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Sample records for administration human cells

  1. Effect of hormonal manipulation and doxorubicin administration on cell cycle kinetics of human breast cancer cells.

    PubMed Central

    Bontenbal, M.; Sieuwerts, A. M.; Klijn, J. G.; Peters, H. A.; Krijnen, H. L.; Sonneveld, P.; Foekens, J. A.

    1989-01-01

    Dual-parameter flow cytometry, following bromodeoxyuridine (BrdUrd) incorporation and propidium iodide (PI) uptake into DNA, was used to study the effects of oestradiol and/or insulin on cell cycle kinetics of human breast cancer cells in vitro. After a lag-period of 6-12 h, an optimum in the percentage of S-phase cells was reached between 18 and 24 h after hormone administration. A 1 h pulse of oestradiol was as effective as the continuous presence of oestradiol in pushing the cells from quiescent growing cultures into the cell cycle. A 1 h pulse of insulin was less effective than continuous administration. The addition of doxorubicin resulted in an accumulation of the cells in the late S/G2M-phases. It is concluded that dual-parameter flow cytometry allows accurate assessment of the effects of hormones and chemotherapy on the cell cycle. Therefore this method is very suitable for studying the interaction of hormones and chemotherapy on cell growth. Images Figure 1 Figure 2 Figure 4 PMID:2679851

  2. Cre-inducible human CD59 mediates rapid cell ablation after intermedilysin administration

    PubMed Central

    Feng, Dechun; Dai, Shen; Liu, Fengming; Ohtake, Yosuke; Zhou, Zhou; Wang, Hua; Zhang, Yonggang; Kearns, Alison; Peng, Xiao; Zhu, Faliang; Hayat, Umar; Li, Man; He, Yong; Xu, Mingjiang; Zhao, Chunling; Cheng, Min; Zhang, Lining; Wang, Hong; Yang, Xiaofeng; Ju, Cynthia; Bryda, Elizabeth C.; Gordon, Jennifer; Khalili, Kamel; Hu, Wenhui; Li, Shuxin; Qin, Xuebin

    2016-01-01

    Cell ablation is a powerful tool for studying cell lineage and/or function; however, current cell-ablation models have limitations. Intermedilysin (ILY), a cytolytic pore-forming toxin that is secreted by Streptococcus intermedius, lyses human cells exclusively by binding to the human complement regulator CD59 (hCD59), but does not react with CD59 from nonprimates. Here, we took advantage of this feature of ILY and developed a model of conditional and targeted cell ablation by generating floxed STOP-CD59 knockin mice (ihCD59), in which expression of human CD59 only occurs after Cre-mediated recombination. The administration of ILY to ihCD59+ mice crossed with various Cre-driver lines resulted in the rapid and specific ablation of immune, epithelial, or neural cells without off-target effects. ILY had a large pharmacological window, which allowed us to perform dose-dependent studies. Finally, the ILY/ihCD59-mediated cell-ablation method was tested in several disease models to study immune cell functionalities, hepatocyte and/or biliary epithelial damage and regeneration, and neural cell damage. Together, the results of this study demonstrate the utility of the ihCD59 mouse model for studying the effects of cell ablation in specific organ systems in a variety of developmental and disease states. PMID:27159394

  3. Administrative Aspects of Human Experimentation.

    ERIC Educational Resources Information Center

    Irvine, George W.

    1992-01-01

    The following administrative aspects of scientific experimentation with human subjects are discussed: the definition of human experimentation; the distinction between experimentation and treatment; investigator responsibility; documentation; the elements and principles of informed consent; and the administrator's role in establishing and…

  4. Multiple Intravenous Administrations of Human Umbilical Cord Blood Cells Benefit in a Mouse Model of ALS

    PubMed Central

    Garbuzova-Davis, Svitlana; Rodrigues, Maria C. O.; Mirtyl, Santhia; Turner, Shanna; Mitha, Shazia; Sodhi, Jasmine; Suthakaran, Subatha; Eve, David J.; Sanberg, Cyndy D.; Kuzmin-Nichols, Nicole; Sanberg, Paul R.

    2012-01-01

    Background A promising therapeutic strategy for amyotrophic lateral sclerosis (ALS) is the use of cell-based therapies that can protect motor neurons and thereby retard disease progression. We recently showed that a single large dose (25×106 cells) of mononuclear cells from human umbilical cord blood (MNC hUCB) administered intravenously to pre-symptomatic G93A SOD1 mice is optimal in delaying disease progression and increasing lifespan. However, this single high cell dose is impractical for clinical use. The aim of the present pre-clinical translation study was therefore to evaluate the effects of multiple low dose systemic injections of MNC hUCB cell into G93A SOD1 mice at different disease stages. Methodology/Principal Findings Mice received weekly intravenous injections of MNC hUCB or media. Symptomatic mice received 106 or 2.5×106 cells from 13 weeks of age. A third, pre-symptomatic, group received 106 cells from 9 weeks of age. Control groups were media-injected G93A and mice carrying the normal hSOD1 gene. Motor function tests and various assays determined cell effects. Administered cell distribution, motor neuron counts, and glial cell densities were analyzed in mouse spinal cords. Results showed that mice receiving 106 cells pre-symptomatically or 2.5×106 cells symptomatically significantly delayed functional deterioration, increased lifespan and had higher motor neuron counts than media mice. Astrocytes and microglia were significantly reduced in all cell-treated groups. Conclusions/Significance These results demonstrate that multiple injections of MNC hUCB cells, even beginning at the symptomatic disease stage, could benefit disease outcomes by protecting motor neurons from inflammatory effectors. This multiple cell infusion approach may promote future clinical studies. PMID:22319620

  5. B-Cell Responses to Intramuscular Administration of a Bivalent Virus-like Particle Human Norovirus Vaccine.

    PubMed

    Ramani, Sasirekha; Neill, Frederick H; Ferreira, Jennifer; Treanor, John J; Frey, Sharon E; Topham, David J; Goodwin, Robert R; Borkowski, Astrid; Baehner, Frank; Mendelman, Paul M; Estes, Mary K; Atmar, Robert L

    2017-03-01

    Human noroviruses (HuNoVs) are a leading cause of acute gastroenteritis worldwide. A virus-like particle (VLP) candidate vaccine induces the production of serum histo-blood group antigen (HBGA) blocking antibodies, the first identified correlate of protection from HuNoV gastroenteritis. Recently, virus-specific IgG memory B-cells were identified as another potential correlate of protection against HuNoV gastroenteritis. We assessed B-cell responses following intramuscular administration of a bivalent (GI.1/GII.4) VLP vaccine using protocols identical to those used to evaluate cellular immunity following experimental HuNoV infection. The kinetics and magnitude of cellular immunity to G1.1 infection versus VLP vaccination were compared. Intramuscular immunization with bivalent VLP vaccine induces the production of antibody-secreting cells (ASCs) and memory B-cells. ASC responses peaked at day 7 post 1st dose of vaccine and returned to nearly baseline levels by day 28. Minimal increases in ASCs were seen after a second vaccine dose at day 28. Antigen-specific IgG memory B-cells persist at day 180 post-vaccination for both GI.1 and GII.4 VLPs. The overall trends in B-cell responses to vaccination were similar to infection, where there was a greater bias of ASC response towards IgA and memory B-cell response to IgG. The magnitude of ASC and memory B-cell responses to the GI.1 VLP component of the vaccine were also comparable to responses following GI.1 infection. The production of IgG memory B-cells and persistence at day 180 is a key finding and underscores the need for future studies to determine if IgG memory B-cells are a correlate of protection following vaccination.

  6. Prevention and treatment of colon cancer by peroral administration of HAMLET (human α-lactalbumin made lethal to tumour cells)

    PubMed Central

    Puthia, Manoj; Storm, Petter; Nadeem, Aftab; Hsiung, Sabrina; Svanborg, Catharina

    2014-01-01

    Background Most colon cancers start with dysregulated Wnt/β-catenin signalling and remain a major therapeutic challenge. Examining whether HAMLET (human α-lactalbumin made lethal to tumour cells) may be used for colon cancer treatment is logical, based on the properties of the complex and its biological context. Objective To investigate if HAMLET can be used for colon cancer treatment and prevention. ApcMin/+ mice, which carry mutations relevant to hereditary and sporadic human colorectal tumours, were used as a model for human disease. Method HAMLET was given perorally in therapeutic and prophylactic regimens. Tumour burden and animal survival of HAMLET-treated and sham-fed mice were compared. Tissue analysis focused on Wnt/β-catenin signalling, proliferation markers and gene expression, using microarrays, immunoblotting, immunohistochemistry and ELISA. Confocal microscopy, reporter assay, immunoprecipitation, immunoblotting, ion flux assays and holographic imaging were used to determine effects on colon cancer cells. Results Peroral HAMLET administration reduced tumour progression and mortality in ApcMin/+ mice. HAMLET accumulated specifically in tumour tissue, reduced β-catenin and related tumour markers. Gene expression analysis detected inhibition of Wnt signalling and a shift to a more differentiated phenotype. In colon cancer cells with APC mutations, HAMLET altered β-catenin integrity and localisation through an ion channel-dependent pathway, defining a new mechanism for controlling β-catenin signalling. Remarkably, supplying HAMLET to the drinking water from the time of weaning also significantly prevented tumour development. Conclusions These data identify HAMLET as a new, peroral agent for colon cancer prevention and treatment, especially needed in people carrying APC mutations, where colon cancer remains a leading cause of death. PMID:23348960

  7. Human cord blood cells and myocardial infarction: effect of dose and route of administration on infarct size.

    PubMed

    Henning, Robert J; Burgos, Jose D; Vasko, Mark; Alvarado, Felipe; Sanberg, Cyndy D; Sanberg, Paul R; Morgan, Michael B

    2007-01-01

    There is no consensus regarding the optimal dose of stem cells or the optimal route of administration for the treatment of acute myocardial infarction. Bone marrow cells, containing hematopoietic and mesenchymal stem cells, in doses of 0.5 x 10(6) to >30 x 10(6) have been directly injected into the myocardium or into coronary arteries or infused intravenously in subjects with myocardial infarctions to reduce infarct size and improve heart function. Therefore, we determined the specific effects of different doses of human umbilical cord blood mononuclear cells (HUCBC), which contain hematopoietic and mesenchymal stem cells, on infarct size. In order to determine the optimal technique for stem cell administration, HUCBC were injected directly into the myocardium (IM), or into the LV cavity with the ascending aorta transiently clamped to facilitate coronary artery perfusion (IA), or injected intravenously (IV) in rats 1-2 h after the left anterior coronary artery was permanently ligated. Immune suppressive therapy was not given to any rat. One month later, the infarct size in control rat hearts treated with only Isolyte averaged 23.7 +/- 1.7% of the LV muscle area. Intramyocardial injection of HUCBC reduced the infarct size by 71% with 0.5 x 10(6) HUCBC and by 93% with 4 x 10(6) HUCBC in comparison with the controls (p < 0.001). Intracoronary injection reduced the infarction size by 47% with 0.5 x 10(6) HUCBC and by 80% with 4 x 10(6) HUCBC (p < 0.001), and IV HUCBC reduced infarct size by 51% with 0.5 x 10(6) and by 75-77% with 16-32 million HUCBC (p < 0.001) in comparison with control hearts. With 4 x 10(6) HUCBC, infarction size was 65% smaller with IM HUCBC than with IA HUCBC and 78% smaller than with IV HUCBC (p < 0.05). Nevertheless, IM, IA, and IV HUCBC all produced significant reductions in infarct size in comparison with Isolyte-treated infarcted hearts without requirements for host immune suppression. The present experiments demonstrate that the optimal dose

  8. Pathways to the Humanities in Educational Administration.

    ERIC Educational Resources Information Center

    Popper, Samuel H.

    Based on the author's own experience as a professor of educational administration, this monograph is an argument for establishing a link between the humanities and instruction in school administration. Part I discusses the instrumental value of the humanities in administrative preparation and recounts the limitations of past attempts by the…

  9. Increased epidermal cell proliferation in normal human skin in vivo following local administration of interferon-gamma.

    PubMed Central

    Barker, J. N.; Goodlad, J. R.; Ross, E. L.; Yu, C. C.; Groves, R. W.; MacDonald, D. M.

    1993-01-01

    Recombinant human interferon-gamma was administered intradermally (10 micrograms in 0.1 ml) to healthy adult human volunteers from day 1 to day 3, and epidermal cell proliferation was measured on whole skin biopsies at day 6. Three independent parameters were assessed, namely, a) epidermal keratin-16 expression, b) keratinocyte proliferating cell nuclear antigen expression, and c) keratinocyte silver nucleolar organizer region counts. Significantly increased scores for each parameter were observed after interferon-gamma injection (P < 0.01 in each case) compared to site-matched controls. Keratin-16 expression was confined to suprabasal epidermis, whereas proliferating cell nuclear antigen and silver nucleolar organizer region counts were particularly elevated in the basal epidermis. Taken together with previous findings, these studies indicate both proinflammatory and growth regulatory roles for interferon-gamma in human skin. These data are likely to be of particular importance to pathophysiological mechanisms of psoriasis and related cutaneous inflammatory diseases. Images Figure 1 Figure 2 Figure 3 PMID:7682760

  10. Effects of Intravenous Administration of Human Umbilical Cord Blood Stem Cells in 3-Acetylpyridine-Lesioned Rats

    PubMed Central

    Calatrava-Ferreras, Lucía; Gonzalo-Gobernado, Rafael; Herranz, Antonio S.; Reimers, Diana; Montero Vega, Teresa; Jiménez-Escrig, Adriano; Richart López, Luis Alberto; Bazán, Eulalia

    2012-01-01

    Cerebellar ataxias include a heterogeneous group of infrequent diseases characterized by lack of motor coordination caused by disturbances in the cerebellum and its associated circuits. Current therapies are based on the use of drugs that correct some of the molecular processes involved in their pathogenesis. Although these treatments yielded promising results, there is not yet an effective therapy for these diseases. Cell replacement strategies using human umbilical cord blood mononuclear cells (HuUCBMCs) have emerged as a promising approach for restoration of function in neurodegenerative diseases. The aim of this work was to investigate the potential therapeutic activity of HuUCBMCs in the 3-acetylpyridine (3-AP) rat model of cerebellar ataxia. Intravenous administered HuUCBMCs reached the cerebellum and brain stem of 3-AP ataxic rats. Grafted cells reduced 3-AP-induced neuronal loss promoted the activation of microglia in the brain stem, and prevented the overexpression of GFAP elicited by 3-AP in the cerebellum. In addition, HuUCBMCs upregulated the expression of proteins that are critical for cell survival, such as phospho-Akt and Bcl-2, in the cerebellum and brain stem of 3-AP ataxic rats. As all these effects were accompanied by a temporal but significant improvement in motor coordination, HuUCBMCs grafts can be considered as an effective cell replacement therapy for cerebellar disorders. PMID:23150735

  11. Repeated administrations of human umbilical cord blood cells improve disease outcomes in a mouse model of Sanfilippo syndrome type III B.

    PubMed

    Willing, Alison E; Garbuzova-Davis, Svitlana N; Zayko, Olga; Derasari, Hiranya M; Rawls, Ashley E; James, Chris R; Mervis, Ron F; Sanberg, Cyndy D; Kuzmin-Nichols, Nicole; Sanberg, Paul R

    2014-01-01

    Sanfilippo syndrome type III B (MPS III B) is an inherited disorder characterized by a deficiency of α-N-acetylglucosaminidase (Naglu) enzyme leading to accumulation of heparan sulfate in lysosomes and severe neurological deficits. We have previously shown that a single administration of human umbilical cord mononuclear cells (hUCB MNCs) into Naglu knockout mice decreased behavioral abnormalities and tissue pathology. In this study, we tested whether repeated doses of hUCB MNCs would be more beneficial than a single dose of cells. Naglu mice at 3 months of age were randomly assigned to either a Media-only group or one of three hUCB MNC treatment groups--single low dose (3 × 10(6) cells), single high dose (1.8 × 10(7) cells), or multiple doses (3 × 10(6) cells monthly for 6 months) delivered intravenously; cyclosporine was injected intraperitoneally to immune suppress the mice for the duration of the study. An additional control group of wild-type mice was also used. We measured anxiety in an open field test and cognition in an active avoidance test prior to treatment and then at monthly intervals for 6 months. hUCB MNCs restored normal anxiety-like behavior in these mice (p < 0.001). The repeated cell administrations also restored hippocampal cytoarchitecture, protected the dendritic tree, decreased GM3 ganglioside accumulation, and decreased microglial activation, particularly in the hippocampus and cortex. These data suggest that the neuroprotective effect of hUCB MNCs can be enhanced by repeated cell administrations.

  12. Repeated Administrations of Human Umbilical Cord Blood Cells Improve Disease Outcomes in a Mouse Model of Sanfilippo Syndrome Type III B.

    PubMed

    Willing, Alison E; Garbuzova-Davis, Svitlana N; Zayko, Olga; Derasari, Hiranya M; Rawls, Ashley E; James, Chris R; Mervis, Ron F; Sanberg, Cyndy D; Kuzmin-Nichols, Nicole; Sanberg, Paul R

    2013-12-30

    Sanfilippo syndrome type III B (MPS III B) is an inherited disorder characterized by a deficiency of ?-N-acetylglucosaminidase (Naglu) enzyme leading to accumulation of heparan sulfate in lysosomes and severe neurological deficits. We have previously shown that a single administration of human umbilical cord mononuclear cells (hUCB MNC) into Nagluknockout mice decreased behavioral abnormalities and tissue pathology. In this study, we tested whether repeated doses of hUCB MNCs would be more beneficial than a single dose of cells. Naglumice at 3 months of age were randomly assigned to either a Media only group, or one of three hUCB MNC treatment groups - single low dose (3x10(6) cells), single high dose (1.8x10(7) cells) or multiple doses (3x10(6) cells monthly for 6 months) delivered intravenously (i.v.); cyclosporine was injected i.p. to immune suppress the mice for the duration of the study. An additional control group of wild type mice was also used. We measured anxiety in an open field test and cognition inactive avoidance test prior to treatment and then at monthly intervals for 6 months. hUCB MNCs restored normal anxiety-like behavior in these mice (p < 0.001). The repeated cell administrations also restored hippocampal cytoarchitecture, protected the dendritic tree, decreased GM3 ganglioside accumulation and decreased microglial activation, particularly in hippocampus and cortex. These data suggest that the neuroprotective effect of hUCB MNCs can be enhanced by repeated cell administrations.

  13. Murine hypothalamic destruction with vascular cell apoptosis subsequent to combined administration of human papilloma virus vaccine and pertussis toxin

    PubMed Central

    Aratani, Satoko; Fujita, Hidetoshi; Kuroiwa, Yoshiyuki; Usui, Chie; Yokota, Shumpei; Nakamura, Ikuro; Nishioka, Kusuki; Nakajima, Toshihiro

    2016-01-01

    Vaccination is the most powerful way to prevent human beings from contracting infectious diseases including viruses. In the case of the human papillomavirus (HPV) vaccine, an unexpectedly novel disease entity, HPV vaccination associated neuro-immunopathetic syndrome (HANS), has been reported and remains to be carefully verified. To elucidate the mechanism of HANS, we applied a strategy similar to the active experimental autoimmune encephalitis (EAE) model - one of the most popular animal models used to induce maximum immunological change in the central nervous system. Surprisingly, mice vaccinated with pertussis toxin showed neurological phenotypes that include low responsiveness of the tail reflex and locomotive mobility. Pathological analyses revealed the damage to the hypothalamus and circumventricular regions around the third ventricle, and these regions contained apoptotic vascular endothelial cells. These data suggested that HPV-vaccinated donners that are susceptible to the HPV vaccine might develop HANS under certain environmental factors. These results will give us the new insight into the murine pathological model of HANS and help us to find a way to treat of patients suffering from HANS. PMID:27833142

  14. Different routes of administration of human umbilical tissue-derived cells improve functional recovery in the rat after focal cerebral ischemia.

    PubMed

    Zhang, Li; Li, Yi; Romanko, Michael; Kramer, Brian C; Gosiewska, Anna; Chopp, Michael; Hong, Klaudyne

    2012-12-13

    Human umbilical tissue-derived cells (hUTC) are a potential neurorestorative candidate for stroke treatment. Here, we test the effects of hUTC treatment in a rat model of stroke via various routes of administration. Rats were treated with hUTC or phosphate-buffered saline (PBS) via different routes including intraarterial (IA), intravenous (IV), intra-cisterna magna (ICM), lumber intrathecal (IT), or intracerebral injection (IC) at 24h after stroke onset. Treatment with hUTC via IV and IC route led to significant functional improvements starting at day 14, which persisted to day 60 compared with respective PBS-treated rats. HUTC administered via IA, ICM, and IT significantly improved neurological functional recovery starting at day 14 and persisted up to day 49 compared with PBS-treated rats. Although IA administration resulted in the highest donor cell number detected within the ischemic brain compared to the other routes, hUTC treatments significantly increased ipsilateral bromodeoxyuridine incorporating subventricular zone (SVZ) cells and vascular density in the ischemic boundary compared with PBS-treated rats regardless of the route of administration. While rats received hUTC treatment via IA, IV, IC, and ICM routes showed greater synaptophysin immunoreactivity, significant reductions in TUNEL-positive cells in the ipsilateral hemisphere were observed in IA, IV, and IC routes compared with PBS-treated rats. hUTC treatments did not reduce infarct volume when compared to the PBS groups. Our data indicate that hUTC administered via multiple routes provide therapeutic benefit after stroke. The enhancement of neurorestorative events in the host brain may contribute to the therapeutic benefits of hUTC in the treatment of stroke.

  15. Effect of Single and Double Administration of Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Following Focal Cerebral Ischemia in Rats.

    PubMed

    Park, Hyung Woo; Kim, Yona; Chang, Jong Wook; Yang, Yoon Sun; Oh, Wonil; Lee, Jae Min; Park, Hye Ran; Kim, Dong Gyu; Paek, Sun Ha

    2017-02-01

    Stem cell therapies are administered during the acute phase of stroke to preserve the penumbral tissues from ischemic injury. However, the effect of repeated cell therapy during the acute phase remains unclear. In this study, we investigated and compared the functional outcome of single (two days post-injury) and repeated (two and nine days post-injury) treatment with human umbilical cord derived mesenchymal stem cells (hUCB-MSCs) after middle cerebral artery occlusion (MCAO). The rotarod and limb placement tests were utilized to investigate functional outcomes, while infarct volume and tissue damage were measured by immunofluorescent staining for neovascularization, neurogenesis, apoptosis, and inflammation in the penumbral zones. We observed notable motor dysfunction and a significant decrease in infarcted brain volume, as well as increases in neurons and vessels in both single and repeated hUCB-MSC treatments compared to the control group. Interestingly, repeated administration of hUCB-MSCs was not found to elicit additional or synergistic improvements over monotherapy. This study suggests that a clearer understanding of the therapeutic window after stroke will facilitate the development of more efficient treatment protocols in the clinical application of stem cell therapy.

  16. Effect of Single and Double Administration of Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells Following Focal Cerebral Ischemia in Rats

    PubMed Central

    Park, Hyung Woo; Kim, Yona; Chang, Jong Wook; Yang, Yoon Sun; Oh, Wonil; Lee, Jae Min; Park, Hye Ran; Kim, Dong Gyu

    2017-01-01

    Stem cell therapies are administered during the acute phase of stroke to preserve the penumbral tissues from ischemic injury. However, the effect of repeated cell therapy during the acute phase remains unclear. In this study, we investigated and compared the functional outcome of single (two days post-injury) and repeated (two and nine days post-injury) treatment with human umbilical cord derived mesenchymal stem cells (hUCB-MSCs) after middle cerebral artery occlusion (MCAO). The rotarod and limb placement tests were utilized to investigate functional outcomes, while infarct volume and tissue damage were measured by immunofluorescent staining for neovascularization, neurogenesis, apoptosis, and inflammation in the penumbral zones. We observed notable motor dysfunction and a significant decrease in infarcted brain volume, as well as increases in neurons and vessels in both single and repeated hUCB-MSC treatments compared to the control group. Interestingly, repeated administration of hUCB-MSCs was not found to elicit additional or synergistic improvements over monotherapy. This study suggests that a clearer understanding of the therapeutic window after stroke will facilitate the development of more efficient treatment protocols in the clinical application of stem cell therapy. PMID:28243167

  17. Kinetics of human hemopoietic cells after in vivo administration of granulocyte-macrophage colony-stimulating factor.

    PubMed Central

    Aglietta, M; Piacibello, W; Sanavio, F; Stacchini, A; Aprá, F; Schena, M; Mossetti, C; Carnino, F; Caligaris-Cappio, F; Gavosto, F

    1989-01-01

    The kinetic changes induced by granulocyte-macrophage colony-stimulating factor (GM-CSF) on hemopoietic cells were assessed in physiological conditions by administering GM-CSF (8 micrograms/kg per d) for 3 d to nine patients with solid tumors and normal bone marrow (BM), before chemotherapy. GM-CSF increased the number of circulating granulocytes and monocytes; platelets, erythrocytes, lymphocyte number, and subsets were unmodified. GM-CSF increased the percentage of BM S phase BFU-E (from 32 +/- 7 to 79 +/- 16%), day 14 colony-forming unit granulocyte-macrophage (CFU-GM) (from 43 +/- 20 to 82 +/- 11%) and day 7 CFU-GM (from 41 +/- 14 to 56 +/- 20%). The percentage of BM myeloblasts, promyelocytes, and myelocytes in S phase increased from 26 +/- 14 to 41 +/- 6%, and that of erythroblasts increased from 25 +/- 12 to 30 +/- 12%. This suggests that GM-CSF activates both erythroid and granulomonopoietic progenitors but that, among the morphologically recognizable BM precursors, only the granulomonopoietic lineage is a direct target of the molecule. GM-CSF increased the birth rate of cycling cells from 1.3 to 3.4 cells %/h and decreased the duration of the S phase from 14.3 to 9.1 h and the cell cycle time from 86 to 26 h. After treatment discontinuation, the number of circulating granulocytes and monocytes rapidly fell. The proportion of S phase BM cells dropped to values lower than pretreatment levels, suggesting a period of relative refractoriness to cell cycle-active antineoplastic agents. PMID:2643633

  18. High incidence of HAM/TSP-like symptoms in WKA rats after administration of human T-cell leukemia virus type 1-producing cells.

    PubMed Central

    Kushida, S; Mizusawa, H; Matsumura, M; Tanaka, H; Ami, Y; Hori, M; Yagami, K; Kameyama, T; Tanaka, Y; Yoshida, A

    1994-01-01

    We demonstrate a significantly high incidence of human T-cell leukemia virus type 1 (HTLV-1)-associated myelopathy (HAM)-or tropical spastic paraparesis (TSP)-like symptoms in WKA rats after injection with HTLV-1-producing MT-2 cells, while no symptoms were observed in F344 rats injected with MT-2 cells or in control WKA rats. Five of the eight (63%) WKA rats injected with MT-2 cells showed HAM/TSP-like paraparesis at 105 weeks of age, but none of seven MT-2-injected F344 rats or eight control WKA rats showed symptoms. This high incidence of HAM/TSP-like symptoms in WKA rats was statistically significant (P < 0.05). Six of the eight (75%) WKA rats injected with MT-2 cells showed HAM/TSP-like paraparesis at 108 weeks of age. HAM/TSP-like symptoms were also observed in one of the two WKA rats injected with HTLV-1-producing Ra-1 cells at 128 weeks of age. HTLV-1 provirus was detected in peripheral blood mononuclear cells in both WKA and F344 rats. The provirus was detected in the spinal cords of the HAM/TSP-like WKA rats that had severe neuropathological changes. WKA and F344 rats showed no significant difference in antibody response against HTLV-1 Gag antigen. However, the antibody response against the C-terminal half of gp46 HTLV-1 envelope protein was lower in WKA rats than in F344 rats. Pathological analysis of the HAM/TSP-like rats showed degeneration of the white matter of the spinal cord and peripheral nerves. These findings suggest that both the genetic background of the host and HTLV-1 infection are important in neuropathogenesis of HAM/TSP-like paraparesis in rats. Images PMID:7933104

  19. The Human Experience With Ghrelin Administration

    PubMed Central

    Garin, Margaret C.; Burns, Carrie M.; Kaul, Shailja

    2013-01-01

    Context: Ghrelin is an endogenous stimulator of GH and is implicated in a number of physiological processes. Clinical trials have been performed in a variety of patient populations, but there is no comprehensive review of the beneficial and adverse consequences of ghrelin administration to humans. Evidence Acquisition: PubMed was utilized, and the reference list of each article was screened. We included 121 published articles in which ghrelin was administered to humans. Evidence Synthesis: Ghrelin has been administered as an infusion or a bolus in a variety of doses to 1850 study participants, including healthy participants and patients with obesity, prior gastrectomy, cancer, pituitary disease, diabetes mellitus, eating disorders, and other conditions. There is strong evidence that ghrelin stimulates appetite and increases circulating GH, ACTH, cortisol, prolactin, and glucose across varied patient populations. There is a paucity of evidence regarding the effects of ghrelin on LH, FSH, TSH, insulin, lipolysis, body composition, cardiac function, pulmonary function, the vasculature, and sleep. Adverse effects occurred in 20% of participants, with a predominance of flushing and gastric rumbles and a mild degree of severity. The few serious adverse events occurred in patients with advanced illness and were not clearly attributable to ghrelin. Route of administration may affect the pattern of adverse effects. Conclusions: Existing literature supports the short-term safety of ghrelin administration and its efficacy as an appetite stimulant in diverse patient populations. There is some evidence to suggest that ghrelin has wider ranging therapeutic effects, although these areas require further investigation. PMID:23533240

  20. Withholding gonadotropins until human chorionic gonadotropin administration.

    PubMed

    Abdallah, Rony; Kligman, Isaac; Davis, Owen; Rosenwaks, Zev

    2010-11-01

    Withholding gonadotropins in women who exhibit high estradiol responses before follicles reach full maturation is called "coasting." Coasting, or suspending gonadotropin administration, can be an effective strategy for decreasing the risk of ovarian hyperstimulation syndrome (OHSS) while reducing cancelation rates. In in vitro fertilization cycles, mechanistically it is believed that withholding gonadotropins starves smaller follicles, induces apoptosis, and decreases the potential for these follicles to elaborate vascular endothelial growth factor, a known mediator of OHSS. It is generally accepted that coasting should be initiated when the estradiol (E₂) level is >3000 pg/mL in the setting of immature follicles. The human chorionic gonadotropin (hCG) trigger should be administered when the E₂ level subsequently drops to a "safe" level. Cycle cancellation should be considered if, after 3 to 4 days of coasting, the E₂ level remains excessively elevated. Oocyte retrieval may also be cancelled if the E₂ level on the day after hCG trigger drops precipitously. In gonadotropin-releasing hormone agonist (GnRHa)-based protocols, one can consider withholding GnRHa administration if the E₂ level continues to increase after a few days of coasting. Current data seem to show that the coasting period is short and/or is less likely to be required in GnRH-antagonist protocols as compared with GnRHa-based protocols. Large randomized control trials are still needed to establish the relative efficacy of coasting versus embryo cryopreservation in the context of OHSS prevention.

  1. Pathways to the Humanities in School Administration. Third Edition.

    ERIC Educational Resources Information Center

    Popper, Samuel H.

    Based on the author's experience as a professor of educational administration, this monograph is an argument for establishing a link between the humanities and instruction in school administration. Part 1 discusses the instrumental value of the humanities in administrative preparation and recounts the limitations of past attempts by the University…

  2. Impact of inhomogeneous static magnetic field (31.7-232.0 mT) exposure on human neuroblastoma SH-SY5Y cells during cisplatin administration.

    PubMed

    Vergallo, Cristian; Ahmadi, Meysam; Mobasheri, Hamid; Dini, Luciana

    2014-01-01

    Beneficial or adverse effects of Static Magnetic Fields (SMFs) are a large concern for the scientific community. In particular, the effect of SMF exposure during anticancer therapies still needs to be fully elucidated. Here, we evaluate the effects of SMF at induction levels that cisPt-treated cancer patients experience during the imaging process conducted in Low field (200-500 mT), Open field (300-700 mT) and/or inhomogeneous High field (1.5-3 T) Magnetic Resonance Imaging (MRI) machines. Human adrenergic neuroblastoma SH-SY5Y cells treated with 0.1 µM cisPt (i.e. the lowest concentration capable of inducing apoptosis) were exposed to SMF and their response was studied in vitro. Exposure of 0.1 µM cisPt-treated cells to SMF for 2 h decreased cell viability (30%) and caused overexpression of the apoptosis-related cleaved caspase-3 protein (46%). Furthermore, increase in ROS (Reactive Oxygen Species) production (23%) and reduction in the number of mitochondria vs controls were seen. The sole exposure of SMF for up to 24 h had no effect on cell viability but increased ROS production and modified cellular shape. On the other hand, the toxicity of cisPt was significantly prevented during 24 h exposure to SMF as shown by the levels of cell viability, cleaved caspase-3 and ROS production. In conclusion, due to the cytoprotective effect of 31.7-232.0 mT SMF on low-cisPt-concentration-treated SH-SY5Y cells, our data suggest that exposure to various sources of SMF in cancer patients under a cisPt regimen should be strictly controlled.

  3. Humoral, Mucosal, and Cell-Mediated Immunity Against Vaccine and Nonvaccine Genotypes After Administration of Quadrivalent Human Papillomavirus Vaccine to HIV-Infected Children

    PubMed Central

    Weinberg, Adriana; Song, Lin-Ye; Saah, Alfred; Brown, Martha; Moscicki, Anna B.; Meyer, William A.; Bryan, Janine; Levin, Myron J.

    2012-01-01

    Objectives. To characterize the immunogenicity of a quadrivalent human papillomavirus vaccine (QHPV) in human immunodeficiency virus (HIV)–infected children, we studied their immune responses to 3 or 4 doses. Methods. HIV-infected children aged 7–12 years with a CD4 cell percentage of ≥15% of lymphocytes, received 3 doses of QHPV with or without a fourth dose after 72 weeks. Type-specific and cross-reactive antibodies and cell-mediated immunity were measured. Results. Type-specific antibodies to HPV6, 11, and 16 were detected in 100% and ≥94% of children at 4 and 72 weeks, respectively, after the third QHPV dose. Corresponding numbers for HPV18 were 97% and 76%, respectively. A fourth QHPV dose increased seropositivity to ≥96% for all vaccine genotypes. Four weeks after the third QHPV dose, 67% of vaccinees seroconverted to HPV31, an HPV16-related genotype not in the vaccine; 69% and 39% of vaccinees developed mucosal HPV16 and 18 immunoglobulin G antibodies, respectively; and 60% and 52% of vaccinees developed cytotoxic T lymphocytes (CTLs) for HPV16 and 31, respectively. Conclusions. Three QHPV doses generated robust and persistent antibodies to HPV6, 11, and 16 but comparatively weaker responses to HPV18. A fourth dose increased antibodies against all vaccine genotypes in an anamnestic fashion. CTLs and mucosal antibodies against vaccine genotypes, as well as cross-reactive antibodies and CTL against nonvaccine genotypes, were detected. PMID:22859825

  4. Impact of CD4+ T Cell Responses on Clinical Outcome following Oral Administration of Wild-Type Enterotoxigenic Escherichia coli in Humans

    PubMed Central

    Chen, Wilbur H.; Magder, Laurence; Levine, Myron M.; Sztein, Marcelo B.

    2017-01-01

    Enterotoxigenic Escherichia coli (ETEC) is a non-invasive enteric pathogen of considerable public health importance, being one of the most common attributable causes of diarrheal illness in infants and young children in developing countries and the most common cause of traveler’s diarrhea. To enhance study-to-study consistency of our experimental challenge model of ETEC in volunteers, and to allow concomitant multi-site trials to evaluate anti-ETEC immunoprophylactic products, hundreds of vials, each containing a standardized inoculum of virulent wild-type (wt) ETEC strain H10407 (serotype O78:H11 expressing colonization factor antigen I and heat-labile and heat-stable enterotoxins), were prepared under current Good Manufacturing Practices (cGMP) and frozen. Following thawing, the contents of each vial can be used (diluted as necessary) to prepare consistent challenge inoculum, even at different study sites. A preliminary human experimental challenge study using this cGMP inoculum was conducted on a research isolation ward and the clinical and cell-mediated immune responses evaluated. Of the 6 healthy adult volunteers challenged 83% (5/6) developed diarrhea and 50% developed moderate-to-severe diarrhea (MSD). Moderate and severe diarrhea were defined as passage of ≥ 1 liter or ≥ 3 liters of diarrheal stool respectively. We compared the CD4+ T cell responses of volunteers who developed MSD against those who did not and identified significant differences in ETEC-specific cytokine production and gut homing potential. We furthermore demonstrated that increased expression of the gut-homing molecule integrin α4β7 by peripheral T follicular helper cells (pTfh) correlated with decreased stool volume and increased ETEC-specific IgA B memory cell (BM) development. Collectively, despite small numbers of volunteers, our results indicate a potential role for CD4+ T cells, in particular pTfh, in modulating disease outcome following exposure to wt ETEC in a volunteer

  5. Effects of Intermittent Administration of Parathyroid Hormone (1-34) on Bone Differentiation in Stromal Precursor Antigen-1 Positive Human Periodontal Ligament Stem Cells

    PubMed Central

    Wang, Xiaoxiao; Wang, Yanlan; Dai, Xubin; Chen, Tianyu; Yang, Fanqiao; Dai, Shuangye; Ou, Qianmin; Wang, Yan; Lin, Xuefeng

    2016-01-01

    Periodontitis is the most common cause of tooth loss and bone destruction in adults worldwide. Human periodontal ligament stem cells (hPDLSCs) may represent promising new therapeutic biomaterials for tissue engineering applications. Stromal precursor antigen-1 (STRO-1) has been shown to have roles in adherence, proliferation, and multipotency. Parathyroid hormone (PTH) has been shown to enhance proliferation in osteoblasts. Therefore, in this study, we aimed to compare the functions of STRO-1(+) and STRO-1(−) hPDLSCs and to investigate the effects of PTH on the osteogenic capacity of STRO-1(+) hPDLSCs in order to evaluate their potential applications in the treatment of periodontitis. Our data showed that STRO-1(+) hPDLSCs expressed higher levels of the PTH-1 receptor (PTH1R) than STRO-1(−) hPDLSCs. In addition, intermittent PTH treatment enhanced the expression of PTH1R and osteogenesis-related genes in STRO-1(+) hPDLSCs. PTH-treated cells also exhibited increased alkaline phosphatase activity and mineralization ability. Therefore, STRO-1(+) hPDLSCs represented a more promising cell resource for biomaterials and tissue engineering applications. Intermittent PTH treatment improved the capacity for STRO-1(+) hPDLSCs to repair damaged tissue and ameliorate the symptoms of periodontitis. PMID:27069479

  6. Human Factors in Library Administration. Revised Edition.

    ERIC Educational Resources Information Center

    Barnhard, Neil

    Intended for the beginning or inexperienced supervisor, this continuing education course syllabus presents basic information on the development of human relations skills, particularly in the areas of leadership, communication, conflict, and motivation. Role playing situations set in various types of medical libraries are also outlined to provide…

  7. Protection of Human Subjects: A Primer for the New Administrator.

    ERIC Educational Resources Information Center

    Steinert, Bruce W.

