increasing use of cellular phones called recent attention to the possible health risks of RF/MW exposures. In 2011, a group of international experts organized by IARC (International Agency for Research on Cancer in Lyon) concluded that RF/MW radiations should be listed as a possible carcinogen (group 2B) for humans. Three meta-analyses of case-control studies have concluded that using cell phones for more than ten years was associated with an increase in the overall risk of developing a brain tumor. The Interphone Study, the largest health-related case-control international study of use of cell phones and head and neck tumors, showed no statistically significant increases in brain cancers related to higher amounts of cell phone use, but excess risk in a small subgroup of more heavily exposed users associated with latency and laterality was reported. So far, the published studies do not show that mobile phones could for sure increase the risk of cancer. This conclusion is based on the lack of a solid biological mechanism, and the fact that brain cancer rates are not going up significantly. However, all of the studies so far have weaknesses, which make it impossible to entirely rule out a risk. Mobile phones are still a new technology and there is little evidence about effects of long-term use. For this reason, bioelectromagnetic experts advise application of a precautionary resources. It suggests that if people want to use a cell phone, they can choose to minimize their exposure by keeping calls short and preferably using hand-held sets. It also advises discouraging children from making non essential calls as well as also keeping their calls short.

  9. Reduced Toxicity Fuel Satellite Propulsion System Including Fuel Cell Reformer with Alcohols Such as Methanol

    NASA Technical Reports Server (NTRS)

    Schneider, Steven J. (Inventor)

    2001-01-01

    A reduced toxicity fuel satellite propulsion system including a reduced toxicity propellant supply for consumption in an axial class thruster and an ACS class thruster. The system includes suitable valves and conduits for supplying the reduced toxicity propellant to the ACS decomposing element of an ACS thruster. The ACS decomposing element is operative to decompose the reduced toxicity propellant into hot propulsive gases. In addition the system includes suitable valves and conduits for supplying the reduced toxicity propellant to an axial decomposing element of the axial thruster. The axial decomposing element is operative to decompose the reduced toxicity propellant into hot gases. The system further includes suitable valves and conduits for supplying a second propellant to a combustion chamber of the axial thruster, whereby the hot gases and the second propellant auto-ignite and begin the combustion process for producing thrust.

  10. Polarization birefringence measurements for characterizing the myocardium, including healthy, infarcted, and stem-cell-regenerated tissues

    NASA Astrophysics Data System (ADS)

    Wood, Michael F. G.; Ghosh, Nirmalya; Wallenburg, Marika A.; Li, Shu-Hong; Weisel, Richard D.; Wilson, Brian C.; Li, Ren-Ke; Vitkin, I. Alex

    2010-07-01

    Myocardial infarction leads to structural remodeling of the myocardium, in particular to the loss of cardiomyocytes due to necrosis and an increase in collagen with scar formation. Stem cell regenerative treatments have been shown to alter this remodeling process, resulting in improved cardiac function. As healthy myocardial tissue is highly fibrous and anisotropic, it exhibits optical linear birefringence due to the different refractive indices parallel and perpendicular to the fibers. Accordingly, changes in myocardial structure associated with infarction and treatment-induced remodeling will alter the anisotropy exhibited by the tissue. Polarization-based linear birefringence is measured on the myocardium of adult rat hearts after myocardial infarction and compared with hearts that had received mesenchymal stem cell treatment. Both point measurement and imaging data show a decrease in birefringence in the region of infarction, with a partial rebound back toward the healthy values following regenerative treatment with stem cells. These results demonstrate the ability of optical polarimetry to characterize the micro-organizational state of the myocardium via its measured anisotropy, and the potential of this approach for monitoring regenerative treatments of myocardial infarction.

  11. Dehydroepiandrosterone inhibits cell proliferation and improves viability by regulating S phase and mitochondrial permeability in primary rat Leydig cells.

    PubMed

    Liu, Lin; Wang, Dian; Li, Longlong; Ding, Xiao; Ma, Haitian

    2016-07-01

    Dehydroepiandrosterone (DHEA) is widely used as a nutritional supplement and exhibits putative anti‑aging properties. However, the molecular basis of the actions of DHEA, particularly on the biological characteristics of target cells, remain unclear. The aim of the current study was to investigate the effects of DHEA on cell viability, cell proliferation, cell cycle and mitochondrial function in primary rat Leydig cells. Adult Leydig cells were purified by Percoll gradient centrifugation, and cell proliferation was detected using a Click-iT® EdU Assay kit and cell cycle assessment performed using flow cytometry. Mitochondrial membrane potential was detected using JC-1 staining assay. The results of the current study demonstrate that DHEA decreased cell proliferation in a dose‑dependent manner, whereas it improved cell viability in a time‑dependent and dose‑dependent manner. Flow cytometry analysis demonstrated that DHEA treatment increased the S phase cell population and decreased the G2/M cell population. Cyclin A and CDK2 mRNA levels were decreased in primary rat Leydig cells following DHEA treatment. DHEA treatment decreased the transmembrane electrical gradient in primary Leydig cells, whereas treatment significantly increased succinate dehydrogenase activity. These results indicated that DHEA inhibits primary rat Leydig cell proliferation by decreasing cyclin mRNA level, whereas it improves cells viability by modulating the permeability of the mitochondrial membrane and succinate dehydrogenase activity. These findings may demonstrate an important molecular mechanism by which DHEA activity is mediated.

  12. A Review of the Physiological and Immunological Functions of Biliary Epithelial Cells: Targets for Primary Biliary Cirrhosis, Primary Sclerosing Cholangitis and Drug-induced Ductopenias

    PubMed Central

    Wu, Chih-Te; Davis, Paul A.; Luketic, VelImir A.; Gershwin, M. Eric

    2004-01-01

    Our understanding of biliary epithelial cells (BEC) in physiobiology and immunology has steadily expanded. BEC transports IgA as well as IgM into bile, synthesizes and secretes various chemokines, cytokines, and expresses adhesion molecules involved in cell interaction and signal transduction. These then suggest a myriad of potential roles for BEC in defense from invading microorganisms as well as the pathogenesis of diverse immunologically driven diseases such as primary biliary cirrhosis (PBC), graft-versus-host disease, and primary sclerosing cholangitis (PSC). Despite the progress, there still remain many areas of BEC biology that require further investigation. Most importantly, it remains to be clarified that the extent to which the immunologic activities observed in BEC represent a BEC response to tissue injury or whether BEC themselves are the active participants in the pathogenesis of various cholestatic immunological diseases, including PBC and PSC. PMID:15559365

  13. Evaluation of primary mediastinal large B cell lymphoma by flow cytometry.

    PubMed

    Cherian, Sindhu; Fromm, Jonathan R

    2017-07-21

    Primary mediastinal large B cell lymphoma (PMLBCL) is a B cell non-Hodgkin lymphoma (B-NHL) that shows morphologic, immunophenotypic, and genetic similarities to classical Hodgkin lymphoma (CHL). We evaluated the neoplastic and reactive infiltrate of PMLBCL by flow cytometry (FC). A total of 24 cases of PMLBCL were retrospectively characterized using FC combinations for B-NHL, T-NHL, and CHL. The CHL assay identified the neoplastic population (NP) in all cases, while the B-NHL assay identified the NP in 18 of 24 cases. In four cases, the neoplastic population was retrospectively identified in the B-NHL tube, given the CHL tube-derived immunophenotype. Neoplastic cells of PMLBCL displayed a B cell immunophenotype (CD19/CD20(+) ), with CD40 consistently and brightly expressed. Most cases lacked immunoglobulin light chains, CD10, and CD15; CD30 was frequently expressed. All cases showed expression of CD71 and CD95. The reactive infiltrate in PMLBCL showed increases in T cells relative to reactive tissues. Lower CD4/CD8 ratios in PMLBCL relative to CHL and reactive tissues were also observed. Although not seen as commonly as with CHL, 41% of cases showed a reactive CD3/CD4/CD7(bright) /CD45(bright+) T cell population. Reactive populations seen in PMLBCL were similar to that seen in diffuse large B cell lymphoma, not otherwise specified. FC can detect the NP of PMLBCL and a FC assay for CHL performed better than a B-NHL assay in this regard. The neoplastic cells showed a B cell immunophenotype with over-expression of CD40. The reactive infiltrate in PMLBCL shows unique features including a prominent CD8(+) T cell infiltrate. © 2017 International Clinical Cytometry Society. © 2017 International Clinical Cytometry Society.

  14. Role of non-coding RNAs in maintaining primary airway smooth muscle cells

    PubMed Central

    2014-01-01

    Background The airway smooth muscle (ASM) cell maintains its own proliferative rate and contributes to the inflammatory response in the airways, effects that are inhibited by corticosteroids, used in the treatment of airways diseases. Objective We determined the differential expression of mRNAs, microRNAs (miRNAs) and long noncoding RNA species (lncRNAs) in primary ASM cells following treatment with a corticosteroid, dexamethasone, and fetal calf serum (FCS). Methods mRNA, miRNA and lncRNA expression was measured by microarray and quantitative real-time PCR. Results A small number of miRNAs (including miR-150, −371-5p, −718, −940, −1181, −1207-5p, −1915, and −3663-3p) were decreased following exposure to dexamethasone and FCS. The mRNA targets of these miRNAs were increased in expression. The changes in mRNA expression were associated with regulation of ASM actin cytoskeleton. We also observed changes in expression of lncRNAs, including natural antisense, pseudogenes, intronic lncRNAs, and intergenic lncRNAs following dexamethasone and FCS. We confirmed the change in expression of three of these, LINC00882, LINC00883, PVT1, and its transcriptional activator, c-MYC. We propose that four of these lincRNAs (RP11-46A10.4, LINC00883, BCYRN1, and LINC00882) act as miRNA ‘sponges’ for 4 miRNAs (miR-150, −371-5p, −940, −1207-5p). Conclusion This in-vitro model of primary ASM cell phenotype was associated with the regulation of several ncRNAs. Their identification allows for in-vitro functional experimentation to establish causality with the primary ASM phenotype, and in airway diseases such as asthma and chronic obstructive pulmonary disease (COPD). PMID:24886442

  15. SDF-1/CXCR4 axis in Tie2-lineage cells including endothelial progenitor cells contributes to bone fracture healing.

    PubMed

    Kawakami, Yohei; Ii, Masaaki; Matsumoto, Tomoyuki; Kuroda, Ryosuke; Kuroda, Tomoya; Kwon, Sang-Mo; Kawamoto, Atsuhiko; Akimaru, Hiroshi; Mifune, Yutaka; Shoji, Taro; Fukui, Tomoaki; Kurosaka, Masahiro; Asahara, Takayuki

    2015-01-01

    CXC chemokine receptor 4 (CXCR4) is a specific receptor for stromal-derived-factor 1 (SDF-1). SDF-1/CXCR4 interaction is reported to play an important role in vascular development. On the other hand, the therapeutic potential of endothelial progenitor cells (EPCs) in fracture healing has been demonstrated with mechanistic insight of vasculogenesis/angiogenesis and osteogenesis enhancement at sites of fracture. The purpose of this study was to investigate the influence of the SDF-1/CXCR4 pathway in Tie2-lineage cells (including EPCs) in bone formation. We created CXCR4 gene conditional knockout mice using the Cre/loxP system and set two groups of mice: Tie2-Cre(ER) CXCR4 knockout mice (CXCR4(-/-) ) and wild-type mice (WT). We report here that in vitro, EPCs derived from of CXCR4(-/-) mouse bone marrow demonstrated severe reduction of migration activity and EPC colony-forming activity when compared with those derived from WT mouse bone marrow. In vivo, radiological and morphological examinations showed fracture healing delayed in the CXCR4(-/-) group and the relative callus area at weeks 2 and 3 was significantly smaller in CXCR4(-/-) group mice. Quantitative analysis of capillary density at perifracture sites also showed a significant decrease in the CXCR4(-/-) group. Especially, CXCR4(-/-) group mice demonstrated significant early reduction of blood flow recovery at fracture sites compared with the WT group in laser Doppler perfusion imaging analysis. Real-time RT-PCR analysis showed that the gene expressions of angiogenic markers (CD31, VE-cadherin, vascular endothelial growth factor [VEGF]) and osteogenic markers (osteocalcin, collagen 1A1, bone morphogenetic protein 2 [BMP2]) were lower in the CXCR4(-/-) group. In the gain-of-function study, the fracture in the SDF-1 intraperitoneally injected WT group healed significantly faster with enough callus formation compared with the SDF-1 injected CXCR4(-/-) group. We demonstrated that an EPC SDF-1/CXCR4 axis plays an

  16. An imbalance in progenitor cell populations reflects tumour progression in breast cancer primary culture models

    PubMed Central

    2011-01-01

    Background Many factors influence breast cancer progression, including the ability of progenitor cells to sustain or increase net tumour cell numbers. Our aim was to define whether alterations in putative progenitor populations could predict clinicopathological factors of prognostic importance for cancer progression. Methods Primary cultures were established from human breast tumour and adjacent non-tumour tissue. Putative progenitor cell populations were isolated based on co-expression or concomitant absence of the epithelial and myoepithelial markers EPCAM and CALLA respectively. Results Significant reductions in cellular senescence were observed in tumour versus non-tumour cultures, accompanied by a stepwise increase in proliferation:senescence ratios. A novel correlation between tumour aggressiveness and an imbalance of putative progenitor subpopulations was also observed. Specifically, an increased double-negative (DN) to double-positive (DP) ratio distinguished aggressive tumours of high grade, estrogen receptor-negativity or HER2-positivity. The DN:DP ratio was also higher in malignant MDA-MB-231 cells relative to non-tumourogenic MCF-10A cells. Ultrastructural analysis of the DN subpopulation in an invasive tumour culture revealed enrichment in lipofuscin bodies, markers of ageing or senescent cells. Conclusions Our results suggest that an imbalance in tumour progenitor subpopulations imbalances the functional relationship between proliferation and senescence, creating a microenvironment favouring tumour progression. PMID:21521500

  17. Host-pathogen interactions during coronavirus infection of primary alveolar epithelial cells

    PubMed Central

    Miura, Tanya A.; Holmes, Kathryn V.

    2009-01-01

    Viruses that infect the lung are a significant cause of morbidity and mortality in animals and humans worldwide. Coronaviruses are being associated increasingly with severe diseases in the lower respiratory tract. Alveolar epithelial cells are an important target for coronavirus infection in the lung, and infected cells can initiate innate immune responses to viral infection. In this overview, we describe in vitro models of highly differentiated alveolar epithelial cells that are currently being used to study the innate immune response to coronavirus infection. We have shown that rat coronavirus infection of rat alveolar type I epithelial cells in vitro induces expression of CXC chemokines, which may recruit and activate neutrophils. Although neutrophils are recruited early in infection in several coronavirus models including rat coronavirus. However, their role in viral clearance and/or immune-mediated tissue damage is not understood. Primary cultures of differentiated alveolar epithelial cells will be useful for identifying the interactions between coronaviruses and alveolar epithelial cells that influence the innate immune responses to infection in the lung. Understanding the molecular details of these interactions will be critical for the design of effective strategies to prevent and treat coronavirus infections in the lung. PMID:19638499

  18. Echinococcus multilocularis primary cells: improved isolation, small-scale cultivation and RNA interference.

    PubMed

    Spiliotis, Markus; Mizukami, Chiaki; Oku, Yuzaburo; Kiss, Ferenc; Brehm, Klaus; Gottstein, Bruno

    2010-11-01

    In this study we demonstrate RNA interference mediated knock-down of target gene expression in Echinococcus multilocularis primary cells on both the transcriptional and translational level. In addition, we report on an improved method for generating E. multilocularis primary cell mini-aggregates from in vitro cultivated metacestode vesicles, and on the cultivation of small numbers of small interfering RNA-transfected cells in vitro over an extended period of time. This allows assessments on the effects of RNA interference performed on Echinococcus primary cells with regard to growth, proliferation, differentiation of the parasite and the formation of novel metacestode vesicles in vitro.

  19. Proteasome inhibitors induce apoptosis and reduce viral replication in primary effusion lymphoma cells

    SciTech Connect

    Saji, Chiaki; Higashi, Chizuka; Niinaka, Yasufumi; Yamada, Koji; Noguchi, Kohji; Fujimuro, Masahiro

    2011-12-02

    Highlights: Black-Right-Pointing-Pointer Constitutive NF-{kappa}B signaling is essential for the survival and growth of PEL cells. Black-Right-Pointing-Pointer NF-{kappa}B signaling is upregulated by the proteasome-dependent degradation of I{kappa}B{alpha}. Black-Right-Pointing-Pointer Proteasome inhibitors suppress NF-{kappa}B signaling and induce apoptosis in PEL cells through stabilization of I{kappa}B{alpha}. Black-Right-Pointing-Pointer Proteasome inhibitors suppress viral replication in PEL cells during lytic KSHV infection. -- Abstract: Primary effusion lymphoma (PEL) is an aggressive neoplasm caused by Kaposi's sarcoma-associated herpesvirus (KSHV). This study provides evidence that proteasomal activity is required for both survival of PEL cells stably harboring the KSHV genome and viral replication of KSHV. We evaluated the cytotoxic effects of proteasome inhibitors on PEL cells. The proteasome inhibitors MG132, lactacystin, and proteasome inhibitor I dramatically inhibited cell proliferation and induced apoptosis of PEL cells through the accumulation of p21 and p27. Furthermore, proteasome inhibitors induced the stabilization of NF-{kappa}B inhibitory molecule (I{kappa}B{alpha}) and suppressed the transcriptional activity of NF-{kappa}B in PEL cells. The NF-{kappa}B specific inhibitor BAY11-7082 also induced apoptosis in PEL cells. The constitutive activation of NF-{kappa}B signaling is essential for the survival and growth of B cell lymphoma cells, including PEL cells. NF-{kappa}B signaling is upregulated by proteasome-dependent degradation of I{kappa}B{alpha}. The suppression of NF-{kappa}B signaling by proteasome inhibitors may contribute to the induction of apoptosis in PEL cells. In addition, proteasome activity is required for KSHV replication in KSHV latently infected PEL cells. MG132 reduced the production of progeny virus from PEL cells at low concentrations, which do not affect PEL cell growth. These findings suggest that proteasome inhibitors

  20. Regulation of CDPKs, including identification of PAL kinase, in biotically stressed cells of French bean.

    PubMed

    Allwood, Ellen G; Davies, Dewi R; Gerrish, Chris; Bolwell, G Paul

    2002-07-01

    Changes in protein kinase activity have been investigated during the early response of suspension cultured cells of French bean to fungal elicitor. One of the kinases activated has a known target, phenylalanine ammonia-lyase (PAL), which has an important role in plant defence responses, and was purified. Kinase acivity during purification was monitored for both the PAL-derived peptide and syntide-2, which it also phosphorylated. The kinase had an Mr of 55,000 on the basis of gel migration, 45Ca2+ binding, autophosphorylation and phosphorylation of various substrates using in-gel assays. The kinase has been characterised with respect to kinetics and other properties in vitro and appears to be a CDPK. In-gel assays were also used to show that this kinase and a number of other CDPKs of similar Mr showed complex changes in elicitor-treated suspension-cultured cells of French bean. An activation was observed within 10 min and was maintained for up to 4 h. The time course of activation was different from MAP kinase and casein kinase assayed in the same extracts. However, at 5 min after addition of elicitor there is a transient inactivation of the CDPKs before activation. This inactivation can be mimicked by adding forskolin to the cells 30 min before elicitation, which brings about changes in the cellular pH. Forskolin potentiates the oxidative burst when elicitor is subsequently added while the CDPK cannot be activated by elicitor upon forskolin treatment. In contrast, intracellular acidification brought about by forskolin brings about slight activation of MAPkinase.