    2002-01-01

    Discusses clinical trials in general, protecting human research subjects, and the role the research administrator can play in facilitating the process. The discussion is not intended to be a thorough treatise on clinical trials, but is an overview of new and lesser known human protection issues that are better handled prospectively. (EV)

  8. 21 CFR 864.2280 - Cultured animal and human cells.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Cultured animal and human cells. 864.2280 Section 864.2280 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES... Cultured animal and human cells. (a) Identification. Cultured animal and human cells are in...

  9. 21 CFR 864.2280 - Cultured animal and human cells.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Cultured animal and human cells. 864.2280 Section 864.2280 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES... Cultured animal and human cells. (a) Identification. Cultured animal and human cells are in...

  10. Particle-based transcutaneous administration of HIV-1 p24 protein to human skin explants and targeting of epidermal antigen presenting cells.

    PubMed

    Rancan, Fiorenza; Amselgruber, Sarah; Hadam, Sabrina; Munier, Sevérine; Pavot, Vincent; Verrier, Bernard; Hackbarth, Steffen; Combadiere, Behazine; Blume-Peytavi, Ulrike; Vogt, Annika

    2014-02-28

    Transcutaneous immunization is a promising vaccination strategy for the treatment of infectious diseases and cancer. In this study, we investigate the combination of cyanoacrylate skin surface stripping (CSSS) and particle-based antigen delivery to target the HIV-1 p24 protein to skin antigen presenting cells (APC). The CSSS treatment pre-activates skin APC and opens hair follicles, where protein-loaded particles accumulate and allow for sustained delivery of the loaded antigen to perifollicular APC. We found that poly-lactic acid (PLA) and polystyrene (PS) particles targeted the adsorbed HIV-1 p24 protein to the hair follicles. Small amounts of PS and PLA particles were found to translocate to the epidermis and be internalized by skin cells, whereas most of the particles aggregated in the hair follicle canal, where they released the loaded antigen. The p24 protein diffused to the epidermis and dermis and was detected in skin cells, especially in Langerhans cells and dermal dendritic cells. Furthermore, the combination of CSSS and particle-based delivery resulted in activation and maturation of Langerhans cells (HLA-DR, CD80 and CD83). We conclude that particle-based antigen delivery across partially disrupted skin barrier is a feasible and effective approach to needle-free transcutaneous vaccination.

  11. Administration of Adult Human Bone Marrow-Derived, Cultured, Pooled, Allogeneic Mesenchymal Stromal Cells in Critical Limb Ischemia Due to Buerger's Disease: Phase II Study Report Suggests Clinical Efficacy.

    PubMed

    Gupta, Pawan K; Krishna, Murali; Chullikana, Anoop; Desai, Sanjay; Murugesan, Rajkumar; Dutta, Santanu; Sarkar, Uday; Raju, Radhakrishnan; Dhar, Anita; Parakh, Rajiv; Jeyaseelan, Lakshmanan; Viswanathan, Pachaiyappan; Vellotare, Prasanth Kulapurathu; Seetharam, Raviraja N; Thej, Charan; Rengasamy, Mathiyazhagan; Balasubramanian, Sudha; Majumdar, Anish S

    2017-03-01

    Critical limb ischemia (CLI) due to Buerger's disease is a major unmet medical need with a high incidence of morbidity. This phase II, prospective, nonrandomized, open-label, multicentric, dose-ranging study was conducted to assess the efficacy and safety of i.m. injection of adult human bone marrow-derived, cultured, pooled, allogeneic mesenchymal stromal cells (BMMSC) in CLI due to Buerger's disease. Patients were allocated to three groups: 1 and 2 million cells/kg body weight (36 patients each) and standard of care (SOC) (18 patients). BMMSCs were administered as 40-60 injections in the calf muscle and locally, around the ulcer. Most patients were young (age range, 38-42 years) and ex-smokers, and all patients had at least one ulcer. Both the primary endpoints-reduction in rest pain (0.3 units per month [SE, 0.13]) and healing of ulcers (11% decrease in size per month [SE, 0.05])-were significantly better in the group receiving 2 million cells/kg body weight than in the SOC arm. Improvement in secondary endpoints, such as ankle brachial pressure index (0.03 [SE, 0.01] unit increase per month) and total walking distance (1.03 [SE, 0.02] times higher per month), were also significant in the group receiving 2 million cells/kg as compared with the SOC arm. Adverse events reported were remotely related or unrelated to BMMSCs. In conclusion, i.m. administration of BMMSC at a dose of 2 million cells/kg showed clinical benefit and may be the best regimen in patients with CLI due to Buerger's disease. However, further randomized controlled trials are required to confirm the most appropriate dose. Stem Cells Translational Medicine 2017;6:689-699.

  12. A review of human drug self-administration procedures

    PubMed Central

    Jones, Jermaine D.; Comer, Sandra D.

    2014-01-01

    Drug self-administration procedures in laboratory settings allow us to closely model drug-taking behavior in real-world settings. This review provides an overview of many of the common self-administration methods used in human laboratory research. Typically, self-administration studies provide a quantifiable measure of the reinforcing effect of a drug, which is believed to be predictive of its potential for abuse. Several adaptations of the self-administration paradigm exist, the simplest of which allows participants free access to the drug under investigation. Free-access procedures allow investigators to observe patterns of drug self-administration and drug effects in a controlled setting. Allowing participants to choose between two simultaneously available reinforcers (choice procedures) is another well-established method of assessing the reinforcing effects of a drug. Offering a choice between two reinforcers (e.g. two different doses of the same drug, two different drugs, or drug and nondrug reinforcers) provides researchers with a point of comparison (e.g. between a drug of known abuse potential and a novel drug). When combined with other endpoints, such as subjective effects ratings, physiological responses, and cognitive performance, human self-administration paradigms have contributed significantly to our understanding of the factors that contribute to, maintain, and alter drug-taking behavior including: craving, positive subjective effects, toxicity, drug interactions and abstinence. This area of research has also begun to incorporate other techniques such as imaging and genetics to further understand the multifaceted nature of substance abuse. The present paper summarizes the different self-administration techniques that are commonly used today and the application of other procedures that may complement interpretation of the drug PMID:23839027

  13. Human mast cell transcriptome project.

    PubMed

    Saito, H; Nakajima, T; Matsumoto, K

    2001-05-01

    After draft reading of the human genome sequence, systemic analysis of the transcriptome (the whole transcripts present in a cell) is progressing especially in commonly available cell types. Until recently, human mast cells were not commonly available. We have succeeded to generate a substantial number of human mast cells from umbilical cord blood and from adult peripheral blood progenitors. Then, we have examined messenger RNA selectively transcribed in these mast cells using high-density oligonucleotide probe arrays. Many unexpected but important transcripts were selectively expressed in human mast cells. We discuss the results obtained from transcriptome screening by introducing our data regarding mast-cell-specific genes.

  14. Systemic IL-12 Administration Alters Hepatic Dendritic Cell Stimulation Capabilities

    PubMed Central

    Chan, Tim; Back, Timothy C.; Subleski, Jeffrey J.; Weiss, Jonathan M.; Ortaldo, John R.; Wiltrout, Robert H.

    2012-01-01

    The liver is an immunologically unique organ containing tolerogenic dendritic cells (DC) that maintain an immunosuppressive microenvironment. Although systemic IL-12 administration can improve responses to tumors, the effects of IL-12-based treatments on DC, in particular hepatic DC, remain incompletely understood. In this study, we demonstrate systemic IL-12 administration induces a 2–3 fold increase in conventional, but not plasmacytoid, DC subsets in the liver. Following IL-12 administration, hepatic DC became more phenotypically and functionally mature, resembling the function of splenic DC, but differed as compared to their splenic counterparts in the production of IL-12 following co-stimulation with toll-like receptor (TLR) agonists. Hepatic DCs from IL-12 treated mice acquired enhanced T cell proliferative capabilities similar to levels observed using splenic DCs. Furthermore, IL-12 administration preferentially increased hepatic T cell activation and IFNγ expression in the RENCA mouse model of renal cell carcinoma. Collectively, the data shows systemic IL-12 administration enables hepatic DCs to overcome at least some aspects of the inherently suppressive milieu of the hepatic environment that could have important implications for the design of IL-12-based immunotherapeutic strategies targeting hepatic malignancies and infections. PMID:22428016

  15. 78 FR 42381 - Administrative Detention of Drugs Intended for Human or Animal Use

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-07-15

    ... July 15, 2013 Part IV Department of Health and Human Services Food and Drug Administration 21 CFR Parts... / Proposed Rules#0;#0; ] DEPARTMENT OF HEALTH AND HUMAN SERVICES Food and Drug Administration 21 CFR Parts 1 and 16 Administrative Detention of Drugs Intended for Human or Animal Use AGENCY: Food and...

  16. 76 FR 25538 - Criteria Used To Order Administrative Detention of Food for Human or Animal Consumption

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-05-05

    ... Administrative Detention of Food for Human or Animal Consumption AGENCY: Food and Drug Administration, HHS... amending its regulations on administrative detention of food for human or animal consumption. As required... detention of food for human or animal consumption under the Bioterrorism Act (68 FR 25242 at 25250)....

  17. Intranasal administration of oxytocin increases human aggressive behavior.

    PubMed

    Ne'eman, R; Perach-Barzilay, N; Fischer-Shofty, M; Atias, A; Shamay-Tsoory, S G

    2016-04-01

    Considering its role in prosocial behaviors, oxytocin (OT) has been suggested to diminish levels of aggression. Nevertheless, recent findings indicate that oxytocin may have a broader influence on increasing the salience of social stimuli and may therefore, under certain circumstances, increase antisocial behaviors such as aggression. This controversy led to the following speculations: If indeed oxytocin promotes primarily prosocial behavior, administration of OT is expected to diminish levels of aggression. However, if oxytocin mainly acts to increase the salience of social stimuli, it is expected to elevate levels of aggression following provocation. In order to test this assumption we used the Social Orientation Paradigm (SOP), a monetary game played against a fictitious partner that allows measuring three types of responses in the context of provocation: an aggressive response - reducing a point from the fictitious partner, an individualistic response - adding a point to oneself, and a collaborative response - adding half a point to the partner and half a point to oneself. In the current double-blind, placebo-controlled, within-subject study design, 45 participants completed the SOP task following the administration of oxytocin or placebo. The results indicated that among subjects naïve to the procedure oxytocin increased aggressive responses in comparison with placebo. These results support the saliency hypothesis of oxytocin and suggest that oxytocin plays a complex role in the modulation of human behavior.

  18. Imaging robust microglial activation after lipopolysaccharide administration in humans with PET

    PubMed Central

    Sandiego, Christine M.; Gallezot, Jean-Dominique; Pittman, Brian; Nabulsi, Nabeel; Lim, Keunpoong; Lin, Shu-Fei; Matuskey, David; Lee, Jae-Yun; O’Connor, Kevin C.; Huang, Yiyun; Carson, Richard E.; Hannestad, Jonas; Cosgrove, Kelly P.

    2015-01-01

    Neuroinflammation is associated with a broad spectrum of neurodegenerative and psychiatric diseases. The core process in neuroinflammation is activation of microglia, the innate immune cells of the brain. We measured the neuroinflammatory response produced by a systemic administration of the Escherichia coli lipopolysaccharide (LPS; also called endotoxin) in humans with the positron emission tomography (PET) radiotracer [11C]PBR28, which binds to translocator protein, a molecular marker that is up-regulated by microglial activation. In addition, inflammatory cytokines in serum and sickness behavior profiles were measured before and after LPS administration to relate brain microglial activation with systemic inflammation and behavior. Eight healthy male subjects each had two 120-min [11C]PBR28 PET scans in 1 d, before and after an LPS challenge. LPS (1.0 ng/kg, i.v.) was administered 180 min before the second [11C]PBR28 scan. LPS administration significantly increased [11C]PBR28 binding 30–60%, demonstrating microglial activation throughout the brain. This increase was accompanied by an increase in blood levels of inflammatory cytokines, vital sign changes, and sickness symptoms, well-established consequences of LPS administration. To our knowledge, this is the first demonstration in humans that a systemic LPS challenge induces robust increases in microglial activation in the brain. This imaging paradigm to measure brain microglial activation with [11C]PBR28 PET provides an approach to test new medications in humans for their putative antiinflammatory effects. PMID:26385967

  19. Elimination of progressive mammary cancer by repeated administrations of chimeric antigen receptor-modified T cells.

    PubMed

    Globerson-Levin, Anat; Waks, Tova; Eshhar, Zelig

    2014-05-01

    Continuous oncogenic processes that generate cancer require an on-going treatment approach to eliminate the transformed cells, and prevent their further development. Here, we studied the ability of T cells expressing a chimeric antibody-based receptor (CAR) to offer a therapeutic benefit for breast cancer induced by erbB-2. We tested CAR-modified T cells (T-bodies) specific to erbB-2 for their antitumor potential in a mouse model overexpressing a human erbB-2 transgene that develops mammary tumors. Comparing the antitumor reactivity of CAR-modified T cells under various therapeutic settings, either prophylactic, prior to tumor development, or therapeutically. We found that repeated administration of CAR-modified T cells is required to eliminate spontaneously developing mammary cancer. Systemic, as well as intratumoral administered CAR-modified T cells accumulated at tumor sites and eventually eliminated the malignant cells. Interestingly, within a few weeks after a single CAR T cells' administration, and rejection of primary lesion, tumors usually relapsed both in treated mammary gland and at remote sites; however, repeated injections of CAR-modified T cells were able to control the secondary tumors. Since spontaneous tumors can arise repeatedly, especially in the case of syndromes characterized by specific susceptibility to cancer, multiple administrations of CAR-modified T cells can serve to control relapsing disease.

  20. 21 CFR 864.2280 - Cultured animal and human cells.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Cultured animal and human cells. 864.2280 Section 864.2280 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Cell And Tissue Culture Products §...

  1. 21 CFR 864.2280 - Cultured animal and human cells.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Cultured animal and human cells. 864.2280 Section 864.2280 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Cell And Tissue Culture Products §...

  2. 21 CFR 864.2280 - Cultured animal and human cells.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Cultured animal and human cells. 864.2280 Section 864.2280 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Cell And Tissue Culture Products §...

  3. Genome engineering in human cells.

    PubMed

    Song, Minjung; Kim, Young-Hoon; Kim, Jin-Soo; Kim, Hyongbum

    2014-01-01

    Genome editing in human cells is of great value in research, medicine, and biotechnology. Programmable nucleases including zinc-finger nucleases, transcription activator-like effector nucleases, and RNA-guided engineered nucleases recognize a specific target sequence and make a double-strand break at that site, which can result in gene disruption, gene insertion, gene correction, or chromosomal rearrangements. The target sequence complexities of these programmable nucleases are higher than 3.2 mega base pairs, the size of the haploid human genome. Here, we briefly introduce the structure of the human genome and the characteristics of each programmable nuclease, and review their applications in human cells including pluripotent stem cells. In addition, we discuss various delivery methods for nucleases, programmable nickases, and enrichment of gene-edited human cells, all of which facilitate efficient and precise genome editing in human cells.

  4. 78 FR 7994 - Criteria Used To Order Administrative Detention of Food for Human or Animal Consumption

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-02-05

    ... Administrative Detention of Food for Human or Animal Consumption AGENCY: Food and Drug Administration, HHS... Detention of Food for Human or Animal Consumption'' that published in the Federal Register on May 5, 2011... food for human or animal consumption which were established to implement changes to the FD&C Act...

  5. Carbon Nanotubes and Human Cells?

    ERIC Educational Resources Information Center

    King, G. Angela

    2005-01-01

    Single-walled carbon nanotubes that were chemically altered to be water soluble are shown to enter fibroblasts, T cells, and HL60 cells. Nanoparticles adversely affect immortalized HaCaT human keratinocyte cultures, indicating that they may enter cells.

  6. Identification of cells initiating human melanomas

    PubMed Central

    Schatton, Tobias; Murphy, George F.; Frank, Natasha Y.; Yamaura, Kazuhiro; Waaga-Gasser, Ana Maria; Gasser, Martin; Zhan, Qian; Jordan, Stefan; Duncan, Lyn M.; Weishaupt, Carsten; Fuhlbrigge, Robert C.; Kupper, Thomas S.; Sayegh, Mohamed H.; Frank, Markus H.

    2012-01-01

    Tumour-initiating cells capable of self-renewal and differentiation, which are responsible for tumour growth, have been identified in human haematological malignancies1,2 and solid cancers3–6. If such minority populations are associated with tumour progression in human patients, specific targeting of tumour-initiating cells could be a strategy to eradicate cancers currently resistant to systemic therapy. Here we identify a subpopulation enriched for human malignant-melanoma-initiating cells (MMIC) defined by expression of the chemoresistance mediator ABCB5 (refs 7, 8) and show that specific targeting of this tumorigenic minority population inhibits tumour growth. ABCB5+ tumour cells detected in human melanoma patients show a primitive molecular phenotype and correlate with clinical melanoma progression. In serial human-to-mouse xenotransplantation experiments, ABCB5+ melanoma cells possess greater tumorigenic capacity than ABCB5− bulk populations and re-establish clinical tumour heterogeneity. In vivo genetic lineage tracking demonstrates a specific capacity of ABCB5+ sub-populations for self-renewal and differentiation, because ABCB5+ cancer cells generate both ABCB5+ and ABCB5− progeny, whereas ABCB5− tumour populations give rise, at lower rates, exclusively to ABCB5− cells. In an initial proof-of-principle analysis, designed to test the hypothesis that MMIC are also required for growth of established tumours, systemic administration of a monoclonal antibody directed at ABCB5, shown to be capable of inducing antibody-dependent cell-mediated cytotoxicity in ABCB5+ MMIC, exerted tumour-inhibitory effects. Identification of tumour-initiating cells with enhanced abundance in more advanced disease but susceptibility to specific targeting through a defining chemoresistance determinant has important implications for cancer therapy. PMID:18202660

  7. Stem Cell Therapy and Administration Routes After Stroke.

    PubMed

    Rodríguez-Frutos, Berta; Otero-Ortega, Laura; Gutiérrez-Fernández, María; Fuentes, Blanca; Ramos-Cejudo, Jaime; Díez-Tejedor, Exuperio

    2016-10-01

    Cell-based therapy has demonstrated safety and efficacy in experimental animal models of stroke, as well as safety in stroke patients. However, various questions remain regarding the therapeutic window, dosage, route of administration, and the most appropriate cell type and source, as well as mechanisms of action and immune-modulation to optimize treatment based on stem cell therapy. Various delivery routes have been used in experimental stroke models, including intracerebral, intraventricular, subarachnoid, intra-arterial, intraperitoneal, intravenous, and intranasal routes. From a clinical point of view, it is necessary to demonstrate which is the most feasible, safest, and most effective for use with stroke patients. Therefore, further experimental studies concerning the safety, efficacy, and mechanisms of action involved in these therapeutic effects are required to determine their optimal clinical use.

  8. Human natural killer cell development.

    PubMed

    Freud, Aharon G; Caligiuri, Michael A

    2006-12-01

    Our understanding of human natural killer (NK) cell development lags far behind that of human B- or T-cell development. Much of our recent knowledge of this incomplete picture comes from experimental animal models that have aided in identifying fundamental in vivo processes, including those controlling NK cell homeostasis, self-tolerance, and the generation of a diverse NK cell repertoire. However, it has been difficult to fully understand the mechanistic details of NK cell development in humans, primarily because the in vivo cellular intermediates and microenvironments of this developmental pathway have remained elusive. Although there is general consensus that NK cell development occurs primarily within the bone marrow (BM), recent data implicate secondary lymphoid tissues as principal sites of NK cell development in humans. The strongest evidence stems from the observation that the newly described stages of human NK cell development are naturally and selectively enriched within lymph nodes and tonsils compared with blood and BM. In the current review, we provide an overview of these recent findings and discuss these in the context of existing tenets in the field of lymphocyte development.

  9. Congratulations or Condolences? The Role of Human Capital in the Cultivation of a University Administrator

    ERIC Educational Resources Information Center

    McDowell, John; Singell, Larry D., Jr.; Stater, Mark

    2009-01-01

    Administrative skill is essential to organizational effectiveness. Yet, few studies examine how human capital investments over a career affect selection into administration. We use panel data for economists to estimate the probability of choosing administration over a pure academic track. The results show that, while research-specific human…

  10. Impact of Inhomogeneous Static Magnetic Field (31.7–232.0 mT) Exposure on Human Neuroblastoma SH-SY5Y Cells during Cisplatin Administration

    PubMed Central

    Mobasheri, Hamid; Dini, Luciana

    2014-01-01

    Beneficial or adverse effects of Static Magnetic Fields (SMFs) are a large concern for the scientific community. In particular, the effect of SMF exposure during anticancer therapies still needs to be fully elucidated. Here, we evaluate the effects of SMF at induction levels that cisPt-treated cancer patients experience during the imaging process conducted in Low field (200–500 mT), Open field (300–700 mT) and/or inhomogeneous High field (1.5–3 T) Magnetic Resonance Imaging (MRI) machines. Human adrenergic neuroblastoma SH-SY5Y cells treated with 0.1 µM cisPt (i.e. the lowest concentration capable of inducing apoptosis) were exposed to SMF and their response was studied in vitro. Exposure of 0.1 µM cisPt-treated cells to SMF for 2 h decreased cell viability (30%) and caused overexpression of the apoptosis-related cleaved caspase-3 protein (46%). Furthermore, increase in ROS (Reactive Oxygen Species) production (23%) and reduction in the number of mitochondria vs controls were seen. The sole exposure of SMF for up to 24 h had no effect on cell viability but increased ROS production and modified cellular shape. On the other hand, the toxicity of cisPt was significantly prevented during 24 h exposure to SMF as shown by the levels of cell viability, cleaved caspase-3 and ROS production. In conclusion, due to the cytoprotective effect of 31.7–232.0 mT SMF on low-cisPt-concentration-treated SH-SY5Y cells, our data suggest that exposure to various sources of SMF in cancer patients under a cisPt regimen should be strictly controlled. PMID:25423171

  11. Relevance Revisited: Curriculum Development in the Humanities. No. II: Administrative Strategies for Curriculum Change.

    ERIC Educational Resources Information Center

    Mondale, Clarence C., Ed.

    Papers presented in advance of a workshop on "administrative strategies" for curricular change in the humanities and brief summaries of discussions taking place at the workshop are provided. Background papers include: "Curricular Change and the Humanities," by Edward A. Lindell; "Developing Administrative Strategies for…

  12. Human Relations Skills in Administrator Training: A Counselor Education Perspective and Contribution.

    ERIC Educational Resources Information Center

    Kelly, Eugene W., Jr.

    The success of schools depends in large measure on the effectiveness of school administrators. A major component of administrator effectiveness is human relations skills. Over the past 20 years, a large and systematic body of knowledge and training methods in human relations skills has been developed in counselor education. Given the collegial…

  13. Intranasal oxytocin administration is reflected in human saliva.

    PubMed

    Weisman, Omri; Zagoory-Sharon, Orna; Feldman, Ruth

    2012-09-01

    Following the discovery that intranasal administration of neuropeptides can reach the central nervous system, a growing number of studies applied intranasal oxytocin (OT) paradigms to demonstrate the positive effects of OT on social and emotional processes. The three-step paradigm typically included: OT administration, a 45-min waiting period, and approximately 1-h period of active drug effects when experimental manipulations are applied. Yet, this schedule has not been put to systematic validation. Utilizing a double-blind placebo-control within-subject design, ten individuals were administered OT or placebo and salivary OT was measured ten times, at baseline and nine times over four consecutive hours. OT administration induced substantial increases in salivary OT across the entire period. OT rose dramatically 15 min after administration (from 6.9 pg/ml at baseline to 1265.4 pg/ml), reached plateau at 45-120 min (range=131.6 and 105.3 pg/ml), and did not return to baseline by 4h. Results contribute to discussion on brain-periphery coordination of OT and highlight the need for further research on the temporal dynamics and durations of OT administration effects.

  14. 76 FR 36543 - Draft Guidance for Industry and Food and Drug Administration Staff: Applying Human Factors and...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-06-22

    ... HUMAN SERVICES Food and Drug Administration Draft Guidance for Industry and Food and Drug Administration... AGENCY: Food and Drug Administration, HHS. ACTION: Notice. SUMMARY: The Food and Drug Administration (FDA... and Food and Drug Administration Staff: Applying Human Factors and Usability Engineering to...

  15. Human Methadone Self-Administration and the Generalized Matching Law

    ERIC Educational Resources Information Center

    Spiga, Ralph; Maxwell, R. Stockton; Meisch, Richard A.; Grabowski, John

    2005-01-01

    The present study examined whether in humans the generalized matching law described the relation between relative responding and relative drug intake by humans under concurrent variable interval variable interval (conc VI VI) schedules of drug reinforcement. Methadone-maintained patients, stabilized on 80 mg per day of methadone, were recruited…

  16. Human Resources Administration in Education: A Management Approach. Sixth Edition.

    ERIC Educational Resources Information Center

    Rebore, Ronald W.

    This book reflects the changing aspects of school human-resources management. Current concerns include the impact of new laws related to disabilities, civil rights, family and medical leave, and the testing of school bus drivers for alcohol and controlled substances. Also examined are human resources' responsibilities to military reservists and…

  17. Human Thermoregulation After Atropine and/or Pralidoxime Administration

    DTIC Science & Technology

    1987-06-01

    regulation and performance eccrine sweat glands. Am. J. Physiol. 1981; 240:R44-R51. during prolonged exercise. Eur. J. Appl. Physiol. 1978; 38:225 15...atropine flush" accompanies these inhibitory effects on the sweat and/or pralidoxime administration. Aviat. Space Environ. Med. gland (4), but whether the...been was meoasureI hIwIce each minute by venous occlusion pieth- undertaken. ysmogrophy.->Wholo body sweating was calculated from weight Pralidoxime

  18. Intermittent parathyroid hormone administration converts quiescent lining cells to active osteoblasts.

    PubMed

    Kim, Sang Wan; Pajevic, Paola Divieti; Selig, Martin; Barry, Kevin J; Yang, Jae-Yeon; Shin, Chan Soo; Baek, Wook-Young; Kim, Jung-Eun; Kronenberg, Henry M

    2012-10-01

    Intermittent administration of parathyroid hormone (PTH) increases bone mass, at least in part, by increasing the number of osteoblasts. One possible source of osteoblasts might be conversion of inactive lining cells to osteoblasts, and indirect evidence is consistent with this hypothesis. To better understand the possible effect of PTH on lining cell activation, a lineage tracing study was conducted using an inducible gene system. Dmp1-CreERt2 mice were crossed with ROSA26R reporter mice to render targeted mature osteoblasts and their descendents, lining cells and osteocytes, detectable by 5-bromo-4-chloro-3-indolyl-β-d-galactopyranoside (X-gal) staining. Dmp1-CreERt2(+):ROSA26R mice were injected with 0.25 mg 4-OH-tamoxifen (4-OHTam) on postnatal days 3, 5, 7, 14, and 21. The animals were euthanized on postnatal day 23, 33, or 43 (2, 12, or 22 days after the last 4-OHTam injection). On day 43, mice were challenged with a subcutaneous injection of human PTH (1-34, 80 µg/kg) or vehicle once daily for 3 days. By 22 days after the last 4-OHTam injection, most X-gal (+) cells on the periosteal surfaces of the calvaria and the tibia were flat. Moreover, bone formation rate and collagen I(α1) mRNA expression were decreased at day 43 compared to day 23. After 3 days of PTH injections, the thickness of X-gal (+) cells increased, as did their expression of osteocalcin and collagen I(α1) mRNA. Electron microscopy revealed X-gal-associated chromogen particles in thin cells prior to PTH administration and in cuboidal cells following PTH administration. These data support the hypothesis that intermittent PTH treatment can increase osteoblast number by converting lining cells to mature osteoblasts in vivo.

  19. Nasal submucosal administration of antigen-presenting cells induces effective immunological responses in cancer immunotherapy.

    PubMed

    Okamoto, Yoshitaka; Fujikawa, Akira; Kurosaki, Motoyoshi; Yamasaki, Kazuki; Sakurai, Daijyu; Horiguchi, Shigetoshi; Nakayama, Toshinori

    2011-01-01

    Human NKT cells are known to have strong antitumor activities and to be activated by specific ligand, α-galactosylceramide (αGelCer). We examined the migration pattern of αGalCer-pulsed DCs and the immune responses after administration by different routes. DCs injected into nasal submucosa quickly migrated to the lateral neck lymph rather than the lateral lymph nodes. The absolute number of NKT cells and the IFN-γ-producing cells increased in peripheral blood after injection of the DCs into nasal submucosa. We conducted a phase I study with αGalCer-pulsed DCs administered in nasal submucosa of patients with head and neck cancer, and evaluated safety and feasibility. The results showed that nasal submucosal administration of α-GalCer-pulsed DCs was safe and a smaller number of these DCs could exhibit significant immune responses and some positive clinical effects. In additional study, the use of the intra-arterial infusion of activated NKT cells and the submucosal injection of α-GalCer-pulsed DCs has been shown to induce significant antitumor immunity and had beneficial clinical effects in the management of advanced head and neck squamous cell carcinoma. The NKT cell-based cancer immunotherapy may be helpful in management of head and neck cancer and needs to be explored in further detail.

  20. Human Resources Administration: A School-Based Perspective. Fourth Edition

    ERIC Educational Resources Information Center

    Smith, Richard

    2009-01-01

    Enhanced and updated, this Fourth Edition of Richard E. Smith's highly successful text examines the growing role of the principal in planning, hiring, staff development, supervision, and other human resource functions. The Fourth Edition includes new sections on ethics, induction, and the role of the mentor teacher. This edition also introduces…

  1. Annotated Bibliography on Human Relations--Teachers and Administrators.

    ERIC Educational Resources Information Center

    1987

    This annotated bibliography is a listing of 286 papers and journal articles on human relations in public education published between 1982 and 1986. Nearly 50 entries concern the principal's role as manager and instructional leader. Another 50 address teacher supervision and evaluation, while 30 others treat teacher burnout, stress, morale, and…

  2. Administration route-dependent vaccine efficiency of murine dendritic cells pulsed with antigens

    PubMed Central

    Okada, N; Tsujino, M; Hagiwara, Y; Tada, A; Tamura, Y; Mori, K; Saito, T; Nakagawa, S; Mayumi, T; Fujita, T; Yamamoto, A

    2001-01-01

     Dendritic cells (DCs) loaded with tumour antigens have been successfully used to induce protective tumour immunity in murine models and human trials. However, it is still unclear which DC administration route elicits a superior therapeutic effect. Herein, we investigated the vaccine efficiency of DC2.4 cells, a murine dendritic cell line, pulsed with ovalbumin (OVA) in the murine E.G7-OVA tumour model after immunization via various routes. After a single vaccination using 1 × 106OVA-pulsed DC2.4 cells, tumour was completely rejected in the intradermally (i.d.; three of four mice), subcutaneously (s.c.; three of four mice), and intraperitoneally (i.p.; one of four mice) immunized groups. Double vaccinations enhanced the anti-tumour effect in all groups except the intravenous (i.v.) group, which failed to achieve complete rejection. The anti-tumour efficacy of each immunization route was correlated with the OVA-specific cytotoxic T lymphocyte (CTL) activity evaluated on day 7 post-vaccination. Furthermore, the accumulation of DC2.4 cells in the regional lymph nodes was detected only in the i.d.-and s.c.-injected groups. These results demonstrate that the administration route of antigen-loaded DCs affects the migration of DCs to lymphoid tissues and the magnitude of antigen-specific CTL response. Furthermore, the immunization route affects vaccine efficiency. © 2001 Cancer Research Campaign http://www.bjcancer.com PMID:11384109

  3. Changes of human plasma dopamine-beta-hydroxylase activity after intravenous administration of theophylline.

    PubMed

    Aunis, D; Mandel, P; Miras-Portugal, M T; Coquillat, G; Rohmer, F; Warter, J M

    1975-03-01

    The intravenous administration of theophylline to ten healthy human subjects produced either an increase of circulating plasma dopamine-beta-hydroxylase or no change. The rise of plasma enzyme activity may reflect the increased peripheral catecholamine release induced by theophylline.

  4. [Improvement of light and color perception in humans upon prolonged administration of eleutherococcus].

    PubMed

    Arushanian, E B; Shikina, I B

    2004-01-01

    The chronic administration of a liquid eleuterococcus extract significantly improves light and color perception in healthy humans. Significant positive changes in eye sensitivity were observed in both morning and evening hours.

  5. Human Temperature Regulation during Exercise after Oral Pyridostigmine Administration

    DTIC Science & Technology

    1990-03-01

    or vasomotor elements) which may or during exercise . Tsk was different in the three conditions by result from increased cholinergic activity after...of cholines- oration during steady-state exercise in control experi- terase enzyme activity units by the manual delta pH method ments. Thus, the small...Reprint & Copyright © by Aerospace Medical Association, Washington. DC D T IC ELECTF 0 Human Temperature Regulation * ~During Exercise After N Oral

  6. Effect of puerarin on human choriocarcinoma cells

    PubMed Central

    Lidao, Bao; Yi, Wang; Ruilian, Ma; Xianhua, Ren; Agula, B

    2015-01-01

    Objective To discuss the effect of puerarin on human choriocarcinoma cells. Methods Survival rates under puerarin monotherapy, fluorouracil (5-FU) monotherapy and puerarin in combination with 5-FU were detected by MTT assay. Apoptotic morphology was observed with Hoechst 33258 staining. Apoptosis rates were detected with flow cytometry. Expressions of AKT, mechanistic target of rapamycin (mTOR), and P70S6K mRNAs and phosphorylated proteins were detected by RT-PCR and Western blot. Tumor-bearing mice were administered puerarin and puerarin+5-FU, and serum levels of β-human chorionic gonadotropin (β-HCG) were measured. Results Proliferation inhibition and apoptosis rates of JEG-3 cells were positively correlated with puerarin concentration, which increased in the puerarin+5-FU group. Expression levels of AKT, mTOR, P70S6K mRNAs, and phosphorylated proteins decreased significantly after action of puerarin at different concentrations. With increasing puerarin concentration, expression of cleaved-caspase-3 in JEG-3 cells increased, whereas that of Bcl-2 decreased. Puerarin significantly inhibited tumor growth in choriocarcinoma-bearing SCID mice. Serum β-HCG levels were significantly lower than those of control group after administration. Magnitude of β-HCG decline was positively correlated with concentration.. Conclusion Puerarin+5-FU inhibited proliferation of JEG-3 choriocarcinoma cells and promoted their apoptosis, being associated with the mTOR signaling pathway. PMID:28352705

  7. Cultured Human Renal Cortical Cells

    NASA Technical Reports Server (NTRS)

    1998-01-01

    During the STS-90 shuttle flight in April 1998, cultured renal cortical cells revealed new information about genes. Timothy Hammond, an investigator in NASA's microgravity biotechnology program was interested in culturing kidney tissue to study the expression of proteins useful in the treatment of kidney diseases. Protein expression is linked to the level of differentiation of the kidney cells, and Hammond had difficulty maintaining differentiated cells in vitro. Intrigued by the improvement in cell differentiation that he observed in rat renal cells cultured in NASA's rotating wall vessel (a bioreactor that simulates some aspects of microgravity) and during an experiment performed on the Russian Space Station Mir, Hammond decided to sleuth out which genes were responsible for controlling differentiation of kidney cells. To do this, he compared the gene activity of human renal cells in a variety of gravitational environments, including the microgravity of the space shuttle and the high-gravity environment of a centrifuge. Hammond found that 1,632 genes out of 10,000 analyzed changed their activity level in microgravity, more than in any of the other environments. These results have important implications for kidney research as well as for understanding the basic mechanism for controlling cell differentiation.

  8. Plasma proteomic alterations in non-human primates and humans after chronic alcohol self-administration.

    PubMed

    Freeman, Willard M; Vanguilder, Heather D; Guidone, Elizabeth; Krystal, John H; Grant, Kathleen A; Vrana, Kent E

    2011-08-01

    Objective diagnostics of excessive alcohol use are valuable tools in the identification and monitoring of subjects with alcohol use disorders. A number of potential biomarkers of alcohol intake have been proposed, but none have reached widespread clinical usage, often due to limited diagnostic sensitivity and specificity. In order to identify novel potential biomarkers, we performed proteomic biomarker target discovery in plasma samples from non-human primates that chronically self-administer high levels of ethanol. Two-dimensional difference in-gel electrophoresis (2D-DIGE) was used to quantify plasma proteins from within-subject samples collected before exposure to ethanol and after 3 months of excessive ethanol self-administration. Highly abundant plasma proteins were depleted from plasma samples to increase proteomic coverage. Altered plasma levels of serum amyloid A4 (SAA4), retinol-binding protein, inter-alpha inhibitor H4, clusterin, and fibronectin, identified by 2D-DIGE analysis, were confirmed in unmanipulated, whole plasma from these animals by immunoblotting. Examination of these target plasma proteins in human subjects with excessive alcohol consumption (and control subjects) revealed increased levels of SAA4 and clusterin and decreased levels of fibronectin compared to controls. These proteins not only serve as targets for further development as biomarker candidates or components of biomarker panels, but also add to the growing understanding of dysregulated immune function and lipoprotein metabolism with chronic, excessive alcohol consumption.

  9. Plasma proteomic alterations in non-human primates and humans after chronic alcohol self-administration

    PubMed Central

    Freeman, Willard M.; VanGuilder, Heather D.; Guidone, Elizabeth; Krystal, John H.; Grant, Kathleen A.; Vrana, Kent E.

    2011-01-01

    Objective diagnostics of excessive alcohol use are valuable tools in the identification and monitoring of subjects with alcohol use disorders. A number of potential biomarkers of alcohol intake have been proposed, but none have reached widespread clinical usage, often due to limited diagnostic sensitivity and specificity. In order to identify novel potential biomarkers, we performed proteomic biomarker target discovery in plasma samples from non-human primates that chronically self-administer high levels of ethanol. 2-dimensional in-gel electrophoresis (2D-DIGE) was used to quantify plasma proteins from within subject samples collected before exposure to ethanol and after three months of excessive ethanol self-administration. Highly abundant plasma proteins were depleted from plasma samples to increase proteomic coverage. Altered plasma levels of SAA4, RBP, ITIH4, clusterin, and fibronectin, identified by 2D-DIGE analysis, were confirmed in unmanipulated, whole plasma from these animals by immunoblotting. Examination of these target plasma proteins in human subjects with excessive alcohol consumption (and control subjects) revealed increased levels of SAA4 and clusterin and decreased levels of fibronectin compared to controls. These proteins not only serve as targets for further development as biomarker candidates or components of biomarker panels, but also add to the growing understanding of dysregulated immune function and lipoprotein metabolism with chronic, excessive alcohol consumption. PMID:21303580

  10. Human fetal mesenchymal stem cells.

    PubMed

    O'Donoghue, Keelin; Chan, Jerry

    2006-09-01

    Stem cells have been isolated at all stages of development from the early developing embryo to the post-reproductive adult organism. However, the fetal environment is unique as it is the only time in ontogeny that there is migration of stem cells in large numbers into different organ compartments. While fetal neural and haemopoietic stem cells (HSC) have been well characterised, only recently have mesenchymal stem cells from the human fetus been isolated and evaluated. Our group have characterised in human fetal blood, liver and bone marrow a population of non-haemopoietic, non-endothelial cells with an immunophenotype similar to adult bone marrow-derived mesenchymal stem cells (MSC). These cells, human fetal mesenchymal stem cells (hfMSC), are true multipotent stem cells with greater self-renewal and differentiation capacity than their adult counterparts. They circulate in first trimester fetal blood and have been found to traffic into the maternal circulation, engrafting in bone marrow, where they remain microchimeric for decades after pregnancy. Though fetal microchimerism has been implicated in the pathogenesis of autoimmune disease, the biological role of hfMSC microchimerism is unknown. Potential downstream applications of hfMSC include their use as a target cell for non-invasive pre-natal diagnosis from maternal blood, and for fetal cellular and gene therapy. Using hfMSC in fetal therapy offers the theoretical advantages of avoidance of immune rejection, increased engraftment, and treatment before disease pathology sets in. Aside from allogeneic hfMSC in utero transplantation, the use of autologous hfMSC has been brought a step forward with the development of early blood sampling techniques, efficient viral transduction and clonal expansion. Work is ongoing to determine hfMSC fate post-transplantation in murine models of genetic disease. In this review we will examine what is known about hfMSC biology, as well as discussing areas for future research. The

  11. Differentiated human stem cells resemble fetal, not adult, β cells.

    PubMed

    Hrvatin, Sinisa; O'Donnell, Charles W; Deng, Francis; Millman, Jeffrey R; Pagliuca, Felicia Walton; DiIorio, Philip; Rezania, Alireza; Gifford, David K; Melton, Douglas A

    2014-02-25

    Human pluripotent stem cells (hPSCs) have the potential to generate any human cell type, and one widely recognized goal is to make pancreatic β cells. To this end, comparisons between differentiated cell types produced in vitro and their in vivo counterparts are essential to validate hPSC-derived cells. Genome-wide transcriptional analysis of sorted insulin-expressing (INS(+)) cells derived from three independent hPSC lines, human fetal pancreata, and adult human islets points to two major conclusions: (i) Different hPSC lines produce highly similar INS(+) cells and (ii) hPSC-derived INS(+) (hPSC-INS(+)) cells more closely resemble human fetal β cells than adult β cells. This study provides a direct comparison of transcriptional programs between pure hPSC-INS(+) cells and true β cells and provides a catalog of genes whose manipulation may convert hPSC-INS(+) cells into functional β cells.

  12. Embryonic stem cell patents and human dignity.

    PubMed

    Resnik, David B

    2007-09-01

    This article examines the assertion that human embryonic stem cells patents are immoral because they violate human dignity. After analyzing the concept of human dignity and its role in bioethics debates, this article argues that patents on human embryos or totipotent embryonic stem cells violate human dignity, but that patents on pluripotent or multipotent stem cells do not. Since patents on pluripotent or multipotent stem cells may still threaten human dignity by encouraging people to treat embryos as property, patent agencies should carefully monitor and control these patents to ensure that patents are not inadvertently awarded on embryos or totipotent stem cells.

  13. Embryonic Stem Cell Patents and Human Dignity

    PubMed Central

    Resnik, David B.

    2009-01-01

    This article examines the assertion that human embryonic stem cells patents are immoral because they violate human dignity. After analyzing the concept of human dignity and its role in bioethics debates, this article argues that patents on human embryos or totipotent embryonic stem cells violate human dignity, but that patents on pluripotent or multipotent stem cells do not. Since patents on pluripotent or multipotent stem cells may still threaten human dignity by encouraging people to treat embryos as property, patent agencies should carefully monitor and control these patents to ensure that patents are not inadvertently awarded on embryos or totipotent stem cells. PMID:17922198

  14. Intranasal Neuropeptide Administration To Target the Human Brain in Health and Disease.

    PubMed

    Spetter, Maartje S; Hallschmid, Manfred

    2015-08-03

    Central nervous system control of metabolic function relies on the input of endocrine messengers from the periphery, including the pancreatic hormone insulin and the adipokine leptin. This concept primarily derives from experiments in animals where substances can be directly applied to the brain. A feasible approach to study the impact of peptidergic messengers on brain function in humans is the intranasal (IN) route of administration, which bypasses the blood-brain barrier and delivers neuropeptides to the brain compartment, but induces considerably less, if any, peripheral uptake than other administration modes. Experimental IN insulin administration has been extensively used to delineate the role of brain insulin signaling in the control of energy homeostasis, but also cognitive function in healthy humans. Clinical pilot studies have found beneficial effects of IN insulin in patients with memory deficits, suggesting that the IN delivery of this and other peptides bears some promise for new, selectively brain-targeted pharmaceutical approaches in the treatment of metabolic and cognitive disorders. More recently, experiments relying on the IN delivery of the hypothalamic hormone oxytocin, which is primarily known for its involvement in psychosocial processes, have provided evidence that oxytocin influences metabolic control in humans. The IN administration of leptin has been successfully tested in animal models but remains to be investigated in the human setting. We briefly summarize the literature on the IN administration of insulin, leptin, and oxytocin, with a particular focus on metabolic effects, and address limitations and perspectives of IN neuropeptide administration.

  15. Long-term administration of inulin-type fructans has no significant lipid-lowering effect in normolipidemic humans.

    PubMed

    Forcheron, Fabien; Beylot, Michel

    2007-08-01

    Short-term studies have shown that the addition to diet of inulin-type fructans, a nondigestible carbohydrate, may have a plasma lipid-lowering effect in humans. Whether this beneficial effect persists during long-term administration has not been determined. The study was aimed at determining whether a prolonged (6 months) administration of inulin-type fructans to healthy subjects has a lipid-lowering action. In a double-blind, randomized, placebo-controlled study, 17 healthy subjects were studied before and after 6 months of daily administration of placebo (8 subjects) or 10 g of a mix of inulin and oligofructose (9 subjects). During this 6-month period, they consumed their usual diet and did not modify their everyday way of life. We measured plasma lipid concentrations; cholesterol synthesis and hepatic lipogenesis; and adipose tissue and circulating mononuclear cell messenger RNA concentrations of key regulatory genes of cholesterol metabolism. Compared with the administration of placebo, the administration of inulin-type fructans had no effect on plasma triacylglycerol concentrations and hepatic lipogenesis and induced only a nonsignificant trend for decreased plasma total and low-density lipoprotein cholesterol levels and increased high-density lipoprotein cholesterol concentration. Cholesterol synthesis was not significantly modified. Of all the messenger RNA concentrations measured, none was significantly modified by the administration of inulin-type fructans. In conclusion, contrary to what was observed in short-term studies, we observed no significant beneficial effect of a long-term (6-month) administration of inulin-type fructans on plasma lipids in healthy human subjects.

  16. Intravesical administration of exogenous microRNA-145 as a therapy for mouse orthotopic human bladder cancer xenograft

    PubMed Central

    Inamoto, Teruo; Taniguchi, Kohei; Takahara, Kiyoshi; Iwatsuki, Ayako; Takai, Tomoaki; Komura, Kazumasa; Yoshikawa, Yuki; Uchimoto, Taizo; Saito, Kenkichi; Tanda, Naoki; Kouno, Junko; Minami, Koichiro; Uehara, Hirofumi; Hirano, Hajime; Nomi, Hayahito; Kiyama, Satoshi; Akao, Yukihiro; Azuma, Haruhito

    2015-01-01

    We previously reported that the level of microRNA (miR)-145 is attenuated in human bladder cancer cells. In this current study, we investigated whether intravesical administration of miR-145 could be a potential therapeutic strategy for controlling bladder cancer by using an orthotopic human bladder cancer xenograft model. Following transfection of 253J B-V cells with miR-145, the effects of the ectopic expression of miR-145 were examined by performing MTT, Western blotting analysis, Hoechst33342 staining, and wound healing assay in vitro. Also, a mouse orthotopic human bladder cancer model was established by inoculating 253J B-V cells into the bladder wall of mice. The anti-cancer effects of intravesical injections of miR-145 into these mice were then assessed. Transfection of 253J B-V cells with miR-145 induced apoptosis and suppression of cell migration in vitro. Western blotting showed that the levels of c-Myc, socs7, FSCN1, E-cadherin, β-catenin, and catenin δ-1 were decreased and that the PI3K/Akt and Erk1/2 signaling pathways were increased in compensatory fashion. In vivo, mice treated with miR-145 showed 76% inhibition of tumor growth, with a significant prolongation of animal survival (p = 0.0183 vs. control). Western blotting showed that both apoptosis and cell motility-related genes were significantly decreased as seen in vitro. Furthermore, PI3k/Akt and Erk1/2 signaling pathways, which were activated in a compensatory manner in vitro, were decreased in vivo. Intravesical administration of exogenous miR-145 was thus concluded to be a valid therapy for bladder cancer in this human bladder cancer xenograft model. PMID:26036261

  17. Administrative detention of drugs intended for human or animal use. Final rule.

    PubMed

    2014-05-29

    The Food and Drug Administration (FDA or the Agency) is implementing administrative detention authority with respect to drugs intended for human or animal use as authorized by amendments made to the Federal Food, Drug, and Cosmetic Act (the FD&C Act) by the Food and Drug Administration Safety and Innovation Act (FDASIA). FDA's administrative detention authority with respect to drugs allows FDA to better protect the integrity of the drug supply chain. Specifically, FDA is able to administratively detain drugs encountered during an inspection that an authorized FDA representative conducting an inspection has reason to believe are adulterated or misbranded. This authority is intended to protect the public by preventing distribution or subsequent use of drugs encountered during inspections that are believed to be adulterated or misbranded, until FDA has had time to consider what action it should take concerning the drugs, and to initiate legal action, if appropriate.

  18. Neoplastic transformation of human cells in vitro.

    PubMed

    Rhim, J S

    1993-01-01

    Efforts to investigate the progression of events that lead normal human cells in culture to become neoplastic in response to carcinogenic agents have been aided by the development of the suitable in vitro model systems. For initial human cell transformation studies, a flat, nontumorigenic clonal line, originally derived from a human osteosarcoma (HOS), was used. When treated with chemical carcinogens such as N-methyl-N-nitro-N-nitrosoguanidine (MNNG) and 3-methyl-cholanthrene (3MC), the HOS cells underwent morphological alterations and acquired tumorigenic properties. These cell lines were very useful inasmuch as a non-ras cellular transforming gene, met, and an activated H-ras oncogene have been isolated from MNNG-transformed and 3MC-transformed HOS lines, respectively, by DNA transfection procedure. Alteration of p53 gene in chemically transformed HOS cell lines has recently been shown. Although carcinogens cause human cancer, normal human cells in culture have proven difficult to achieve. Neoplastic transformation of human cells in culture has recently been achieved by a stepwise fashion-immortalization and conversion of the immortalized cells to tumorigenic cells. One of the critical initial events in the progression of normal human cells to tumor cells is the escape from cellular senescence. With few exceptions, normal human cells require immortalization to provide a practical system for transformation studies. Thus, the role of carcinogenic agents in the development of human cancers is now being defined using a variety of human cells. The neoplastic transformation in human cell cultures is reviewed. In doing so, this author attempts to put into perspective the history of human cell transformation by carcinogenic agents, and to discuss the current state of the art in transformation of human cells in culture; thus providing insight into the molecular and cellular mechanisms involved in the conversion of normal cells to a neoplastic state of growth.

  19. A critical review of human endotoxin administration as an experimental paradigm of depression

    PubMed Central

    DellaGioia, Nicole; Hannestad, Jonas

    2009-01-01

    The syndrome called depression may represent the common final pathway at which different aetiopathogenic processes converge. One such atiopathogenic process is innate immune system activation. Some depressed patients increased levels of inflammatory cytokines and other immunologic abnormalities. It is not known whether immune system activation contributes to the pathogenesis of depressive symptoms. Supporting this possibility is the observation that in both rodents and humans, exogenous immune stimuli such as endotoxin can produce symptoms that resemble depression. A new approach to depression research would be to use immune stimuli to elicit depressive symptoms in humans. Here we review each of the symptoms elicited in humans by endotoxin administration, and compare this model to two other immune depression paradigms: interferon-alpha treatment and typhoid vaccine administration, to assess to what degree endotoxin administration represents a valid model of immune depression. We also review corresponding behavioral changes in rodents and the potential molecular pathways through which immune system activation produces each symptom. PMID:19666048

  20. Self-Administration of Cannabinoids by Experimental Animals and Human Marijuana Smokers

    PubMed Central

    Justinova, Zuzana; Goldberg, Steven R.; Heishman, Stephen J.; Tanda, Gianluigi

    2009-01-01

    Drug self-administration behavior has been one of the most direct and productive approaches for studying the reinforcing effects of psychoactive drugs, which are critical in determining their abuse potential. Cannabinoids, which are usually abused by humans in the form of marijuana, have become the most frequently abused illicit class of drugs in the United States. The early elucidation of the structure and stereochemistry of delta-9-tetrahydrocannabinol (THC) in 1964, which is now recognized as the principal psychoactive ingredient in marijuana, activated cannabinoid research worldwide. This review examines advances in research on cannabinoid self-administration behavior by humans and laboratory animals. There have been numerous laboratory demonstrations of the reinforcing effects of cannabinoids in human subjects, but reliable self-administration of cannabinoids by laboratory animals has only recently been demonstrated. It has now been shown that strong and persistent self-administration behavior can be maintained in experimentally and drug-naïve squirrel monkeys by doses of THC comparable to those in marijuana smoke inhaled by humans. Furthermore, reinforcing effects of some synthetic CB1 cannabinoid agonists have been recently reported using intravenous and intracerebroventricular self-administration procedures in rats and mice. These findings support previous conclusions that THC has a pronounced abuse liability comparable to other drugs of abuse under certain experimental conditions. Self-administration of THC by squirrel monkeys provides the most reliable animal model for human marijuana abuse available to date. This animal model now makes it possible to study the relative abuse liability of other natural and synthetic cannabinoids and to preclinically assess new therapeutic strategies for the treatment or prevention of marijuana abuse in humans. PMID:15932767

  1. Neoplastic transformation of human cells

    NASA Technical Reports Server (NTRS)

    Goth-Goldstein, Regine

    1995-01-01

    The goal of this project was to gain a better understanding of the cellular mechanisms of cancer induction by ionizing radiation as a risk assessment for workers subjected to high LET irradiation such as that found in space. The following ions were used for irradiation: Iron, Argon, Neon, and Lanthanum. Two tests were performed: growth in low serum and growth in agar were used as indicators of cell transformation. The specific aims of this project were to: (1) compare the effectiveness of various ions on degree of transformation of a single dose of the same RBE; (2) determine if successive irradiations with the same ion (Ge 600 MeV/u) increases the degree of transformation; (3) test if clones with the greatest degree of transformation produce tumors in nude mice; and (4) construct a cell hybrid of a transformed and control (non-transformed) clone. The cells used for this work are human mammary epithelial cells with an extended lifespan and selected for growth in MEM + 10% serum.

  2. Study of mouse spleen "natural cytotoxic" (NC) cell activity related to in vivo administration of B4 coxsackievirus.

    PubMed

    Voiculescu, C; Roşu, L; Rogoz, S

    1987-01-01

    By using A2G 40-day-old mice as responders, the influence of in vivo infection with a human strain of B4 coxsackievirus on "natural cytotoxic" (NC) cell activity was assayed, in relation to other immunomodulating treatments (beta-interferon, interleukin-1, or prostaglandin-E2). A significant increase of NC cell cytolysis was noticed in virus-infected mouse group, as compared to the control. The NC cell stimulation exerted by in vivo administration of beta-interferon or of interleukin-1 was not modified in virus-infected groups, but coxsackievirus was able to prevent the inhibitory effects of prostaglandin-E2 at the NC cell activity level when the cytotoxic assays were performed with 20:1 and 10:1 effector/"target" cell ratios, respectively. A possible extrapolation of results in the human medical practice is discussed.

  3. Human melanoma immunotherapy using tumor antigen-specific T cells generated in humanized mice

    PubMed Central

    Hu, Zheng; Xia, Jinxing; Fan, Wei; Wargo, Jennifer; Yang, Yong-Guang

    2016-01-01

    A major factor hindering the exploration of adoptive immunotherapy in preclinical settings is the limited availability of tumor-reactive human T cells. Here we developed a humanized mouse model that permits large-scale production of human T cells expressing the engineered melanoma antigen MART-1-specific TCR. Humanized mice, made by transplantation of human fetal thymic tissue and CD34+ cells virally-transduced with HLA class I-restricted melanoma antigen (MART-1)-specific TCR gene, showed efficient development of MART-1-TCR+ human T cells with predominantly CD8+ cells. Importantly, MART-1-TCR+CD8+ T cells developing in these mice were capable of mounting antigen-specific responses in vivo, as evidenced by their proliferation, phenotypic conversion and IFN-γ production following MART-1 peptide immunization. Moreover, these MART-1-TCR+CD8+ T cells mediated efficient killing of melanoma cells in an HLA/antigen-dependent manner. Adoptive transfer of in vitro expanded MART-1-TCR+CD8+ T cells induced potent antitumor responses that were further enhanced by IL-15 treatment in melanoma-bearing recipients. Finally, a short incubation of MART-1-specific T cells with rapamycin acted synergistically with IL-15, leading to significantly improved tumor-free survival in recipients with metastatic melanoma. These data demonstrate the practicality of using humanized mice to produce potentially unlimited source of tumor-specific human T cells for experimental and preclinical exploration of cancer immunotherapy. This study also suggests that pretreatment of tumor-reactive T cells with rapamycin in combination with IL-15 administration may be a novel strategy to improve the efficacy of adoptive T cell therapy. PMID:26824989

  4. Human immunodeficiency virus can productively infect cultured human glial cells.

    PubMed

    Cheng-Mayer, C; Rutka, J T; Rosenblum, M L; McHugh, T; Stites, D P; Levy, J A

    1987-05-01

    Six isolates of the human immunodeficiency virus (HIV) showed differences in their ability to productively infect glioma-derived cell lines and early-passage human brain cell cultures. Susceptibility to HIV infection correlated well with the expression of the astrocyte marker glial fibrillary acidic protein. The CD4 molecule was expressed on some, but not all, of the brain-derived cells; however, no correlation was observed between CD4 protein expression and susceptibility to virus infection. The results show that HIV can productively infect human brain cells, particularly those of glial origin, and suggest that these cell types in the brain can harbor the virus.

  5. Ultrastructural Changes in Murine Peritoneal Cells Following Cyclophosphamide Administration

    PubMed Central

    Chin, K. N.; Hudson, G.

    1974-01-01

    Peritoneal cells were studied at intervals of between 6 h and 30 days following a single intravenous injection of a sublethal dose of cyclophosphamide. With the electron microscope, evidence of cell damage and death could be seen at 6 h, and by 12 h large numbers of dead cells were noted, either lying free or within the cytoplasm of macrophages. Most of the damaged cells were lymphocytes but degenerating blast cells, eosinophils, neutrophils and mast cells were also identified. Nuclei were seen in which margination of chromatin had occurred but nuclei of uniform density were also prominent and showed irregular shape and lobulation. Macrophages exhibited all stages of phagocytosis and digestion and a few phagocytes of atypical appearance were noted. By 24 h most of the dead cells lay within the cytoplasm of macrophages which showed many phagocytic inclusions as well as lipid droplets. By 6 days, the peritoneal cells had regained normal appearances although the proportion of lymphoid cells was still reduced. By the 18th day, all features were indistinguishable from normal. The changes observed showed a general similarity to those noted previously in the lymphoreticular cells of the Peyer's patch; they provide no evidence that the environment of the peritoneal cavity protects cells against the action of cyclophosphamide. ImagesFigs. 4-6Figs. 7-9Figs. 10-12Figs. 1-3 PMID:4447790

  6. Administrative Coordination in Non-Profit Human Service Delivery Networks: The Role of Competition and Trust

    PubMed Central

    Bunger, Alicia C.

    2014-01-01

    Non-profit human service organizations operating within the same regional network are often faced with dual pressure to compete as well as coordinate administrative operations (by sharing funding, staff or space) to enhance efficiency. Emerging evidence has demonstrated that competing organizations coordinate, despite the risks. Trust, or perceived trustworthiness between two organizations may mitigate the negative influence of competition on coordination, however there have been few explicit tests of this hypothesis among non-profit organizations. Drawing on quantitative data collected from a network of 36 non-profit children’s behavioral health organizations, this paper empirically tests how competition and perceived trustworthiness interact to influence administrative coordination. Results support the hypothesis that trustworthiness moderates the influence of competition on administrative coordination. Findings suggest that as competing non-profit leaders build trust, the more their agencies coordinate their administrative functions. This study highlights the importance of leaders’ perceptions for organizational strategy. PMID:25349468

  7. Changes of human plasma dopamine-beta-hydroxylase activity after intravenous administration of theophylline.

    PubMed Central

    Aunis, D; Mandel, P; Miras-Portugal, M T; Coquillat, G; Rohmer, F; Warter, J M

    1975-01-01

    The intravenous administration of theophylline to ten healthy human subjects produced either an increase of circulating plasma dopamine-beta-hydroxylase or no change. The rise of plasma enzyme activity may reflect the increased peripheral catecholamine release induced by theophylline. PMID:1137731

  8. The role of human drug self-administration procedures in the development of medications.

    PubMed

    Comer, S D; Ashworth, J B; Foltin, R W; Johanson, C E; Zacny, J P; Walsh, S L

    2008-07-01

    The purpose of this review is to illustrate the utility and value of employing human self-administration procedures in medication development, including abuse liability assessments of novel medications and evaluation of potential pharmacotherapies for substance use disorders. Traditionally, human abuse liability testing has relied primarily on subjective reports describing drug action by use of questionnaires; similarly, drug interactions between putative treatment agents and the drugs of abuse have relied on these measures. Subjective reports are highly valued because they provide qualitative and quantitative information about the characteristics of central and peripheral pharmacodynamic effects as well as safety and tolerability. However, self-administration procedures directly examine the behavior of interest-that is, drug taking. The present paper (1) reviews the most commonly used human self-administration procedures, (2) discusses the concordance of subjective reports and self-administration within the context of medications development for substance use disorders, focusing primarily on illustrative examples from development efforts with opioid and cocaine dependence, and (3) explores the utility of applying self-administration procedures to assess the abuse liability of novel compounds, including "abuse-deterrent" formulations (ADFs). The review will focus on opioid and cocaine dependence because a rich database from both clinical laboratory and clinical trial research exists for these two drug classes. The data reviewed suggest that drug-induced changes in self-administration and subjective effects are not always concordant. Therefore, assessment of self-administration in combination with subjective effects provides a more comprehensive picture that may have improved predictive validity for translating to the clinical setting.

  9. The role of human drug self-administration procedures in the development of medications

    PubMed Central

    Comer, SD; Ashworth, JB; Foltin, RW; Johanson, CE; Zacny, JP; Walsh, SL

    2008-01-01

    The purpose of this review is to illustrate the utility and value of employing human self-administration procedures in medication development, including abuse liability assessments of novel medications and evaluation of potential pharmacotherapies for substance use disorders. Traditionally, human abuse liability testing has relied primarily on subjective reports describing drug action by use of questionnaires; similarly, drug interactions between putative treatment agents and the drugs of abuse have relied on these measures. Subjective reports are highly valued because they provide qualitative and quantitative information about the characteristics of central and peripheral pharmacodynamic effects as well as safety and tolerability. However, self-administration procedures directly examine the behavior of interest – that is, drug taking. The present paper 1) reviews the most commonly used human self-administration procedures, 2) discusses the concordance of subjective reports and self-administration within the context of medications development for substance use disorders, focusing primarily on illustrative examples from development efforts with opioid and cocaine dependence, and 3) explores the utility of applying self-administration procedures to assess the abuse liability of novel compounds, including “abuse deterrent” formulations (ADFs). The review will focus on opioid and cocaine dependence because a rich database from both clinical laboratory and clinical trial research exists for these two drug classes. The data reviewed suggest that drug-induced changes in self-administration and subjective effects are not always concordant. Therefore, assessment of self-administration in combination with subjective effects provides a more comprehensive picture that may have improved predictive validity for translating to the clinical setting. PMID:18436394

  10. Sclerostin Antibody Administration Converts Bone Lining Cells Into Active Osteoblasts.

    PubMed

    Kim, Sang Wan; Lu, Yanhui; Williams, Elizabeth A; Lai, Forest; Lee, Ji Yeon; Enishi, Tetsuya; Balani, Deepak H; Ominsky, Michael S; Ke, Hua Zhu; Kronenberg, Henry M; Wein, Marc N

    2016-11-14

    Sclerostin antibody (Scl-Ab) increases osteoblast activity, in part through increasing modeling-based bone formation on previously quiescent surfaces. Histomorphometric studies have suggested that this might occur through conversion of bone lining cells into active osteoblasts. However, direct data demonstrating Scl-Ab-induced conversion of lining cells into active osteoblasts are lacking. Here, we used in vivo lineage tracing to determine if Scl-Ab promotes the conversion of lining cells into osteoblasts on periosteal and endocortical bone surfaces in mice. Two independent, tamoxifen-inducible lineage-tracing strategies were used to label mature osteoblasts and their progeny using the DMP1 and osteocalcin promoters. After a prolonged "chase" period, the majority of labeled cells on bone surfaces assumed a thin, quiescent morphology. Then, mice were treated with either vehicle or Scl-Ab (25 mg/kg) twice over the course of the subsequent week. After euthanization, marked cells were enumerated, their thickness quantified, and proliferation and apoptosis examined. Scl-Ab led to a significant increase in the average thickness of labeled cells on periosteal and endocortical bone surfaces, consistent with osteoblast activation. Scl-Ab did not induce proliferation of labeled cells, and Scl-Ab did not regulate apoptosis of labeled cells. Therefore, direct reactivation of quiescent bone lining cells contributes to the acute increase in osteoblast numbers after Scl-Ab treatment in mice. © 2017 American Society for Bone and Mineral Research.

  11. Satellite cells in human skeletal muscle plasticity.

    PubMed

    Snijders, Tim; Nederveen, Joshua P; McKay, Bryon R; Joanisse, Sophie; Verdijk, Lex B; van Loon, Luc J C; Parise, Gianni

    2015-01-01

    Skeletal muscle satellite cells are considered to play a crucial role in muscle fiber maintenance, repair and remodeling. Our knowledge of the role of satellite cells in muscle fiber adaptation has traditionally relied on in vitro cell and in vivo animal models. Over the past decade, a genuine effort has been made to translate these results to humans under physiological conditions. Findings from in vivo human studies suggest that satellite cells play a key role in skeletal muscle fiber repair/remodeling in response to exercise. Mounting evidence indicates that aging has a profound impact on the regulation of satellite cells in human skeletal muscle. Yet, the precise role of satellite cells in the development of muscle fiber atrophy with age remains unresolved. This review seeks to integrate recent results from in vivo human studies on satellite cell function in muscle fiber repair/remodeling in the wider context of satellite cell biology whose literature is largely based on animal and cell models.

  12. Sequence dependence of administration of human recombinant tumor necrosis factor and interleukin-2 in murine tumor therapy.

    PubMed

    Zimmerman, R J; Gauny, S; Chan, A; Landre, P; Winkelhake, J L

    1989-02-01

    Simultaneous administration of recombinant human tumor necrosis factor (rhTNF) and interleukin-2 (rhIL-2) has been shown to block tumor take in murine models. We investigated the effects of sequence and schedule of administration as a function of tumor burden with two tumor models (B16 and Meth A). rhTNF followed by rhIL-2 had extraordinary antitumor efficacy, but rhIL-2 followed by rhTNF was much less effective. Sequential rhTNF/rhIL-2 therapy resulted in complete tumor regression, whereas simultaneous therapy resulted in complete tumor regression, whereas simultaneous therapy resulted in only reduced growth rate. Experiments with genetically immunodeficient mice suggested that T cell factors may be required for synergistic antitumor activity.

  13. Interferon Production by Human Cells In Vitro

    PubMed Central

    Spina, Celsa A.; Chang, R. Shihman; Mishra, L.; Golden, H. Dean

    1972-01-01

    The relative capacity of several types of human cells and tissue to produce interferon was studied. Types of cells and tissue included were fibroblasts from embryos, foreskins, and biopsied skins; amnion cells; peripheral leukocytes; established lymphoid cell lines; established heteroploid cell lines; and chorioamniotic membrane. When Newcastle disease virus was used as the inducer, fibroblasts and amnion cells produced more interferon per 106 cells than leukocytes, lymphoid cells, and heteroploid cells. Only minor variations in interferon-producing capacity were observed among fibroblasts from 36 persons. Culture passage level, cell concentration, and inducer were factors that significantly affected interferon production. PMID:4344957

  14. Human umbilical cord blood cells and diabetes mellitus: recent advances.

    PubMed

    Reddi, Alluru S; Kothari, Neil; Kuppasani, Kishore; Ende, Norman

    2015-01-01

    Stem cell therapy for patients with diabetes is an area of great interest to both scientists and clinicians. Human umbilical cord blood cells (HUCBCs) are being increasingly used as a source of stem cells for cell-based therapy for diabetes because these cells can differentiate into pancreatic islet β-cells. Administration of HUCBCs has been shown to lower blood glucose levels in diabetic animal models. The use of autologous HUCBC transfusion in type 1 diabetic children has not shown any benefit. However, "Stem Cell Educator" therapy has shown promise in long term lowering of blood glucose levels in both type 1 and type 2 diabetic patients. In this review, we will briefly discuss recent advances in HUCBC therapy in the treatment of diabetes and some of its complications.

  15. Endothelial cells derived from human embryonic stem cells

    NASA Astrophysics Data System (ADS)

    Levenberg, Shulamit; Golub, Justin S.; Amit, Michal; Itskovitz-Eldor, Joseph; Langer, Robert

    2002-04-01

    Human embryonic stem cells have the potential to differentiate into various cell types and, thus, may be useful as a source of cells for transplantation or tissue engineering. We describe here the differentiation steps of human embryonic stem cells into endothelial cells forming vascular-like structures. The human embryonic-derived endothelial cells were isolated by using platelet endothelial cell-adhesion molecule-1 (PECAM1) antibodies, their behavior was characterized in vitro and in vivo, and their potential in tissue engineering was examined. We show that the isolated embryonic PECAM1+ cells, grown in culture, display characteristics similar to vessel endothelium. The cells express endothelial cell markers in a pattern similar to human umbilical vein endothelial cells, their junctions are correctly organized, and they have high metabolism of acetylated low-density lipoprotein. In addition, the cells are able to differentiate and form tube-like structures when cultured on matrigel. In vivo, when transplanted into SCID mice, the cells appeared to form microvessels containing mouse blood cells. With further studies, these cells could provide a source of human endothelial cells that could be beneficial for potential applications such as engineering new blood vessels, endothelial cell transplantation into the heart for myocardial regeneration, and induction of angiogenesis for treatment of regional ischemia.

  16. Contraceptive gossypol blocks cell-to-cell communication in human and rat cells.

    PubMed

    Hervé, J C; Pluciennik, F; Bastide, B; Cronier, L; Verrecchia, F; Malassiné, A; Joffre, M; Délèze, J

    1996-10-17

    Gossypol (a polycyclic lipophilic agent naturally present in cottonseed, known as a potent non-steroid antifertility agent and a non-specific enzyme inhibitor) irreversibly impaired the intercellular communication between homologous pairs of various cultured cells, from man or rat, involved (Sertoli or trophoblastic cells) or not involved (ventricular myocytes) in steroidogenesis, in a dose-dependent manner. In serum-free assays, a rapid junctional uncoupling occurred in non-cytotoxic conditions. At 5 microM (approximately twice the peak plasma concentration measured in human patients during chronic administration), gap junctional communication was interrupted within 4 to 10 min, without concomitant rise in the intracellular Ca2+ concentration. The latter importantly increased when gossypol treatment was prolonged (cytotoxic effect). The short term uncoupling effect of gossypol was prevented by serum proteins, but long-lasting treatments (48 h) with moderate concentrations (3 microM) elicited junctional uncoupling and impeded the in vitro differentiation of human trophoblasts.

  17. Evidence for Human Lung Stem Cells

    PubMed Central

    Kajstura, Jan; Rota, Marcello; Hall, Sean R.; Hosoda, Toru; D’Amario, Domenico; Sanada, Fumihiro; Zheng, Hanqiao; Ogórek, Barbara; Rondon-Clavo, Carlos; Ferreira-Martins, João; Matsuda, Alex; Arranto, Christian; Goichberg, Polina; Giordano, Giovanna; Haley, Kathleen J.; Bardelli, Silvana; Rayatzadeh, Hussein; Liu, Xiaoli; Quaini, Federico; Liao, Ronglih; Leri, Annarosa; Perrella, Mark A.; Loscalzo, Joseph; Anversa, Piero

    2011-01-01

    BACKGROUND Although progenitor cells have been described in distinct anatomical regions of the lung, description of resident stem cells has remained elusive. METHODS Surgical lung-tissue specimens were studied in situ to identify and characterize human lung stem cells. We defined their phenotype and functional properties in vitro and in vivo. RESULTS Human lungs contain undifferentiated human lung stem cells nested in niches in the distal airways. These cells are self-renewing, clonogenic, and multipotent in vitro. After injection into damaged mouse lung in vivo, human lung stem cells form human bronchioles, alveoli, and pulmonary vessels integrated structurally and functionally with the damaged organ. The formation of a chimeric lung was confirmed by detection of human transcripts for epithelial and vascular genes. In addition, the self-renewal and long-term proliferation of human lung stem cells was shown in serial-transplantation assays. CONCLUSIONS Human lungs contain identifiable stem cells. In animal models, these cells participate in tissue homeostasis and regeneration. They have the undemonstrated potential to promote tissue restoration in patients with lung disease. (Funded by the National Institutes of Health.) PMID:21561345

  18. Trophoblast lineage cells derived from human induced pluripotent stem cells

    SciTech Connect

    Chen, Ying; Wang, Kai; Chandramouli, Gadisetti V.R.; Knott, Jason G.; Leach, Richard

    2013-07-12

    Highlights: •Epithelial-like phenotype of trophoblast lineage cells derived from human iPS cells. •Trophoblast lineage cells derived from human iPS cells exhibit trophoblast function. •Trophoblasts from iPS cells provides a proof-of-concept in regenerative medicine. -- Abstract: Background: During implantation, the blastocyst trophectoderm attaches to the endometrial epithelium and continues to differentiate into all trophoblast subtypes, which are the major components of a placenta. Aberrant trophoblast proliferation and differentiation are associated with placental diseases. However, due to ethical and practical issues, there is almost no available cell or tissue source to study the molecular mechanism of human trophoblast differentiation, which further becomes a barrier to the study of the pathogenesis of trophoblast-associated diseases of pregnancy. In this study, our goal was to generate a proof-of-concept model for deriving trophoblast lineage cells from induced pluripotency stem (iPS) cells from human fibroblasts. In future studies the generation of trophoblast lineage cells from iPS cells established from patient’s placenta will be extremely useful for studying the pathogenesis of individual trophoblast-associated diseases and for drug testing. Methods and results: Combining iPS cell technology with BMP4 induction, we derived trophoblast lineage cells from human iPS cells. The gene expression profile of these trophoblast lineage cells was distinct from fibroblasts and iPS cells. These cells expressed markers of human trophoblasts. Furthermore, when these cells were differentiated they exhibited invasive capacity and placental hormone secretive capacity, suggesting extravillous trophoblasts and syncytiotrophoblasts. Conclusion: Trophoblast lineage cells can be successfully derived from human iPS cells, which provide a proof-of-concept tool to recapitulate pathogenesis of patient placental trophoblasts in vitro.

  19. Isolation and generation of human dendritic cells.

    PubMed

    Nair, Smita; Archer, Gerald E; Tedder, Thomas F

    2012-11-01

    Dendritic cells are highly specialized antigen-presenting cells (APC), which may be isolated or generated from human blood mononuclear cells. Although mature blood dendritic cells normally represent ∼0.2% of human blood mononuclear cells, their frequency can be greatly increased using the cell enrichment methods described in this unit. More highly purified dendritic cell preparations can be obtained from these populations by sorting of fluorescence-labeled cells. Alternatively, dendritic cells can be generated from monocytes by culture with the appropriate cytokines, as described here. In addition, a negative selection approach is provided that may be employed to generate cell preparations that have been depleted of dendritic cells to be used for comparison in functional studies.

  20. A Child with Local Lipohypertrophy following Recombinant Human Growth Hormone Administration

    PubMed Central

    Koppen, Ilan J. N.; de Kruiff, Chris C.

    2016-01-01

    Local lipohypertrophy due to recombinant human growth hormone (rhGH) administration is a rare phenomenon. Here, we report a case of an 11-year-old girl who presented with a paraumbilical swelling, approximately one year after the start of rhGH treatment for short stature due to the presumed diagnosis of partial growth hormone insensitivity. Ultrasound imaging revealed an asymmetric distribution of subcutaneous fat tissue at the rhGH administration site, indicating local lipohypertrophy. After sparing her routine injection site and alternating other sites, the swelling disappeared within 6 months. Although the precise cause of local lipohypertrophy resulting from rhGH administration is still unclear, it might be related to the presumed diagnosis of partial growth hormone insensitivity. PMID:27803832

  1. Effects of vasopressin administration on diuresis of water immersion in normal humans

    NASA Technical Reports Server (NTRS)

    Epstein, M.; Denunzio, A. G.; Loutzenhiser, R. D.

    1981-01-01

    The influence of vasopressin suppression on the diuresis encountered during water immersion is investigated in studies on normal humans immersed to the neck. Six hydrated male subjects were studied on two occasions while undergoing 6 h of immersion without or during the administration of aqueous vasopressin for the initial 4 h. Neck immersion is found to result in a significant increase in urinary flow rate beginning in the first hour and persisting throughout the immersion. The administration of vasopressin markedly attenuated the diuretic response throughout the period of infusion, while cessation of vasopressin administration during the final 2 h of immersion resulted in a marked offset of the antidiuresis. Results thus support the view that the suppression of antidiuretic hormone contributes to the immersion diuresis of hydrated subjects.

  2. IL-7 enhances thymic human T cell development in "human immune system" Rag2-/-IL-2Rgammac-/- mice without affecting peripheral T cell homeostasis.