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

    PubMed

    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

  2. Current topics of functional links between primary cilia and cell cycle.

    PubMed

    Izawa, Ichiro; Goto, Hidemasa; Kasahara, Kousuke; Inagaki, Masaki

    2015-01-01

    Primary cilia, microtubule-based sensory structures, orchestrate various critical signals during development and tissue homeostasis. In view of the rising interest into the reciprocal link between ciliogenesis and cell cycle, we discuss here several recent advances to understand the molecular link between the individual step of ciliogenesis and cell cycle control. At the onset of ciliogenesis (the transition from centrosome to basal body), distal appendage proteins have been established as components indispensable for the docking of vesicles at the mother centriole. In the initial step of axonemal extension, CP110, Ofd1, and trichoplein, key negative regulators of ciliogenesis, are found to be removed by a kinase-dependent mechanism, autophagy, and ubiquitin-proteasome system, respectively. Of note, their disposal functions as a restriction point to decide that the axonemal nucleation and extension begin. In the elongation step, Nde1, a negative regulator of ciliary length, is revealed to be ubiquitylated and degraded by CDK5-SCF(Fbw7) in a cell cycle-dependent manner. With regard to ciliary length control, it has been uncovered in flagellar shortening of Chlamydomonas that cilia itself transmit a ciliary length signal to cytoplasm. At the ciliary resorption step upon cell cycle re-entry, cilia are found to be disassembled not only by Aurora A-HDAC6 pathway but also by Nek2-Kif24 and Plk1-Kif2A pathways through their microtubule-depolymerizing activity. On the other hand, it is becoming evident that the presence of primary cilia itself functions as a structural checkpoint for cell cycle re-entry. These data suggest that ciliogenesis and cell cycle intimately link each other, and further elucidation of these mechanisms will contribute to understanding the pathology of cilia-related disease including cancer and discovering targets of therapeutic interventions.

  3. Helium Generated Cold Plasma Finely Regulates Activation of Human Fibroblast-Like Primary Cells

    PubMed Central

    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

  4. Study origin of germ cells and formation of new primary follicles in adult human and rat ovaries.

    PubMed

    Bukovsky, Antonin; Gupta, Satish K; Virant-Klun, Irma; Upadhyaya, Nirmala B; Copas, Pleas; Van Meter, Stuart E; Svetlikova, Marta; Ayala, Maria E; Dominguez, Roberto

    2008-01-01

    The central thesis regarding the human ovaries is that, although primordial germ cells in embryonal ovaries are of extraovarian origin, those generated during the fetal period and in postnatal life are derived from the ovarian surface epithelium (OSE) bipotent cells. With the assistance of immune system-related cells, secondary germ cells and primitive granulosa cells originate from OSE stem cells in the fetal and adult human gonads. Fetal primary follicles are formed during the second trimester of intrauterine life, prior to the end of immune adaptation, possibly to be recognized as self-structures and renewed later. With the onset of menarche, a periodical oocyte and follicular renewal emerges to replace aging primary follicles and ensure that fresh eggs for healthy babies are always available during the prime reproductive period. The periodical follicular renewal ceases between 35 and 40 yr of age, and the remaining primary follicles are utilized during the premenopausal period until exhausted. However, the persisting oocytes accumulate genetic alterations and may become unsuitable for ovulation and fertilization. The human OSE stem cells preserve the character of embryonic stem cells, and they may produce distinct cell types, including new eggs in vitro, particularly when derived from patients with premature ovarian failure or aging and postmenopausal ovaries. Our observations also indicate that there are substantial differences in follicular renewal between adult human and rat ovaries. As part of this chapter, we present in detail protocols utilized to analyze oogenesis in humans and to study interspecies differences when compared to the ovaries of rat females.

  5. Primary cutaneous mantle cell lymphoma of the leg with blastoid morphology and aberrant immunophenotype: a diagnostic challenge.

    PubMed

    Cesinaro, Anna Maria; Bettelli, Stefania; Maccio, Livia; Milani, Marina

    2014-02-01

    Mantle cell lymphoma rarely affects the skin and is usually a secondary involvement. The present case illustrates a primary cutaneous mantle cell lymphoma of the leg, with blastoid morphology and aberrant expression of CD10 and bcl-6, which was misinterpreted at the beginning as diffuse large B-cell lymphoma. A larger panel of immunohistochemical markers, including cyclin-D1, and molecular investigation showing the typical translocation (t11;14), pointed toward the correct diagnosis. Cutaneous diffuse B-cell lymphomas with unusual morphology should be interpreted cautiously, and the diagnosis made on the basis of an appropriate panel of antibodies and molecular studies.

  6. Isolation and purification of proteasomes from primary cells.

    PubMed

    Steers, Nicholas J; Peachman, Kristina K; Alving, Carl R; Rao, Mangala

    2014-11-03

    Proteasomes play an important role in cell homeostasis and in orchestrating the immune response by systematically degrading foreign proteins and misfolded or damaged host cell proteins. We describe a protocol to purify functionally active proteasomes from human CD4(+) T cells and dendritic cells derived from peripheral blood mononuclear cells. The purification is a three-step process involving ion-exchange chromatography, ammonium sulfate precipitation, and sucrose density gradient ultracentrifugation. This method can be easily adapted to purify proteasomes from cell lines or from organs. Methods to characterize and visualize the purified proteasomes are also described.

  7. Analysis of cell identity, morphology, apoptosis and mitotic activity in a primary neural cell culture system in Drosophila

    PubMed Central

    2012-01-01

    In Drosophila, most neurogenetic research is carried out in vivo. Mammalian research demonstrates that primary cell culture techniques provide a powerful model to address cell autonomous and non-autonomous processes outside their endogenous environment. We developed a cell culture system in Drosophila using wildtype and genetically manipulated primary neural tissue for long-term observations. We assessed the molecular identity of distinct neural cell types by immunolabeling and genetically expressed fluorescent cell markers. We monitored mitotic activity of cell cultures derived from wildtype and tumorous larval brains. Our system provides a powerful approach to unveil developmental processes in the nervous system and to complement studies in vivo. PMID:22554060

  8. Cortical granule exocytosis in C. elegans is regulated by cell cycle components including separase.

    PubMed

    Bembenek, Joshua N; Richie, Christopher T; Squirrell, Jayne M; Campbell, Jay M; Eliceiri, Kevin W; Poteryaev, Dmitry; Spang, Anne; Golden, Andy; White, John G

    2007-11-01

    In many organisms, cortical granules undergo exocytosis following fertilization, releasing cargo proteins that modify the extracellular covering of the zygote. We identified cortical granules in Caenorhabditis elegans and have found that degranulation occurs in a wave that initiates in the vicinity of the meiotic spindle during anaphase I. Previous studies identified genes that confer an embryonic osmotic sensitivity phenotype, thought to result from abnormal eggshell formation. Many of these genes are components of the cell cycle machinery. When we suppressed expression of several of these genes by RNAi, we observed that cortical granule trafficking was disrupted and the eggshell did not form properly. We conclude that osmotic sensitivity phenotypes occur because of defects in trafficking of cortical granules and the subsequent formation of an impermeable eggshell. We identified separase as a key cell cycle component that is required for degranulation. Separase localized to cortically located filamentous structures in prometaphase I upon oocyte maturation. After fertilization, separase disappeared from these structures and appeared on cortical granules by anaphase I. RNAi of sep-1 inhibited degranulation in addition to causing extensive chromosomal segregation failures. Although the temperature-sensitive sep-1(e2406) allele exhibited similar inhibition of degranulation, it had minimal effects on chromosome segregation. These observations lead us to speculate that SEP-1 has two separable yet coordinated functions: to regulate cortical granule exocytosis and to mediate chromosome separation.

  9. Accurate expressions for solar cell fill factors including series and shunt resistances

    NASA Astrophysics Data System (ADS)

    Green, Martin A.

    2016-02-01

    Together with open-circuit voltage and short-circuit current, fill factor is a key solar cell parameter. In their classic paper on limiting efficiency, Shockley and Queisser first investigated this factor's analytical properties showing, for ideal cells, it could be expressed implicitly in terms of the maximum power point voltage. Subsequently, fill factors usually have been calculated iteratively from such implicit expressions or from analytical approximations. In the absence of detrimental series and shunt resistances, analytical fill factor expressions have recently been published in terms of the Lambert W function available in most mathematical computing software. Using a recently identified perturbative relationship, exact expressions in terms of this function are derived in technically interesting cases when both series and shunt resistances are present but have limited impact, allowing a better understanding of their effect individually and in combination. Approximate expressions for arbitrary shunt and series resistances are then deduced, which are significantly more accurate than any previously published. A method based on the insights developed is also reported for deducing one-diode fits to experimental data.

  10. Differentially expressed epigenome modifiers, including Aurora kinase A and B, in immune cells of rheumatoid arthritis

    PubMed Central

    Glant, Tibor T.; Besenyei, Timea; Kádár, András; Kurkó, Júlia; Tryniszewska, Beata; Gál, János; Soós, Györgyi; Szekanecz, Zoltán; Hoffmann, Gyula; Block, Joel A.; Katz, Robert S.; Mikecz, Katalin; Rauch, Tibor A.