    PubMed

    van Lent, Anja U; Dontje, Wendy; Nagasawa, Maho; Siamari, Rachida; Bakker, Arjen Q; Pouw, Stephan M; Maijoor, Kelly A; Weijer, Kees; Cornelissen, Jan J; Blom, Bianca; Di Santo, James P; Spits, Hergen; Legrand, Nicolas

    2009-12-15

    IL-7 is a central cytokine in the development of hematopoietic cells, although interspecies discrepancies have been reported. By coculturing human postnatal thymus hematopoietic progenitors and OP9-huDL1 stromal cells, we found that murine IL-7 is approximately 100-fold less potent than human IL-7 for supporting human T cell development in vitro. We investigated the role of human IL-7 in newborn BALB/c Rag2(-/-)gamma(c)(-/-) mice transplanted with human hematopoietic stem cells (HSC) as an in vivo model of human hematopoiesis using three approaches to improve IL-7 signaling: administration of human IL-7, ectopic expression of human IL-7 by the transplanted human HSC, or enforced expression of a murine/human chimeric IL-7 receptor binding murine IL-7. We show that premature IL-7 signaling at the HSC stage, before entrance in the thymus, impeded T cell development, whereas increased intrathymic IL-7 signaling significantly enhanced the maintenance of immature thymocytes. Increased thymopoiesis was also observed when we transplanted BCL-2- or BCL-x(L)-transduced human HSC. Homeostasis of peripheral mature T cells in this humanized mouse model was not improved by any of these strategies. Overall, our results provide evidence for an important role of IL-7 in human T cell development in vivo and highlight the notion that IL-7 availability is but one of many signals that condition peripheral T cell homeostasis.

  3. The Joy of Social Work Administration: An Exploratory Qualitative Study of Human Service Administrators' Positive Perceptions of Their Work

    ERIC Educational Resources Information Center

    Watson, Larry D.; Hoefer, Richard A.

    2016-01-01

    Positive organizational psychology suggests that researchers should focus on the rewarding elements of work life, yet those in the fields of social work and nonprofit administration have not conducted research in line with this admonition. Indeed, the current focus on administrative challenges and problems may be part of the reason there is…

  4. The effects of acute alcohol administration on the human brain: insights from neuroimaging.

    PubMed

    Bjork, James M; Gilman, Jodi M

    2014-09-01

    Over the last quarter century, researchers have peered into the living human brain to develop and refine mechanistic accounts of alcohol-induced behavior, as well as neurobiological mechanisms for development and maintenance of addiction. These in vivo neuroimaging studies generally show that acute alcohol administration affects brain structures implicated in motivation and behavior control, and that chronic intoxication is correlated with structural and functional abnormalities in these same structures, where some elements of these decrements normalize with extended sobriety. In this review, we will summarize recent findings about acute human brain responses to alcohol using neuroimaging techniques, and how they might explain behavioral effects of alcohol intoxication. We then briefly address how chronic alcohol intoxication (as inferred from cross-sectional differences between various drinking populations and controls) may yield individual brain differences between drinking subjects that may confound interpretation of acute alcohol administration effects. This article is part of the Special Issue Section entitled 'Neuroimaging in Neuropharmacology'.

  5. Tolerability, usability and acceptability of dissolving microneedle patch administration in human subjects.

    PubMed

    Arya, Jaya; Henry, Sebastien; Kalluri, Haripriya; McAllister, Devin V; Pewin, Winston P; Prausnitz, Mark R

    2017-03-02

    To support translation of microneedle patches from pre-clinical development into clinical trials, this study examined the effect of microneedle patch application on local skin reactions, reliability of use and acceptability to patients. Placebo patches containing dissolving microneedles were administered to fifteen human participants. Microneedle patches were well tolerated in the skin with no pain or swelling and only mild erythema localized to the site of patch administration that resolved fully within seven days. Microneedle patches could be administered by hand without the need of an applicator and delivery efficiencies were similar for investigator-administration and self-administration. Microneedle patch administration was not considered painful and the large majority of subjects were somewhat or fully confident that they self-administered patches correctly. Microneedle patches were overwhelmingly preferred over conventional needle and syringe injection. Altogether, these results demonstrate that dissolving microneedle patches were well tolerated, easily usable and strongly accepted by human subjects, which will facilitate further clinical translation of this technology.

  6. Both acute and prolonged administration of EPO reduce cerebral and systemic vascular conductance in humans.

    PubMed

    Rasmussen, Peter; Kim, Yu-Sok; Krogh-Madsen, Rikke; Lundby, Carsten; Olsen, Niels V; Secher, Niels H; van Lieshout, Johannes J

    2012-03-01

    Administration of erythropoietin (EPO) has been linked to cerebrovascular events. EPO reduces vascular conductance, possibly because of the increase in hematocrit. Whether EPO in itself affects the vasculature remains unknown; here it was evaluated in healthy males by determining systemic and cerebrovascular variables following acute (30,000 IU/d for 3 d; n=8) and chronic (5000 IU/week for 13 wk; n=8) administration of EPO, while the responsiveness of the vasculature was challenged during cycling exercise, with and without hypoxia. Prolonged administration of EPO increased hematocrit from 42.5 ± 3.7 to 47.6 ± 4.1% (P<0.01), whereas hematocrit was unaffected following acute EPO administration. Yet, the two EPO regimes increased arterial pressure similarly (by 8±4 and 7±3 mmHg, respectively; P=0.01) through reduced vascular conductance (by 7±3 and 5±2%; P<0.05). Also, both EPO regimes widened the arterial-to-jugular O(2) differences at rest as well as during normoxic and hypoxic exercise (P<0.01), which indicated reduced cerebral blood flow despite preserved dynamic cerebral autoregulation, and an increase in middle cerebral artery mean blood flow velocity (P<0.05), therefore, reflected vasoconstriction. Thus, administration of EPO to healthy humans lowers systemic and cerebral conductance independent of its effect on hematocrit.

  7. Improved memory for reward cues following acute buprenorphine administration in humans.

    PubMed

    Syal, Supriya; Ipser, Jonathan; Terburg, David; Solms, Mark; Panksepp, Jaak; Malcolm-Smith, Susan; Bos, Peter A; Montoya, Estrella R; Stein, Dan J; van Honk, Jack

    2015-03-01

    In rodents, there is abundant evidence for the involvement of the opioid system in the processing of reward cues, but this system has remained understudied in humans. In humans, the happy facial expression is a pivotal reward cue. Happy facial expressions activate the brain's reward system and are disregarded by subjects scoring high on depressive mood who are low in reward drive. We investigated whether a single 0.2mg administration of the mixed mu-opioid agonist/kappa-antagonist, buprenorphine, would influence short-term memory for happy, angry or fearful expressions relative to neutral faces. Healthy human subjects (n38) participated in a randomized placebo-controlled within-subject design, and performed an emotional face relocation task after administration of buprenorphine and placebo. We show that, compared to placebo, buprenorphine administration results in a significant improvement of memory for happy faces. Our data demonstrate that acute manipulation of the opioid system by buprenorphine increases short-term memory for social reward cues.

  8. Nucleotide Analog Prodrug, Tenofovir Disoproxil, Enhances Lymphoid Cell Loading Following Oral Administration in Monkeys

    PubMed Central

    Durand-Gasselin, Lucie; Van Rompay, Koen K.A.; Vela, Jennifer E.; Henne, Ilana N.; Lee, William A.; Rhodes, Gerry R.; Ray, Adrian S.

    2009-01-01

    The antiviral drug tenofovir (TFV) is orally administered as the fumarate salt of its disoproxil prodrug (TFV disoproxil fumarate (TDF)). TFV is a di-anion at physiological pH and, as a result, has poor lipid membrane permeability. Administration of the lipophilic and cell permeable prodrug, TFV disoproxil, enhances the oral absorption of TFV. In order to determine if oral administration of TDF also increases distribution to sites of viral infection, the plasma and circulating lymphoid cell pharmacokinetics of TFV and its phosphorylated metabolites were assessed following a single oral TDF or subcutaneous TFV administration at doses yielding equivalent plasma exposures to TFV in macaques. Despite TFV disoproxil’s lack of plasma stability and undetectable levels in the first plasma samples taken, oral administration of TDF resulted in 7.9-fold higher peripheral blood mononuclear cell exposures to the active metabolite, TFV-diphosphate. The apparent plasma terminal half-life (t1/2) of TFV was also longer following oral TDF relative to subcutaneous TFV administration (median t1/2 of 15.3 and 3.9 h, respectively), suggesting broader distribution to cells and tissues outside of the central plasma compartment. In conclusion, the disoproxil pro-moiety not only enhances the oral absorption of TFV but also tissue and lymphoid cell loading. These results illustrate that administration of even a fleeting prodrug can increase target tissue loading and gives valuable insight for future prodrug development. PMID:19545170

  9. Hepatic Differentiation from Human Ips Cells Using M15 Cells.

    PubMed

    Umeda, Kahoko; Shiraki, Nobuaki; Kume, Shoen

    2016-01-01

    Here, we describe a procedure of human iPS cells differentiation into the definitive endoderm, further into albumin-expressing and albumin-secreting hepatocyte, using M15, a mesonephros- derived cell line. Approximately 90 % of human iPS cells differentiated into SOX17-positive definitive endoderm then approximately 50 % of cells became albumin-positive cells, and secreted ALB protein. This M15 feeder system for endoderm and hepatic differentiation is a simple and efficient method, and useful for elucidating molecular mechanisms for hepatic fate decision, and could represent an attractive approach for a surrogate cell source for pharmaceutical studies.

  10. Apoptosis of rat kidney cells after 241-americium administration.

    PubMed

    Labéjof, L; Berry, J P; Duchambon, P; Poncy, J L; Galle, P

    1998-01-01

    Tumors induction by americium is well known but there are no data on the biological effects of this radionucleide at subcellular level. In order to study the possible ultrastructural lesions induced by this element, a group of rats were injected with 241-Americium-citrate (9 kBq), once a week for five weeks and sacrificed 7 days after the last injection. We describe the alterations observed in the cortex kidney using cytochemical (TUNEL reaction) and histochemical (PAS staining) methods for light microscopy as well as electron microscopy techniques. Various types of lesions were detected: condensation of nuclear chromatine, fragmentation of the nuclei, swollen mitochondria, disappearance of mitochondrial crests and skrinking of the cytoplasm. This study clearly demonstrated the induction of apoptosis by americium in rat cortex kidney cells.

  11. Activin A programs human TFH cell differentiation

    PubMed Central

    Locci, Michela; Wu, Jennifer; Arumemi, Fortuna; Mikulski, Zbigniew; Dahlberg, Carol; Miller, Andrew T.; Crotty, Shane

    2016-01-01

    SUMMARY Follicular helper T (TFH) cells are CD4+ T cells specialized in helping B cells and are associated both with protective antibody responses and autoimmune diseases. The promise of targeting TFH cells therapeutically has been limited by fragmentary understanding of extrinsic signals regulating human TFH cell differentiation. A screen of a human protein library identified activin A as new regulator of TFH cell differentiation. Activin A orchestrated expression of multiple TFH-associated genes, independently or in concert with additional signals. TFH programming by activin A was antagonized by the cytokine IL-2. Activin A’s capacity to drive TFH cell differentiation in vitro was conserved for non-human primates but not mice. Finally, activin A-induced TFH programming was dependent on SMAD2 and SMAD3 signaling and blocked by pharmacological inhibitors. PMID:27376469

  12. Human Kidney-Derived Cells Ameliorate Acute Kidney Injury Without Engrafting into Renal Tissue.

    PubMed

    Santeramo, Ilaria; Herrera Perez, Zeneida; Illera, Ana; Taylor, Arthur; Kenny, Simon; Murray, Patricia; Wilm, Bettina; Gretz, Norbert

    2017-04-04

    Previous studies have suggested that CD133(+) cells isolated from human kidney biopsies have the potential to ameliorate injury following intravenous (IV) administration in rodent models of kidney disease by integrating into damaged renal tissue and generating specialized renal cells. However, whether renal engraftment of CD133(+) cells is a prerequisite for ameliorating injury has not yet been unequivocally resolved. Here, we have established a cisplatin-induced nephropathy model in immunodeficient rats to assess the efficacy of CD133(+) human kidney cells in restoring renal health, and to determine the fate of these cells after systemic administration. Specifically, following IV administration, we evaluated the impact of the CD133(+) cells on renal function by undertaking longitudinal measurements of the glomerular filtration rate using a novel transcutaneous device. Using histological assays, we assessed whether the human kidney cells could promote renal regeneration, and if this was related to their ability to integrate into the damaged kidneys. Our results show that both CD133(+) and CD133(-) cells improve renal function and promote renal regeneration to a similar degree. However, this was not associated with engraftment of the cells into the kidneys. Instead, after IV administration, both cell types were exclusively located in the lungs, and had disappeared by 24 hours. Our data therefore indicate that renal repair is not mediated by CD133(+) cells homing to the kidneys and generating specialized renal cells. Instead, renal repair is likely to be mediated by paracrine or endocrine factors. © Stem Cells Translational Medicine 2017.

  13. Human embryonic stem cells derived by somatic cell nuclear transfer.

    PubMed

    Tachibana, Masahito; Amato, Paula; Sparman, Michelle; Gutierrez, Nuria Marti; Tippner-Hedges, Rebecca; Ma, Hong; Kang, Eunju; Fulati, Alimujiang; Lee, Hyo-Sang; Sritanaudomchai, Hathaitip; Masterson, Keith; Larson, Janine; Eaton, Deborah; Sadler-Fredd, Karen; Battaglia, David; Lee, David; Wu, Diana; Jensen, Jeffrey; Patton, Phillip; Gokhale, Sumita; Stouffer, Richard L; Wolf, Don; Mitalipov, Shoukhrat

    2013-06-06

    Reprogramming somatic cells into pluripotent embryonic stem cells (ESCs) by somatic cell nuclear transfer (SCNT) has been envisioned as an approach for generating patient-matched nuclear transfer (NT)-ESCs for studies of disease mechanisms and for developing specific therapies. Past attempts to produce human NT-ESCs have failed secondary to early embryonic arrest of SCNT embryos. Here, we identified premature exit from meiosis in human oocytes and suboptimal activation as key factors that are responsible for these outcomes. Optimized SCNT approaches designed to circumvent these limitations allowed derivation of human NT-ESCs. When applied to premium quality human oocytes, NT-ESC lines were derived from as few as two oocytes. NT-ESCs displayed normal diploid karyotypes and inherited their nuclear genome exclusively from parental somatic cells. Gene expression and differentiation profiles in human NT-ESCs were similar to embryo-derived ESCs, suggesting efficient reprogramming of somatic cells to a pluripotent state.

  14. Development of human mast cells in vitro.

    PubMed Central

    Furitsu, T; Saito, H; Dvorak, A M; Schwartz, L B; Irani, A M; Burdick, J F; Ishizaka, K; Ishizaka, T

    1989-01-01

    Nucleated cells of human umbilical cord blood were cocultured with mouse skin-derived 3T3 fibroblasts. After 7-8 weeks in culture, when the number of the other hematopoietic cells declined, metachromatic granule-containing mononuclear cells appeared in the cultures, and the number of the cells increased up to 12 weeks. After 11-14 weeks in culture, the metachromatic mononuclear cells comprised a substantial portion of the cultured cells. These cells contained 1.8-2 micrograms of histamine per 10(6) cells and bore receptors for IgE. All of the cells contained tryptase in their granules. Electron microscopic analysis showed that these cells were mature human mast cells, clearly different from the basophilic granulocytes or eosinophils that arise in a variety of circumstances in cord blood cell cultures. Most of the cultured mast cells expressed some granules with regular crystalline arrays and contained both tryptase and chymase, and thus resembled human skin mast cells. Images PMID:2532357

  15. Expression cloning of human B cell immunoglobulins.

    PubMed

    Wardemann, Hedda; Kofer, Juliane

    2013-01-01

    The majority of lymphomas originate from B cells at the germinal center stage or beyond. Preferential selection of B cell clones by a limited set of antigens has been suggested to drive lymphoma development. However, little is known about the specificity of the antibodies expressed by lymphoma cells, and the role of antibody-specificity in lymphomagenesis remains elusive. Here, we describe a strategy to characterize the antibody reactivity of human B cells. The approach allows the unbiased characterization of the human antibody repertoire on a single cell level through the generation of recombinant monoclonal antibodies from single primary human B cells of defined origin. This protocol offers a detailed description of the method starting from the flow cytometric isolation of single human B cells, to the RT-PCR-based amplification of the expressed Igh, Igκ, and Igλ chain genes, and Ig gene expression vector cloning for the in vitro production of monoclonal antibodies. The strategy may be used to obtain information on the clonal evolution of B cell lymphomas by single cell Ig gene sequencing and on the antibody reactivity of human lymphoma B cells.

  16. Antiproliferative effects on colon adenocarcinoma cells induced by co-administration of vitamin K1 and Lactobacillus rhamnosus GG.

    PubMed

    Orlando, Antonella; Linsalata, Michele; Russo, Francesco

    2016-06-01

    Vitamin K (VK), an essential nutrient associated with the clotting cascade, has also been demonstrated to have anticancer properties in various cancer cells including colon cancer cells. Also probiotics have gained interest as potential anticancer agents. Among them, Lactobacillus rhamnosus GG (L.GG) has been shown to inhibit cell proliferation and polyamine biosynthesis as well as to induce apoptosis in different human gastrointestinal cancer cells. Nevertheless, the exact mechanisms involved in these actions are not completely elucidated. Therefore, the aims of the present study were to evaluate in three differently graded human colon cancer cells (namely Caco-2, HT-29 and SW480) the effects of increasing VK1 concentrations, administered alone or in combination with viable L.GG, on the cell proliferation evaluated by MTT test, apoptosis investigated by Bax/Bcl-2 ratio and the percentage of the apoptotic cells, and the cell cycle evaluated by MUSE cell analyzer. Both VK1 and L.GG administered alone up to 72 h, caused inhibition of proliferation, induction of apoptosis and the cell cycle arrest in all the tested colon cancer cells. When VK1 and L.GG were co-administered, the addition of increasing VK1 concentrations potentiated the probiotic antiproliferative effect in a dose-dependent manner, being also related to the individual features of each cell line. The effect was more evident in Caco-2 and HT-29 cells compared to the less differentiated SW480. The enhanced antiproliferative efficacy due to co-administration of L.GG and VK1 could represent a suitable option in a functional food strategy for cancer growth inhibition and chemoprevention.

  17. Reprogramming of human exocrine pancreas cells to beta cells.

    PubMed

    Staels, Willem; Heremans, Yves; Heimberg, Harry

    2015-12-01

    One of the key promises of regenerative medicine is providing a cure for diabetes. Cell-based therapies are proving their safety and efficiency, but donor beta cell shortages and immunological issues remain major hurdles. Reprogramming of human pancreatic exocrine cells towards beta cells would offer a major advantage by providing an abundant and autologous source of beta cells. Over the past decade our understanding of transdifferentiation processes greatly increased allowing us to design reprogramming protocols that fairly aim for clinical trials.

  18. Novel APC-like properties of human NK cells directly regulate T cell activation

    PubMed Central

    Hanna, Jacob; Gonen-Gross, Tsufit; Fitchett, Jonathan; Rowe, Tony; Daniels, Mark; Arnon, Tal I.; Gazit, Roi; Joseph, Aviva; Schjetne, Karoline W.; Steinle, Alexander; Porgador, Angel; Mevorach, Dror; Goldman-Wohl, Debra; Yagel, Simcha; LaBarre, Michael J.; Buckner, Jane H.; Mandelboim, Ofer

    2004-01-01

    Initiation of the adaptive immune response is dependent on the priming of naive T cells by APCs. Proteomic analysis of unactivated and activated human NK cell membrane–enriched fractions demonstrated that activated NK cells can efficiently stimulate T cells, since they upregulate MHC class II molecules and multiple ligands for TCR costimulatory molecules. Furthermore, by manipulating antigen administration, we show that NK cells possess multiple independent unique pathways for antigen uptake. These results highlight NK cell–mediated cytotoxicity and specific ligand recognition by cell surface–activating receptors on NK cells as unique mechanisms for antigen capturing and presentation. In addition, we analyzed the T cell–activating potential of human NK cells derived from different clinical conditions, such as inflamed tonsils and noninfected and CMV-infected uterine decidual samples, and from transporter-associated processing antigen 2–deficient patients. This in vivo analysis revealed that proinflammatory, but not immune-suppressive, microenvironmental requirements can selectively dictate upregulation of T cell–activating molecules on NK cells. Taken together, these observations offer new and unexpected insights into the direct interactions between NK and T cells and suggest novel APC-like activating functions for human NK cells. PMID:15578093

  19. Relaxation and immunity enhancement effects of gamma-aminobutyric acid (GABA) administration in humans.

    PubMed

    Abdou, Adham M; Higashiguchi, S; Horie, K; Kim, Mujo; Hatta, H; Yokogoshi, H

    2006-01-01

    The effect of orally administrated gamma-aminobutyric acid (GABA) on relaxation and immunity during stress has been investigated in humans. Two studies were conducted. The first evaluated the effect of GABA intake by 13 subjects on their brain waves. Electroencephalograms (EEG) were obtained after 3 tests on each volunteer as follows: intake only water, GABA, or L-theanine. After 60 minutes of administration, GABA significantly increases alpha waves and decreases beta waves compared to water or L-theanine. These findings denote that GABA not only induces relaxation but also reduces anxiety. The second study was conducted to see the role of relaxant and anxiolytic effects of GABA intake on immunity in stressed volunteers. Eight acrophobic subjects were divided into 2 groups (placebo and GABA). All subjects were crossing a suspended bridge as a stressful stimulus. Immunoglobulin A (IgA) levels in their saliva were monitored during bridge crossing. Placebo group showed marked decrease of their IgA levels, while GABA group showed significantly higher levels. In conclusion, GABA could work effectively as a natural relaxant and its effects could be seen within 1 hour of its administration to induce relaxation and diminish anxiety. Moreover, GABA administration could enhance immunity under stress conditions.

  20. Cell proliferation in human coronary arteries.

    PubMed Central

    Gordon, D; Reidy, M A; Benditt, E P; Schwartz, S M

    1990-01-01

    Despite the lack of direct evidence for cell multiplication, proliferation of smooth muscle cells in human atherosclerotic lesions has been assumed to play a central role in ontogeny of the plaque. We used antibodies to cell cycle-related proteins on tissue sections of human arteries and coronary atherosclerotic plaques. Specific cell types were identified by immunochemical reagents for smooth muscle, monocyte-macrophages, and other blood cells. Low rates of smooth muscle cell proliferation were observed. Macrophages were also observed with rates of proliferation comparable to that of the smooth muscle. Additional replicating cells could not be defined as belonging to specific cell types with the reagents used in this study. These findings imply that smooth muscle replication in advanced plaques is indolent and raise the possibility of a role for proliferating leukocytes. Images PMID:1972277

  1. Human genome project and sickle cell disease.

    PubMed

    Norman, Brenda J; Miller, Sheila D

    2011-01-01

    Sickle cell disease is one of the most common genetic blood disorders in the United States that affects 1 in every 375 African Americans. Sickle cell disease is an inherited condition caused by abnormal hemoglobin in the red blood cells. The Human Genome Project has provided valuable insight and extensive research advances in the understanding of the human genome and sickle cell disease. Significant progress in genetic knowledge has led to an increase in the ability for researchers to map and sequence genes for diagnosis, treatment, and prevention of sickle cell disease and other chronic illnesses. This article explores some of the recent knowledge and advances about sickle cell disease and the Human Genome Project.

  2. Production of hepatocyte like cells from human pluripotent stem cells

    PubMed Central

    Hannan, Nicholas R.F; Segeritz, Charis-Patricia; Touboul, Thomas; Vallier, Ludovic

    2013-01-01

    Large scale production of hepatocytes from a variety of genetic backgrounds would be beneficial for drug screening and to provide a source of cells to be used as a substitute for liver transplantation. However, fully functional primary hepatocytes remain difficult to expand in vitro and circumventing this problem by using an alternative source of cells is desirable. Here, we describe a 25 day protocol to direct the differentiation of human pluripotent stem cells into a near homogenous population of hepatocyte-like cells. As cells progress through this protocol they express genes in a chronological manner similar to that described during in-vivo hepatic development. The protocol relies on culture systems devoid of serum, feeders or complex extra-cellular matrices enabling molecular analyses without interference from unknown factors. This approach works efficiently with human embryonic stem cells and human induced pluripotent stem cells and was recently used to model liver diseases in vitro. PMID:23424751

  3. Clinical concerns of immunogenicity produced at cellular levels by biopharmaceuticals following their parenteral administration into human body.

    PubMed

    Tamilvanan, Shunmugaperumal; Raja, Natarajan Livingston; Sa, Biswanath; Basu, Sanat Kumar

    2010-08-01

    Similar to the low molecular weight traditional drugs, biopharmaceuticals are capable of producing not only therapeutic effects but also side effects provided if the dose of these compounds exceeds certain concentration and/or if the exposure duration of these compounds at subtoxic doses is being lengthened. In addition, a major drawback of biopharmaceuticals is the risk of antibody formation. Following the administration of biopharmaceuticals into human body, the formation of antidrug-antibody (ADA) or neutralizing antibody and other general immune system effects (including allergy, anaphylaxis, or serum sickness) are of clinical concern regarding therapeutic efficacy and patient safety. For example, drug-induced neutralizing antibodies to erythropoietin (EPO) result in pure red cell aplasia, whereas drug-induced acquired anti-factor VIII antibodies worsen the pathology associated with hemophilia. Since most of the already developed or under development biopharmaceuticals are to some extent immunogenic, the regulatory agencies insist to conduct potential ADA formation during the drug development process itself. This review encompasses a short overview on the clinical concerns of immunogenicity produced at cellular levels by growth hormone, interferon-alpha, EPO, factor VIII, and factor IX following their parenteral administration into human body. Clinical concerns related to immunogenicity produced by the biosimilar versions of these drugs are also presented wherever possible.

  4. Cell Cycle Progression of Human Cells Cultured in Rotating Bioreactor

    NASA Technical Reports Server (NTRS)

    Parks, Kelsey

    2009-01-01

    Space flight has been shown to alter the astronauts immune systems. Because immune performance is complex and reflects the influence of multiple organ systems within the host, scientists sought to understand the potential impact of microgravity alone on the cellular mechanisms critical to immunity. Lymphocytes and their differentiated immature form, lymphoblasts, play an important and integral role in the body's defense system. T cells, one of the three major types of lymphocytes, play a central role in cell-mediated immunity. They can be distinguished from other lymphocyte types, such as B cells and natural killer cells by the presence of a special receptor on their cell surface called T cell receptors. Reported studies have shown that spaceflight can affect the expression of cell surface markers. Cell surface markers play an important role in the ability of cells to interact and to pass signals between different cells of the same phenotype and cells of different phenotypes. Recent evidence suggests that cell-cycle regulators are essential for T-cell function. To trigger an effective immune response, lymphocytes must proliferate. The objective of this project is to investigate the changes in growth of human cells cultured in rotating bioreactors and to measure the growth rate and the cell cycle distribution for different human cell types. Human lymphocytes and lymphoblasts will be cultured in a bioreactor to simulate aspects of microgravity. The bioreactor is a cylindrical culture vessel that incorporates the aspects of clinostatic rotation of a solid fluid body around a horizontal axis at a constant speed, and compensates gravity by rotation and places cells within the fluid body into a sustained free-fall. Cell cycle progression and cell proliferation of the lymphocytes will be measured for a number of days. In addition, RNA from the cells will be isolated for expression of genes related in cell cycle regulations.

  5. [Effect of combined administration of Angelica polysaccharide and cytarabine on liver of human leukemia NOD/SCID mouse model].

    PubMed

    Zhu, Jia-Hong; Xu, Chun-Yan; Mu, Xin-Yi; Liu, Jun; Zhang, Meng-Si; Jia, Dao-Yong; Zhang, Yan-Yan; Huang, Guo-Ning; Wang, Ya-Ping

    2014-01-01

    Leukemia is a type of malignant tumors of hematopoietic system with the abnormal increased immature leukemia cells showing metastasis and invasion ability. Liver is one of the main targets of the leukemia cells spread to, where they may continue to proliferate and differentiate and cause liver function damage, even liver failure. Our previous studies showed that Angelica polysscharides (APS), the main effective components in Angelica sinensis of Chinese traditional medicine, was able to inhibit the proliferation and induced differentiation of the leukemia cells, however, its effect on the liver during the treatment remains elucidated. In the present study, the human leukemia NOD/SCID mouse model were established by implantation human leukemia K562 cells line, then the leukemia mouse were treated with APS, Ara-c or APS + Ara-c respectively by peritoneal injection for 14 days, to explore the effect and mechanism of the chemicals on the mouse liver. Compared to the human leukemia NOD/SCID mouse model group with the treatments of APS, Ara-c and APS + Ara-c, We found that severe liver damage and pathological changes of the liver were able to alleviate: First, the number of white blood cells in the peripheral blood was significantly lower and with less transplanted K562 leukemia cells; Second, liver function damage was alleviated as liver function tests showed that alanine aminotransferase (ALT), aspartate aminotransferase (AST) and total bilirubin (TBiL) were significantly reduced, while the albumin (Alb) was notably increased; Third, liver antioxidant ability was improved as the activities of the antioxidant enzymes glutathione peroxidase (GSH-Px) and superoxide dismutase (SOD) were significantly increased, and the contents of GSH and malonaldehyde (MDA) were decreased significantly in the liver; Fourth, the inflammation of the liver was relieved as the level of IL-1beta and IL-6, the inflammatory cytokines, were decreased significantly in the liver. Fifth, liver index

  6. The Human Natural Killer Cell Immune Synapse

    NASA Astrophysics Data System (ADS)

    Davis, Daniel M.; Chiu, Isaac; Fassett, Marlys; Cohen, George B.; Mandelboim, Ofer; Strominger, Jack L.

    1999-12-01

    Inhibitory killer Ig-like receptors (KIR) at the surface of natural killer (NK) cells induced clustering of HLA-C at the contacting surface of target cells. In this manner, inhibitory immune synapses were formed as human NK cells surveyed target cells. At target/NK cell synapses, HLA-C/KIR distributed into rings around central patches of intercellular adhesion molecule-1/lymphocyte function-associated antigen-1, the opposite orientation to mature murine T cell-activating synapses. This organization of protein was stable for at least 20 min. Cells could support multiple synapses simultaneously, and clusters of HLA-C moved as NK cells crawled over target cells. Clustering required a divalent metal cation, explaining how metal chelators inhibit KIR function. Surprisingly, however, formation of inhibitory synapses was unaffected by ATP depletion and the cytoskeletal inhibitors, colchicine and cytochalsins B and D. Clearly, supramolecular organization within plasma membranes is critical for NK cell immunosurveillance.

  7. Treatment of Uveitis by In Situ Administration of Ex Vivo-Activated Polyclonal Regulatory T Cells.

    PubMed

    Grégoire, Sylvie; Terrada, Céline; Martin, Gaelle H; Fourcade, Gwladys; Baeyens, Audrey; Marodon, Gilles; Fisson, Sylvain; Billiard, Fabienne; Lucas, Bruno; Tadayoni, Ramin; Béhar-Cohen, Francine; Levacher, Béatrice; Galy, Anne; LeHoang, Phuc; Klatzmann, David; Bodaghi, Bahram; Salomon, Benoît L

    2016-03-01

    CD4(+)CD25(+)Foxp3(+) regulatory T (Treg) cell therapy is a promising approach for the treatment of autoimmune diseases. To be effective, Treg cells should be in an activated state in the target tissue. This can be achieved by systemic administration of Ag-specific Treg cells, which are difficult to produce in conditions that can be translated to the clinic. In this paper, we propose an alternative approach consisting of in situ injection of preactivated polyclonal Treg cells that would exert bystander suppression in the target tissue. We show that polyclonal Treg cells suppressed uveitis in mice as efficiently as Ag-specific Treg cells but only when preactivated and administered in the vitreous. Uveitis control was correlated with an increase of IL-10 and a decrease of reactive oxygen species produced by immune cell infiltrates in the eye. Thus, our results reveal a new mechanism of Treg cell-mediated suppression and a new Treg cell therapy approach.

  8. Nucleofection of human embryonic stem cells.

    PubMed

    Siemen, Henrike; Nix, Michael; Endl, Elmar; Koch, Philipp; Itskovitz-Eldor, Joseph; Brüstle, Oliver

    2005-08-01

    Human embryonic stem (hES) cells provide an important tool for the study of human development, disease, and tissue regeneration. Technologies for efficient genetic modification are required to exploit hES cells fully for these applications. Here we present a customized protocol for the transfection of hES cells with the Nucleofector technology and compare its efficiency with conventional electroporation and lipofection. Cell survival and transfection efficiency were quantified using an enhanced green fluorescent protein (EGFP) reporter construct. Our optimized nucleofection parameters yielded survival rates >70%. Under these conditions, 66% of the surviving cells showed transgene expression 24 h after nucleofection. Transfected cells maintained expression of the pluripotency- associated markers Tra-1-60, Tra-1-81, and Oct4 and could be expanded to stably transgene-expressing clones. The low quantities of hES cells and DNA required for nucleofection could make this method an attractive tool for miniaturized high throughput screening (HTS) applications.

  9. Satellite cells in human skeletal muscle plasticity

    PubMed Central

    Snijders, Tim; Nederveen, Joshua P.; McKay, Bryon R.; Joanisse, Sophie; Verdijk, Lex B.; van Loon, Luc J. C.; Parise, Gianni

    2015-01-01

    Skeletal muscle satellite cells are considered to play a crucial role in muscle fiber maintenance, repair and remodeling. Our knowledge of the role of satellite cells in muscle fiber adaptation has traditionally relied on in vitro cell and in vivo animal models. Over the past decade, a genuine effort has been made to translate these results to humans under physiological conditions. Findings from in vivo human studies suggest that satellite cells play a key role in skeletal muscle fiber repair/remodeling in response to exercise. Mounting evidence indicates that aging has a profound impact on the regulation of satellite cells in human skeletal muscle. Yet, the precise role of satellite cells in the development of muscle fiber atrophy with age remains unresolved. This review seeks to integrate recent results from in vivo human studies on satellite cell function in muscle fiber repair/remodeling in the wider context of satellite cell biology whose literature is largely based on animal and cell models. PMID:26557092

  10. Human progenitor cells for bone engineering applications.

    PubMed

    de Peppo, G M; Thomsen, P; Karlsson, C; Strehl, R; Lindahl, A; Hyllner, J

    2013-06-01

    In this report, the authors review the human skeleton and the increasing burden of bone deficiencies, the limitations encountered with the current treatments and the opportunities provided by the emerging field of cell-based bone engineering. Special emphasis is placed on different sources of human progenitor cells, as well as their pros and cons in relation to their utilization for the large-scale construction of functional bone-engineered substitutes for clinical applications. It is concluded that, human pluripotent stem cells represent a valuable source for the derivation of progenitor cells, which combine the advantages of both embryonic and adult stem cells, and indeed display high potential for the construction of functional substitutes for bone replacement therapies.

  11. Apoptotic Effects of Chrysin in Human Cancer Cell Lines

    PubMed Central

    Khoo, Boon Yin; Chua, Siang Ling; Balaram, Prabha

    2010-01-01

    Chrysin is a natural flavonoid currently under investigation due to its important biological anti-cancer properties. In most of the cancer cells tested, chrysin has shown to inhibit proliferation and induce apoptosis, and is more potent than other tested flavonoids in leukemia cells, where chrysin is likely to act via activation of caspases and inactivation of Akt signaling in the cells. Moreover, structure-activity relationships have revealed that the chemical structure of chrysin meets the key structural requirements of flavonoids for potent cytotoxicity in leukemia cells. It is possible that combination therapy or modified chrysin could be more potent than single-agent use or administration of unmodified chrysin. This study may help to develop ways of improving the effectiveness of chrysin in the treatment of leukemia and other human cancers in vitro. PMID:20559509

  12. Mesenchymal stem cell like (MSCl) cells generated from human embryonic stem cells support pluripotent cell growth

    SciTech Connect

    Varga, Nora; Vereb, Zoltan; Rajnavoelgyi, Eva; Nemet, Katalin; Uher, Ferenc; Sarkadi, Balazs; Apati, Agota

    2011-10-28

    Highlights: Black-Right-Pointing-Pointer MSC like cells were derived from hESC by a simple and reproducible method. Black-Right-Pointing-Pointer Differentiation and immunosuppressive features of MSCl cells were similar to bmMSC. Black-Right-Pointing-Pointer MSCl cells as feeder cells support the undifferentiated growth of hESC. -- Abstract: Mesenchymal stem cell like (MSCl) cells were generated from human embryonic stem cells (hESC) through embryoid body formation, and isolated by adherence to plastic surface. MSCl cell lines could be propagated without changes in morphological or functional characteristics for more than 15 passages. These cells, as well as their fluorescent protein expressing stable derivatives, efficiently supported the growth of undifferentiated human embryonic stem cells as feeder cells. The MSCl cells did not express the embryonic (Oct4, Nanog, ABCG2, PODXL, or SSEA4), or hematopoietic (CD34, CD45, CD14, CD133, HLA-DR) stem cell markers, while were positive for the characteristic cell surface markers of MSCs (CD44, CD73, CD90, CD105). MSCl cells could be differentiated toward osteogenic, chondrogenic or adipogenic directions and exhibited significant inhibition of mitogen-activated lymphocyte proliferation, and thus presented immunosuppressive features. We suggest that cultured MSCl cells can properly model human MSCs and be applied as efficient feeders in hESC cultures.

  13. Increased oxytocin concentrations and prosocial feelings in humans after ecstasy (3,4-methylenedioxymethamphetamine) administration.

    PubMed

    Dumont, G J H; Sweep, F C G J; van der Steen, R; Hermsen, R; Donders, A R T; Touw, D J; van Gerven, J M A; Buitelaar, J K; Verkes, R J

    2009-01-01

    MDMA (3,4-methylenedioxymethamphetamine or "ecstasy") is a recreationally used drug with remarkable and characteristic prosocial effects. In spite of abundant attention in the scientific literature, the mechanism of its prosocial effects has not been elucidated in humans. Recently, research in animals has suggested that the neuropeptide oxytocin may induce these effects. In a double blind, randomized, crossover, and placebo-controlled study in 15 healthy volunteers we assessed blood oxytocin and MDMA concentrations and subjective prosocial effects after oral administration of 100 mg MDMA or placebo. MDMA induced a robust increase of blood oxytocin concentrations and an increase of subjective prosocial feelings. Within subjects, the variations in these feelings were significantly and positively correlated with variation in oxytocin levels, and the correlations between these feelings and oxytocin were significantly stronger than those between these feelings and blood MDMA levels. MDMA induces oxytocin release in humans, which may be involved in the characteristic prosocial effects of ecstasy.

  14. Retrospective birth dating of cells in humans.

    PubMed

    Spalding, Kirsty L; Bhardwaj, Ratan D; Buchholz, Bruce A; Druid, Henrik; Frisén, Jonas

    2005-07-15

    The generation of cells in the human body has been difficult to study, and our understanding of cell turnover is limited. Testing of nuclear weapons resulted in a dramatic global increase in the levels of the isotope 14C in the atmosphere, followed by an exponential decrease after 1963. We show that the level of 14C in genomic DNA closely parallels atmospheric levels and can be used to establish the time point when the DNA was synthesized and cells were born. We use this strategy to determine the age of cells in the cortex of the adult human brain and show that whereas nonneuronal cells are exchanged, occipital neurons are as old as the individual, supporting the view that postnatal neurogenesis does not take place in this region. Retrospective birth dating is a generally applicable strategy that can be used to measure cell turnover in man under physiological and pathological conditions.

  15. Human embryonic stem cells and lung regeneration.

    PubMed

    Varanou, A; Page, C P; Minger, S L

    2008-10-01

    Human embryonic stem cells are pluripotent cells derived from the inner cell mass of preimplantation stage embryos. Their unique potential to give rise to all differentiated cell types has generated great interest in stem cell research and the potential that it may have in developmental biology, medicine and pharmacology. The main focus of stem cell research has been on cell therapy for pathological conditions with no current methods of treatment, such as neurodegenerative diseases, cardiac pathology, retinal dysfunction and lung and liver disease. The overall aim is to develop methods of application either of pure cell populations or of whole tissue parts to the diseased organ under investigation. In the field of pulmonary research, studies using human embryonic stem cells have succeeded in generating enriched cultures of type II pneumocytes in vitro. On account of their potential of indefinite proliferation in vitro, embryonic stem cells could be a source of an unlimited supply of cells available for transplantation and for use in gene therapy. Uncovering the ability to generate such cell types will expand our understanding of biological processes to such a degree that disease understanding and management could change dramatically.

  16. Human brain glial cells synthesize thrombospondin.

    PubMed Central

    Asch, A S; Leung, L L; Shapiro, J; Nachman, R L

    1986-01-01

    Thrombospondin, a 450-kDa multinodular glycoprotein with lectin-type activity, is found in human platelets, endothelial cells, fibroblasts, smooth muscle cells, monocytes, and granular pneumocytes. Thrombospondin interacts with heparin, fibrinogen, fibronectin, collagen, histidine-rich glycoprotein, and plasminogen. Recently, thrombospondin synthesis by smooth muscle cells has been reported to be augmented by platelet-derived growth factor. We present evidence that thrombospondin is present within and synthesized by astrocytic neuroglial cells. Heparin-Sepharose affinity chromatography of material derived from a human brain homogenate yielded a protein that, when reduced, had an apparent size of 180 kDa and comigrated with reduced platelet thrombospondin on NaDodSO4/PAGE. Immunoblot analysis with monospecific anti-thrombospondin confirmed the presence of immunoreactive thrombospondin. Indirect immunofluorescence of cultured human glial cells indicated the presence of thrombospondin. Metabolic labeling of glial cell cultures with [35S]methionine followed by immunoprecipitation with monospecific anti-thrombospondin revealed synthesis of a 180-kDa polypeptide that comigrated with platelet thrombospondin on NaDodSO4/PAGE. Cultured human glial cells were incubated for 48 hr in serum-free medium with purified platelet-derived growth factor at concentrations up to 50 ng/ml. Aliquots taken at intervals were analyzed by a quantitative double-antibody ELISA. The growth factor stimulated the release of thrombospondin into the culture medium by as much as 10-fold over control cultures. The presence of thrombospondin within glial cells of the central nervous system and the augmentation of its synthesis by platelet-derived growth factor suggest that thrombospondin may play an important role in regulating cell-cell and cell-matrix interactions during periods of cell division and growth. Images PMID:2939460

  17. Myeloid derived suppressor cells in human diseases

    PubMed Central

    Greten, Tim F.; Manns, Michael P.; Korangy, Firouzeh

    2012-01-01

    Myeloid derived suppressor cells (MDSC) have been described as a heterogeneous cell population with potent immune suppressor function in mice. Limited data are available on MDSC in human diseases. Interpretation of these data is complicated by the fact that different markers have been used to analyze human MDSC subtypes in various clinical settings. Human MDSC are CD11b+, CD33+, HLA-DRneg/low and can be divided into granulocytic CD14− and monocytic CD14+ subtypes. Interleukin 4Rα, VEGFR, CD15 and CD66b have been suggested to be more specific markers for human MDSC, however these markers can only be found on some MDSC subsets. Until today the best marker for human MDSC remains their suppressor function, which can be either direct or indirect through the induction of regulatory T cells. Immune suppressor activity has been associated with high arginase 1 and iNOS activity as well as ROS production by MDSC. Not only in murine models, but even more importantly in patients with cancer, different drugs have been shown to either reverse the immune suppressor function of MDSC or directly target these cells. Systemic treatment with all-trans-retinoic acid has been shown to mature human MDSC and reverse their immune suppressor function. Alternatively, MDSC can be targeted by treatment with the multi-targeted receptor tyrosine kinase inhibitor sunitinib. In this review will provide a comprehensive summary of the recent literature on human MDSC. PMID:21237299

  18. Pancreastatin producing cell line from human pancreatic islet cell tumor.

    PubMed

    Funakoshi, A; Tateishi, K; Tsuru, M; Jimi, A; Wakasugi, H; Ikeda, Y; Kono, A

    1990-04-30

    It has been characterized that cell line QGP-1 derived from human non-functioning pancreatic islet cell tumor produces human pancreastatin. Exponentially growing cultures produced 5.7 fmol of pancreastatin/10(6) cells/hr. Human pancreastatin immunoreactivities in plasma and tumor after xenografting with QGP-1 into nude mouse were 92.7 fmol/ml and 160.2 pmol/g wet weight, respectively. Immunocytochemical study revealed both chromogranin A and pancreastatin immunoreactive cells in the tumor. Gel filtrations of culture medium and tumor extract identified heterogenous molecular forms of PST-LI which eluted as large and smaller molecular species. These results suggest that plasma pancreastatin levels may be useful as a tumor marker of endocrine tumor of the pancreas, and the pancreastatin producing cell line may be useful for studies of the mechanism of secretions and processing of chromogranin A and pancreastatin.

  19. Interaction of Staphylococci with Human B cells

    PubMed Central

    Nygaard, Tyler K.; Kobayashi, Scott D.; Freedman, Brett; Porter, Adeline R.; Voyich, Jovanka M.; Otto, Michael; Schneewind, Olaf; DeLeo, Frank R.

    2016-01-01

    Staphylococcus aureus is a leading cause of human infections worldwide. The pathogen produces numerous molecules that can interfere with recognition and binding by host innate immune cells, an initial step required for the ingestion and subsequent destruction of microbes by phagocytes. To better understand the interaction of this pathogen with human immune cells, we compared the association of S. aureus and S. epidermidis with leukocytes in human blood. We found that a significantly greater proportion of B cells associated with S. epidermidis relative to S. aureus. Complement components and complement receptors were important for the binding of B cells with S. epidermidis. Experiments using staphylococci inactivated by ultraviolet radiation and S. aureus isogenic deletion mutants indicated that S. aureus secretes molecules regulated by the SaeR/S two-component system that interfere with the ability of human B cells to bind this bacterium. We hypothesize that the relative inability of B cells to bind S. aureus contributes to the microbe’s success as a human pathogen. PMID:27711145

  20. Intrinsic radiation resistance in human chondrosarcoma cells

    SciTech Connect

    Moussavi-Harami, Farid; Mollano, Anthony; Martin, James A.; Ayoob, Andrew; Domann, Frederick E.; Gitelis, Steven; Buckwalter, Joseph A. . E-mail: joseph-buckwalter@uiowa.edu

    2006-07-28

    Human chondrosarcomas rarely respond to radiation treatment, limiting the options for eradication of these tumors. The basis of radiation resistance in chondrosarcomas remains obscure. In normal cells radiation induces DNA damage that leads to growth arrest or death. However, cells that lack cell cycle control mechanisms needed for these responses show intrinsic radiation resistance. In previous work, we identified immortalized human chondrosarcoma cell lines that lacked p16{sup ink4a}, one of the major tumor suppressor proteins that regulate the cell cycle. We hypothesized that the absence of p16{sup ink4a} contributes to the intrinsic radiation resistance of chondrosarcomas and that restoring p16{sup ink4a} expression would increase their radiation sensitivity. To test this we determined the effects of ectopic p16{sup ink4a} expression on chondrosarcoma cell resistance to low-dose {gamma}-irradiation (1-5 Gy). p16{sup ink4a} expression significantly increased radiation sensitivity in clonogenic assays. Apoptosis did not increase significantly with radiation and was unaffected by p16{sup ink4a} transduction of chondrosarcoma cells, indicating that mitotic catastrophe, rather than programmed cell death, was the predominant radiation effect. These results support the hypothesis that p16{sup ink4a} plays a role in the radiation resistance of chondrosarcoma cell lines and suggests that restoring p16 expression will improve the radiation sensitivity of human chondrosarcomas.

  1. Promotion or suppression of experimental metastasis of B16 melanoma cells after oral administration of lapachol

    SciTech Connect

    Maeda, Masayo; Murakami, Manabu; Takegami, Tsutomu; Ota, Takahide

    2008-06-01

    Lapachol [2-hydroxy-3-(3-methyl-2-butenyl)-1,4-naphthoquinone] is a vitamin K antagonist with antitumor activity. The effect of lapachol on the experimental metastasis of murine B16BL6 melanoma cells was examined. A single oral administration of a high toxic dose of lapachol (80-100 mg/kg) 6 h before iv injection of tumor cells drastically promoted metastasis. This promotion of metastasis was also observed in T-cell-deficient mice and NK-suppressed mice. In vitro treatment of B16BL6 cells with lapachol promoted metastasis only slightly, indicating that lapachol promotes metastasis primarily by affecting host factors other than T cells and NK cells. A single oral administration of warfarin, the most commonly used vitamin K antagonist, 6 h before iv injection of tumor cells also drastically promoted the metastasis of B16BL6 cells. The promotion of metastasis by lapachol and warfarin was almost completely suppressed by preadministration of vitamin K3, indicating that the promotion of metastasis by lapachol was derived from vitamin K antagonism. Six hours after oral administration of lapachol or warfarin, the protein C level was reduced maximally, without elongation of prothrombin time. These observations suggest that a high toxic dose of lapachol promotes metastasis by inducing a hypercoagulable state as a result of vitamin K-dependent pathway inhibition. On the other hand, serial oral administration of low non-toxic doses of lapachol (5-20 mg/kg) weakly but significantly suppressed metastasis by an unknown mechanism, suggesting the possible use of lapachol as an anti-metastatic agent.

  2. Local anesthetics induce human renal cell apoptosis.

    PubMed

    Lee, H Thomas; Xu, Hua; Siegel, Cory D; Krichevsky, Igor E

    2003-01-01

    Renal cell apoptosis contributes significantly to the pathogenesis of acute renal failure. Local anesthetics induce apoptosis in neuronal and lymphocytic cell lines. We examined the effects of chronic (48 h) local anesthetic treatment (lidocaine, bupivacaine and tetracaine) on human proximal tubular (HK-2) cells. Apoptosis induction was assessed by detecting poly(ADP)-ribose polymerase fragmentation, caspase activation, terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) staining, DNA laddering and by cellular morphology. Cell death was quantified by measuring neutral red dye uptake and lactate dehydrogenase released into the cell culture medium. All 3 local anesthetics caused concentration-dependent cell death, induced HK-2 cell apoptosis and potentiated TNF-alpha induced apoptosis. Local anesthetics induced HK-2 cell apoptosis by activation of caspases 3, 6, 7, 8 and 9. ZVAD-fmk, a pan-caspase inhibitor, blocked the local anesthetic induced HK-2 cell apoptosis. Local anesthetics also inhibited the activities of anti-apoptotic kinases protein kinase B (Akt) and extracellular signal regulated mitrogen-activated protein kinase. Local anesthetic's pro-apoptotic effects are independent of sodium channel inhibition as tetrodotoxin, a selective voltage-gated sodium channel blocker, failed to mimic local anesthetic-mediated induction or potentiation of HK-2 cell apoptosis. We conclude that local anesthetics induce human renal cell apoptotic signaling by caspase activation and via inhibition of pro-survival signaling pathways.

  3. Human spleen and red blood cells

    NASA Astrophysics Data System (ADS)

    Pivkin, Igor; Peng, Zhangli; Karniadakis, George; Buffet, Pierre; Dao, Ming

    2016-11-01

    Spleen plays multiple roles in the human body. Among them is removal of old and altered red blood cells (RBCs), which is done by filtering cells through the endothelial slits, small micron-sized openings. There is currently no experimental technique available that allows us to observe RBC passage through the slits. It was previously noticed that people without a spleen have less deformable red blood cells, indicating that the spleen may play a role in defining the size and shape of red blood cells. We used detailed RBC model implemented within the Dissipative Particle Dynamics (DPD) simulation framework to study the filter function of the spleen. Our results demonstrate that spleen indeed plays major role in defining the size and shape of the healthy human red blood cells.

  4. Generation of pluripotent stem cells from adult human testis.

    PubMed

    Conrad, Sabine; Renninger, Markus; Hennenlotter, Jörg; Wiesner, Tina; Just, Lothar; Bonin, Michael; Aicher, Wilhelm; Bühring, Hans-Jörg; Mattheus, Ulrich; Mack, Andreas; Wagner, Hans-Joachim; Minger, Stephen; Matzkies, Matthias; Reppel, Michael; Hescheler, Jürgen; Sievert, Karl-Dietrich; Stenzl, Arnulf; Skutella, Thomas

    2008-11-20

    Human primordial germ cells and mouse neonatal and adult germline stem cells are pluripotent and show similar properties to embryonic stem cells. Here we report the successful establishment of human adult germline stem cells derived from spermatogonial cells of adult human testis. Cellular and molecular characterization of these cells revealed many similarities to human embryonic stem cells, and the germline stem cells produced teratomas after transplantation into immunodeficient mice. The human adult germline stem cells differentiated into various types of somatic cells of all three germ layers when grown under conditions used to induce the differentiation of human embryonic stem cells. We conclude that the generation of human adult germline stem cells from testicular biopsies may provide simple and non-controversial access to individual cell-based therapy without the ethical and immunological problems associated with human embryonic stem cells.

  5. Cell mechanics and human disease states

    NASA Astrophysics Data System (ADS)

    Suresh, Subra

    2006-03-01

    This presentation will provide summary of our very recent studies exploring the effects of biochemical factors, influenced by foreign organisms or in vivo processes, on intracellular structural reorganization, single-cell mechanical response and motility of a population of cells in the context of two human diseases: malaria induced by Plasmodium falciparum merozoites that invade red blood cells, and gastrointestinal cancer metastasis involving epithelial cells. In both cases, particular attention will be devoted to systematic changes induced in specific molecular species in response to controlled alterations in disease state. The role of critical proteins in influencing the mechanical response of human red bloods during the intra-erythrocytic development of P. falciparum merozoites has also been assessed quantitatively using specific protein knock-out experiments by recourse to gene inactivation methods. Single-cell mechanical response characterization entails such tools as optical tweezers and mechanical plate stretchers whereas cell motility assays and cell-population biorheology characterization involves microfluidic channels. The experimental studies are accompanied by three-dimensional computational simulations at the continuum and mesoscopic scales of cell deformation. An outcome of such combined experimental and computational biophysical studies is the realization of how chemical factors influence single-cell mechanical response, cytoadherence, the biorheology of a large population of cells through microchannels representative of in vivo conditions, and the onset and progression of disease states.

  6. Mast cells in human health and disease.

    PubMed

    DeBruin, Erin J; Gold, Matthew; Lo, Bernard C; Snyder, Kimberly; Cait, Alissa; Lasic, Nikola; Lopez, Martin; McNagny, Kelly M; Hughes, Michael R

    2015-01-01

    Mast cells are primarily known for their role in defense against pathogens, particularly bacteria; neutralization of venom toxins; and for triggering allergic responses and anaphylaxis. In addition to these direct effector functions, activated mast cells rapidly recruit other innate and adaptive immune cells and can participate in "tuning" the immune response. In this review we touch briefly on these important functions and then focus on some of the less-appreciated roles of mast cells in human disease including cancer, autoimmune inflammation, organ transplant, and fibrosis. Although it is difficult to formally assign causal roles to mast cells in human disease, we offer a general review of data that correlate the presence and activation of mast cells with exacerbated inflammation and disease progression. Conversely, in some restricted contexts, mast cells may offer protective roles. For example, the presence of mast cells in some malignant or cardiovascular diseases is associated with favorable prognosis. In these cases, specific localization of mast cells within the tissue and whether they express chymase or tryptase (or both) are diagnostically important considerations. Finally, we review experimental animal models that imply a causal role for mast cells in disease and discuss important caveats and controversies of these findings.

  7. Repetitive busulfan administration after hematopoietic stem cell gene therapy associated with a dominant HDAC7 clone in a nonhuman primate.

    PubMed

    Xie, Jianjun; Larochelle, Andre; Maric, Irina; Faulhaber, Marion; Donahue, Robert E; Dunbar, Cynthia E

    2010-06-01

    The risk of genotoxicity of retroviral vector-delivered gene therapy targeting hematopoietic stem cells (HSCs) has been highlighted by the development of clonal dominance and malignancies in human and animal gene therapy trials. Large-animal models have proven invaluable to test the safety of retroviral vectors, but the detection of clonal dominance may require years of follow-up. We hypothesized that hematopoietic stress may accelerate the proliferation and therefore the detection of abnormal clones in these models. We administered four monthly busulfan (Bu) infusions to induce hematopoietic stress in a healthy rhesus macaque previously transplanted with CD34+ cells transduced with retroviral vectors carrying a simple marker gene. Busulfan administration resulted in significant cytopenias with each cycle, and prolonged pancytopenia after the final cycle with eventual recovery. Before busulfan treatment there was highly polyclonal marking in all lineages. After Bu administration clonal diversity was markedly decreased in all lineages. Unexpectedly, we found no evidence of selection of the MDS1/EVI1 clones present before Bu administration, but a clone with a vector integration in intron 1 of the histone deacetylase-7 (HDAC7) gene became dominant in granulocytes over time after Bu administration. The overall marking level in the animal was increased significantly after Bu treatment and coincident with expansion of the HDAC7 clone, suggesting an in vivo advantage for this clone under stress. HDAC7 expression was upregulated in marrow progenitors containing the vector. Almost 5 years after Bu administration, the animal developed progressive cytopenias, and at autopsy the marrow showed complete lack of neutrophil or platelet maturation, with a new population of approximately 20% undifferentiated blasts. These data suggest that chemotherapeutic stress may accelerate vector-related clonal dominance, even in the absence of drug resistance genes expressed by the vector

  8. Recommendations for the standardisation of oxytocin nasal administration and guidelines for its reporting in human research.

    PubMed

    Guastella, Adam J; Hickie, Ian B; McGuinness, Margaret M; Otis, Melissa; Woods, Elizabeth A; Disinger, Hannah M; Chan, Hak-Kim; Chen, Timothy F; Banati, Richard B

    2013-05-01

    A series of studies have reported on the salubrious effects of oxytocin nasal spray on social cognition and behavior in humans, across physiology (e.g., eye gaze, heart rate variability), social cognition (e.g., attention, memory, and appraisal), and behavior (e.g., trust, generosity). Findings suggest the potential of oxytocin nasal spray as a treatment for various psychopathologies, including autism and schizophrenia. There are, however, increasing reports of variability of response to oxytocin nasal spray between experiments and individuals. In this review, we provide a summary of factors that influence transmucosal nasal drug delivery, deposition, and their impact on bioavailability. These include variations in anatomy and resultant airflow dynamic, vascularisation, status of blood vessels, mode of spray application, gallenic formulation (including presence of uptake enhancers, control release formulation), and amount and method of administration. These key variables are generally poorly described and controlled in scientific reports, in spite of their potential to alter the course of treatment outcome studies. Based on this review, it should be of no surprise that differences emerge across individuals and experiments when nasal drug delivery methods are employed. We present recommendations for researchers to use when developing and administering the spray, and guidelines for reporting on peptide nasal spray studies in humans. We hope that these recommendations assist in establishing a scientific standard that can improve the rigor and subsequent reliability of reported effects of oxytocin nasal spray in humans.

  9. Intrapericardial administration of mesenchymal stem cells in a large animal model: a bio-distribution analysis.

    PubMed

    Blázquez, Rebeca; Sánchez-Margallo, Francisco Miguel; Crisóstomo, Verónica; Báez, Claudia; Maestre, Juan; García-Lindo, Mónica; Usón, Alejandra; Álvarez, Verónica; Casado, Javier G

    2015-01-01

    The appropriate administration route for cardiovascular cell therapy is essential to ensure the viability, proliferative potential, homing capacity and implantation of transferred cells. At the present, the intrapericardial administration of pharmacological agents is considered an efficient method for the treatment of cardiovascular diseases. However, only a few reports have addressed the question whether the intrapericardial delivery of Mesenchymal Stem Cells (MSCs) could be an optimal administration route. This work firstly aimed to analyze the pericardial fluid as a cell-delivery vehicle. Moreover, the in vivo biodistribution pattern of intrapericardially administered MSCs was evaluated in a clinically relevant large animal model. Our in vitro results firstly showed that, MSCs viability, proliferative behavior and phenotypic profile were unaffected by exposure to pericardial fluid. Secondly, in vivo cell tracking by magnetic resonance imaging, histological examination and Y-chromosome amplification clearly demonstrated the presence of MSCs in pericardium, ventricles (left and right) and atrium (left and right) when MSCs were administered into the pericardial space. In conclusion, here we demonstrate that pericardial fluid is a suitable vehicle for MSCs and intrapericardial route provides an optimal retention and implantation of MSCs.

  10. Intrapericardial Administration of Mesenchymal Stem Cells in a Large Animal Model: A Bio-Distribution Analysis

    PubMed Central

    Crisóstomo, Verónica; Báez, Claudia; Maestre, Juan; García-Lindo, Mónica; Usón, Alejandra; Álvarez, Verónica

    2015-01-01

    The appropriate administration route for cardiovascular cell therapy is essential to ensure the viability, proliferative potential, homing capacity and implantation of transferred cells. At the present, the intrapericardial administration of pharmacological agents is considered an efficient method for the treatment of cardiovascular diseases. However, only a few reports have addressed the question whether the intrapericardial delivery of Mesenchymal Stem Cells (MSCs) could be an optimal administration route. This work firstly aimed to analyze the pericardial fluid as a cell-delivery vehicle. Moreover, the in vivo biodistribution pattern of intrapericardially administered MSCs was evaluated in a clinically relevant large animal model. Our in vitro results firstly showed that, MSCs viability, proliferative behavior and phenotypic profile were unaffected by exposure to pericardial fluid. Secondly, in vivo cell tracking by magnetic resonance imaging, histological examination and Y-chromosome amplification clearly demonstrated the presence of MSCs in pericardium, ventricles (left and right) and atrium (left and right) when MSCs were administered into the pericardial space. In conclusion, here we demonstrate that pericardial fluid is a suitable vehicle for MSCs and intrapericardial route provides an optimal retention and implantation of MSCs. PMID:25816232

  11. 3 CFR - Guidelines for Human Stem Cell Research

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 3 The President 1 2010-01-01 2010-01-01 false Guidelines for Human Stem Cell Research Presidential Documents Other Presidential Documents Memorandum of July 30, 2009 Guidelines for Human Stem Cell Research..., scientifically worthy human stem cell research, including human embryonic stem cell research, to the...

  12. Myristoylation profiling in human cells and zebrafish

    PubMed Central

    Broncel, Malgorzata; Serwa, Remigiusz A; Ciepla, Paulina; Krause, Eberhard; Dallman, Margaret J.; Magee, Anthony I.; Tate, Edward W.

    2015-01-01

    Human cells (HEK 293, HeLa, MCF-7) and zebrafish embryos were metabolically tagged with an alkynyl myristic acid probe, lysed with an SDS buffer and tagged proteomes ligated to multifunctional capture reagents via copper-catalyzed alkyne azide cycloaddition (CuAAC). This allowed for affinity enrichment and high-confidence identification, by delivering direct MS/MS evidence for the modification site, of 87 and 61 co-translationally myristoylated proteins in human cells and zebrafish, respectively. The data have been deposited to ProteomeXchange Consortium (Vizcaíno et al., 2014 Nat. Biotechnol., 32, 223–6) (PXD001863 and PXD001876) and are described in detail in Multifunctional reagents for quantitative proteome-wide analysis of protein modification in human cells and dynamic protein lipidation during vertebrate development׳ by Broncel et al., Angew. Chem. Int. Ed. PMID:26217820

  13. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins

    PubMed Central

    Fukusumi, Hayato; Shofuda, Tomoko; Bamba, Yohei; Yamamoto, Atsuyo; Kanematsu, Daisuke; Handa, Yukako; Okita, Keisuke; Nakamura, Masaya; Yamanaka, Shinya; Okano, Hideyuki; Kanemura, Yonehiro

    2016-01-01

    Human neural progenitor cells (hNPCs) have previously been generated from limited numbers of human induced pluripotent stem cell (hiPSC) clones. Here, 21 hiPSC clones derived from human dermal fibroblasts, cord blood cells, and peripheral blood mononuclear cells were differentiated using two neural induction methods, an embryoid body (EB) formation-based method and an EB formation method using dual SMAD inhibitors (dSMADi). Our results showed that expandable hNPCs could be generated from hiPSC clones with diverse somatic tissue origins. The established hNPCs exhibited a mid/hindbrain-type neural identity and uniform expression of neural progenitor genes. PMID:27212953

  14. The human mast cell: an overview.

    PubMed

    Krishnaswamy, Guha; Ajitawi, Omar; Chi, David S

    2006-01-01

    Mast cells are fascinating, multifunctional, tissue-dwelling cells that have been traditionally associated with the allergic response. However, recent studies suggest these cells may be capable of regulating inflammation, host defense, and innate immunity. The purpose of this review is to present salient aspects of mast cell biology in the context of mast cell function in physiology and disease. After their development from bone marrow-derived progenitor cells that are primed with stem cell factor, mast cells continue their maturation and differentiation in peripheral tissue, developing into two well-described subsets of cells, MC(T) and MC(TC) cells. These cells can be distinguished on the basis of their tissue location, dependence on T lymphocytes, and their granule contents. Mast cells can undergo activation by antigens/allergens, superoxides, complement proteins, neuropeptides, and lipoproteins. After activation, mast cells express histamine, leukotrienes, and prostanoids, as well as proteases, and many cytokines and chemokines. These mediators may be pivotal to the genesis of an inflammatory response. By virtue of their location and mediator expression, mast cells may play an active role in many diseases, such as allergy, parasitic diseases, atherosclerosis, malignancy, asthma, pulmonary fibrosis, and arthritis. Recent data also suggest that mast cells play a vital role in host defense against pathogens by elaboration of tumor necrosis factor alpha. Mast cells also express the Toll-like receptor, which may further accentuate their role in the immune-inflammatory response. This chapter summarizes the many well-known and novel functional aspects of human mast cell biology and emphasizes their unique role in the inflammatory response.

  15. METHAMPHETAMINE SELF-ADMINISTRATION IN HUMANS DURING D-AMPHETAMINE MAINTENANCE

    PubMed Central

    Pike, Erika; Stoops, William W.; Hays, Lon R.; Glaser, Paul E. A.; Rush, Craig R.

    2014-01-01

    Agonist replacement may be a viable treatment approach for managing stimulant use disorders. This study sought to determine the effects of d-amphetamine maintenance on methamphetamine self-administration in stimulant using human participants. We predicted d-amphetamine maintenance would reduce methamphetamine self-administration. Eight participants completed the protocol, which tested two d-amphetamine maintenance conditions in counter-balanced order (0 and 40 mg/day). Participants completed 4 experimental sessions under each maintenance condition in which they first sampled one of four doses of intranasal methamphetamine (0, 10, 20, or 30 mg). Participants then had the opportunity to respond on a computerized progressive ratio task to earn portions of the sampled methamphetamine dose. Subject-rated drug-effect and physiological measures were completed at regular intervals prior to and after sampling methamphetamine. Methamphetamine was self-administered as an orderly function of dose regardless of the maintenance condition. Methamphetamine produced prototypical subject-rated effects on 13 items of the drug-effects questionnaires, 10 of which were attenuated by d-amphetamine maintenance (e.g., increased ratings were attenuated on items such as Any Effect, Like Drug, and Willing to Take Again on the Drug Effect Questionnaire). Methamphetamine produced significant increases in systolic blood pressure, which were attenuated by d-amphetamine maintenance compared to placebo maintenance. Methamphetamine was well tolerated during d-amphetamine maintenance and no adverse events occurred. Although d-amphetamine attenuated some subject-rated effects of methamphetamine, the self-administration results are concordant with those of clinical trials showing that d-amphetamine did not reduce methamphetamine use. Unique pharmacological approaches may be needed for treating amphetamine use disorders. PMID:25154010

  16. Metabolites of protoberberine alkaloids in human urine following oral administration of Coptidis Rhizoma decoction.

    PubMed

    Yang, Yihui; Kang, Ning; Xia, Hongjun; Li, Jun; Chen, Lixia; Qiu, Feng

    2010-11-01

    Coptidis Rhizoma has been used as a traditional Chinese herbal medicine to treat typhoid, pharyngolaryngitis, diabetes mellitus, gastroenteritis and secretory diarrhea for more than a thousand years in China. However, there is little information on the IN VIVO chemical constituents of Coptidis Rhizoma following oral administration. In this paper, the alkaloid constituents in urine were studied in humans following oral administration of Coptidis Rhizoma decoction. Using macroporous adsorption resin chromatography, open ODS column chromatography, and preparative high-performance liquid chromatography, twelve protoberberine alkaloid constituents were isolated. Their structures were elucidated by chemical evidence, enzymatic deconjugation and analyses of mass, (1)H-NMR and NOESY spectra. The identified alkaloid constituents include berberine ( P1), groenlandicine 3-O- β-D-glucuronide (M1), dehydrocheilanthifoline 2-O-β-D-glucuronide (M2), thalifendine 10-O-β-D-glucuronide (M3), jatrorrhizine 3-O-β-D-glucuronide (M4), columbamine 2-O-β-D-glucuronide (M5), berberrubine 9-O-β-D-glucuronide (M6), jatrorrhizine 3-O-sulfate (M7), demethyleneberberine 2-O-sulfate (M8), dehydrocorydalmine 10-O-sulfate (M9), 3,10-demethylpalmatine 10-O-sulfate (M10) and 2,3,10-trihydroxyberberine 2-O-sulfate ( M11). No other parent protoberberine alkaloids from Coptidis Rhizoma except for a trace of berberine were found in the urine. These findings suggested that the protoberberine alkaloids, which were absorbed in vivo following oral administration of Coptidis Rhizoma decoction, were mainly conjugated with glucuronic acid or sulfuric acid to form phase II metabolites directly or after biotransformation to phase I metabolites, and finally excreted in urine.

  17. Plasma prolactin changes during the administration of human menopausal gonadotropins in nonovulatory women.

    PubMed

    Kemmann, E; GEMZELL, C A; Beinert, W C; Beling, C B; Jones, J R

    1977-09-15

    Plasma prolactin concentrations were determined in 16 nonovulatory women during treatment with human meonpausal gonadotropins (hMG). In eight patients with initially normal prolactin levels of less than 20 ng. per milliliter, a significant rise was noted at the end of hMG administration, this is thought to be a response to increased endogenous estrogen concentrations. A similar rise in plasma prolactin levels was seen in some but not all of the eight patients with initially elevated "basal" prolactin concentrations. Three of these hyperprolactinemic patients had radiographic evidence of a pituitary lesion--either a pituitary adenoma or a "microadenoma"--but the variance in prolactin response could not be explained on this basis. The two groups of normo- and hyper-prolactinemic patients showed no significant difference in the required dosage and duration or hMG treatment, plasma estradiol-17 beta response, and ovulatory and pregnancy outcome.

  18. Acute d-Amphetamine Pretreatment Does Not Alter Stimulant Self-Administration in Humans

    PubMed Central

    Stoops, William W.; Vansickel, Andrea R.; Lile, Joshua A.; Rush, Craig R.

    2007-01-01

    Recent clinical research indicates that d-amphetamine is effective in treating cocaine and methamphetamine dependence. There is concern, however, with the use of d-amphetamine as a pharmacotherapy because acute administration of d-amphetamine decreases inhibition in cocaine-using individuals and may increase drug-taking behavior. The purpose of the present experiment was to determine whether acute d-amphetamine pretreatment would alter the reinforcing, subject-rated, and cardiovascular effects of d-amphetamine. To this end, 7 human volunteers first sampled doses of oral d-amphetamine (0, 8, and 16 mg). These doses engender moderate drug taking and were selected to avoid a ceiling or floor effect. Volunteers were then allowed to self-administer these sampled doses using a modified-progressive ratio procedure in two sessions in which they received pretreatments with either 0 or 15 mg oral d-amphetamine 2 hours prior to completing the modified progressive-ratio procedure. d-Amphetamine produced prototypical stimulant-like effects (e.g., increased ratings of stimulated, elevated blood pressure) and maintained responding on the modified progressive-ratio schedule. Pretreatment with 15 mg oral d-amphetamine also produced prototypical stimulant-like effects, but failed to alter break points for d-amphetamine on the modified progressive-ratio procedure relative to placebo pretreatment. These results indicate that acute d-amphetamine pretreatment does not increase stimulant self-administration. PMID:17490738

  19. A new paradigm for the closed-loop intraoperative administration of analgesics in humans.

    PubMed

    Gentilini, Andrea; Schaniel, Christoph; Morari, Manfred; Bieniok, Christian; Wymann, Rolf; Schnider, Thomas

    2002-04-01

    We present a new paradigm for the closed-loop administration of analgesics during general anesthesia. The manipulated variable in the control system is the infusion rate of the opiate alfentanil, administered intravenously through a computer-controlled infusion pump (CCIP). The outputs to be controlled are the patient's mean arterial pressure (MAP) and the drug concentration in the plasma. Maintaining MAP within appropriate ranges provides optimal treatment of the patient's reactions to surgical stimuli. Maintaining plasma drug concentrations close to a reference value specified by the anesthesiologist allows to titrate analgesic administration to qualitative clinical end-points of insufficient analgesia. MAP is acquired invasively through a catheter cannula. Since plasma drug concentrations cannot be measured on-line, they are estimated via a pharmacokinetic model. We describe an explicit model-predictive controller which achieves the above-mentioned objectives. An upper constraint on drug concentrations is maintained to avoid overdosing. Constraints on the MAP are introduced to trigger a prompt controller reaction during hypertensive and hypotensive periods. Measurement artifacts in the MAP signal are rejected to prevent harmful misbehavior of the controller. We discuss the results of the clinical validation of the controller on humans.

  20. Rapid intravenous administration of amino acids prevents biliary sludge induced by total parenteral nutrition in humans.

    PubMed

    Wu, Z S; Yu, L; Lin, Y J; Jun, Z J; Min, W S; Jun, Y; Hua, Z B

    2000-01-01

    The aim of this study was to evaluate whether daily rapid intravenous administration of amino acids (IVAA) prevented the formation of biliary sludge in humans receiving long-term total parenteral nutrition (TPN). Thirty adult patients receiving TPN for more than 28 consecutive days were studied. They were randomized to receive either saline solution (placebo) intravenously (15 patients) or 6.9% branched chain amino acid (BCAA)-enriched amino acid (15 synthetic amino acids; Freamine HBC) solution given by administration rapid intravenous (15 patients). The groups were similar with respect to age, sex, diagnosis, liver function test results, amylase levels, TPN time, and time of study. All patients underwent weekly ultrasound studies. Volume and emptying studies of the gallbladder in response to the study drug were performed after 1 week. As a result, none of the patients receiving rapid IVAA had sludge, whereas 11 of the 15 patients receiving placebo had sludge (P < 0.01). Results of emptying studies showed significant contraction of the gallbladder in those in the rapid IVAA group, but not in the placebo group. Consequently, the data suggest that rapid IVAA given daily prevents TPN-induced stasis and sludge in the gallbladder. We conclude that rapid IVAA should be used as routine prophylaxis against biliary sludge and formation of gallstones in patients receiving long-term TPN.

  1. Infrastructure development for human cell therapy translation.

    PubMed

    Dietz, A B; Padley, D J; Gastineau, D A

    2007-09-01

    The common conception of a drug is that of a chemical with defined medicinal effect. However, cells used as drugs remain critical to patient care. Cell therapy's origins began with the realization that complex tissues such as blood can retain function when transplanted to the patient. More complex transplantation followed, culminating with the understanding that transplantation of some tissues such as bone marrow may act medicinally. Administration of cells with an intended therapeutic effect is a hallmark of cellular therapy. While cells have been used as drugs for decades, testing a specific therapeutic effect of cells has begun clinical testing relatively recently. Lessons learned during the establishment of blood banking (including the importance of quality control, process control, sterility, and product tracking) are key components in the assurance of the safety and potency of cell therapy preparations. As more academic medical centers and private companies move toward exploiting the full potential of cells as drugs, needs arise for the development of the infrastructure necessary to support these investigations. Careful consideration of the design of the structure used to manufacture is important in terms of the significant capital outlay involved and the facility's role in achieving regulatory compliance. This development perspective describes the regulatory environment surrounding the infrastructure support for cell therapy and practical aspects for design consideration with particular focus on those activities associated with early clinical trials.

  2. Toxicokinetics of acrylamide in rats and humans following single oral administration of low doses

    SciTech Connect

    Kopp, Eva Katharina; Dekant, Wolfgang

    2009-03-01

    The rodent carcinogen acrylamide (AA) is formed during preparation of starch-containing foods. AA is partly metabolized to the genotoxic epoxide glycidamide (GA). After metabolic processing, the mercapturic acids N-acetyl-S-(2-carbamoylethyl)-L-cysteine (AAMA), rac-N-acetyl-S-(2-carbamoyl-2-hydroxyethyl)-L-cysteine (GAMA) and rac-N-acetyl-S-(1-carbamoyl-moyl-2-hydroxyethyl)-L-cysteine (iso-GAMA) are excreted with urine. In humans, AAMA can be sulfoxidized to AAMA-sulfoxide. The aim of this study was to assess potential species-differences in AA-toxicokinetics in rats and humans after single oral administration of doses similar to the daily human dietary exposure. Male Fischer 344 rats (n = 5/dose group) were administered 20 and 100 {mu}g/kg b.w. {sup 13}C{sub 3}-AA in deionized water via oral gavage. Human subjects (n = 3/gender) were orally administered 0.5 and 20 {mu}g/kg b.w. {sup 13}C{sub 3}-AA with drinking water. Urine samples were collected in intervals for 96 and 94 h, respectively. Urinary concentrations of {sup 13}C{sub 3}-AAMA, {sup 13}C{sub 3}-GAMA and {sup 13}C{sub 3}-AAMA-sulfoxide were monitored by liquid chromatography-tandem mass spectrometry. The recovered urinary metabolites accounted for 66.3% and 70.5% of the 20 and 100 {mu}g/kg b.w. doses in rats and for 71.3% and 70.0% of the 0.5 and 20 {mu}g/kg b.w. doses in humans. In rats, {sup 13}C{sub 3}-AAMA accounted for 33.6% and 38.8% of dose and 32.7% and 31.7% of dose was recovered as {sup 13}C{sub 3}-GAMA; {sup 13}C{sub 3}-AAMA-sulfoxide was not detected in rat urine. In humans, {sup 13}C{sub 3}-AAMA, {sup 13}C{sub 3}-GAMA and {sup 13}C{sub 3}-AAMA-sulfoxide accounted for 51.7% and 49.2%, 6.3% and 6.4% and 13.2% and 14.5% of the applied dose, respectively. The obtained results suggest that the extent of AA bioactivation to GA in humans is lower than in rodents.