    2014-01-01

    Objective The aim of this study was to identify epigenetic factors that are implicated in the pathogenesis of rheumatoid arthritis (RA) and to explore the therapeutic potential of the targeted inhibition of these factors. Methods PCR arrays were utilized to investigate the expression profile of genes that encod key epigenetic regulator enzymes. Mononuclear cells from RA patients and mice were monitored for gene expression changes, in association with arthritis development in murine models of RA. Selected genes were further characterized by quantitative real-time PCR, Western blot and flow cytometry methods. The targeted inhibition of the upregulated enzymes was studied in arthritic mice. Results A set of genes with arthritis-specific expression was identified by the PCR arrays. Aurora kinase A and B, both of which were highly expressed in arthritic mice and treatment naïve RA patients, were selected for detailed analysis. Elevated Aurora kinase expression was accompanied with an increased phosphorylation of histone H3, which promotes proliferation of T lymphocytes. Treatment with VX-680, a pan-Aurora kinase inhibitor, promoted B cell apoptosis, provided significant protection against the onset, and attenuated the inflammatory reactions in arthritic mice. Conclusions Arthritis development is accompanied the changes in the expression of a number of epigenome-modifying enzymes. Drug-induced downregulation of the Aurora kinases, among other targets, seems to be sufficient to treat experimental arthritis. Development of new therapeutics that target the Aurora kinases can potentially improve RA management. PMID:23653330

  11. Review of isolated ascending aortitis: differential diagnosis, including syphilitic, Takayasu's and giant cell aortitis.

    PubMed

    Tavora, Fabio; Burke, Allen

    2006-08-01

    The image of tree-barking and proximal aortic root dilatation is firmly entrenched in the minds of practising pathologists as representing syphilis until proven otherwise. We discuss the differential diagnosis of syphilitic aortitis, Takayasu's disease, and giant cell aortitis, with a review of the literature and brief overview of other types of aortitis. As a starting point, we report a case of non-specific, or idiopathic, aortitis with aneurysm that was initially misdiagnosed as syphilitic aortitis. We then review the literature and emphasise the lack of histological data and histopathological criteria for the diagnosis of non-infectious aortitis and the implications for treatment in cases of isolated aortitis. Tree-barking is a non-specific finding in aortitis of any aetiology, and syphilitic aortitis in developed countries is rare. It is still unclear if there are histological features that separate Takayasu's disease and giant cell arteritis. In the majority of patients presenting with aortic root aneurysms, aortitis is an isolated finding not associated with autoimmune disease. Despite a plethora of literature, a histological classification of aortitis has yet to be attempted.

  12. Consensus for nonmelanoma skin cancer treatment: basal cell carcinoma, including a cost analysis of treatment methods.

    PubMed

    Kauvar, Arielle N B; Cronin, Terrence; Roenigk, Randall; Hruza, George; Bennett, Richard

    2015-05-01

    Basal cell carcinoma (BCC) is the most common cancer in the US population affecting approximately 2.8 million people per year. Basal cell carcinomas are usually slow-growing and rarely metastasize, but they do cause localized tissue destruction, compromised function, and cosmetic disfigurement. To provide clinicians with guidelines for the management of BCC based on evidence from a comprehensive literature review, and consensus among the authors. An extensive review of the medical literature was conducted to evaluate the optimal treatment methods for cutaneous BCC, taking into consideration cure rates, recurrence rates, aesthetic and functional outcomes, and cost-effectiveness of the procedures. Surgical approaches provide the best outcomes for BCCs. Mohs micrographic surgery provides the highest cure rates while maximizing tissue preservation, maintenance of function, and cosmesis. Mohs micrographic surgery is an efficient and cost-effective procedure and remains the treatment of choice for high-risk BCCs and for those in cosmetically sensitive locations. Nonsurgical modalities may be used for low-risk BCCs when surgery is contraindicated or impractical, but the cure rates are lower.

  13. T cell reactions of Eimeria bovis primary and challenge-infected calves.

    PubMed

    Sühwold, Anke; Hermosilla, Carlos; Seeger, Torsten; Zahner, Horst; Taubert, Anja

    2010-02-01

    Eimeria bovis infections commonly have clinical impact only on young animals, as homologous reinfections generally are under immunological control. So far, the nature of the immune responses delivering protection to calves has not been investigated. In this study we therefore analysed local and peripheral proliferative T cell activities of primary and challenge-infected calves and investigated the occurrence of T cell phenotypes in the peripheral blood and in mucosal gut segments isolated either by bioptic means or by necropsies.We show that lymphocytes of E. bovis-infected calves exhibit effective, transient antigen-specific proliferative responses in the course of prepatency of primary infection but fail to react after homologous reinfection suggesting early abrogation of parasite development. Whilst in primary infection an expansion of peripheral CD4+ T cells was observed, reinfection had no effect on the proportions of CD4+, CD8+ subsets or gammadeltaTCR+ T cells. In contrast, both E. bovis primary and challenge infections had an impact on local tissue T cell distribution. Primary infection was characterised by a CD4+ T cell infiltration early in prepatency in ileum and later in colon mucosa, whereas CD8+ T cells were only found accumulating in the latter gut segment. Challenge infection led to infiltration of both CD4+ and CD8+ T cells in small intestine and large intestine segments indicating protective functions of both cell types. In contrast, infiltration of ileum and colon mucosa with gammadeltaTCR+ T cells was restricted to primary infection.

  14. Interactions between MUR10/CesA7-dependent secondary cellulose biosynthesis and primary cell wall structure.

    PubMed

    Bosca, Sonia; Barton, Christopher J; Taylor, Neil G; Ryden, Peter; Neumetzler, Lutz; Pauly, Markus; Roberts, Keith; Seifert, Georg J

    2006-12-01

    Primary cell walls are deposited and remodeled during cell division and expansion. Secondary cell walls are deposited in specialized cells after the expansion phase. It is presently unknown whether and how these processes are interrelated. The Arabidopsis (Arabidopsis thaliana) MUR10 gene is required for normal primary cell wall carbohydrate composition in mature leaves as well as for normal plant growth, hypocotyl strength, and fertility. The overall sugar composition of young mur10 seedlings is not significantly altered; however, the relative proportion of pectin side chains is shifted toward an increase in 1 --> 5-alpha-arabinan relative to 1 --> 4-beta-galactan. mur10 seedlings display reduced fucogalactosylation of tightly cell wall-bound xyloglucan. Expression levels of genes encoding either nucleotide sugar interconversion enzymes or glycosyl transferases, known to be involved in primary and secondary cell wall biosynthesis, are generally unaffected; however, the CesA7 transcript is specifically suppressed in the mur10-1 allele. The MUR10 locus is identical with the CesA7 gene, which encodes a cellulose catalytic subunit previously thought to be specifically involved in secondary cell wall formation. The xylem vessels in young mur10 hypocotyls are collapsed and their birefringence is lost. Moreover, a fucogalactosylated xyloglucan epitope is reduced and a 1 --> 5-alpha-arabinan epitope increased in every cell type in mur10 hypocotyls, including cells that do not deposit secondary walls. mur10 also displays altered distribution of an arabinogalactan-protein epitope previously associated with xylem differentiation and secondary wall thickening. This work indicates the existence of a mechanism that senses secondary cell wall integrity and controls biosynthesis or structural remodeling of primary cell walls and cellular differentiation.

  15. Method of preparing a negative electrode including lithium alloy for use within a secondary electrochemical cell

    DOEpatents

    Tomczuk, Zygmunt; Olszanski, Theodore W.; Battles, James E.

    1977-03-08

    A negative electrode that includes a lithium alloy as active material is prepared by briefly submerging a porous, electrically conductive substrate within a melt of the alloy. Prior to solidification, excess melt can be removed by vibrating or otherwise manipulating the filled substrate to expose interstitial surfaces. Electrodes of such as solid lithium-aluminum filled within a substrate of metal foam are provided.

  16. Primary signet ring cell carcinoma of gall bladder: report of an extremely rare histological type of primary gall bladder carcinoma.

    PubMed

    Ahmad, Zubair; Qureshi, Asim

    2010-09-07

    Signet ring cell carcinoma is an extremely rare type of gall bladder carcinoma composed overwhelmingly (90%) of signet ring cells. It is necessary to exclude a gastric or colonic signet ring cell carcinoma secondarily involving the gall bladder. The primary aim of this case report is to describe the histopathological aspects of this tumour. Primary signet ring cell carcinoma of gall bladder shows dysplastic surface gall bladder epithelium with infiltration of gall bladder wall. It is also necessary to exclude benign signet ring change, which sometimes occurs in the gall bladder. However, it is always confined to the mucosa and does not infiltrate the wall. This case showed grossly diffuse thickening of the gall bladder wall and dysplastic surface epithelium of the gall bladder on histology, with sheets of signet ring cells infiltrating full thickness of the wall. It is also necessary to exclude benign signet ring cell change, which sometimes occurs in the gall bladder. However it is always confined to the mucosa and does not infiltrate the wall.

  17. T cell targeting and phagocytosis of apoptotic biliary epithelial cells in primary biliary cirrhosis.

    PubMed

    Allina, Jorge; Hu, Bin; Sullivan, Daniel M; Fiel, Maria Isabel; Thung, Swan N; Bronk, Steven F; Huebert, Robert C; van de Water, Judy; LaRusso, Nicholas F; Gershwin, M E; Gores, Gregory J; Odin, Joseph A

    2006-12-01

    Primary biliary cirrhosis (PBC) is characterized by loss of tolerance against ubiquitously expressed mitochondrial autoantigens followed by biliary and salivary gland epithelial cell (BEC and SGEC) destruction by autoreactive T cells. It is unclear why BECs and SGECs are targeted. Previous work demonstrated that the reduced form of the major PBC autoantigen predominated in apoptotic BECs and SGECs as opposed to an oxidized form in other apoptotic cells. This led to the hypothesis that presentation of novel self-peptides from phagocytosed apoptotic BECs might contribute to BEC targeting by autoreactive T cells. The effect of autoantigen redox status on self-peptide formation was examined along with the phagocytic ability of BECs. Oxidation of PBC autoantigens first was shown to be due to protein S-glutathionylation of lipoyllysine residues. Absence of protein S-glutathionylation generated novel self-peptides and affected T cell recognition of a lipoyllysine containing peptide. Liver biopsy staining revealed BEC phagocytosis of apoptotic BECs (3.74+/-2.90% of BEC) was present in PBC (7 of 7 cases) but not in normal livers (0 of 3). BECs have the ability to present novel mitochondrial self-peptides derived from phagocytosed apoptotic BECs. Apoptotic cell phagocytosis by non-professional phagocytes may influence the tissue specificity of autoimmune diseases.