  3. CLOSTRIDIUM SPORE ATTACHMENT TO HUMAN CELLS

    SciTech Connect

    PANESSA-WARREN,B.; TORTORA,G.; WARREN,J.

    1997-08-10

    This paper uses high resolution scanning electron microscopy (SEM) with a LaB6 gun and the newest commercial field emission guns, to obtain high magnification images of intact clostridial spores throughout the activation/germination/outgrowth process. By high resolution SEM, the clostridial exosporial membrane can be seen to produce numerous delicate projections (following activation), that extend from the exosporial surface to a nutritive substrate (agar), or cell surface when anaerobically incubated in the presence of human cells (embryonic fibroblasts and colon carcinoma cells). Magnifications of 20,000 to 200,000Xs at accelerating voltages low enough to minimize or eliminate specimen damage (1--5 kV) have permitted the entire surface of C.sporogenes and C.difficile endospores to be examined during all stages of germination. The relationships between the spore and the agar or human cell surface were also clearly visible.

  4. Human Colon Cancer Cells Cultivated in Space

    NASA Technical Reports Server (NTRS)

    1995-01-01

    Within five days, bioreactor cultivated human colon cancer cells (shown) grown in Microgravity on the STS-70 mission in 1995, had grown 30 times the volume of the control specimens on Earth. The samples grown in space had a higher level of cellular organization and specialization. Because they more closely resemble tumors found in the body, microgravity grown cell cultures are ideal for research purposes.

  5. Human somatic cell nuclear transfer and cloning.

    PubMed

    2012-10-01

    This document presents arguments that conclude that it is unethical to use somatic cell nuclear transfer (SCNT) for infertility treatment due to concerns about safety; the unknown impact of SCNT on children, families, and society; and the availability of other ethically acceptable means of assisted reproduction. This document replaces the ASRM Ethics Committee report titled, "Human somatic cell nuclear transfer (cloning)," last published in Fertil Steril 2000;74:873-6.

  6. 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.

  7. Intranasal Administration of Maleic Anhydride-Modified Human Serum Albumin for Pre-Exposure Prophylaxis of Respiratory Syncytial Virus Infection

    PubMed Central

    Sun, Zhiwu; Wang, Qian; Jia, Ran; Xia, Shuai; Li, Yuan; Liu, Qi; Xu, Wei; Xu, Jin; Du, Lanying; Lu, Lu; Jiang, Shibo

    2015-01-01

    Respiratory syncytial virus (RSV) is the leading cause of pediatric viral respiratory tract infections. Neither vaccine nor effective antiviral therapy is available to prevent and treat RSV infection. Palivizumab, a humanized monoclonal antibody, is the only product approved to prevent serious RSV infection, but its high cost is prohibitive in low-income countries. Here, we aimed to identify an effective, safe, and affordable antiviral agent for pre-exposure prophylaxis (PrEP) of RSV infection in children at high risk. We found that maleic anhydride (ML)-modified human serum albumin (HSA), designated ML-HSA, exhibited potent antiviral activity against RSV and that the percentages of the modified lysines and arginies in ML- are correlated with such anti-RSV activity. ML-HSA inhibited RSV entry and replication by interacting with viral G protein and blocking RSV attachment to the target cells, while ML-HAS neither bound to F protein, nor inhibited F protein-mediated membrane fusion. Intranasal administration of ML-HSA before RSV infection resulted in significant decrease of the viral titers in the lungs of mice. ML-HSA shows promise for further development into an effective, safe, affordable, and easy-to-use intranasal regimen for pre-exposure prophylaxis of RSV infection in children at high risk in both low- and high-income countries. PMID:25690799

  8. Human Mammary Luminal Epithelial Cells Contain Progenitors to Myoepithelial Cells

    SciTech Connect

    Pechoux, Christine; Gudjonsson, Thorarinn; Ronnov-Jessen, Lone; Bissell, Mina J; Petersen, Ole

    1999-02-01

    The origin of the epithelial and myoepithelial cells in the human breast has not been delineated. In this study we have addressed whether luminal epithelial cells and myoepithelial cells are vertically connected, i.e., whether one is the precursor for the other. We used a primary culture assay allowing preservation of basic phenotypic traits of luminal epithelial and myoepithelial cells in culture. The two cell types were then separated immunomagnetically using antibodies directed against lineage-specific cell surface antigens into at best 100% purity. The cellular identity was ascertained by cytochemistry, immunoblotting, and 2-D gel electrophoresis. Luminal epithelial cells were identified by strong expression of cytokeratins 18 and 19 while myoepithelial cells were recognized by expression of vimentin and {alpha}-smooth muscle actin. We used a previously devised culture medium (CDM4) that allows vigorous expansion of proliferative myoepithelial cells and also devised a medium (CDM6) that allowed sufficient expansion of differentiated luminal epithelial cells based on addition of hepatocyte growth factor/scatter factor. The two different culture media supported each lineage for at least five passages without signs of interconversion. We used parallel cultures where we switched culture media, thus testing the ability of each lineage to convert to the other. Whereas the myoepithelial lineage showed no signs of interconversion, a subset of luminal epithelial cells, gradually, but distinctly, converted to myoepithelial cells. We propose that in the mature human breast, it is the luminal epithelial cell compartment that gives rise to myoepithelial cells rather than the other way around.

  9. Intravenous Administration of Endothelial Colony-Forming Cells Overexpressing Integrin β1 Augments Angiogenesis in Ischemic Legs.

    PubMed

    Goto, Kazuko; Takemura, Genzou; Takahashi, Tomoyuki; Okada, Hideshi; Kanamori, Hiromitsu; Kawamura, Itta; Watanabe, Takatomo; Morishita, Kentaro; Tsujimoto, Akiko; Miyazaki, Nagisa; Ushikoshi, Hiroaki; Kawasaki, Masanori; Mikami, Atsushi; Kosai, Ken-ichiro; Minatoguchi, Shinya

    2016-02-01

    When injected directly into ischemic tissue in patients with peripheral artery disease, the reparative capacity of endothelial progenitor cells (EPCs) appears to be limited by their poor survival. We, therefore, attempted to improve the survival of transplanted EPCs through intravenous injection and gene modification. We anticipated that overexpression of integrin β1 will enable injected EPCs to home to ischemic tissue, which abundantly express extracellular matrix proteins, the ligands for integrins. In addition, integrin β1 has an independent angiogenesis-stimulating function. Human endothelial colony-forming cells (ECFCs; late-outgrowth EPCs) were transduced using a lentiviral vector encoding integrin β1 (ITGB1) or enhanced green fluorescent protein (GFP). We then locally or systemically injected phosphate-buffered saline or the genetically modified ECFCs (GFP-ECFCs or ITGB1-ECFCs; 1 × 10(5) cells each) into NOD/Shi-scid, IL-2Rγnull mice whose right femoral arteries had been occluded 24 hours earlier. Upregulation of extracellular matrix proteins, including fibronectin, was apparent in the ischemic legs. Four weeks later, blood perfusion of the ischemic limb was significantly augmented only in the ITGB1-ECFC group. Scanning electron microscopy of vascular casts revealed increases in the perfused blood vessels in the ischemic legs of mice in the ITGB1-ECFC group and significant increases in the density of both capillaries and arterioles. Transplanted ECFC-derived vessels accounted for 28% ± 4.2% of the vessels in the ITGB1-ECFC group, with no cell fusion. Intravenous administration of ECFCs engineered to home to ischemic tissue appears to efficiently mediate therapeutic angiogenesis in a mouse model of peripheral artery disease. Significance: The intravenous administration of endothelial colony-forming cells (ECFCs) genetically modified to overexpress integrin β1 effectively stimulated angiogenesis in ischemic mouse hindlimbs. Transplanted ECFCs were

  10. Intravenous Administration of Endothelial Colony-Forming Cells Overexpressing Integrin β1 Augments Angiogenesis in Ischemic Legs

    PubMed Central

    Goto, Kazuko; Takahashi, Tomoyuki; Okada, Hideshi; Kanamori, Hiromitsu; Kawamura, Itta; Watanabe, Takatomo; Morishita, Kentaro; Tsujimoto, Akiko; Miyazaki, Nagisa; Ushikoshi, Hiroaki; Kawasaki, Masanori; Mikami, Atsushi; Kosai, Ken-ichiro; Minatoguchi, Shinya

    2016-01-01

    When injected directly into ischemic tissue in patients with peripheral artery disease, the reparative capacity of endothelial progenitor cells (EPCs) appears to be limited by their poor survival. We, therefore, attempted to improve the survival of transplanted EPCs through intravenous injection and gene modification. We anticipated that overexpression of integrin β1 will enable injected EPCs to home to ischemic tissue, which abundantly express extracellular matrix proteins, the ligands for integrins. In addition, integrin β1 has an independent angiogenesis-stimulating function. Human endothelial colony-forming cells (ECFCs; late-outgrowth EPCs) were transduced using a lentiviral vector encoding integrin β1 (ITGB1) or enhanced green fluorescent protein (GFP). We then locally or systemically injected phosphate-buffered saline or the genetically modified ECFCs (GFP-ECFCs or ITGB1-ECFCs; 1 × 105 cells each) into NOD/Shi-scid, IL-2Rγnull mice whose right femoral arteries had been occluded 24 hours earlier. Upregulation of extracellular matrix proteins, including fibronectin, was apparent in the ischemic legs. Four weeks later, blood perfusion of the ischemic limb was significantly augmented only in the ITGB1-ECFC group. Scanning electron microscopy of vascular casts revealed increases in the perfused blood vessels in the ischemic legs of mice in the ITGB1-ECFC group and significant increases in the density of both capillaries and arterioles. Transplanted ECFC-derived vessels accounted for 28% ± 4.2% of the vessels in the ITGB1-ECFC group, with no cell fusion. Intravenous administration of ECFCs engineered to home to ischemic tissue appears to efficiently mediate therapeutic angiogenesis in a mouse model of peripheral artery disease. Significance The intravenous administration of endothelial colony-forming cells (ECFCs) genetically modified to overexpress integrin β1 effectively stimulated angiogenesis in ischemic mouse hindlimbs. Transplanted ECFCs were observed

  11. Human embryonic stem cell lines model experimental human cytomegalovirus latency.

    PubMed

    Penkert, Rhiannon R; Kalejta, Robert F

    2013-05-28

    Herpesviruses are highly successful pathogens that persist for the lifetime of their hosts primarily because of their ability to establish and maintain latent infections from which the virus is capable of productively reactivating. Human cytomegalovirus (HCMV), a betaherpesvirus, establishes latency in CD34(+) hematopoietic progenitor cells during natural infections in the body. Experimental infection of CD34(+) cells ex vivo has demonstrated that expression of the viral gene products that drive productive infection is silenced by an intrinsic immune defense mediated by Daxx and histone deacetylases through heterochromatinization of the viral genome during the establishment of latency. Additional mechanistic details about the establishment, let alone maintenance and reactivation, of HCMV latency remain scarce. This is partly due to the technical challenges of CD34(+) cell culture, most notably, the difficulty in preventing spontaneous differentiation that drives reactivation and renders them permissive for productive infection. Here we demonstrate that HCMV can establish, maintain, and reactivate in vitro from experimental latency in cultures of human embryonic stem cells (ESCs), for which spurious differentiation can be prevented or controlled. Furthermore, we show that known molecular aspects of HCMV latency are faithfully recapitulated in these cells. In total, we present ESCs as a novel, tractable model for studies of HCMV latency.

  12. Single administration of butylparaben induces spermatogenic cell apoptosis in prepubertal rats.

    PubMed

    Alam, Mohammad Shah; Ohsako, Seiichiroh; Kanai, Yoshiakira; Kurohmaru, Masamichi

    2014-04-01

    Parabens are p-hydroxybenzoic acid ester compounds widely used as preservatives in foods, cosmetics, toiletries and pharmaceuticals. Some parabens, including butylparaben, exert an estrogenic activity as determined by in vitro estrogen receptor assay and in vivo uterotrophic assay, and adversely affect endocrine secretion and male reproductive function. We conducted a research study to evaluate the acute effects of butylparaben on testicular tissues of prepubertal rats. Three-week-old male rats (n=8) were given a single dose of 1000mg/kg butylparaben. The rats were sacrificed under anesthesia at 3, 6 and 24h after administration, and their testes were collected for histopathological examination. The study revealed progressive detachment and sloughing of spermatogenic cells into the lumen of the seminiferous tubules at 3h, and this effect was enhanced at 6h after administration. Thin seminiferous epithelia and wide tubular lumina were seen at 24h in the butylparaben-treated group, compared to the control. In order to clarify whether sloughed spermatogenic cells underwent apoptosis, TUNEL assay was carried out. We found a significant increase in the number of apoptotic spermatogenic cells in all the treated groups, compared to the controls and a maximal number of apoptotic cells were detected at 6h after administration. In semithin sections, apoptotic cells were easily detected by their prominent basophilia and condensed chromatin, mainly found in spermatocytes. Ultrastructurally, the condensed chromatin and shrunken cytoplasm and nucleus, hallmarks of apoptotic cell death, were observed in butylparaben-treated groups. These observations lead us to postulate that butylparaben, similar to other estrogenic compounds, also induces spermatogenic cell apoptosis.

  13. Enriched retinal ganglion cells derived from human embryonic stem cells

    PubMed Central

    Gill, Katherine P.; Hung, Sandy S. C.; Sharov, Alexei; Lo, Camden Y.; Needham, Karina; Lidgerwood, Grace E.; Jackson, Stacey; Crombie, Duncan E.; Nayagam, Bryony A.; Cook, Anthony L.; Hewitt, Alex W.; Pébay, Alice; Wong, Raymond C. B.

    2016-01-01

    Optic neuropathies are characterised by a loss of retinal ganglion cells (RGCs) that lead to vision impairment. Development of cell therapy requires a better understanding of the signals that direct stem cells into RGCs. Human embryonic stem cells (hESCs) represent an unlimited cellular source for generation of human RGCs in vitro. In this study, we present a 45-day protocol that utilises magnetic activated cell sorting to generate enriched population of RGCs via stepwise retinal differentiation using hESCs. We performed an extensive characterization of these stem cell-derived RGCs by examining the gene and protein expressions of a panel of neural/RGC markers. Furthermore, whole transcriptome analysis demonstrated similarity of the hESC-derived RGCs to human adult RGCs. The enriched hESC-RGCs possess long axons, functional electrophysiological profiles and axonal transport of mitochondria, suggestive of maturity. In summary, this RGC differentiation protocol can generate an enriched population of functional RGCs from hESCs, allowing future studies on disease modeling of optic neuropathies and development of cell therapies. PMID:27506453

  14. Generation of regulatory dendritic cells and CD4+Foxp3+ T cells by probiotics administration suppresses immune disorders.

    PubMed

    Kwon, Ho-Keun; Lee, Choong-Gu; So, Jae-Seon; Chae, Chang-Suk; Hwang, Ji-Sun; Sahoo, Anupama; Nam, Jong Hee; Rhee, Joon Haeng; Hwang, Ki-Chul; Im, Sin-Hyeog

    2010-02-02

    The beneficial effects of probiotics have been described in many diseases, but the mechanism by which they modulate the immune system is poorly understood. In this study, we identified a mixture of probiotics that up-regulates CD4(+)Foxp3(+) regulatory T cells (Tregs). Administration of the probiotics mixture induced both T-cell and B-cell hyporesponsiveness and down-regulated T helper (Th) 1, Th2, and Th17 cytokines without apoptosis induction. It also induced generation of CD4(+)Foxp3(+) Tregs from the CD4(+)CD25(-) population and increased the suppressor activity of naturally occurring CD4(+)CD25(+) Tregs. Conversion of T cells into Foxp3(+) Tregs is directly mediated by regulatory dendritic cells (rDCs) that express high levels of IL-10, TGF-beta, COX-2, and indoleamine 2,3-dioxygenase. Administration of probiotics had therapeutical effects in experimental inflammatory bowel disease, atopic dermatitis, and rheumatoid arthritis. The therapeutical effect of the probiotics is associated with enrichment of CD4(+)Foxp3(+) Tregs in the inflamed regions. Collectively, the administration of probiotics that enhance the generation of rDCs and Tregs represents an applicable treatment of inflammatory immune disorders.

  15. Human mesenchymal stem cell homing induced by SKOV3 cells

    PubMed Central

    Fan, Dongmei; Xie, Xiaojuan; Qi, Pengwei; Yang, Xianan; Jin, Ximeng

    2017-01-01

    Human mesenchymal stem cell (hMSC) homing is the migration of endogenous and exogenous hMSCS to the target organs and the subsequent colonization under the action chemotaxic factors. This is an important process involved in the repair of damaged tissues. However, we know little about the mechanism of hMSC homing. Stromal cell derived factor-1 (SDF-1) is a cytokine secreted by stromal cells. Its only receptor CXCR4 is widely expressed in blood cells, immune cells and cells in the central nervous system. SDF-1/CXCR4 signaling pathway plays an important role in hMSC homing and tissue repair. Human cbll1 gene encodes E3 ubiquitin-protein ligase Hakai (also known as CBLL1) consisting of RING-finger domain that is involved in ubiquitination, endocytosis and degradation of epithelial cadherin (E-cadherin) as well as in the regulation of cell proliferation. We successfully constructed LV3-CXCR4 siRNA lentiviral vector, LV3-CBLL1 RNAi lentiviral vector and the corresponding cell systems which were used to induce hMSC homing in the presence of SKOV3 cells. Thus the mechanism of hMSC homing was studied. PMID:28337256

  16. Surface display of human growth hormone on Bacillus subtilis spores for oral administration.

    PubMed

    Lian, Chaoqun; Zhou, Yang; Feng, Fan; Chen, Liang; Tang, Qi; Yao, Qin; Chen, Keping

    2014-04-01

    Human growth hormone (hGH) is the major and important hormone component of human being. At present, hGH for clinical uses is mostly produced in Escherichia coli, which requires costly denaturation and refolding to recover functionality. To obtain long-term bioactive hormone, we used hGH as a foreign gene and constructed a recombinant plasmid pJS700-hGH which carries a recombinant gene cotC-hgh with an enterokinase site under the control of cotC promoter. Plasmid pJS700-hGH was transformed into Bacillus subtilis by double crossover and an amylase-inactivated mutant was produced. After spore formation, Western blot and fluorescence immunoassay were used to monitor hGH surface expression on spores. Oral administration to silkworm with spores displaying hGH further showed that the recombinant spores may have potential ability to be digested and absorbed into the silkworm's hemolymph due to both the resistant characters of spores and the addition of enterokinase site.

  17. GABAergic modulation of human social interaction in a prisoner's dilemma model by acute administration of alprazolam.

    PubMed

    Lane, Scott D; Gowin, Joshua L

    2009-10-01

    Recent work in neuroeconomics has used game theory paradigms to examine neural systems that subserve human social interaction and decision making. Attempts to modify social interaction through pharmacological manipulation have been less common. Here we show dose-dependent modification of human social behavior in a prisoner's dilemma model after acute administration of the γ-aminobutyric acid (GABA)-A modulating benzodiazepine alprazolam. Nine healthy adults received doses of placebo, 0.5, 1.0, and 2.0 mg alprazolam in a counterbalanced within-subject design, while completing multiple test blocks per day on an iterated prisoner's dilemma game. During test blocks in which peak subjective effects of alprazolam were reported, cooperative choices were significantly decreased as a function of dose. Consistent with previous reports showing that high acute doses of GABA-modulating drugs are associated with violence and other antisocial behavior, our data suggest that at sufficiently high doses, alprazolam can decrease cooperation. These behavioral changes may be facilitated by changes in inhibitory control facilitated by GABA. Game theory paradigms may prove useful in behavioral pharmacology studies seeking to measure social interaction, and may help inform the emerging field of neuroeconomics.

  18. Human embryonic stem cells and cardiac repair.

    PubMed

    Zhu, Wei-Zhong; Hauch, Kip D; Xu, Chunhui; Laflamme, Michael A

    2009-01-01

    The muscle lost after a myocardial infarction is replaced with noncontractile scar tissue, often initiating heart failure. Whole-organ cardiac transplantation is the only currently available clinical means of replacing the lost muscle, but this option is limited by the inadequate supply of donor hearts. Thus, cell-based cardiac repair has attracted considerable interest as an alternative means of ameliorating cardiac injury. Because of their tremendous capacity for expansion and unquestioned cardiac potential, pluripotent human embryonic stem cells (hESCs) represent an attractive candidate cell source for obtaining cardiomyocytes and other useful mesenchymal cell types for such therapies. Human embryonic stem cell-derived cardiomyocytes exhibit a committed cardiac phenotype and robust proliferative capacity, and recent testing in rodent infarct models indicates that they can partially remuscularize injured hearts and improve contractile function. Although the latter successes give good reason for optimism, considerable challenges remain in the successful application of hESCs to cardiac repair, including the need for preparations of high cardiac purity, improved methods of delivery, and approaches to overcome immune rejection and other causes of graft cell death. This review will describe the phenotype of hESC-derived cardiomyocytes and preclinical experience with these cells and will consider strategies to overcoming the aforementioned challenges.

  19. Human norovirus culture in B cells

    PubMed Central

    Jones, Melissa K; Grau, Katrina R; Costantini, Veronica; Kolawole, Abimbola O; de Graaf, Miranda; Freiden, Pamela; Graves, Christina L; Koopmans, Marion; Wallet, Shannon M; Tibbetts, Scott A; Schultz-Cherry, Stacey; Wobus, Christiane E; Vinjé, Jan; Karst, Stephanie M

    2015-01-01

    Human noroviruses (HunoVs) are a leading cause of foodborne disease and severe childhood diarrhea, and they cause a majority of the gastroenteritis outbreaks worldwide. However, the development of effective and long-lasting HunoV vaccines and therapeutics has been greatly hindered by their uncultivability. We recently demonstrated that a HunoV replicates in human B cells, and that commensal bacteria serve as a cofactor for this infection. In this protocol, we provide detailed methods for culturing the GII.4-sydney HunoV strain directly in human B cells, and in a coculture system in which the virus must cross a confluent epithelial barrier to access underlying B cells. We also describe methods for bacterial stimulation of HunoV B cell infection and for measuring viral attachment to the surface of B cells. Finally, we highlight variables that contribute to the efficiency of viral replication in this system. Infection assays require 3 d and attachment assays require 3 h. analysis of infection or attachment samples, including rna extraction and rt-qpcr, requires ~6 h. PMID:26513671

  20. Phagocytosis of dying tumor cells by human peritoneal mesothelial cells.

    PubMed

    Wagner, Britta Janina; Lindau, Dennis; Ripper, Dagmar; Stierhof, York-Dieter; Glatzle, Jörg; Witte, Maria; Beck, Henning; Keppeler, Hildegard; Lauber, Kirsten; Rammensee, Hans-Georg; Königsrainer, Alfred

    2011-05-15

    Peritoneal carcinomatosis is an advanced form of metastatic disease characterized by cancer cell dissemination onto the peritoneum. It is commonly observed in ovarian and colorectal cancers and is associated with poor patient survival. Novel therapies consist of cytoreductive surgery in combination with intraperitoneal chemotherapy, aiming at tumor cell death induction. The resulting dying tumor cells are considered to be eliminated by professional as well as semi-professional phagocytes. In the present study, we have identified a hitherto unknown type of 'amateur' phagocyte in this environment: human peritoneal mesothelial cells (HMCs). We demonstrate that HMCs engulf corpses of dying ovarian and colorectal cancer cells, as well as other types of apoptotic cells. Flow cytometric, confocal and electron microscopical analyses revealed that HMCs ingest dying cell fragments in a dose- and time-dependent manner and the internalized material subsequently traffics into late phagolysosomes. Regarding the mechanisms of prey cell recognition, our results show that HMCs engulf apoptotic corpses in a serum-dependent and -independent fashion and quantitative real-time PCR (qRT-PCR) analyses revealed that diverse opsonin receptor systems orchestrating dying cell clearance are expressed in HMCs at high levels. Our data strongly suggest that HMCs contribute to dying cell removal in the peritoneum, and future studies will elucidate in what manner this influences tumor cell dissemination and the antitumor immune response.

  1. Harnessing Human Dendritic Cell Subsets for Medicine

    PubMed Central

    Ueno, Hideki; Schmitt, Nathalie; Klechevsky, Eynav; Pedroza-Gonzales, Alexander; Matsui, Toshimichi; Zurawski, Gerard; Oh, SangKon; Fay, Joseph; Pascual, Virginia; Banchereau, Jacques; Palucka, Karolina

    2010-01-01

    Summary Immunity results from a complex interplay between the antigen-nonspecific innate immune system and the antigen-specific adaptive immune system. The cells and molecules of the innate system employ non-clonal recognition receptors including lectins, Toll-like receptors, NOD-like receptors and helicases. B and T lymphocytes of the adaptive immune system employ clonal receptors recognizing antigens or their derived peptides in a highly specific manner. An essential link between innate and adaptive immunity is provided by dendritic cells (DCs). DCs can induce such contrasting states as immunity and tolerance. The recent years have brought a wealth of information on the biology of DCs revealing the complexity of this cell system. Indeed, DC plasticity and subsets are prominent determinants of the type and quality of elicited immune responses. Here we summarize our recent studies aimed at a better understanding of the DC system to unravel the pathophysiology of human diseases and design novel human vaccines. PMID:20193020

  2. Androgen receptor in human endothelial cells

    PubMed Central

    Torres-Estay, Verónica; Carreño, Daniela V; San Francisco, Ignacio F; Sotomayor, Paula; Godoy, Alejandro S; Smith, Gary J

    2015-01-01

    Androgen receptor (AR) is a ligand-inducible transcription factor, and a member of the steroid-thyroid-retinoid receptor superfamily, that mediates the biological effects of androgens in a wide range of physiological and pathological processes. AR expression was identified in vascular cells nearly 20 years ago, and recent research has shown that AR mediates a variety of actions of androgens in endothelial and vascular smooth muscle cells. In this mini-review, we review evidence indicating the importance of AR in human endothelial cell (HUVEC) homeostatic and pathogenic processes. Although a role for AR in the modulation of HUVEC biology is evident, the molecular mechanisms by which AR regulates HUVEC homeostasis and disease processes are not fully understood. Understanding these mechanisms could provide critical insights into the processes of pathogenesis of diseases ranging from cardiovascular disease to cancer that are major causes of human morbidity and mortality. PMID:25563353

  3. SWCNTs induced autophagic cell death in human bronchial epithelial cells.

    PubMed

    Park, Eun-Jung; Zahari, Nur Elida M; Lee, Eun-Woo; Song, Jaewhan; Lee, Jae-Hyeok; Cho, Myung-Haing; Kim, Jae-Ho

    2014-04-01

    Carbon nanotubes are being actively introduced in electronics, computer science, aerospace, and other industries. Thus, the urgent need for toxicological studies on CNTs is mounting. In this study, we investigated the alterations in cellular response with morphological changes induced by single-walled carbon nanotubes (SWCNTs) in BEAS-2B cells, a human bronchial epithelial cell line. At 24h after exposure, SWCNTs rapidly decreased ATP production and cell viability as well a slight increase in the number of cells in the subG1 and G1 phases. In addition, SWCNTs increased the expression of superoxide dismutase (SOD)-1, but not SOD-2, and the number of cells generating ROS. The concentration of Cu and Zn ions also increased in a dose-dependent manner in cells exposed to SWCNTs. SWCNTs significantly enhanced the release of nitric oxide, interleukin (IL)-6, and IL-8 and up-regulated the expression of chemokine- and cytokine-related genes. Furthermore, the levels of autophagy-related genes, especially the DRAM1 gene, and the autophagosome formation-related proteins, were clearly up-regulated together with an increase of autophagosome-like vacuoles. Based on these results, we suggest that SWCNTs induce autophagic cell death through mitochondrial dysfunction and cytosolic damage in human bronchial epithelial cells.

  4. Human errors and violations in computer and information security: the viewpoint of network administrators and security specialists.

    PubMed

    Kraemer, Sara; Carayon, Pascale

    2007-03-01

    This paper describes human errors and violations of end users and network administration in computer and information security. This information is summarized in a conceptual framework for examining the human and organizational factors contributing to computer and information security. This framework includes human error taxonomies to describe the work conditions that contribute adversely to computer and information security, i.e. to security vulnerabilities and breaches. The issue of human error and violation in computer and information security was explored through a series of 16 interviews with network administrators and security specialists. The interviews were audio taped, transcribed, and analyzed by coding specific themes in a node structure. The result is an expanded framework that classifies types of human error and identifies specific human and organizational factors that contribute to computer and information security. Network administrators tended to view errors created by end users as more intentional than unintentional, while errors created by network administrators as more unintentional than intentional. Organizational factors, such as communication, security culture, policy, and organizational structure, were the most frequently cited factors associated with computer and information security.

  5. Reporter gene expression in dendritic cells after gene gun administration of plasmid DNA.

    PubMed

    Watkins, Craig; Hopkins, John; Harkiss, Gordon

    2005-07-21

    Dendritic cells (DC) play an integral role in plasmid DNA vaccination. However, the interaction between plasmid DNA and DC in vivo is incompletely understood. In this report, we utilise the sheep pseudoafferent cannulation model to examine the interaction between plasmid DNA encoding enhanced green fluorescent protein (pEGFP) and afferent lymph DC (ALDC) following gene gun administration. The results show that peaks of fluorescent ALDC tended to appear around days 1-4 and 9-13, then erratically thereafter for up to 2 months. Phenotypic analysis showed that EGFP+ ALDC expressed MHC class II, WC6, CD1b, and SIRPalpha markers. Plasmid, detected by PCR, was found in lymph cells and cell-free plasma on a daily basis, and was present variably for up to 2 months. Plasmid was also detected in purified CD1b+ ALDC, but the presence of plasmid did not correlate with EGFP expression by ALDC. Free EGFP in afferent lymph plasma was detectable by luminometry only after three administrations of the plasmid. The results show that gene gun administered pEGFP persisted for extended periods after a single administration, leeching out of skin on a daily basis. The plasmid was associated with both the cellular and fluid components of afferent lymph. EGFP protein appeared in afferent lymph in a pulsatile manner, but associated only with ALDC.

  6. Bone marrow hypoplasia and intestinal crypt cell necrosis associated with fenbendazole administration in five painted storks.

    PubMed

    Weber, Martha A; Terrell, Scott P; Neiffer, Donald L; Miller, Michele A; Mangold, Barbara J

    2002-08-01

    Five painted storks were treated with fenbendazole for 5 days for internal parasitism. Four birds died following treatment. Profound heteropenia was a consistent finding in all samples evaluated; additionally, the 1 surviving bird had progressive anemia. Consistent necropsy findings in the 4 birds that died were small intestinal crypt cell necrosis and severe bone marrow depletion and necrosis. Fenbendazole has been associated with bone marrow hypoplasia and enteric damage in mammals and other species of birds. The dosages of fenbendazole used in birds are often substantially higher than those recommended for mammals, which may contribute to bone marrow hypoplasia and intestinal crypt cell necrosis associated with fenbendazole administration in birds.

  7. United States Food and Drug Administration Regulation of Gene and Cell Therapies.

    PubMed

    Bailey, Alexander M; Arcidiacono, Judith; Benton, Kimberly A; Taraporewala, Zenobia; Winitsky, Steve

    2015-01-01

    The United States (US) Food and Drug Administration (FDA) is a regulatory agency that has oversight for a wide range of products entering the US market, including gene and cell therapies. The regulatory approach for these products is similar to other medical products within the United States and consists of a multitiered framework of statutes, regulations, and guidance documents. Within this framework, there is considerable flexibility which is necessary due to the biological and technical complexity of these products in general. This chapter provides an overview of the US FDA regulatory oversight of gene and cell therapy products.

  8. Calprotectin induces cell death in human prostate cancer cell (LNCaP) through survivin protein alteration.

    PubMed

    Sattari, Mina; Pazhang, Yaghub; Imani, Mehdi

    2014-11-01

    Calprotectin (CP), an abundant heterodimeric cytosolic protein of neutrophils, conveys a variety of functions such as tumor cell growth arrest and antimicrobial activity. We investigated CP activity and its possible apoptosis-inducing mechanism of action against an antiandrogen therapy-resistance prostate cancer cell line LNCaP. Cell viability and Annexin V FITC assays were performed in order to investigate its cell death activity and apoptosis, respectively. In order to address cell death inducing mechanism(s), immunocytochemistry and immunobloting analysis, reactive oxygen species (ROS) and nitric oxide (NO) measurements were performed. The effective concentration of CP against LNCaP promoting LNCaP cell death was 200 µg/mL. ROS and NO levels of cells remarkably were enhanced following treatment with 50 and 100 µg/mL of CP, respectively. Protein expression of anti-apoptotic protein survivin was significantly decreased after administration of tumor cells with CP. Our data indicate that CP regulates the LNCaP cells viability via survivin-mediated pathway and ROS and NO enhancement. Thus, inhibition of survivin expression, enhancement of ROS and NO level by CP or other similar pharmaceutical agents might be effective in lowering the malignant proliferation of human prostate cancer cells.

  9. Henipavirus pathogenesis in human respiratory epithelial cells.

    PubMed

    Escaffre, Olivier; Borisevich, Viktoriya; Carmical, J Russ; Prusak, Deborah; Prescott, Joseph; Feldmann, Heinz; Rockx, Barry

    2013-03-01

    Hendra virus (HeV) and Nipah virus (NiV) are deadly zoonotic viruses for which no vaccines or therapeutics are licensed for human use. Henipavirus infection causes severe respiratory illness and encephalitis. Although the exact route of transmission in human is unknown, epidemiological studies and in vivo studies suggest that the respiratory tract is important for virus replication. However, the target cells in the respiratory tract are unknown, as are the mechanisms by which henipaviruses can cause disease. In this study, we characterized henipavirus pathogenesis using primary cells derived from the human respiratory tract. The growth kinetics of NiV-Malaysia, NiV-Bangladesh, and HeV were determined in bronchial/tracheal epithelial cells (NHBE) and small airway epithelial cells (SAEC). In addition, host responses to infection were assessed by gene expression analysis and immunoassays. Viruses replicated efficiently in both cell types and induced large syncytia. The host response to henipavirus infection in NHBE and SAEC highlighted a difference in the inflammatory response between HeV and NiV strains as well as intrinsic differences in the ability to mount an inflammatory response between NHBE and SAEC. These responses were highest during HeV infection in SAEC, as characterized by the levels of key cytokines (interleukin 6 [IL-6], IL-8, IL-1α, monocyte chemoattractant protein 1 [MCP-1], and colony-stimulating factors) responsible for immune cell recruitment. Finally, we identified virus strain-dependent variability in type I interferon antagonism in NHBE and SAEC: NiV-Malaysia counteracted this pathway more efficiently than NiV-Bangladesh and HeV. These results provide crucial new information in the understanding of henipavirus pathogenesis in the human respiratory tract at an early stage of infection.

  10. Clinical potentials of human pluripotent stem cells.

    PubMed

    Mora, Cristina; Serzanti, Marialaura; Consiglio, Antonella; Memo, Maurizio; Dell'Era, Patrizia

    2017-02-08

    Aging, injuries, and diseases can be considered as the result of malfunctioning or damaged cells. Regenerative medicine aims to restore tissue homeostasis by repairing or replacing cells, tissues, or damaged organs, by linking and combining different disciplines including engineering, technology, biology, and medicine. To pursue these goals, the discipline is taking advantage of pluripotent stem cells (PSCs), a peculiar type of cell possessing the ability to differentiate into every cell type of the body. Human PSCs can be isolated from the blastocysts and maintained in culture indefinitely, giving rise to the so-called embryonic stem cells (ESCs). However, since 2006, it is possible to restore in an adult cell a pluripotent ESC-like condition by forcing the expression of four transcription factors with the rejuvenating reprogramming technology invented by Yamanaka. Then the two types of PSC can be differentiated, using standardized protocols, towards the cell type necessary for the regeneration. Although the use of these derivatives for therapeutic transplantation is still in the preliminary phase of safety and efficacy studies, a lot of efforts are presently taking place to discover the biological mechanisms underlying genetic pathologies, by differentiating induced PSCs derived from patients, and new therapies by challenging PSC-derived cells in drug screening.

  11. Central Administration of Galanin Receptor 1 Agonist Boosted Insulin Sensitivity in Adipose Cells of Diabetic Rats

    PubMed Central

    Zhang, Zhenwen; Fang, Penghua; He, Biao; Guo, Lili; Runesson, Johan; Langel, Ülo; Shi, Mingyi; Zhu, Yan; Bo, Ping

    2016-01-01

    Our previous studies testified the beneficial effect of central galanin on insulin sensitivity of type 2 diabetic rats. The aim of the study was further to investigate whether central M617, a galanin receptor 1 agonist, can benefit insulin sensitivity. The effects of intracerebroventricular administration of M617 on insulin sensitivity and insulin signaling were evaluated in adipose tissues of type 2 diabetic rats. The results showed that central injection of M617 significantly increased plasma adiponectin contents, glucose infusion rates in hyperinsulinemic-euglycemic clamp tests, GLUT4 mRNA expression levels, GLUT4 contents in plasma membranes, and total cell membranes of the adipose cells but reduced the plasma C-reactive protein concentration in nondiabetic and diabetic rats. The ratios of GLUT4 contents were higher in plasma membranes to total cell membranes in both nondiabetic and diabetic M617 groups than each control. In addition, the central administration of M617 enhanced the ratios of pAkt/Akt and pAS160/AS160, but not phosphorylative cAMP response element-binding protein (pCREB)/CREB in the adipose cells of nondiabetic and diabetic rats. These results suggest that excitation of central galanin receptor 1 facilitates insulin sensitivity via activation of the Akt/AS160 signaling pathway in the fat cells of type 2 diabetic rats. PMID:27127795

  12. 77 FR 5489 - Identification of Human Cell Lines Project

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-03

    ... National Institute of Standards and Technology Identification of Human Cell Lines Project AGENCY: National... tandem repeat (STR) profiling up to 1500 human cell line samples as part of the Identification of Human... its intent to unambiguously identify by short tandem repeat (STR) profiling up to 1500 human cell...