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

    PubMed

    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.

  19. Generation of mouse and human induced pluripotent stem cells (iPSC) from primary somatic cells.

    PubMed

    Lorenzo, I M; Fleischer, A; Bachiller, D

    2013-08-01

    Cellular reprogramming consists of the conversion of differentiated cells into pluripotent cells; the so-called induced Pluripotent Stem Cells. iPSC are amenable to in vitro manipulation and, in theory, direct production of any differentiated cell type. Furthermore, iPSC can be obtained from sick individuals and subsequently used for disease modeling, drug discovery and regenerative treatments. iPSC production was first achieved by transducing, with the use of retroviral vectors, four specific transcription factors: Oct4, Klf4, Sox2 and c-Myc (OKSM), into primary cells in culture Takahashi and Yamanaka, (Cell 126(4):663-676, 2006). Many alternative protocols have since been proposed: repeated transfections of expression plasmids containing the four pluripotency-associated genes Okita et al. (Science 322(5903):949-953, 2008), lentiviral delivery of the four factors Sommer et al. (Stem Cells 27(3):543-549, 2009), Sendai virus delivery Fusaki et al. (Proceedings of the Japan Academy. Series B, Physical and Biological Sciences 85(8):348-362, 2009), removal of the reprogramming vectors by 'piggyBac' transposition Woltjen et al. (Nature 458(7239):766-770, 2009); Kaji et al. (Nature 458(7239):771-775, 2009), Cre-recombinase excisable viruses Soldner et al. (Cell 136(5):964-977, 2009), episomal vectors Yu et al. (Science 324(5928):797-801, 2009), cell-penetrating reprogramming proteins Zhou et al. (Stem Cells 4(5):381-384, 2009), mammalian artificial chromosomes Hiratsuka et al. (PLoS One 6(10):e25961, 2011) synthetically modified mRNAs Warren et al. (Scientific Reports 2:657, 2012), miRNA Anokye-Danso et al. (Cell Stem Cell 8(4):376-388, 2009); however, although some of these methods are commercially available, in general they still need to attain the reproducibility and reprogramming efficiency required for routine applications Mochiduki and Okita (Biotechnol Journal 7(6):789-797, 2012). Herein we explain, in four detailed protocols, the isolation of mouse and human

  20. Carcinogens induce loss of the primary cilium in human renal proximal tubular epithelial cells independently of effects on the cell cycle

    PubMed Central

    Radford, Robert; Slattery, Craig; Jennings, Paul; Blacque, Oliver; Pfaller, Walter; Gmuender, Hans; Van Delft, Joost; Ryan, Michael P.

    2012-01-01

    The primary cilium is an immotile sensory and signaling organelle found on the majority of mammalian cell types. Of the multitude of roles that the primary cilium performs, perhaps some of the most important include maintenance of differentiation, quiescence, and cellular polarity. Given that the progression of cancer requires disruption of all of these processes, we have investigated the effects of several carcinogens on the primary cilium of the RPTEC/TERT1 human proximal tubular epithelial cell line. Using both scanning electron microscopy and immunofluorescent labeling of the ciliary markers acetylated tubulin and Arl13b, we confirmed that RPTEC/TERT1 cells express primary cilium upon reaching confluence. Treatment with the carcinogens ochratoxin A (OTA) and potassium bromate (KBrO3) caused a significant reduction in the number of ciliated cells, while exposure to nifedipine, a noncarcinogenic renal toxin, had no effect on primary cilium expression. Flow cytometric analysis of the effects of all three compounds on the cell cycle revealed that only KBrO3 resulted in an increase in the proportion of cells entering the cell cycle. Microarray analysis revealed dysregulation of multiple pathways affecting ciliogenesis and ciliary maintenance following OTA and KBrO3 exposure, which were unaffected by nifedipine exposure. The primary cilium represents a unique physical checkpoint with relevance to carcinogenesis. We have shown that the renal carcinogens OTA and KBrO3 cause significant deciliation in a model of the proximal tubule. With KBrO3, this was followed by reentry into the cell cycle; however, deciliation was not found to be associated with reentry into the cell cycle following OTA exposure. Transcriptomic analysis identified dysregulation of Wnt signaling and ciliary trafficking in response to OTA and KBrO3 exposure. PMID:22262483

  1. Carcinogens induce loss of the primary cilium in human renal proximal tubular epithelial cells independently of effects on the cell cycle.

    PubMed

    Radford, Robert; Slattery, Craig; Jennings, Paul; Blacque, Oliver; Blaque, Oliver; Pfaller, Walter; Gmuender, Hans; Van Delft, Joost; Ryan, Michael P; McMorrow, Tara

    2012-04-15

    The primary cilium is an immotile sensory and signaling organelle found on the majority of mammalian cell types. Of the multitude of roles that the primary cilium performs, perhaps some of the most important include maintenance of differentiation, quiescence, and cellular polarity. Given that the progression of cancer requires disruption of all of these processes, we have investigated the effects of several carcinogens on the primary cilium of the RPTEC/TERT1 human proximal tubular epithelial cell line. Using both scanning electron microscopy and immunofluorescent labeling of the ciliary markers acetylated tubulin and Arl13b, we confirmed that RPTEC/TERT1 cells express primary cilium upon reaching confluence. Treatment with the carcinogens ochratoxin A (OTA) and potassium bromate (KBrO(3)) caused a significant reduction in the number of ciliated cells, while exposure to nifedipine, a noncarcinogenic renal toxin, had no effect on primary cilium expression. Flow cytometric analysis of the effects of all three compounds on the cell cycle revealed that only KBrO(3) resulted in an increase in the proportion of cells entering the cell cycle. Microarray analysis revealed dysregulation of multiple pathways affecting ciliogenesis and ciliary maintenance following OTA and KBrO(3) exposure, which were unaffected by nifedipine exposure. The primary cilium represents a unique physical checkpoint with relevance to carcinogenesis. We have shown that the renal carcinogens OTA and KBrO(3) cause significant deciliation in a model of the proximal tubule. With KBrO(3), this was followed by reentry into the cell cycle; however, deciliation was not found to be associated with reentry into the cell cycle following OTA exposure. Transcriptomic analysis identified dysregulation of Wnt signaling and ciliary trafficking in response to OTA and KBrO(3) exposure.

  2. Primary Cutaneous Lymphoma-Associated Pseudoepitheliomatous Hyperplasia Masquerading as Squamous Cell Carcinoma in a Young Adult.

    PubMed

    Ansari, Mahsa; Azmoodeh Ardalan, Farid; Najafi, Masoumeh; Goodarzi, Azadeh; Ghanadan, Alireza

    2015-12-01

    Primary cutaneous anaplastic large cell lymphoma is a T-cell malignancy with atypical CD30 positive lymphocytes. Pseudoepitheliomatous hyperplasia is an uncommon finding in primary cutaneous anaplastic large cell lymphoma, and may mimic squamous cell carcinoma as pseudomalignancy. Careful attention of a pathologist to correct diagnosis of pseudoepitheliomatous hyperplasia and its underlying causes will help physicians to avoid inappropriate management. Here, we present a 22-year-old man referred to our hospital with a solitary nodule persistent on his forearm which was diagnosed as squamous cell carcinoma in the first biopsy. The lesion recurred after two months and histopathologic and immunohistochemistry examination revealed anaplastic large cell lymphoma with florid pseudoepitheliomatous hyperplasia which masquerading as well-differentiated squamous cell carcinoma. Diagnosis of pseudoepitheliomatous hyperplasia must guide the pathologist to search for underlying causes, such as primary cutaneous lymphoma. Pseudoepitheliomatous hyperplasia may mimic squamous cell carcinoma and this can result in inappropriate diagnosis and management.

  3. Intracellular trafficking mechanism of cationic phospholipids including cationic liposomes in HeLa cells.

    PubMed

    Un, K; Sakai-Kato, K; Goda, Y

    2014-07-01

    The development of gene delivery methods is essential for the achievement of effective gene therapy. Elucidation of the intracellular transfer mechanism for cationic carriers is in progress, but there are few reports regarding the intracellular trafficking processes of the cationic phospholipids taken up into cells. In the present work, the trafficking processes of a cationic phospholipid (1,2-dioleoyl-3-trimethylammonium-propane, DOTAP) were investigated from intracellular uptake to extracellular efflux using cationic liposomes in vitro. Following intracellular transport of liposomes via endocytosis, DOTAP was localized in the endoplasmic reticulum, Golgi apparatus, and mitochondria. Moreover, the proteins involved in DOTAP intracellular trafficking and extracellular efflux were identified. In addition, helper lipids of cationic liposomes were found to partially affect this intracellulartrafficking. These findings might provide valuable information for designing cationic carriers and avoiding unexpected toxic side effects derived from cationic liposomal components.

  4. Electrodes including a polyphosphazene cyclomatrix, methods of forming the electrodes, and related electrochemical cells

    SciTech Connect

    Gering, Kevin L; Stewart, Frederick F; Wilson, Aaron D; Stone, Mark L

    2014-10-28

    An electrode comprising a polyphosphazene cyclomatrix and particles within pores of the polyphosphazene cyclomatrix. The polyphosphazene cyclomatrix comprises a plurality of phosphazene compounds and a plurality of cross-linkages. Each phosphazene compound of the plurality of phosphazene compounds comprises a plurality of phosphorus-nitrogen units, and at least one pendant group bonded to each phosphorus atom of the plurality of phosphorus-nitrogen units. Each phosphorus-nitrogen unit is bonded to an adjacent phosphorus-nitrogen unit. Each cross-linkage of the plurality of cross-linkages bonds at least one pendant group of one phosphazene compound of the plurality of phosphazene compounds with the at least one pendant group of another phosphazene compound of the plurality of phosphazene compounds. A method of forming a negative electrode and an electrochemical cell are also described.

  5. Reciprocal regulation of the primary sodium absorptive pathways in rat intestinal epithelial cells

    PubMed Central

    Coon, Steven; Kekuda, Ramesh; Saha, Prosenjit

    2011-01-01

    Sodium absorption in the mammalian small intestine occurs predominantly by two primary pathways that include Na/H exchange (NHE3) and Na-glucose cotransport (SGLT1) on the brush border membrane (BBM) of villus cells. However, whether NHE3 and SGLT1 function together to regulate intestinal sodium absorption is unknown. Nontransformed small intestinal epithelial cells (IEC-18) were transfected with either NHE3 or SGLT1 small interfering RNAs (siRNAs) and were grown in confluent monolayers on transwell plates to measure the effects on Na absorption. Uptake studies were performed as well as molecular studies to determine the effects on NHE3 and SGLT1 activity. When IEC-18 monolayers were transfected with silencing NHE3 RNA, the cells demonstrated decreased NHE3 activity as well as decreased NHE3 mRNA and protein. However, in NHE3 siRNA-transected cells, SGLT1 activity, mRNA, and protein in the BBM were significantly increased. Thus, inhibition of NHE3 expression regulates the expression and function of SGLT1 in the BBM of intestinal epithelial cells. In addition, IEC-18 cells transected with silencing SGLT1 RNA demonstrated an inhibition of Na-dependent glucose uptake and a decrease in SGLT1 activity, mRNA, and protein levels. However, in these cells, Na/H exchange activity was significantly increased. Furthermore, NHE3 mRNA and protein levels were also increased. Therefore, the inhibition of SGLT1 expression stimulates the transcription and function of NHE3 and vice versa in the BBM of intestinal epithelial cells. Thus this study demonstrates that the major sodium absorptive pathways together function to regulate sodium absorption in epithelial cells. PMID:21148403

  6. Intralesional lymphokine-activated killer cells as adjuvant therapy for primary glioblastoma.

    PubMed

    Dillman, Robert Owen; Duma, Christopher Michael; Ellis, Robin Anne; Cornforth, Andrew Nimitz; Schiltz, Patric Michael; Sharp, Shari Lynn; DePriest, Madeline Carol

    2009-01-01

    Despite recent advances, median survival for patients with resectable glioblastoma multiforme (GBM) is only 12 to 15 months. We previously observed minimal toxicity and a 9.0-month median survival after treatment with intralesional autologous lymphokine-activated killer (LAK) cells in 40 patients with recurrent GBM. In this study, GBM patients were treated with adjuvant intralesional LAK cells. Eligible patients had completed primary therapy for GBM without disease progression. LAK cells were produced by incubating autologous peripheral blood mononuclear cells with interleukin-2 for 3 to 7 days and then placed into the surgically exposed tumor cavity by a neurosurgeon. The 19 men and 14 women had a median age of 57 years. Prior therapy included surgical resection (97%), partial brain irradiation (97%), gamma knife radiosurgery (97%), and temozolomide chemotherapy (70%). Median time from diagnosis to LAK cell therapy was 5.3 months (range: 3.0 to 11.1 mo). LAK cell treatment was well tolerated; average length of hospitalization was 3 days. At the time of this analysis, 27 patients have died; the median survival from the date of original diagnosis is 20.5 months with a 1-year survival rate of 75%. In subset analyses, superior survival was observed for patients who received higher numbers of CD3+/CD16+/CD56+ (T-LAK) cells in the cell products, which was associated with not taking corticosteroids in the month before leukopheresis. Intralesional LAK cell therapy is safe and the survival sufficiently encouraging to warrant further evaluation in a randomized phase 2 trial of intralesional therapies with LAK or carmustine-impregnated wafers.

  7. Pelvic floor muscle training included in a pregnancy exercise program is effective in primary prevention of urinary incontinence: a randomized controlled trial.

    PubMed

    Pelaez, Mireia; Gonzalez-Cerron, Silvia; Montejo, Rocío; Barakat, Rubén

    2014-01-01

    To investigate the effect of pelvic floor muscle training (PFMT) taught in a general exercise class during pregnancy on the prevention of urinary incontinence (UI) in nulliparous continent pregnant women. This was a unicenter two armed randomized controlled trial. One hundred sixty-nine women were randomized by a central computer system to an exercise group (EG) (exercise class including PFMT) (n = 73) or a control group (CG) (n = 96). 10.1% loss to follow-up: 10 from EG and 7 from CG. The intervention consisted of 70-75 sessions (22 weeks, three times per week, 55-60 min/session including 10 min of PFMT). The CG received usual care (which included follow up by midwifes including information about PFMT). Questions on prevalence and degree of UI were posed before (week 10-14) and after intervention (week 36-39) using the International Consultation on Incontinence Questionnaire-Urinary Incontinence Short Form (ICIQ-UI SF). At the end of the intervention, there was a statistically significant difference in favor of the EG. Reported frequency of UI [Never: CG: 54/60.7%, EG: 60/95.2% (P < 0.001)]. Amount of leakage [None: CG: 45/60.7%, EG: 60/95.2% (P < 0.001)]. There was also a statistically significant difference in ICIQ-UI SF Score between groups after the intervention period [CG: 2.7 (SD 4.1), EG: 0.2 (SD 1.2) (P < 0.001)]. The estimated effect size was 0.8. PFMT taught in a general exercise class three times per week for at least 22 weeks, without former assessment of ability to perform a correct contraction was effective in primary prevention of UI in primiparous pregnant women. © 2013 Wiley Periodicals, Inc.

  8. Method of preparing a negative electrode including lithium alloy for use within a secondary electrochemical cell

    DOEpatents

    Tomczuk, Z.; Olszanski, W.; Battles, J.E.

    1975-12-09

    A negative electrode that includes a lithium alloy as active material is prepared by briefly submerging a porous, electrically conductive substrate within a melt of the alloy. Prior to solidification, excess melt can be removed by vibrating or otherwise manipulating the filled substrate to expose interstitial surfaces. Electrodes of such a solid lithium--aluminum filled within a substrate of metal foam are provided. 1 figure, 1 table.

  9. Primary plasma cell leukemia 2.0: advances in biology and clinical management.

    PubMed

    Neri, Antonino; Todoerti, Katia; Lionetti, Marta; Simeon, Vittorio; Barbieri, Marzia; Nozza, Filomena; Vona, Gabriella; Pompa, Alessandra; Baldini, Luca; Musto, Pellegrino

    2016-11-01

    Primary plasma cell leukemia (PPCL) is a rare and aggressive variant of multiple myeloma. The introduction of novel agents and modern technologies has recently partially changed the clinical and biological scenario of this malignancy, allowing limited, but not negligible, progresses. Areas covered: We will discuss: the complex landscape of genetic alterations in PPCL, derived from conventional and high-throughput technologies; the best available treatments for PPCL; the possible future therapeutic perspectives. Expert commentary: PPCL requires an immediate and intensive multi-phase treatment with short therapy-free intervals, which should include novel agents and autologous stem cell transplantation in eligible patients. Allogeneic transplantation should be considered in selected cases. In older and/or frailer individuals, personalized approaches should be applied. Integrated treatments with next generation proteasome inhibitors/IMIDs and monoclonal antibodies are currently planned or under investigation. The identification of novel genomic biomarkers may be potentially helpful for risk stratification and future personalized therapies.

  10. Pediatric medulloblastoma xenografts including molecular subgroup 3 and CD133+ and CD15+ cells are sensitive to killing by oncolytic herpes simplex viruses

    PubMed Central

    Friedman, Gregory K.; Moore, Blake P.; Nan, Li; Kelly, Virginia M.; Etminan, Tina; Langford, Catherine P.; Xu, Hui; Han, Xiaosi; Markert, James M.; Beierle, Elizabeth A.; Gillespie, G. Yancey

    2016-01-01

    Background Childhood medulloblastoma is associated with significant morbidity and mortality that is compounded by neurotoxicity for the developing brain caused by current therapies, including surgery, craniospinal radiation, and chemotherapy. Innate therapeutic resistance of some aggressive pediatric medulloblastoma has been attributed to a subpopulation of cells, termed cancer-initiating cells or cancer stemlike cells (CSCs), marked by the surface protein CD133 or CD15. Brain tumors characteristically contain areas of pathophysiologic hypoxia, which has been shown to drive the CSC phenotype leading to heightened invasiveness, angiogenesis, and metastasis. Novel therapies that target medulloblastoma CSCs are needed to improve outcomes and decrease toxicity. We hypothesized that oncolytic engineered herpes simplex virus (oHSV) therapy could effectively infect and kill pediatric medulloblastoma cells, including CSCs marked by CD133 or CD15. Methods Using 4 human pediatric medulloblastoma xenografts, including 3 molecular subgroup 3 tumors, which portend worse patient outcomes, we determined the expression of CD133, CD15, and the primary HSV-1 entry molecule nectin-1 (CD111) by fluorescence activated cell sorting (FACS) analysis. Infectability and cytotoxicity of clinically relevant oHSVs (G207 and M002) were determined in vitro and in vivo by FACS, immunofluorescent staining, cytotoxicity assays, and murine survival studies. Results We demonstrate that hypoxia increased the CD133+ cell fraction, while having the opposite effect on CD15 expression. We established that all 4 xenografts, including the CSCs, expressed CD111 and were highly sensitive to killing by G207 or M002. Conclusions Pediatric medulloblastoma, including Group 3 tumors, may be an excellent target for oHSV virotherapy, and a clinical trial in medulloblastoma is warranted. PMID:26188016