  13. Safety of administration of human butyrylcholinesterase and its conjugates with soman or VX in rats.

    PubMed

    Genovese, Raymond F; Sun, Wei; Johnson, Christina C; Ditargiani, Robert C; Doctor, Bhupendra P; Saxena, Ashima

    2010-05-01

    We evaluated the effects of conjugated enzyme-nerve agent product resulting from the inhibition of bioscavenger human serum butyrylcholinesterase (Hu BChE) by nerve agents soman or VX. Rats were trained on a multiple Fixed-Ratio 32, Extinction 30 sec. (FR32, Ext30) schedule of food reinforcement and then injected (i.m.) with Hu BChE (30 mg/kg), equivalent amounts of Hu BChE-soman conjugate (GDC), Hu BChE-VX conjugate, oxotremorine (OXO) (0.316 mg/kg) or vehicle (n = 8, each group). On the day of injection and on 10 subsequent daily sessions, performance was evaluated on the FR32, Ext30 schedule. Neither conjugates nor Hu BChE produced a performance deficit under the schedule. OXO produced a substantial decrease in responding on the day of administration, with complete recovery observed on subsequent sessions. None of the treatments affected circulating acetylcholinesterase (AChE) activity when evaluated 24-72 hr after injection. The dose of Hu BChE produced a 20,000-fold increase above baseline in circulating BChE activity. Pathological evaluation of organ systems approximately 2 weeks following administration of conjugates or Hu BChE alone did not show toxicity. Taken together, these results suggest that Hu BChE - nerve agent conjugates produced following bioscavenger protection against nerve agents soman and VX do not appear to be particularly toxic. These results add to the safety assessment of Hu BChE as a bioscavenger countermeasure against nerve agent exposure.

  14. Incorporation of methamphetamine and amphetamine in human hair following controlled oral methamphetamine administration

    PubMed Central

    Polettini, Aldo; Cone, Edward J.; Gorelick, David A.; Huestis, Marilyn A.

    2012-01-01

    Background Although hair testing is well established for the assessment of past drug exposure, uncertainties persist about mechanisms of drug incorporation into hair and interpretation of results. The aim of this study was to administer methamphetamine (MAMP) under controlled conditions as a model drug to investigate drug incorporation into human hair. Material and Methods Seven volunteers with a history of stimulant use received 4×10 mg (low) doses of sustained release S-(+)-MAMP HCl within one week, with weekly head hair samples collected by shaving. 3 weeks later, 4 of them received 4×20 mg (high) doses. After extensive isopropanol/phosphate buffer washing of the hair, MAMP and its metabolite amphetamine (AMP) concentrations were determined in all weekly hair samples by LC-MS-MS in selected reaction monitoring mode with the undeca- and deca-deuterated drugs, respectively, as internal standards (LLOQ, 0.005 ng/mg). Results MAMP Tmax occurred from 1 to 2 weeks after both doses, with Cmax ranging from 0.6–3.5 ng/mg after the low and 1.2–5.3 ng/mg after the high MAMP doses. AMP Cmax in hair was 0.1–0.3 ng/mg and 0.2–0.5 ng/mg, respectively, for low and high doses. Highly dose–related concentrations within subjects, but large variability between subjects were observed. MAMP concentrations were above the 0.2 ng/mg cutoff for at least two weeks following administration of both low and high doses. The overall AMP/MAMP ratio ranged from 0.07 to 0.37 with a mean value of 0.15±0.07, and a median of 0.13. The percentage of MAMP and AMP removed with the washing procedure decreased with time after administration. A strong correlation was found between area under the curve of MAMP (r2=0.90, p=0.00) and AMP (r2=0.94, p=0.00) concentrations calculated for the 3-week period following administration and the total melanin concentration in hair. Significant correlations were observed also between Cmax and melanin. Conclusions This study demonstrated that despite large

  15. Effects of 3-methylcholanthrene and aspirin co-administration on ALDH3A1 in HepG2 cells.

    PubMed

    Sotiropoulou, M; Pappas, P; Marselos, M

    2001-01-30

    The effects of two different protocols of 3-methylcholanthrene (3MC) and aspirin co-administration were studied in a well-established human hepatoma cell line (HepG2). During this work, we have performed toxicity tests for cell viability/cell proliferation as well as studies on the expression of ALDH3A1 after exposure of HepG2 cells to 3MC or/and aspirin. For the evaluation of toxic concentrations of 3MC and aspirin, the WST-1 test was used. WST-1 is a reliable cytotoxicity test which is based on the cleavage of the tetrazolium salt WST-1 to formazan by mitochondrial enzymes of living cells. A broad range of drug concentrations for either 3MC (0.25-50.0 microM) or aspirin (0.05-10.0 mM) were used for cell exposure, in several periods of time. The expression of ALDH3A1 in HepG2 cells showed typical time- and dose-response curves of induction after application of 3MC (1-5 days, 1.5-5.0 microM, respectively). When cells were firstly exposed to 3MC (2.5 and 5.0 microM) and then to aspirin (0.25 mM), the induced ALDH3A1 activity was further enhanced in a statistically significant way (P<0.05). On the contrary, when aspirin application was preceded 3MC exposuring a statistically significant decrease in ALDH3A1 inducibility was observed, as compared with the application of 3MC alone.

  16. Interdigitating reticulum cells in human renal grafts.

    PubMed

    Wakabayashi, T; Onoda, H

    1991-01-01

    Seventeen human renal graft biopsies taken 1 h to 50 days after transplantation and 3 human renal non-graft biopsies (2 minimal change and 1 non-tumour portion of angiomyolipoma) were investigated with immunoelectron microscopy in order to identify interdigitating reticulum cells (IDC) or dendritic cells (DC) in renal tissues. The antibodies used consisted of a rabbit polyclonal antibody of antihuman S100 beta protein, mouse monoclonal antibodies of antihuman HLA-DR, anti-CD3, and anti-CD1a. IDC or DC were identified in 11 renal grafts. They were found both in the glomerular and interstitial (peritubular) capillary lumens but not in the interstitium of 1 case: both were present in the interstitial capillary lumens and interstitium of another case, and in the interstitium only of 9 cases. In the remaining 6 grafts and 3 non-grafts they were not detected. These 6 grafts and 3 non-grafts did not show any pathological change except for foot process fusion of the glomerular epithelia in 2 cases of minimal change. These findings suggest that IDC or DC are not normally present in human renal tissues. The presence of the cell in the glomerular and peritubular capillary lumens of a biopsy taken after 1 h and their presence in the interstitial capillary lumens of another graft biopsy, suggest that the IDC or DC in human renal grafts are derived from recipients, not donors, and that they migrate from the circulating blood toward the interstitium.

  17. DNA repair responses in human skin cells

    SciTech Connect

    Hanawalt, P.C.; Liu, S.C.; Parsons, C.S.

    1981-07-01

    Sunlight and some environmental chemical agents produce lesions in the DNA of human skin cells that if unrepaired may interfere with normal functioning of these cells. The most serious outcome of such interactions may be malignancy. It is therefore important to develop an understanding of mechanisms by which the lesions may be repaired or tolerated without deleterious consequences. Our models for the molecular processing of damaged DNA have been derived largely from the study of bacterial systems. Some similarities but significant differences are revealed when human cell responses are tested against these models. It is also of importance to learn DNA repair responses of epidermal keratinocytes for comparison with the more extensive studies that have been carried out with dermal fibroblasts. Our experimental results thus far indicate similarities for the excision-repair of ultraviolet-induced pyrimidine dimers in human keratinocytes and fibroblasts. Both the monoadducts and the interstrand crosslinks produced in DNA by photoactivated 8-methoxypsoralen (PUVA) can be repaired in normal human fibroblasts but not in those from xeroderma pigmentosum patients. The monoadducts, like pyrimidine dimers, are probably the more mutagenic/carcinogenic lesions while the crosslinks are less easily repaired and probably result in more effective blocking of DNA function. It is suggested that a split-dose protocol that maximizes the production of crosslinks while minimizing the yield of monoadducts may be more effective and potentially less carcinogenic than the single ultraviolet exposure regimen in PUVA therapy for psoriasis.

  18. Chronic hypothyroidism only marginally affects adult-type Leydig cell regeneration after EDS administration.

    PubMed

    Rijntjes, Eddy; van Kesteren-Buiting, Anita; Keijer, Jaap; Teerds, Katja J

    2010-02-01

    Chronic prenatally induced dietary hypothyroidism delays adult-type Leydig cell development, but does not block this process. Using a chemical model to induce hypothyroidism, it was suggested that development of a new population of Leydig cells was completely inhibited following the addition of the cytotoxic compound ethane-1,2-dimethyl sulphonate (EDS). In this study, we used a dietary approach to induce hypothyroidism and reinvestigated the regeneration of the Leydig cell population following EDS administration. Eighty-four day old euthyroid and chronically hypothyroid rats received an injection of EDS and were killed directly before or at regular intervals up to 77 days after EDS. In some control and hypothyroid animals, the first progenitor-type Leydig cells were observed at day 12 after EDS. At day 16, Leydig cell progenitors were present in all rats. The percentage of proliferating Leydig cells peaked in the euthyroid animals at day 21 after EDS. In the hypothyroid testis such a peak was not observed, although the percentage of proliferating regenerating Leydig cells was significantly higher from days 35 to 56 compared with the controls. This suggested that the wave of Leydig cell proliferation was delayed in the hypothyroid animals as compared with the euthyroid controls. On the day of EDS injection, the Leydig/Sertoli cell ratio was 37% lower in the hypothyroid rats compared with the controls. The Leydig/Sertoli cell ratio remained lower in the EDS-treated hypothyroid animals compared with the controls at all time points investigated. At day 77 after EDS, the Leydig cell population had returned to its pre-treatment size in both groups. Plasma testosterone production was reduced to below detectable levels immediately after EDS injection, and started to increase again on day 16, reaching pre-treatment values on day 21 in both groups. Taken together, severely reduced thyroid hormone levels did not block the regeneration of the adult-type Leydig cell population

  19. Social Work Values in Human Services Administration: Implications for Social Work Education

    ERIC Educational Resources Information Center

    Watson, Larry D.; Hoefer, Richard

    2014-01-01

    The perceived wisdom in the social work education community, based on empirical research from the 1990s and the early part of this century, says that the master of social work (MSW) degree is not competitive with the master of business administration or the master of public administration to obtain top-level administration jobs in nonprofit…

  20. Human T Cell Memory: A Dynamic View

    PubMed Central

    Macallan, Derek C.; Borghans, José A. M.; Asquith, Becca

    2017-01-01

    Long-term T cell-mediated protection depends upon the formation of a pool of memory cells to protect against future pathogen challenge. In this review we argue that looking at T cell memory from a dynamic viewpoint can help in understanding how memory populations are maintained following pathogen exposure or vaccination. For example, a dynamic view resolves the apparent paradox between the relatively short lifespans of individual memory cells and very long-lived immunological memory by focussing on the persistence of clonal populations, rather than individual cells. Clonal survival is achieved by balancing proliferation, death and differentiation rates within and between identifiable phenotypic pools; such pools correspond broadly to sequential stages in the linear differentiation pathway. Each pool has its own characteristic kinetics, but only when considered as a population; single cells exhibit considerable heterogeneity. In humans, we tend to concentrate on circulating cells, but memory T cells in non-lymphoid tissues and bone marrow are increasingly recognised as critical for immune defence; their kinetics, however, remain largely unexplored. Considering vaccination from this viewpoint shifts the focus from the size of the primary response to the survival of the clone and enables identification of critical system pinch-points and opportunities to improve vaccine efficacy. PMID:28165397

  1. Human T Cell Memory: A Dynamic View.

    PubMed

    Macallan, Derek C; Borghans, José A M; Asquith, Becca

    2017-02-04

    Long-term T cell-mediated protection depends upon the formation of a pool of memory cells to protect against future pathogen challenge. In this review we argue that looking at T cell memory from a dynamic viewpoint can help in understanding how memory populations are maintained following pathogen exposure or vaccination. For example, a dynamic view resolves the apparent paradox between the relatively short lifespans of individual memory cells and very long-lived immunological memory by focussing on the persistence of clonal populations, rather than individual cells. Clonal survival is achieved by balancing proliferation, death and differentiation rates within and between identifiable phenotypic pools; such pools correspond broadly to sequential stages in the linear differentiation pathway. Each pool has its own characteristic kinetics, but only when considered as a population; single cells exhibit considerable heterogeneity. In humans, we tend to concentrate on circulating cells, but memory T cells in non-lymphoid tissues and bone marrow are increasingly recognised as critical for immune defence; their kinetics, however, remain largely unexplored. Considering vaccination from this viewpoint shifts the focus from the size of the primary response to the survival of the clone and enables identification of critical system pinch-points and opportunities to improve vaccine efficacy.

  2. HUMAN VASCULAR ENDOTHELIAL CELLS IN CULTURE

    PubMed Central

    Gimbrone, Michael A.; Cotran, Ramzi S.; Folkman, Judah

    1974-01-01

    Human endothelial cells, obtained by collagenase treatment of term umbilical cord veins, were cultured using Medium 199 supplemented with 20% fetal calf serum. Small clusters of cells initially spread on plastic or glass, coalesced and grew to form confluent monolayers of polygonal cells by 7 days. Cells in primary and subcultures were identified as endothelium by the presence of Weibel-Palade bodies by electron microscopy. A morphologically distinct subpopulation of cells contaminating some primary endothelial cultures was selectively subcultured, and identified by ultrastructural criteria as vascular smooth muscle. Autoradiography of endothelial cells after exposure to [3H]thymidine showed progressive increases in labeling in growing cultures beginning at 24 h. In recently confluent cultures, labeling indices were 2.4% in central closely packed regions, and 53.2% in peripheral growing regions. 3 days after confluence, labeling was uniform, being 3.5 and 3.9% in central and peripheral areas, respectively. When small areas of confluent cultures were experimentally "denuded," there were localized increases in [3H]thymidine labeling and eventual reconstitution of the monolayer. Liquid scintillation measurements of [3H]thymidine incorporation in primary and secondary endothelial cultures in microwell trays showed a similar correlation of DNA synthesis with cell density. These data indicate that endothelial cell cultures may provide a useful in vitro model for studying pathophysiologic factors in endothelial regeneration. PMID:4363161

  3. Transport and Biodistribution of Dendrimers Across Human Fetal Membranes: Implications for Intravaginal Administration of Dendrimers

    PubMed Central

    Menjoge, Anupa R.; Navath, Raghavendra S.; Asad, Abbas; Kannan, Sujatha; Kim, Chong Jai; Romero, Roberto; Kannan, Rangaramanujam M.

    2010-01-01

    Dendrimers are emerging as promising topical antimicrobial agents, and as targeted nanoscale drug delivery vehicles. Topical intravaginal antimicrobial agents are prescribed to treat the ascending genital infections in pregnant women. The fetal membranes separate the extra-amniotic space and fetus. The purpose of the study is to determine if the dendrimers can be selectively used for local intravaginal application to pregnant women without crossing the membranes into the fetus. In the present study, the transport and permeability of PAMAM (poly(amidoamine)) dendrimers, across human fetal membrane (using a side-by-side diffusion chamber), and its biodistribution (using immunofluorescence) are evaluated ex-vivo. Transport across human fetal membranes (from the maternal side) was evaluated using Fluorescein (FITC), an established transplacental marker (positive control, size~ 400 Da) and fluorophore-tagged G4-PAMAM dendrimers (~ 16 kDa). The fluorophore-tagged G4-PAMAM dendrimers were synthesized and characterized using 1H NMR, MALDI TOF-MS and HPLC analysis. Transfer was measured across the intact fetal membrane (chorioamnion), and the separated chorion and amnion layers. Over a five hour period, the dendrimer transport across all the three membranes was less than < 3 %, whereas the transport of FITC was relatively fast with as much as 49% transport across the amnion. The permeability of FITC (7.9 × 10-7 cm2/s) through the chorioamnion was 7-fold higher than that of the dendrimer (5.8 × 10-8 cm2/s). The biodistribution showed that the dendrimers were largely present in interstitial spaces in the decidual stromal cells and the chorionic trophoblast cells (in 2.5 to 4 h) and surprisingly, to a smaller extent internalized in nuclei of trophoblast cells and nuclei and cytoplasm of stromal cells. Passive diffusion and paracellular transport appear to be the major route for dendrimer transport. The overall findings further suggest that entry of drugs conjugated to

  4. Human fetal liver stromal cells expressing erythropoietin promote hematopoietic development from human embryonic stem cells.

    PubMed

    Yang, Chao; Ji, Lei; Yue, Wen; Shi, Shuang-Shuang; Wang, Ruo-Yong; Li, Yan-Hua; Xie, Xiao-Yan; Xi, Jia-Fei; He, Li-Juan; Nan, Xue; Pei, Xue-Tao

    2012-02-01

    Blood cells transfusion and hematopoietic stem cells (HSCs) transplantation are important methods for cell therapy. They are widely used in the treatment of incurable hematological disorder, infectious diseases, genetic diseases, and immunologic deficiency. However, their availability is limited by quantity, capacity of proliferation and the risk of blood transfusion complications. Recently, human embryonic stem cells (hESCs) have been shown to be an alternative resource for the generation of hematopoietic cells. In the current study, we describe a novel method for the efficient production of hematopoietic cells from hESCs. The stable human fetal liver stromal cell lines (hFLSCs) expressing erythropoietin (EPO) were established using the lentiviral system. We observed that the supernatant from the EPO transfected hFLSCs could induce the hESCs differentiation into hematopoietic cells, especially erythroid cells. They not only expressed fetal and embryonic globins but also expressed the adult-globin chain on further maturation. In addition, these hESCs-derived erythroid cells possess oxygen-transporting capacity, which indicated hESCs could generate terminally mature progenies. This should be useful for ultimately developing an animal-free culture system to generate large numbers of erythroid cells from hESCs and provide an experimental model to study early human erythropoiesis.

  5. Repeated administration of an acetylcholinesterase inhibitor attenuates nicotine taking in rats and smoking behavior in human smokers.

    PubMed

    Ashare, R L; Kimmey, B A; Rupprecht, L E; Bowers, M E; Hayes, M R; Schmidt, H D

    2016-01-19

    Tobacco smoking remains the leading cause of preventable death worldwide and current smoking cessation medications have limited efficacy. Thus, there is a clear need for translational research focused on identifying novel pharmacotherapies for nicotine addiction. Our previous studies demonstrated that acute administration of an acetylcholinesterase inhibitor (AChEI) attenuates nicotine taking and seeking in rats and suggest that AChEIs could be repurposed for smoking cessation. Here, we expand upon these findings with experiments designed to determine the effects of repeated AChEI administration on voluntary nicotine taking in rats as well as smoking behavior in human smokers. Rats were trained to self-administer intravenous infusions of nicotine (0.03 mg kg(-1) per 0.59 ml) on a fixed-ratio-5 schedule of reinforcement. Once rats maintained stable nicotine taking, galantamine or donepezil was administered before 10 consecutive daily nicotine self-administration sessions. Repeated administration of 5.0 mg kg(-1) galantamine and 3.0 mg kg(-1) donepezil attenuated nicotine self-administration in rats. These effects were reinforcer-specific and not due to adverse malaise-like effects of drug treatment as repeated galantamine and donepezil administration had no effects on sucrose self-administration, ad libitum food intake and pica. The effects of repeated galantamine (versus placebo) on cigarette smoking were also tested in human treatment-seeking smokers. Two weeks of daily galantamine treatment (8.0 mg (week 1) and 16.0 mg (week 2)) significantly reduced smoking rate as well as smoking satisfaction and reward compared with placebo. This translational study indicates that repeated AChEI administration reduces nicotine reinforcement in rats and smoking behavior in humans at doses not associated with tolerance and/or adverse effects.

  6. Cell phoney: human cloning after Quintavalle.

    PubMed

    Morgan, Derek; Ford, Mary

    2004-12-01

    Reproductive cloning has thrown up new scientific possibilities, ethical conundrums, and legal challenges. An initial question, considered by the English courts in 2003, was whether the technique presently available, that of cell nucleus replacement, falls outside the provisions of the Human Fertilisation and Embryology Act 1990. If it does, the creation and use, including use in research protocols, of human embryos would be unregulated, disclosing a need to consider remedial legislation. The resolution by the courts of this legal question dramatically engages them in a resolution of fundamental ethical dilemmas, and discloses the possibilities and limitation of negotiating science policy through the processes of litigation.

  7. Characterization of Human Mammary Epithelial Stem Cells

    DTIC Science & Technology

    2009-10-01

    Appendix……………………………………………………………………………… 11 Eirew,P., Stingl,J., Raouf,A., Turashvili,G., Aparicio ,S., Emerman,J.T., and Eaves,C.J. A method for... Aparicio , Joanne Emerman and Connie Eaves. A method for quantifying normal human mammary epithelial stem cells with in vivo regenerative ability...Abstracts Peter Eirew, John Stingl, Afshin Raouf, Gulisa Turshvili, Sam Aparicio , Joanne Emerman and Connie Eaves, “Identification of Human Mammary

  8. Effects of nicotine administration on elemental concentrations in mouse granulosa cells, maturing oocytes and oviduct epithelium studied by X-ray microanalysis.

    PubMed

    Jin, Z; Jin, M; Nilsson, B O; Roomans, G M

    1998-10-01

    A normal maturation of the oocytes is dependent upon, among other things, normally functioning granulosa and corona radiata cells. Analyses performed during human in vitro fertilization programs have revealed that, in smokers, ovarian functions are affected and that smokers have a decreased fertilization rate. Further, animal studies have indicated that nicotine can reach the genital tractus, and that nicotine administration interferes with oocyte maturation, fertilization and early pregnancy. We applied X-ray microanalysis to monitor whether nicotine administration changed the ionic balance of cells in the reproductive tract (granulosa cells, oocytes and oviduct epithelial cells). The animals were given nicotine in the drinking water at a concentration of 108 mumol/l. After 15 days the animals were superovulated, ovaries and oviducts were frozen, and thick cryosections were prepared for energy-dispersive X-ray microanalysis. In the granulosa cells, the concentrations of Na and Cl increased after nicotine treatment, while the K concentrations decreased resulting in an increased Na/K ratio. The treated oocytes had a higher K concentration and a decreased Na/K ratio compared to the controls. In the epithelial cells of the oviduct, the concentrations of Na and K decreased after nicotine treatment without any changes in the Na/K ratio. Thus, heavy nicotine administration to mice causes significant changes in the ionic composition of the granulosa cells, the ovarian oocytes and the oviduct epithelium.

  9. 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

  10. Radioactive excretion in human milk following administration of /sup 99m/Tc macroaggregated albumin

    SciTech Connect

    Pittard, W.B.; Merkatz, R.; Fletcher, B.D.

    1982-08-01

    Albumin-tagged sodium pertechnetate (technetium) is routinely used in nuclear medicine for scanning procedures of the lung. The rate of excretion of this radionuclide into breast milk and the resultant potential radiation hazard to the nursing infant have received little attention. Therefore the milk from a nursing mother who required a lung scan because of suspected pulmonary emboli using an intravenous injection of 4 mCi of /sup 99m/Tc macroaggregated human serum albumin was monitored. Albumin tagging severely limited the entrance of technetium into her milk and the radioactivity of the milk returned to base line by 24 hours. A total of 2.02 muCi of technetium was measured in the 24-hour milk collection after technetium injection and 94% of this amount was excreted by 15.5 hours. This amount of technetium administered orally to a newborn would deliver a total body radiation dose of .3 mrad. Therefore, an infant would receive trivial doses of radiation if breast-feeding were resumed 15.5 hours after administration of the radionuclide to the mother and nursing can clearly be resumed safely 24 hours after injection.

  11. Tissue Distribution of trans-Resveratrol and Its Metabolites after Oral Administration in Human Eyes

    PubMed Central

    Weinreb, Robert N.

    2017-01-01

    Purpose. This study was performed to measure the concentration of trans-resveratrol and its three metabolites in human eyes. Methods. The patients who underwent pars plana vitrectomy for rhegmatogenous retinal detachment were included. The participants were orally given trans-resveratrol-based supplement (Longevinex®). A suitable amount of conjunctiva, aqueous humor, and vitreous humor were obtained during the operation. High-performance liquid chromatography (HPLC) with mass spectrometry (LC/MS/MS) was used to detect the concentration of trans-resveratrol and its three metabolites in the various samples. Results. The average concentration of resveratrol in the conjunctiva was 17.19 ± 15.32 nmol/g (mean ± SD). The concentration of resveratrol in the aqueous humor was close to the limit of detection, but its metabolites could be quantified. The concentrations of resveratrol metabolites in the aqueous humor can be detected. In the vitreous humor, the average concentration of resveratrol-3-O-sulfate was 62.95 ± 41.97 nmol/L. The sulfate conjugations of resveratrol were recovered in the conjunctiva, aqueous humor, and vitreous humor. Conclusions. Resveratrol and its three metabolites can be detected in the ocular tissues after oral administration. Although the concentration of parent resveratrol was low in the eyes, its metabolites could be detected and may have a role in the treatment of ocular diseases.

  12. Osmotic water permeability of human red cells

    PubMed Central

    1981-01-01

    The osmotic water permeability of human red cells has been reexamined with a stopped-flow device and a new perturbation technique. Small osmotic gradients are used to minimize the systematic error caused by nonlinearities in the relationship between cell volume and light scattering. Corrections are then made for residual systematic error. Our results show that the hydraulic conductivity, Lp, is essentially independent of the direction of water flow and of osmolality in the range 184-365 mosM. the mean value of Lp obtained obtained was 1.8 +/- 0.1 (SEM) X 10-11 cm3 dyne -1 s-1. PMID:7229611

  13. Lymphoid Cell-Glioma Cell Interaction Enhances Cell Coat Production by Human Gliomas: Novel Suppressor Mechanism

    NASA Astrophysics Data System (ADS)

    Dick, Steven J.; Macchi, Beatrice; Papazoglou, Savvas; Oldfield, Edward H.; Kornblith, Paul L.; Smith, Barry H.; Gately, Maurice K.

    1983-05-01

    Certain human glioma lines produce mucopolysaccharide coats that impair the generation of cytolytic lymphocytes in response to these lines in vitro. Coat production is substantially enhanced by the interaction of glioma cells with a macromolecular factor released by human peripheral blood mononuclear cells in culture. This interaction thus constitutes an unusual mechanism by which inflammatory cells may nonspecifically suppress the cellular immune response to at least one class of solid tumors in humans.

  14. High prevalence of side population in human cancer cell lines

    PubMed Central

    Boesch, Maximilian; Zeimet, Alain G.; Fiegl, Heidi; Wolf, Barbara; Huber, Julia; Klocker, Helmut; Gastl, Guenther

    2016-01-01

    Cancer cell lines are essential platforms for performing cancer research on human cells. We here demonstrate that, across tumor entities, human cancer cell lines harbor minority populations of putative stem-like cells, molecularly defined by dye extrusion resulting in the side population phenotype. These findings establish a heterogeneous nature of human cancer cell lines and argue for their stem cell origin. This should be considered when interpreting research involving these model systems. PMID:27226981

  15. Human Olfactory Mucosa Multipotent Mesenchymal Stromal Cells Promote Survival, Proliferation, and Differentiation of Human Hematopoietic Cells

    PubMed Central

    Diaz-Solano, Dylana; Wittig, Olga; Ayala-Grosso, Carlos; Pieruzzini, Rosalinda

    2012-01-01

    Multipotent mesenchymal stromal cells (MSCs) from the human olfactory mucosa (OM) are cells that have been proposed as a niche for neural progenitors. OM-MSCs share phenotypic and functional properties with bone marrow (BM) MSCs, which constitute fundamental components of the hematopoietic niche. In this work, we investigated whether human OM-MSCs may promote the survival, proliferation, and differentiation of human hematopoietic stem cells (HSCs). For this purpose, human bone marrow cells (BMCs) were co-cultured with OM-MSCs in the absence of exogenous cytokines. At different intervals, nonadherent cells (NACs) were harvested from BMC/OM-MSC co-cultures, and examined for the expression of blood cell markers by flow cytometry. OM-MSCs supported the survival (cell viability >90%) and proliferation of BMCs, after 54 days of co-culture. At 20 days of co-culture, flow cytometric and microscopic analyses showed a high percentage (73%) of cells expressing the pan-leukocyte marker CD45, and the presence of cells of myeloid origin, including polymorphonuclear leukocytes, monocytes, basophils, eosinophils, erythroid cells, and megakaryocytes. Likewise, T (CD3), B (CD19), and NK (CD56/CD16) cells were detected in the NAC fraction. Colony-forming unit–granulocyte/macrophage (CFU-GM) progenitors and CD34+ cells were found, at 43 days of co-culture. Reverse transcriptase–polymerase chain reaction (RT-PCR) studies showed that OM-MSCs constitutively express early and late-acting hematopoietic cytokines (i.e., stem cell factor [SCF] and granulocyte- macrophage colony-stimulating factor [GM-CSF]). These results constitute the first evidence that OM-MSCs may provide an in vitro microenvironment for HSCs. The capacity of OM-MSCs to support the survival and differentiation of HSCs may be related with the capacity of OM-MSCs to produce hematopoietic cytokines. PMID:22471939

  16. Programming and reprogramming a human heart cell.

    PubMed

    Sahara, Makoto; Santoro, Federica; Chien, Kenneth R

    2015-03-12

    The latest discoveries and advanced knowledge in the fields of stem cell biology and developmental cardiology hold great promise for cardiac regenerative medicine, enabling researchers to design novel therapeutic tools and approaches to regenerate cardiac muscle for diseased hearts. However, progress in this arena has been hampered by a lack of reproducible and convincing evidence, which at best has yielded modest outcomes and is still far from clinical practice. To address current controversies and move cardiac regenerative therapeutics forward, it is crucial to gain a deeper understanding of the key cellular and molecular programs involved in human cardiogenesis and cardiac regeneration. In this review, we consider the fundamental principles that govern the "programming" and "reprogramming" of a human heart cell and discuss updated therapeutic strategies to regenerate a damaged heart.

  17. Chromium oxidation state mapping in human cells

    NASA Astrophysics Data System (ADS)

    Ortega, R.; Fayard, B.; Salomé, M.; Devès, G.; Susini, J.

    2003-03-01

    The widespread use of chromium in industrial applications such as chemical production of pigments, refractory brick production, tanning, metallurgy, electroplating, and combustion of fuels has lead to human occupational exposure and to its increased introduction into the environment. Hexavalent chromium compounds are established carcinogens but their mechanism of cell transformation is not known. Up to now, no microanalytical technique was sensitive enough to allow the observation of chromium distribution, and oxidation state identification, within isolated cells at carcinogenic concentrations. In this experiment, we used successfully the ID-21 X-ray microscope to map Cr(VI) and total Cr distributions in cells exposed in vitro to soluble, and insoluble, Cr(VI) compounds. Exposure to soluble compounds, weak carcinogens, resulted in a homogeneous intracellular distribution of Cr, confirming by in situ measurement that Cr is present in the cell nucleus. Cr(VI) was never detected in cells which suggests a mechanism of rapid intracellular reducticn. On the other hand, exposure to insoluble compounds, strong carcinogens, also resulted in a homogeneous distribution of reduced forms of Cr in cells, and their nucleus. However, in this case, Cr(VI)-rich structures were observed into the cells suggesting that carcinogenicity is enhanced when oxidation reactions due to Cr(VI) chronic exposure are associated to Cr-DNA alterations.

  18. Characterization of Human Mammary Epithelial Stem Cells

    DTIC Science & Technology

    2007-10-01

    robust and reproducible methodology to detect, quantify and isolate stem cells in normal human mammary tissue, using a xenotransplantation system...covers the first year of the grant, during which substantial progress has been made in the development and validation of the xenotransplantation assay...Subrenal xenotransplantation surgery. The hair on the back of anesthetized mice was shaved, and the skin swabbed with 70% alcohol. An anterior to

  19. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2011-10-11

    types have potential for being physiologically relevant in vitro models for botulinum toxin detection and neuron-glia interactions, respectively...unknown neurotoxicants. (5) We developed an immunoblot based method for detecting botulinum toxin using the mixed neuronal hN2™ cell line, thus creating...a first generation human cellular model for botulinum toxin detection – standard of comparison for existing and future models.neurotoxicity

  20. TALEN-Induced Translocations in Human Cells.

    PubMed

    Piganeau, Marion; Renouf, Benjamin; Ghezraoui, Hind; Brunet, Erika

    2016-01-01

    Induction of chromosomal translocations in human cells is of a great interest to study tumorigenesis and genome instability. Here, we explain in detail a method to induce translocations using the transcription activator-like effector nucleases (TALENs). We describe how to detect translocation formation by PCR, calculate translocation frequency by 96-well PCR screen, and analyze breakpoint junctions. When inducing cancer translocations, it is also possible to detect the fusion gene by FISH analysis or western blot.

  1. Increased human hybridoma formation by electrofusion of human B cells with heteromyeloma SPAM-8 cells.

    PubMed

    Panova, I; Gustafsson, B

    1995-06-01

    A fusion protocol was designed for the optimal production of hybridomas following electrofusion of human B cells with cells of the heteromyeloma fusion partner SPAM-8. Peripheral blood lymphocytes showed an average fusion efficiency of 0.4 x 10(-4) whereas Epstein-Barr virus-transformed B cells showed fusion efficiencies ranging from 6.2 x 10(-4) to 9.0 x 10(-4). Similar results were obtained with bone marrow-derived lymphocytes. Trypsin treatment of the cells prior to electrofusion further increased the fusion efficiency to 12.3 x 10(-4). In comparison, conventional polyethylene glycol-induced fusion resulted in a fusion efficiency of 0.8 x 10(-4). Thus, electrofusion of human B cells with SPAM-8 heteromyeloma cells introduced a 15-fold increase in hybridoma formation as compared to the conventional fusion method.

  2. The core regulatory network in human cells.

    PubMed

    Kim, Man-Sun; Kim, Dongsan; Kang, Nam Sook; Kim, Jeong-Rae

    2017-03-04

    In order to discover the common characteristics of various cell types in the human body, many researches have been conducted to find the set of genes commonly expressed in various cell types and tissues. However, the functional characteristics of a cell is determined by the complex regulatory relationships among the genes rather than by expressed genes themselves. Therefore, it is more important to identify and analyze a core regulatory network where all regulatory relationship between genes are active across all cell types to uncover the common features of various cell types. Here, based on hundreds of tissue-specific gene regulatory networks constructed by recent genome-wide experimental data, we constructed the core regulatory network. Interestingly, we found that the core regulatory network is organized by simple cascade and has few complex regulations such as feedback or feed-forward loops. Moreover, we discovered that the regulatory links from genes in the core regulatory network to genes in the peripheral regulatory network are much more abundant than the reverse direction links. These results suggest that the core regulatory network locates at the top of regulatory network and plays a role as a 'hub' in terms of information flow, and the information that is common to all cells can be modified to achieve the tissue-specific characteristics through various types of feedback and feed-forward loops in the peripheral regulatory networks. We also found that the genes in the core regulatory network are evolutionary conserved, essential and non-disease, non-druggable genes compared to the peripheral genes. Overall, our study provides an insight into how all human cells share a common function and generate tissue-specific functional traits by transmitting and processing information through regulatory network.

  3. Cell Culture Assay for Human Noroviruses [response

    SciTech Connect

    Straub, Tim M.; Honer Zu Bentrup, Kerstin; Orosz Coghlan, Patricia; Dohnalkova, Alice; Mayer, Brooke K.; Bartholomew, Rachel A.; Valdez, Catherine O.; Bruckner-Lea, Cindy J.; Gerba, Charles P.; Abbaszadegan, Morteza A.; Nickerson, Cheryl A.

    2007-07-01

    We appreciate the comments provided by Leung et al., in response to our recently published article “In Vitro Cell Culture Infectivity Assay for Human Noroviruses” by Straub et al. (1). The specific aim of our project was to develop an in vitro cell culture infectivity assay for human noroviruses (hNoV) to enhance risk assessments when they are detected in water supplies. Reverse transcription (RT) qualitative or quantitative PCR are the primary assays for waterborne NoV monitoring. However, these assays cannot distinguish between infectious vs. non-infectious virions. When hNoV is detected in water supplies, information provided by our infectivity assay will significantly improve risk assessment models and protect human health, regardless of whether we are propagating NoV. Indeed, in vitro cell culture infectivity assays for the waterborne pathogen Cryptosporidium parvum that supplement approved fluorescent microscopy assays, do not result in amplification of the environmentally resistant hard-walled oocysts (2). However, identification of life cycle stages in cell culture provides evidence of infectious oocysts in a water supply. Nonetheless, Leung et al.’s assertion regarding the suitability of our method for the in vitro propagation of high titers of NoV is valid for the medical research community. In this case, well-characterized challenge pools of virus would be useful for developing and testing diagnostics, therapeutics, and vaccines. As further validation of our published findings, we have now optimized RT quantitative PCR to assess the level of viral production in cell culture, where we are indeed finding significant increases in viral titer. The magnitude and time course of these increases is dependent on both virus strain and multiplicity of infection. We are currently preparing a manuscript that will discuss these findings in greater detail, and the implications this may have for creating viral challenge pools

  4. Inner Ear Hair Cell-Like Cells from Human Embryonic Stem Cells

    PubMed Central

    Ronaghi, Mohammad; Nasr, Marjan; Ealy, Megan; Durruthy-Durruthy, Robert; Waldhaus, Joerg; Diaz, Giovanni H.; Joubert, Lydia-Marie; Oshima, Kazuo

    2014-01-01

    In mammals, the permanence of many forms of hearing loss is the result of the inner ear's inability to replace lost sensory hair cells. Here, we apply a differentiation strategy to guide human embryonic stem cells (hESCs) into cells of the otic lineage using chemically defined attached-substrate conditions. The generation of human otic progenitor cells was dependent on fibroblast growth factor (FGF) signaling, and protracted culture led to the upregulation of markers indicative of differentiated inner ear sensory epithelia. Using a transgenic ESC reporter line based on a murine Atoh1 enhancer, we show that differentiated hair cell-like cells express multiple hair cell markers simultaneously. Hair cell-like cells displayed protrusions reminiscent of stereociliary bundles, but failed to fully mature into cells with typical hair cell cytoarchitecture. We conclude that optimized defined conditions can be used in vitro to attain otic progenitor specification and sensory cell differentiation. PMID:24512547

  5. Immortalization of human myogenic progenitor cell clone retaining multipotentiality

    SciTech Connect

    Hashimoto, Naohiro . E-mail: nao@nils.go.jp; Kiyono, Tohru; Wada, Michiko R.; Shimizu, Shirabe; Yasumoto, Shigeru; Inagawa, Masayo

    2006-10-06

    Human myogenic cells have limited ability to proliferate in culture. Although forced expression of telomerase can immortalize some cell types, telomerase alone delays senescence of human primary cultured myogenic cells, but fails to immortalize them. In contrast, constitutive expression of both telomerase and the E7 gene from human papillomavirus type 16 immortalizes primary human myogenic cells. We have established an immortalized primary human myogenic cell line preserving multipotentiality by ectopic expression of telomerase and E7. The immortalized human myogenic cells exhibit the phenotypic characteristics of their primary parent, including an ability to undergo myogenic, osteogenic, and adipogenic terminal differentiation under appropriate culture conditions. The immortalized cells will be useful for both basic and applied studies aimed at human muscle disorders. Furthermore, immortalization by transduction of telomerase and E7 represents a useful method by which to expand human myogenic cells in vitro without compromising their ability to differentiate.