  11. Perception of primary care doctors and nurses about care provided to sickle cell disease patients

    PubMed Central

    Xavier Gomes, Ludmila Mourão; de Andrade Barbosa, Thiago Luis; Souza Vieira, Elen Débora; Caldeira, Antônio Prates; de Carvalho Torres, Heloísa; Viana, Marcos Borato

    2015-01-01

    Objective To analyze the perception of primary care physicians and nurses about access to services and routine health care provided to sickle cell disease patients. Methods This descriptive exploratory study took a qualitative approach by surveying thirteen primary care health professionals who participated in a focus group to discuss access to services and assistance provided to sickle cell disease patients. The data were submitted to thematic content analysis. Results Access to primary care services and routine care for sickle cell disease patients were the categories that emerged from the analysis. Interaction between people with sickle cell disease and primary care health clinics was found to be minimal and limited mainly to scheduling appointments. Patients sought care from the primary care health clinics only in some situations, such as for pain episodes and vaccinations. The professionals noted that patients do not recognize primary care as the gateway to the system, and reported that they feel unprepared to assist sickle cell disease patients. Conclusion In the perception of these professionals, there are restrictions to accessing primary care health clinics and the primary care assistance for sickle cell disease patients is affected. PMID:26190428

  12. The Cellulosome System of Acetivibrio cellulolyticus Includes a Novel Type of Adaptor Protein and a Cell Surface Anchoring Protein

    PubMed Central

    Xu, Qi; Gao, Wenchen; Ding, Shi-You; Kenig, Rina; Shoham, Yuval; Bayer, Edward A.; Lamed, Raphael

    2003-01-01

    A scaffoldin gene cluster was identified in the mesophilic cellulolytic anaerobe Acetivibrio cellulolyticus. The previously described scaffoldin gene, cipV, encodes an N-terminal family 9 glycoside hydrolase, a family 3b cellulose-binding domain, seven cohesin domains, and a C-terminal dockerin. The gene immediately downstream of cipV was sequenced and designated scaB. The protein encoded by this gene has 942 amino acid residues and a calculated molecular weight of 100,358 and includes an N-terminal signal peptide, four type II cohesions, and a C-terminal dockerin. ScaB cohesins 1 and 2 are very closely linked. Similar, but not identical, 39-residue Thr-rich linker segments separate cohesin 2 from cohesin 3 and cohesin 3 from cohesin 4, and an 84-residue Thr-rich linker connects the fourth cohesin to a C-terminal dockerin. The scaC gene downstream of scaB codes for a 1,237-residue polypeptide that includes a signal peptide, three cohesins, and a C-terminal S-layer homology (SLH) module. A long, ca. 550-residue linker separates the third cohesin and the SLH module of ScaC and is characterized by an 18-residue Pro-Thr-Ala-Ser-rich segment that is repeated 27 times. The calculated molecular weight of the mature ScaC polypeptide (excluding the signal peptide) is 124,162. The presence of the cohesins and the conserved SLH module implies that ScaC acts as an anchoring protein. The ScaC cohesins are on a separate branch of the phylogenetic tree that is close to, but distinct from, the type I cohesins. Affinity blotting with representative recombinant probes revealed the following specific intermodular interactions: (i) an expressed CipV cohesin binds selectively to an enzyme-borne dockerin, (ii) a representative ScaB cohesin binds to the CipV band of the cell-free supernatant fraction, and (iii) a ScaC cohesin binds to the ScaB dockerin. The experimental evidence thus indicates that CipV acts as a primary (enzyme-recognizing) scaffoldin, and the protein was also

  13. Distinct CD4 T-cell effects on primary versus recall CD8 T-cell responses during viral encephalomyelitis.

    PubMed

    Hwang, Mihyun; Phares, Timothy W; Hinton, David R; Stohlman, Stephen A; Bergmann, Cornelia C; Min, Booki

    2015-03-01

    CD4 T-cell help is not a universal requirement for effective primary CD8 T cells but is essential to generate memory CD8 T cells capable of recall responses. This study examined how CD4 T cells affect primary and secondary anti-viral CD8 T-cell responses within the central nervous system (CNS) during encephalomyelitis induced by sublethal gliatropic coronavirus. CD4 T-cell depletion before infection did not impair peripheral expansion, interferon-γ production, CNS recruitment or initial CNS effector capacity of virus-specific CD8 T cells ex vivo. Nevertheless, impaired virus control in the absence of CD4 T cells was associated with gradually diminished CNS CD8 T-cell interferon-γ production. Furthermore, within the CD8 T-cell population short-lived effector cells were increased and memory precursor effector cells were significantly decreased, consistent with higher T-cell turnover. Transfer of memory CD8 T cells to reduce viral load in CD4-depleted mice reverted the recipient CNS CD8 T-cell phenotype to that in wild-type control mice. However, memory CD8 T cells primed without CD4 T cells and transferred into infected CD4-sufficient recipients expanded less efficiently and were not sustained in the CNS, contrasting with their helped counterparts. These data suggest that CD4 T cells are dispensable for initial expansion, CNS recruitment and differentiation of primary resident memory CD8 T cells as long as the duration of antigen exposure is limited. By contrast, CD4 T cells are essential to prolong primary CD8 T-cell function in the CNS and imprint memory CD8 T cells for recall responses. © 2014 John Wiley & Sons Ltd.

  14. Distinct CD4 T-cell effects on primary versus recall CD8 T-cell responses during viral encephalomyelitis

    PubMed Central

    Hwang, Mihyun; Phares, Timothy W; Hinton, David R; Stohlman, Stephen A; Bergmann, Cornelia C; Min, Booki

    2015-01-01

    CD4 T-cell help is not a universal requirement for effective primary CD8 T cells but is essential to generate memory CD8 T cells capable of recall responses. This study examined how CD4 T cells affect primary and secondary anti-viral CD8 T-cell responses within the central nervous system (CNS) during encephalomyelitis induced by sublethal gliatropic coronavirus. CD4 T-cell depletion before infection did not impair peripheral expansion, interferon-γ production, CNS recruitment or initial CNS effector capacity of virus-specific CD8 T cells ex vivo. Nevertheless, impaired virus control in the absence of CD4 T cells was associated with gradually diminished CNS CD8 T-cell interferon-γ production. Furthermore, within the CD8 T-cell population short-lived effector cells were increased and memory precursor effector cells were significantly decreased, consistent with higher T-cell turnover. Transfer of memory CD8 T cells to reduce viral load in CD4-depleted mice reverted the recipient CNS CD8 T-cell phenotype to that in wild-type control mice. However, memory CD8 T cells primed without CD4 T cells and transferred into infected CD4-sufficient recipients expanded less efficiently and were not sustained in the CNS, contrasting with their helped counterparts. These data suggest that CD4 T cells are dispensable for initial expansion, CNS recruitment and differentiation of primary resident memory CD8 T cells as long as the duration of antigen exposure is limited. By contrast, CD4 T cells are essential to prolong primary CD8 T-cell function in the CNS and imprint memory CD8 T cells for recall responses. PMID:25187405

  15. Proton NMR characterization of intact primary and metastatic melanoma cells in 2D & 3D cultures.

    PubMed

    Ramachandran, Gokula Krishnan; Yeow, Chen Hua

    2017-03-16

    To characterize the differences between the primary and metastatic melanoma cell lines grown in 2D cultures and 3D cultures. Primary melanoma cells (WM115) and metastatic melanoma cells (WM266) extracted from a single donor was cultured in 2D as well as 3D cultures. These cells were characterized using proton NMR spectrometry, and the qualitative chemical shifts markers were identified and discussed. In monolayer culture (2D), we observed one qualitative chemical shift marker for primary melanoma cells. In spheroid cultures (3D), we observed nine significant chemical shifts, of which eight markers were specific for primary melanoma spheroids, whereas the other one marker was specific to metastatic melanoma spheroids. This study suggests that the glucose accumulation and phospholipid composition vary significantly between the primary and metastatic cells lines that are obtained from a single donor and also with the cell culturing methods. 14 qualitative chemical shift markers were obtained in the comparison between monolayer culture and spheroids cultures irrespective of the differences in the cell lines. Among which 4 were unique to monolayer cultures whereas 10 chemical shifts were unique to the spheroid cultures. This study also shows that the method of cell culture would drastically affect the phospholipid composition of the cells and also depicts that the cells in spheroid culture closely resembles the cells in vivo. This study shows the high specificity of proton NMR spectrometry in characterizing cancer cell lines and also shows the variations in the glucose accumulation and phospholipid composition between the primary and metastatic melanoma cell lines from the same donor. Differences in the cell culture method does plays an important role in phospholipid composition of the cells.

  16. Potato Snakin-1 Gene Silencing Affects Cell Division, Primary Metabolism, and Cell Wall Composition1[W

    PubMed Central

    Nahirñak, Vanesa; Almasia, Natalia Inés; Fernandez, Paula Virginia; Hopp, Horacio Esteban; Estevez, José Manuel; Carrari, Fernando; Vazquez-Rovere, Cecilia

    2012-01-01

    Snakin-1 (SN1) is an antimicrobial cysteine-rich peptide isolated from potato (Solanum tuberosum) that was classified as a member of the Snakin/Gibberellic Acid Stimulated in Arabidopsis protein family. In this work, a transgenic approach was used to study the role of SN1 in planta. Even when overexpressing SN1, potato lines did not show remarkable morphological differences from the wild type; SN1 silencing resulted in reduced height, which was accompanied by an overall reduction in leaf size and severe alterations of leaf shape. Analysis of the adaxial epidermis of mature leaves revealed that silenced lines had 70% to 90% increases in mean cell size with respect to wild-type leaves. Consequently, the number of epidermal cells was significantly reduced in these lines. Confocal microscopy analysis after agroinfiltration of Nicotiana benthamiana leaves showed that SN1-green fluorescent protein fusion protein was localized in plasma membrane, and bimolecular fluorescence complementation assays revealed that SN1 self-interacted in vivo. We further focused our study on leaf metabolism by applying a combination of gas chromatography coupled to mass spectrometry, Fourier transform infrared spectroscopy, and spectrophotometric techniques. These targeted analyses allowed a detailed examination of the changes occurring in 46 intermediate compounds from primary metabolic pathways and in seven cell wall constituents. We demonstrated that SN1 silencing affects cell division, leaf primary metabolism, and cell wall composition in potato plants, suggesting that SN1 has additional roles in growth and development beyond its previously assigned role in plant defense. PMID:22080603

  17. Ongoing activation of autoantigen-specific B cells in primary biliary cirrhosis.

    PubMed

    Zhang, Jun; Zhang, Weici; Leung, Patrick S C; Bowlus, Christopher L; Dhaliwal, Sandeep; Coppel, Ross L; Ansari, Aftab A; Yang, Guo-Xiang; Wang, Jinjun; Kenny, Thomas P; He, Xiao-Song; Mackay, Ian R; Gershwin, M Eric

    2014-11-01

    The serologic hallmark of primary biliary cirrhosis (PBC), the antimitochondrial response to the E2 component of the pyruvate dehydrogenase complex (PDC-E2), has unique features, including continuous high titers of immunoglobulin M (IgM) and IgG reactivity throughout all stages of disease, capable not only of target enzyme inhibition, but also crossreactive with chemical xenobiotics that share molecular homology with the inner lipoyl domain of PDC-E2; such chemicals have been proposed as potential etiological agents. We used flow cytometry and enzyme-linked immunospot assay (ELISPOT) to examine B-cell subsets in 59 subjects, including 28 with PBC, 13 with primary sclerosing cholangitis (PSC), and 18 healthy controls. Strikingly, in PBC, although there were no significant differences in B-cell phenotype subpopulations, 10% of the total IgG and IgA plasmablast population and 23% of the IgM plasmablast population were uniquely reactive with PDC-E2, detected in the CXCR7+ CCR10low plasmablast population. In contrast, plasmablast reactivity to a control antigen, tetanus toxoid, was minimal and similar in all groups. Additionally, we isolated plasmablast-derived polyclonal antibodies and compared reactivity with plasma-derived antibodies and noted a distinct noncirculating tissue source of xenobiotic crossreacting antibodies. The high levels of autoantigen specific peripheral plasmablasts indicate recent activation of naive or memory B cells and a continuous and robust activation. The presence of CXCR7+ CCR10low PDC-E2-specific ASCs suggests a mechanistic basis for the migration of circulating antigen specific plasmablasts to the mucosal epithelial ligands CXCL12 and CCL28. Our findings suggest a sustained rigorous B-cell response in PBC, likely activated and perpetuated by cognate autoantigen. © 2014 by the American Association for the Study of Liver Diseases.

  18. Atypical protein kinase C regulates primary dendrite specification of cerebellar Purkinje cells by localizing Golgi apparatus.

    PubMed

    Tanabe, Koji; Kani, Shuichi; Shimizu, Takashi; Bae, Young-Ki; Abe, Takaya; Hibi, Masahiko

    2010-12-15

    Neurons have highly polarized structures that determine what parts of the soma elaborate the axon and dendrites. However, little is known about the mechanisms that establish neuronal polarity in vivo. Cerebellar Purkinje cells extend a single primary dendrite from the soma that ramifies into a highly branched dendritic arbor. We used the zebrafish cerebellum to investigate the mechanisms by which Purkinje cells acquire these characteristics. To examine dendritic morphogenesis in individual Purkinje cells, we marked the cell membrane using a Purkinje cell-specific promoter to drive membrane-targeted fluorescent proteins. We found that zebrafish Purkinje cells initially extend multiple neurites from the soma and subsequently retract all but one, which becomes the primary dendrite. In addition, the Golgi apparatus specifically locates to the root of the primary dendrite, and its localization is already established in immature Purkinje cells that have multiple neurites. Inhibiting secretory trafficking through the Golgi apparatus reduces dendritic growth, suggesting that the Golgi apparatus is involved in the dendritic morphogenesis. We also demonstrated that in a mutant of an atypical protein kinase C (aPKC), Prkci, Purkinje cells retain multiple primary dendrites and show disrupted localization of the Golgi apparatus. Furthermore, a mosaic inhibition of Prkci in Purkinje cells recapitulates the aPKC mutant phenotype. These results suggest that the aPKC cell autonomously controls the Golgi localization and thereby regulates the specification of the primary dendrite of Purkinje cells.

  19. Biocompatibility of magnesium implants in primary human reaming debris-derived cells stem cells in vitro.

    PubMed

    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.

  20. Slow conduction in mixed cultured strands of primary ventricular cells and stem cell-derived cardiomyocytes

    PubMed Central

    Kucera, Jan P.; Prudat, Yann; Marcu, Irene C.; Azzarito, Michela; Ullrich, Nina D.

    2015-01-01

    Modern concepts for the treatment of myocardial diseases focus on novel cell therapeutic strategies involving stem cell-derived cardiomyocytes (SCMs). However, functional integration of SCMs requires similar electrophysiological properties as primary cardiomyocytes (PCMs) and the ability to establish intercellular connections with host myocytes in order to contribute to the electrical and mechanical activity of the heart. The aim of this project was to investigate the properties of cardiac conduction in a co-culture approach using SCMs and PCMs in cultured cell strands. Murine embryonic SCMs were pooled with fetal ventricular cells and seeded in predefined proportions on microelectrode arrays to form patterned strands of mixed cells. Conduction velocity (CV) was measured during steady state pacing. SCM excitability was estimated from action potentials measured in single cells using the patch clamp technique. Experiments were complemented with computer simulations of conduction using a detailed model of cellular architecture in mixed cell strands. CV was significantly lower in strands composed purely of SCMs (5.5 ± 1.5 cm/s, n = 11) as compared to PCMs (34.9 ± 2.9 cm/s, n = 21) at similar refractoriness (100% SCMs: 122 ± 25 ms, n = 9; 100% PCMs: 139 ± 67 ms, n = 14). In mixed strands combining both cell types, CV was higher than in pure SCMs strands, but always lower than in 100% PCM strands. Computer simulations demonstrated that both intercellular coupling and electrical excitability limit CV. These data provide evidence that in cultures of murine ventricular cardiomyocytes, SCMs cannot restore CV to control levels resulting in slow conduction, which may lead to reentry circuits and arrhythmias. PMID:26442264

  1. Linking the Primary Cilium to Cell Migration in Tissue Repair and Brain Development

    PubMed Central

    Veland, Iben Rønn; Lindbæk, Louise; Christensen, Søren Tvorup

    2014-01-01

    Primary cilia are unique sensory organelles that coordinate cellular signaling networks in vertebrates. Inevitably, defects in the formation or function of primary cilia lead to imbalanced regulation of cellular processes that causes multisystemic disorders and diseases, commonly known as ciliopathies. Mounting evidence has demonstrated that primary cilia coordinate multiple activities that are required for cell migration, which, when they are aberrantly regulated, lead to defects in organogenesis and tissue repair, as well as metastasis of tumors. Here, we present an overview on how primary cilia may contribute to the regulation of the cellular signaling pathways that control cyclic processes in directional cell migration. PMID:26955067

  2. Campylobacter-induced interleukin-8 responses in human intestinal epithelial cells and primary intestinal chick cells.

    PubMed

    Borrmann, Erika; Berndt, Angela; Hänel, Ingrid; Köhler, Heike

    2007-09-20

    Campylobacter (C.) jejuni and C. coli can cause gastrointestinal disorders in humans characterized by acute inflammation. Inflammatory signals are initiated during interaction between these pathogens and human intestinal cells, but nothing is known about the stimulation of avian intestinal cells by Campylobacter. Interleukin-8 (IL-8) as a proinflammatory chemokine plays an important role in mobilizing cellular defence mechanism. IL-8 mRNA expression in both human intestinal cells (INT 407) and primary intestinal chick cells (PIC) was determined by quantitative real-time RT-PCR. The secretion of IL-8 protein by INT407 was measured using ELISA. Although C. jejuni and C. coli are considered to be harmless commensals in the gut of birds, the avian Campylobacter isolates investigated were able to induce the proinflammatory IL-8 in PIC as well as in INT407. In an in vitro system, C. jejuni as well as C. coli were able to induce IL-8 mRNA in PIC. Relation between the virulence properties like toxin production, the ability to invade and to survive in Caco-2 cells and the level of IL-8 mRNA produced by INT 407 and PIC after infection with Campylobacter strains was also investigated.

  3. Primary cultures of embryonic chick lens cells as a model system to study lens gap junctions and fiber cell differentiation.

    PubMed

    Musil, Linda S

    2012-07-01

    A major limitation in lens gap junction research has been the lack of experimentally tractable ex vivo systems to study the formation and regulation of fiber-type gap junctions. Although immortalized lens-derived cell lines are amenable to both gene transfection and siRNA-mediated knockdown, to our knowledge none are capable of undergoing appreciable epithelial-to-fiber differentiation. Lens central epithelial explants have the converse limitation. A key advance in the field was the development of a primary embryonic chick lens cell culture system by Drs. Sue Menko and Ross Johnson. Unlike central epithelial explants, these cultures also include cells from the peripheral (preequatorial and equatorial) epithelium, which is the most physiologically relevant population for the study of fiber-type gap junction formation. We have modified the Menko/Johnson system and refer to our cultures as dissociated cell-derived monolayer cultures (DCDMLs). We culture DCDMLs without serum to mimic the avascular lens environment and on laminin, the major matrix component of the lens capsule. Here, I review the features of the DCDML system and how we have used it to study lens gap junctions and fiber cell differentiation. Our results demonstrate the power of DCDMLs to generate new findings germane to the mammalian lens and how these cultures can be exploited to conduct experiments that would be impossible, prohibitively expensive and/or difficult to interpret using transgenic animals in vivo.