  6. Amygdalin induces apoptosis in human cervical cancer cell line HeLa cells.

    PubMed

    Chen, Yu; Ma, Jinshu; Wang, Fang; Hu, Jie; Cui, Ai; Wei, Chengguo; Yang, Qing; Li, Fan

    2013-02-01

    Amygdalin, a naturally occurring substance, has been suggested to be efficacious as an anticancer substance. The effect of amygdalin on cervical cancer cells has never been studied. In this study, we found that the viability of human cervical cancer HeLa cell line was significantly inhibited by amygdalin. 4,6-Diamino-2-phenyl indole (DAPI) staining showed that amygdalin-treated HeLa cells developed typical apoptotic changes. The development of apoptosis in the amygdalin-treated HeLa cells were confirmed by double staining of amygdalin-treated HeLa cells with annexin V-FITC and propidium iodide (PI) along with increase in caspase-3 activity in these cells. Further studies indicated that antiapoptotic protein Bcl-2 was downregulated whereas proapoptotic Bax protein was upregulated in the amygdalin-treated HeLa cells implying involvement of the intrinsic pathway of apoptosis. In vivo, amygdalin administration inhibited the growth of HeLa cell xenografts through a mechanism of apoptosis. The results in the present study suggest that amygdalin may offer a new therapeutic option for patients with cervical cancer.

  7. Antiherpesvirus Activity in Human Sera and Urines after Administration of Adenine Arabinoside

    PubMed Central

    Champney, Keith J.; Lauter, Carl B.; Bailey, Elizabeth J.; Lerner, A. Martin

    1978-01-01

    The minimum inhibitory concentration (MIC) of adenine arabinoside (ara-A) in rabbit kidney microtiter tissue cultures (RK-13) to a prototype strain of herpes simplex virus, type 1 (E115) based upon inhibition of cytopathic effects is 1.5 μg/ml. In this system, the MIC of arabinosylhypoxanthine (ara-Hx), the major in vivo metabolic derivative of ara-A, is 75 μg/ml. Inhibition of cytopathic effects of herpes simplex virus, type 1 (HSV-1) in microtiter wells of RK-13 cells varies directly with the concentrations of ara-A or ara-Hx, and inversely with residual HSV-1. The MIC of ara-A for HSV-1 in RK-13 cells is 5-20 times lower than similar measures with vero renal, mouse embryo, or human foreskin cultures. With RK-13 tissue cultures in microtiter plates, an assay for “ara-A equivalents” in human body fluids was developed which compares in sensitivity with high pressure liquid chromatography and has the advantage of simultaneously measuring combined antiherpesvirus effects of ara-A and its major metabolic derivative, ara-Hx. In vitro checkerboard studies in RK-13 cells confirmed that ara-A and ara-Hx in combination had antiviral effects which are synergistic. The total of the fractional MIC of ara-A plus ara-Hx in combination also varies inversely with residual HSV-1 in microtiter wells. Because virus adsorption is complete at 2 h before specimens to be tested are added in this assay, and because human interferon is not measured in rabbit cells, the antiviral assay is not affected by the presence of type-specific antiherpesvirus antibody or human interferon. Antiviral activity (AVA) was assayed as ara-A equivalents in sera and urines from 10 patients with serious herpesvirus infections who received 2.5-20 mg/kg daily of ara-A by intramuscular or intravenous routes. When a dosage schedule of 10 mg/kg per day or more was used, sustained concentrations of AVA that ranged from 0.8 to 14.4 μg/ml were found. When an inhibitor of adenosine deaminase (covidarabine) was

  8. Nicotinamide extends replicative lifespan of human cells.

    PubMed

    Kang, Hyun Tae; Lee, Hyung Il; Hwang, Eun Seong

    2006-10-01

    We found that an ongoing application of nicotinamide to normal human fibroblasts not only attenuated expression of the aging phenotype but also increased their replicative lifespan, causing a greater than 1.6-fold increase in the number of population doublings. Although nicotinamide by itself does not act as an antioxidant, the cells cultured in the presence of nicotinamide exhibited reduced levels of reactive oxygen species (ROS) and oxidative damage products associated with cellular senescence, and a decelerated telomere shortening rate without a detectable increase in telomerase activity. Furthermore, in the treated cells growing beyond the original Hayflick limit, the levels of p53, p21WAF1, and phospho-Rb proteins were similar to those in actively proliferating cells. The nicotinamide treatment caused a decrease in ATP levels, which was stably maintained until the delayed senescence point. Nicotinamide-treated cells also maintained high mitochondrial membrane potential but a lower respiration rate and superoxide anion level. Taken together, in contrast to its demonstrated pro-aging effect in yeast, nicotinamide extends the lifespan of human fibroblasts, possibly through reduction in mitochondrial activity and ROS production.

  9. Landscape of transcription in human cells.

    PubMed

    Djebali, Sarah; Davis, Carrie A; Merkel, Angelika; Dobin, Alex; Lassmann, Timo; Mortazavi, Ali; Tanzer, Andrea; Lagarde, Julien; Lin, Wei; Schlesinger, Felix; Xue, Chenghai; Marinov, Georgi K; Khatun, Jainab; Williams, Brian A; Zaleski, Chris; Rozowsky, Joel; Röder, Maik; Kokocinski, Felix; Abdelhamid, Rehab F; Alioto, Tyler; Antoshechkin, Igor; Baer, Michael T; Bar, Nadav S; Batut, Philippe; Bell, Kimberly; Bell, Ian; Chakrabortty, Sudipto; Chen, Xian; Chrast, Jacqueline; Curado, Joao; Derrien, Thomas; Drenkow, Jorg; Dumais, Erica; Dumais, Jacqueline; Duttagupta, Radha; Falconnet, Emilie; Fastuca, Meagan; Fejes-Toth, Kata; Ferreira, Pedro; Foissac, Sylvain; Fullwood, Melissa J; Gao, Hui; Gonzalez, David; Gordon, Assaf; Gunawardena, Harsha; Howald, Cedric; Jha, Sonali; Johnson, Rory; Kapranov, Philipp; King, Brandon; Kingswood, Colin; Luo, Oscar J; Park, Eddie; Persaud, Kimberly; Preall, Jonathan B; Ribeca, Paolo; Risk, Brian; Robyr, Daniel; Sammeth, Michael; Schaffer, Lorian; See, Lei-Hoon; Shahab, Atif; Skancke, Jorgen; Suzuki, Ana Maria; Takahashi, Hazuki; Tilgner, Hagen; Trout, Diane; Walters, Nathalie; Wang, Huaien; Wrobel, John; Yu, Yanbao; Ruan, Xiaoan; Hayashizaki, Yoshihide; Harrow, Jennifer; Gerstein, Mark; Hubbard, Tim; Reymond, Alexandre; Antonarakis, Stylianos E; Hannon, Gregory; Giddings, Morgan C; Ruan, Yijun; Wold, Barbara; Carninci, Piero; Guigó, Roderic; Gingeras, Thomas R

    2012-09-06

    Eukaryotic cells make many types of primary and processed RNAs that are found either in specific subcellular compartments or throughout the cells. A complete catalogue of these RNAs is not yet available and their characteristic subcellular localizations are also poorly understood. Because RNA represents the direct output of the genetic information encoded by genomes and a significant proportion of a cell's regulatory capabilities are focused on its synthesis, processing, transport, modification and translation, the generation of such a catalogue is crucial for understanding genome function. Here we report evidence that three-quarters of the human genome is capable of being transcribed, as well as observations about the range and levels of expression, localization, processing fates, regulatory regions and modifications of almost all currently annotated and thousands of previously unannotated RNAs. These observations, taken together, prompt a redefinition of the concept of a gene.

  10. Human cell culture in a space bioreactor

    NASA Technical Reports Server (NTRS)

    Morrison, Dennis R.

    1988-01-01

    Microgravity offers new ways of handling fluids, gases, and growing mammalian cells in efficient suspension cultures. In 1976 bioreactor engineers designed a system using a cylindrical reactor vessel in which the cells and medium are slowly mixed. The reaction chamber is interchangeable and can be used for several types of cell cultures. NASA has methodically developed unique suspension type cell and recovery apparatus culture systems for bioprocess technology experiments and production of biological products in microgravity. The first Space Bioreactor was designed for microprocessor control, no gaseous headspace, circulation and resupply of culture medium, and slow mixing in very low shear regimes. Various ground based bioreactors are being used to test reactor vessel design, on-line sensors, effects of shear, nutrient supply, and waste removal from continuous culture of human cells attached to microcarriers. The small Bioreactor is being constructed for flight experiments in the Shuttle Middeck to verify systems operation under microgravity conditions and to measure the efficiencies of mass transport, gas transfer, oxygen consumption and control of low shear stress on cells.

  11. Osmotic properties of human red cells.

    PubMed

    Solomon, A K; Toon, M R; Dix, J A

    1986-01-01

    When an osmotic pressure gradient is applied to human red cells, the volume changes anomalously, as if there were a significant fraction of "nonosmotic water" which could not serve as solvent for the cell solutes, a finding which has been discussed widely in the literature. In 1968, Gary-Bobo and Solomon (J. Gen. Physiol. 52:825) concluded that the anomalies could not be entirely explained by the colligative properties of hemoglobin (Hb) and proposed that there was an additional concentration dependence of the Hb charge (ZHb). A number of investigators, particularly Freedman and Hoffman (1979, J. Gen. Physiol. 74:157) have been unable to confirm Gary-Bobo and Solomon's experimental evidence for this concentration dependence of ZHb and we now report that we are also unable to repeat the earlier experiments. Nonetheless, there still remains a significant anomaly which amounts to 12.5 +/- 0.8% of the total isosmotic cell water (P much less than 0.0005, t test), even after taking account of the concentration dependence of the Hb osmotic coefficient and all the other known physical chemical constraints, ideal and nonideal. It is suggested that the anomalies at high Hb concentration in shrunken cells may arise from the ionic strength dependence of the Hb osmotic coefficient. In swollen red cells at low ionic strength, solute binding to membrane and intracellular proteins is increased and it is suggested that this factor may account, in part, for the anomalous behavior of these cells.

  12. Tryptophan hydroxylase expression in human skin cells.

    PubMed

    Slominski, Andrzej; Pisarchik, Alexander; Johansson, Olle; Jing, Chen; Semak, Igor; Slugocki, George; Wortsman, Jacobo

    2003-10-15

    We attempted to further characterize cutaneous serotoninergic and melatoninergic pathways evaluating the key biosynthetic enzyme tryptophan hydroxylase (TPH). There was wide expression of TPH mRNA in whole human skin, cultured melanocytes and melanoma cells, dermal fibroblasts, squamous cell carcinoma cells and keratinocytes. Gene expression was associated with detection of TPH immunoreactive species by Western blotting. Characterization of the TPH immunoreactive species performed with two different antibodies showed expression of the expected protein (55-60 kDa), and of forms with higher and lower molecular weights. This pattern of broad spectrum of TPH expression including presumed degradation products suggests rapid turnover of the enzyme, as previously reported in mastocytoma cells. RP-HPLC of skin extracts showed fluorescent species with the retention time of serotonin and N-acetylserotonin. Immunocytochemistry performed in skin biopsies localized TPH immunoreactivity to normal and malignant melanocytes. We conclude that while the TPH mRNA and protein are widely expressed in cultured normal and pathological epidermal and dermal skin cells, in vivo TPH expression is predominantly restricted to cells of melanocytic origin.

  13. Human prostatic cancer cells, PC3, elaborate mitogenic activity which selectively stimulates human bone cells

    SciTech Connect

    Perkel, V.S.; Mohan, S.; Herring, S.J.; Baylink, D.J.; Linkhart, T.A. )

    1990-11-01

    Prostatic cancer typically produces osteoblastic metastases which are not attended by marrow fibrosis. In the present study we sought to test the hypothesis that prostatic cancer cells produce factor(s) which act selectively on human osteoblasts. Such a paracrine mechanism would explain the observed increase in osteoblasts, unaccompanied by an increase in marrow fibroblasts. To test this hypothesis we investigated the mitogenic activity released by the human prostatic tumor cell line, PC3. PC3 cells have been reported previously to produce mitogenic activity for cells that was relatively specific for rat osteoblasts compared to rat fibroblasts. However, the effects of this activity on human cells has not been examined previously. PC3-conditioned medium (CM) (5-50 micrograms CM protein/ml) stimulated human osteoblast proliferation by 200-950% yet did not stimulate human fibroblast proliferation ((3H)thymidine incorporation). PC3 CM also increased cell numbers in human osteoblast but not fibroblast cell cultures. To determine whether the osteoblast-specific mitogenic activity could be attributed to known bone growth factors, specific assays for these growth factors were performed. PC3 CM contained 10 pg insulin-like growth factor (IGF) I, less than 2 pg IGF II, 54 pg basic fibroblast growth factor, and 16 pg transforming growth factor beta/microgram CM protein. None of these growth factors alone or in combination could account for the observed osteoblast-specific PC3 cell-derived mitogenic activity. Furthermore, when 5 micrograms/ml PC3 CM was tested in combination with maximally effective concentrations of either basic fibroblast growth factor, IGF I, IGF II, or transforming growth factor beta, it produced an additive effect suggesting that PC3 CM stimulates osteoblast proliferation by a mechanism independent of these bone mitogens.

  14. The administration of multipotent stromal cells at precancerous stage precludes tumor growth and epithelial dedifferentiation of oral squamous cell carcinoma.

    PubMed

    Bruna, Flavia; Arango-Rodríguez, Martha; Plaza, Anita; Espinoza, Iris; Conget, Paulette

    2017-01-01

    Multipotent stromal cells (MSCs) are envisioned as a powerful therapeutic tool. As they home into tumors, secrete trophic and vasculogenic factors, and suppress immune response their role in carcinogenesis is a matter of controversy. Worldwide oral squamous cell carcinoma (OSCC) is the fifth most common epithelial cancer. Our aim was to determine whether MSC administration at precancerous stage modifies the natural progression of OSCC. OSCC was induced in Syrian hamsters by topical application of DMBA in the buccal pouch. At papilloma stage, the vehicle or 3×10(6) allogenic bone marrow-derived MSCs were locally administered. Four weeks later, the lesions were studied according to: volume, stratification (histology), proliferation (Ki-67), apoptosis (Caspase 3 cleaved), vasculature (ASMA), inflammation (Leukocyte infiltrate), differentiation (CK1 and CK4) and gene expression profile (mRNA). Tumors found in individuals that received MSCs were smaller than those presented in the vehicle group (87±80 versus 54±62mm(3), p<0.05). The rate of proliferation was two times lower and the apoptosis was 2.5 times higher in lesions treated with MSCs than in untreated ones. While the laters presented dedifferentiated cells, the former maintained differentiated cells (cytokeratin and gene expression profile similar to normal tissue). Thus, MSC administration at papilloma stage precludes tumor growth and epithelial dedifferentiation of OSCC.

  15. Dermal Lipogenesis Inhibits Adiponectin Production in Human Dermal Fibroblasts while Exogenous Adiponectin Administration Prevents against UVA-Induced Dermal Matrix Degradation in Human Skin

    PubMed Central

    Fang, Chien-Liang; Huang, Ling-Hung; Tsai, Hung-Yueh; Chang, Hsin-I

    2016-01-01

    Adiponectin is one of the most abundant adipokines from the subcutaneous fat, and regulates multiple activities through endocrine, paracrine, or autocrine mechanisms. However, its expression in adipogenic induced fibroblasts, and the potential role in photoaging has not been determined. Here, human dermal fibroblasts, Hs68, were presented as a cell model of dermal lipogenesis through stimulation of adipogenic differentiation medium (ADM). Similar to other studies in murine pre-adipocyte models (i.e., 3T3-L1), Hs68 fibroblasts showed a tendency to lipogenesis based on lipid accumulation, triglyceride formation, and the expressions of PPAR-γ, lipoprotein lipase (LPL), and FABP4 mRNA. As expected, ADM-treated fibroblasts displayed a reduction on adiponectin expression. Next, we emphasized the photoprotective effects of adiponectin against UVA-induced damage in Hs68 fibroblasts. UVA radiation can downregulate cell adhesion strength and elastic modulus of Hs68 fibroblasts. Moreover, UVA radiation could induce the mRNA expressions of epidermal growth factor receptor (EGFR), adiponectin receptor 1 (AdipoR1), matrix metalloproteinase-1 (MMP-1), MMP-3, and cyclooxygenase-2 (COX-2), but downregulate the mRNA expressions of type I and type III collagen. On the other hand, post-treatment of adiponectin can partially overcome UVA-induced reduction in the cell adhesion strength of Hs68 fibroblasts through the activation of AdipoR1 and the suppression of EGF-R. In addition, post-treatment of adiponectin indicated the increase of type III collagen and elastin mRNA expression and the decrease of MMP-1 and MMP-3 mRNA expression, but a limited degree of recovery of elastic modulus on UVA-irradiated Hs68 fibroblasts. Overall, these results suggest that dermal lipogenesis may inhibit the expression of adiponectin while exogenous adiponectin administration prevents against UVA-induced dermal matrix degradation in Hs68 fibroblasts. PMID:27428951

  16. Dermal Lipogenesis Inhibits Adiponectin Production in Human Dermal Fibroblasts while Exogenous Adiponectin Administration Prevents against UVA-Induced Dermal Matrix Degradation in Human Skin.

    PubMed

    Fang, Chien-Liang; Huang, Ling-Hung; Tsai, Hung-Yueh; Chang, Hsin-I

    2016-07-14

    Adiponectin is one of the most abundant adipokines from the subcutaneous fat, and regulates multiple activities through endocrine, paracrine, or autocrine mechanisms. However, its expression in adipogenic induced fibroblasts, and the potential role in photoaging has not been determined. Here, human dermal fibroblasts, Hs68, were presented as a cell model of dermal lipogenesis through stimulation of adipogenic differentiation medium (ADM). Similar to other studies in murine pre-adipocyte models (i.e., 3T3-L1), Hs68 fibroblasts showed a tendency to lipogenesis based on lipid accumulation, triglyceride formation, and the expressions of PPAR-γ, lipoprotein lipase (LPL), and FABP4 mRNA. As expected, ADM-treated fibroblasts displayed a reduction on adiponectin expression. Next, we emphasized the photoprotective effects of adiponectin against UVA-induced damage in Hs68 fibroblasts. UVA radiation can downregulate cell adhesion strength and elastic modulus of Hs68 fibroblasts. Moreover, UVA radiation could induce the mRNA expressions of epidermal growth factor receptor (EGFR), adiponectin receptor 1 (AdipoR1), matrix metalloproteinase-1 (MMP-1), MMP-3, and cyclooxygenase-2 (COX-2), but downregulate the mRNA expressions of type I and type III collagen. On the other hand, post-treatment of adiponectin can partially overcome UVA-induced reduction in the cell adhesion strength of Hs68 fibroblasts through the activation of AdipoR1 and the suppression of EGF-R. In addition, post-treatment of adiponectin indicated the increase of type III collagen and elastin mRNA expression and the decrease of MMP-1 and MMP-3 mRNA expression, but a limited degree of recovery of elastic modulus on UVA-irradiated Hs68 fibroblasts. Overall, these results suggest that dermal lipogenesis may inhibit the expression of adiponectin while exogenous adiponectin administration prevents against UVA-induced dermal matrix degradation in Hs68 fibroblasts.

  17. Glucocorticoid receptor in human respiratory epithelial cells.

    PubMed

    Pujolsa, Laura; Mullol, Joaquim; Picado, Cèsar

    2009-01-01

    Inhaled and intranasal glucocorticoids (GCs) are the most common and effective drugs for controlling symptoms and airway inflammation in respiratory diseases such as allergic rhinitis, chronic rhinosinusitis with/without nasal polyps, and asthma, and the respiratory epithelium is a primary target of GC anti-inflammatory actions. GC effects are mediated through the GC receptor (GR). In humans, one single GR gene gives rise to two main GR products, namely GRalpha and GRbeta, which are subject to translational and posttranslational modifications. GRalpha is expressed in virtually all human cells and tissues, including respiratory epithelial cells, and - at least in vitro - is downregulated by GC. GRalpha mediates the anti-inflammatory actions of GC by activating transcription of anti-inflammatory genes through binding of GRalpha to glucocorticoid response elements (GRE) located in the promoter region of target genes, repressing transcription of proinflammatory genes through direct interaction between GRalpha and proinflammatory transcription factors, such as AP-1 and NF-kappaB (transrepression), and also by destabilizing the mRNA of proinflammatory mediators. GRbeta acts as a dominant negative inhibitor of GRalpha-mediated transactivation and transrepression in certain in vitro studies with transfected cells. The GRbeta message is expressed at low levels in numerous tissues and its protein is mainly expressed in inflammatory cells, although it has also been detected in airway epithelial cells. Increased GRbeta expression has been reported in bronchial asthma and nasal polyposis, and after incubation of cells with certain proinflammatory stimuli. However, the role of GRbeta in modulating GC sensitivity in vivo has been highly debated and is as yet unclear.

  18. Appearance of Human Plasma Cells Following Differentiation of Human B Cells in NOD/SCID Mouse Spleen

    PubMed Central

    Kikuchi, Kentaro; Lian, Zhe-Xiong; He, Xiao-Song; Ansari, Aftab A.; Ishibashi, Miyuki; Miyakawa, Hiroshi; Shultz, Leonard D.; Ikehara, Susumu; Gershwin, M. Eric

    2003-01-01

    Relatively little is known for the differentiation and maturation process of human B cells to plasma cells. This is particularly important in reconstitution work involving transfer of autoantibodies. To address this issue, we transplanted human peripheral blood mononuclear cells (PBMC) directly into the spleen of irradiated NOD/SCID mice depleted of natural killer cell activity. Within 6 weeks, naïve B cells differentiated into memory B cells and, importantly, the numbers of human CD138+ plasma cells in spleen increased by 100 fold after transplantation. Plasma cell numbers correlated with the detection of human IgM and IgG in serum, indicating that human B cells had differentiated into mature plasma cells in the murine spleen. In addition to CD19+ plasma cells, a distinct CD19- plasma cell population was detected, suggesting that downregulation of CD19 associated with maturation of plasma cells occurred. When purified human B cells were transplanted, those findings were not observed. Our results indicate that differentiation and maturation of human B cells and plasma cells can be investigated by transplantation of human PBMC into the spleen of NOD/SCID mice. The model will be useful for studying the differentiation of human B cells and generation of plasma cells. PMID:14768952

  19. Systemic Administration of Allogeneic Mesenchymal Stem Cells Does Not Halt Osteoporotic Bone Loss in Ovariectomized Rats

    PubMed Central

    Sun, Yuxin; Lin, Sien; Gu, Weidong; Liu, Yamei; Zhang, Jinfang; Chen, Lin; Li, Gang

    2016-01-01

    Mesenchymal stem cells (MSCs) have innate ability to self-renew and immunosuppressive functions, and differentiate into various cell types. They have become a promising cell source for treating many diseases, particular for bone regeneration. Osteoporosis is a common metabolic bone disorder with elevated systemic inflammation which in turn triggers enhanced bone loss. We hypothesize that systemic infusion of MSCs may suppress the elevated inflammation in the osteoporotic subjects and slow down bone loss. The current project was to address the following two questions: (1) Will a single dose systemic administration of allogenic MSCs have any effect on osteoporotic bone loss? (2) Will multiple administration of allogenic MSCs from single or multiple donors have similar effect on osteoporotic bone loss? 18 ovariectomized (OVX) rats were assigned into 3 groups: the PBS control group, MSCs group 1 (receiving 2x106 GFP-MSCs at Day 10, 46, 91 from the same donor following OVX) and MSCs group 2 (receiving 2x106 GFP-MSCs from three different donors at Day 10, 46, 91). Examinations included Micro-CT, serum analysis, mechanical testing, immunofluorescence staining and bone histomorphometry analysis. Results showed that BV/TV at Day 90, 135, BMD of TV and trabecular number at Day 135 in the PBS group were significantly higher than those in the MSCs group 2, whereas trabecular spacing at Day 90, 135 was significantly smaller than that in MSCs group 2. Mechanical testing data didn’t show significant difference among the three groups. In addition, the ELISA assay showed that level of Rantes in serum in MSCs group 2 was significantly higher than that of the PBS group, whereas IL-6 and IL-10 were significantly lower than those of the PBS group. Bone histomorphometry analysis showed that Oc.S/BS and Oc.N/BS in the PBS group were significant lower than those in MSCs group 2; Ob.S/BS and Ob.N/BS did not show significant difference among the three groups. The current study

  20. Intracerebral Stem Cell Administration Inhibits Relapse-like Alcohol Drinking in Rats.

    PubMed

    Israel, Yedy; Ezquer, Fernando; Quintanilla, María Elena; Morales, Paola; Ezquer, Marcelo; Herrera-Marschitz, Mario

    2017-01-01

    Study describes the blockade of relapse-like alcohol drinking by mesenchymal stem cells (MSCs). High alcohol-intake bred rats consumed alcohol for 3 months and were subjected to repeated alcohol deprivations for 7-14 days, followed by alcohol reaccess. Upon reaccess, animals consumed 2.2 g alcohol/kg in 60 minutes. A single intra-cerebroventricular MSC administration inhibited relapse-like drinking up to 80-85% for 40 days (P < 0.001). An alcohol-use-disorder was prevented.

  1. Optimizing Intradermal Administration of Cryopreserved Plasmodium falciparum Sporozoites in Controlled Human Malaria Infection.

    PubMed

    Lyke, Kirsten E; Laurens, Matthew B; Strauss, Kathy; Adams, Matthew; Billingsley, Peter F; James, Eric; Manoj, Anita; Chakravarty, Sumana; Plowe, Christopher V; Li, Ming Lin; Ruben, Adam; Edelman, Robert; Green, Michael; Dube, Tina J; Sim, B Kim Lee; Hoffman, Stephen L

    2015-12-01

    Controlled human malaria infection (CHMI) is a powerful tool to evaluate malaria vaccine and prophylactic drug efficacy. Until recently CHMI was only carried out by the bite of infected mosquitoes. A parenteral method of CHMI would standardize Plasmodium falciparum sporozoite (PfSPZ) administration, eliminate the need for expensive challenge facility infrastructure, and allow for use of many P. falciparum strains. Recently, intradermal (ID) injection of aseptic, purified, cryopreserved PfSPZ was shown to induce P. falciparum malaria; however, 100% infection rates were not achieved by ID injection. To optimize ID PfSPZ dosing so as to achieve 100% infection, 30 adults aged 18-45 years were randomized to one of six groups composed of five volunteers each. The parameters of dose (1 × 10(4) versus 5 × 10(4) PfSPZ total dose per volunteer), number of injections (two versus eight), and aliquot volume per ID injection (10 μL versus 50 μL) were studied. Three groups attained 100% infection: 1 × 10(4) PfSPZ in 50 μL/2 doses, 1 × 10(4) PfSPZ in 10 μL/2 doses, and 5 × 10(4) PfSPZ in 10 μL/8 doses. The group that received 5 × 10(4) PfSPZ total dose in eight 10 μL injections had a 100% infection rate and the shortest prepatent period (mean of 12.7 days), approaching the prepatent period for the current CHMI standard of five infected mosquitoes.

  2. Pharmacokinetics of human chorionic gonadotropin after i.m. administration in goats (Capra hircus).

    PubMed

    Saleh, M; Shahin, M; Wuttke, W; Gauly, M; Holtz, W

    2012-07-01

    The present investigation addresses the pharmacokinetics of human chorionic gonadotropin (hCG), intramuscularly (i.m.) administered to goats. Nine pluriparous does of the Boer goat breed, 2-6 years of age and weighing 45-60 kg, were administered 500 IU hCG (2 ml Chorulon) deep into the thigh musculature 18 h after superovulatory FSH treatment. Blood samples were drawn from the jugular vein at 2  h intervals for the first 24h, at 6 h intervals until 42 h, and at 12 h intervals until 114 h after administration. After centrifugation, plasma hCG concentrations were determined by electrochemiluminescence immunoassay. Pharmacokinetical parameters were as follows: lag time, 0.4 (s.e.m. 0.1) h; absorption rate constant, 0.34 (s.e.m. 0.002) h; absorption half-life, 2.7 (s.e.m. 0.5) h; elimination rate constant, 0.02 (s.e.m. 0.002) h; biological half-life, 39.4 (s.e.m. 5.1) h; and apparent volume of distribution, 16.9 (s.e.m. 4.3) l. The plasma hCG profile was characterized by an absorption phase of 11.6 (s.e.m. 1.8) h and an elimination phase of 70.0 (s.e.m. 9.8) h, with considerable individual variation in bioavailability and pharmacokinetical parameters. Biological half-life was negatively correlated (P<0.05) with peak concentration (r=-0.76), absorption rate constant (r=-0.78), and elimination rate constant (r=-0.87). The results indicate that after rapid absorption, hCG remains in the circulation for an extended period. This has to be taken into account when assessing the stimulatory response to hCG treatment on an ovarian level.

  3. Optimizing Intradermal Administration of Cryopreserved Plasmodium falciparum Sporozoites in Controlled Human Malaria Infection

    PubMed Central

    Lyke, Kirsten E.; Laurens, Matthew B.; Strauss, Kathy; Adams, Matthew; Billingsley, Peter F.; James, Eric; Manoj, Anita; Chakravarty, Sumana; Plowe, Christopher V.; Li, Ming Lin; Ruben, Adam; Edelman, Robert; Green, Michael; Dube, Tina J.; Kim Lee Sim, B.; Hoffman, Stephen L.

    2015-01-01

    Controlled human malaria infection (CHMI) is a powerful tool to evaluate malaria vaccine and prophylactic drug efficacy. Until recently CHMI was only carried out by the bite of infected mosquitoes. A parenteral method of CHMI would standardize Plasmodium falciparum sporozoite (PfSPZ) administration, eliminate the need for expensive challenge facility infrastructure, and allow for use of many P. falciparum strains. Recently, intradermal (ID) injection of aseptic, purified, cryopreserved PfSPZ was shown to induce P. falciparum malaria; however, 100% infection rates were not achieved by ID injection. To optimize ID PfSPZ dosing so as to achieve 100% infection, 30 adults aged 18–45 years were randomized to one of six groups composed of five volunteers each. The parameters of dose (1 × 104 versus 5 × 104 PfSPZ total dose per volunteer), number of injections (two versus eight), and aliquot volume per ID injection (10 μL versus 50 μL) were studied. Three groups attained 100% infection: 1 × 104 PfSPZ in 50 μL/2 doses, 1 × 104 PfSPZ in 10 μL/2 doses, and 5 × 104 PfSPZ in 10 μL/8 doses. The group that received 5 × 104 PfSPZ total dose in eight 10 μL injections had a 100% infection rate and the shortest prepatent period (mean of 12.7 days), approaching the prepatent period for the current CHMI standard of five infected mosquitoes. PMID:26416102

  4. Stereoselective urinary MDMA (ecstasy) and metabolites excretion kinetics following controlled MDMA administration to humans.

    PubMed

    Schwaninger, Andrea E; Meyer, Markus R; Barnes, Allan J; Kolbrich-Spargo, Erin A; Gorelick, David A; Goodwin, Robert S; Huestis, Marilyn A; Maurer, Hans H

    2012-01-01

    The R- and S-enantiomers of racemic 3,4-methylenedioxymethamphetamine (MDMA) exhibit different dose-concentration curves. In plasma, S-MDMA was eliminated at a higher rate, most likely due to stereoselective metabolism. Similar data were shown in various in vitro experiments. The aim of the present study was the in vivo investigation of stereoselective elimination of MDMA's phase I and phase II metabolites in human urine following controlled oral MDMA administration. Urine samples from 10 participants receiving 1.0 and 1.6 mg/kg MDMA separated by at least one week were analyzed blind by liquid chromatography-high resolution-mass spectrometry and gas chromatography-mass spectrometry after chiral derivatization with S-heptafluorobutyrylprolyl chloride. R/S ratios at C(max) were comparable after low and high doses with ratios >1 for MDMA, free DHMA, and HMMA sulfate, and with ratios <1 for MDA, free HMMA, DHMA sulfate and HMMA glucuronide. In the five days after the high MDMA dose, a median of 21% of all evaluated compounds were excreted as R-stereoisomers and 17% as S-stereoisomers. Significantly greater MDMA, DHMA, and HMMA sulfate R-enantiomers and HMMA and HMMA glucuronide S-stereoisomers were excreted. No significant differences were observed for MDA and DHMA sulfate stereoisomers. Changes in R/S ratios could be observed over time for all analytes, with steady increases in the first 48 h. R/S ratios could help to roughly estimate time of MDMA ingestion and therefore, improve interpretation of MDMA and metabolite urinary concentrations in clinical and forensic toxicology.

  5. Effects of methamphetamine administration on information gathering during probabilistic reasoning in healthy humans.

    PubMed

    Ermakova, Anna O; Ramachandra, Pranathi; Corlett, Philip R; Fletcher, Paul C; Murray, Graham K

    2014-01-01

    Jumping to conclusions (JTC) during probabilistic reasoning is a cognitive bias repeatedly demonstrated in people with schizophrenia and shown to be associated with delusions. Little is known about the neurochemical basis of probabilistic reasoning. We tested the hypothesis that catecholamines influence data gathering and probabilistic reasoning by administering intravenous methamphetamine, which is known to cause synaptic release of the catecholamines noradrenaline and dopamine, to healthy humans whilst they undertook a probabilistic inference task. Our study used a randomised, double-blind, cross-over design. Seventeen healthy volunteers on three visits were administered either placebo or methamphetamine or methamphetamine preceded by amisulpride. In all three conditions participants performed the "beads" task in which participants decide how much information to gather before making a probabilistic inference, and which measures the cognitive bias towards jumping to conclusions. Psychotic symptoms triggered by methamphetamine were assessed using Comprehensive Assessment of At-Risk Mental States (CAARMS). Methamphetamine induced mild psychotic symptoms, but there was no effect of drug administration on the number of draws to decision (DTD) on the beads task. DTD was a stable trait that was highly correlated within subjects across visits (intra-class correlation coefficients of 0.86 and 0.91 on two versions of the task). The less information was sampled in the placebo condition, the more psychotic-like symptoms the person had after the methamphetamine plus amisulpride condition (p = 0.028). Our results suggest that information gathering during probabilistic reasoning is a stable trait, not easily modified by dopaminergic or noradrenergic modulation.

  6. Inhibition of effector antigen-specific T cells by intradermal administration of heme oxygenase-1 inducers.

    PubMed

    Simon, Thomas; Pogu, Julien; Rémy, Séverine; Brau, Frédéric; Pogu, Sylvie; Maquigneau, Maud; Fonteneau, Jean-François; Poirier, Nicolas; Vanhove, Bernard; Blancho, Gilles; Piaggio, Eliane; Anegon, Ignacio; Blancou, Philippe

    2017-03-22

    Developing protocols aimed at inhibiting effector T cells would be key for the treatment of T cell-dependent autoimmune diseases including type 1 autoimmune diabetes (T1D) and multiple sclerosis (MS). While heme oxygenase-1 (HO-1) inducers are clinically approved drugs for non-immune-related diseases, they do have immunosuppressive properties when administered systemically in rodents. Here we show that HO-1 inducers inhibit antigen-specific effector T cells when injected intradermally together with the T cell cognate antigens in mice. This phenomenon was observed in both a CD8(+) T cell-mediated model of T1D and in a CD4(+) T cell-dependent MS model. Intradermal injection of HO-1 inducers induced the recruitment of HO-1(+) monocyte-derived dendritic cell (MoDCs) exclusively to the lymph nodes (LN) draining the site of intradermal injection. After encountering HO-1(+)MoDCs, effector T-cells exhibited a lower velocity and a reduced ability to migrate towards chemokine gradients resulting in impaired accumulation to the inflamed organ. Intradermal co-injection of a clinically approved HO-1 inducer and a specific antigen to non-human primates also induced HO-1(+) MoDCs to accumulate in dermal draining LN and to suppress delayed-type hypersensitivity. Therefore, in both mice and non-human primates, HO-1 inducers delivered locally inhibited effector T-cells in an antigen-specific manner, paving the way for repositioning these drugs for the treatment of immune-mediated diseases.

  7. The appetite suppressant d-fenfluramine induces apoptosis in human serotonergic cells.

    PubMed

    Bengel, D; Isaacs, K R; Heils, A; Lesch, K P; Murphy, D L

    1998-09-14

    Fenfluramine is an amphetamine analogue which has been widely used in the treatment of obesity. In rodents, non-human primates, and humans, fenfluramine is associated with some indices of neurotoxicity, as well as pulmonary hypertension and cardiac valve pathology. In the present study, d-fenfluramine was found to be cytotoxic to the serotonin (5-HT) transporter (5-HTT) expressing human placental choriocarcinoma cells. d-Fenfluramine caused DNA fragmentation and apoptosis. Apoptosis was not observed after the 5-HTT had been blocked by fluoxetine, indicating that intact 5-HTT function is required for d-fenfluramine to induce programmed cell death. These observations in a human cell line may reflect a possible mechanism associated with the risks of fenfluramine administration in several species, including humans.

  8. Cell cycle regulation in human embryonic stem cells: links to adaptation to cell culture.

    PubMed

    Barta, Tomas; Dolezalova, Dasa; Holubcova, Zuzana; Hampl, Ales

    2013-03-01

    Cell cycle represents not only a tightly orchestrated mechanism of cell replication and cell division but it also plays an important role in regulation of cell fate decision. Particularly in the context of pluripotent stem cells or multipotent progenitor cells, regulation of cell fate decision is of paramount importance. It has been shown that human embryonic stem cells (hESCs) show unique cell cycle characteristics, such as short doubling time due to abbreviated G1 phase; these properties change with the onset of differentiation. This review summarizes the current understanding of cell cycle regulation in hESCs. We discuss cell cycle properties as well as regulatory machinery governing cell cycle progression of undifferentiated hESCs. Additionally, we provide evidence that long-term culture of hESCs is accompanied by changes in cell cycle properties as well as configuration of several cell cycle regulatory molecules.

  9. Proinflammatory cytokines differentially influence adult hippocampal cell proliferation depending upon the route and chronicity of administration.

    PubMed

    Seguin, Julie Anne; Brennan, Jordan; Mangano, Emily; Hayley, Shawn

    2009-01-01

    Disturbances of hippocampal plasticity, including impaired dendritic branching and reductions of neurogenesis, are provoked by stressful insults and may occur in depression. Although corticoids likely contribute to stressor-induced reductions of neurogenesis, other signaling messengers, including pro-inflammatory cytokines might also be involved. Accordingly, the present investigation assessed whether three proinflammatory cytokines, namely interleukin-1beta (IL-1beta), IL-6, and tumor necrosis factor-alpha (TNF-alpha) (associated with depression) influenced cellular proliferation within the hippocampus. In this regard, systemic administration of TNF-alpha reduced 5-bromo-2-deoxyuridine (BrdU) labeling within the hippocampus, whereas IL-1beta and IL-6 had no such effect. However, repeated but not a single intra-hippocampal infusion of IL-6 and IL-1beta actually increased cellular proliferation and IL-6 infusion also enhanced microglial staining within the hippocampus. Yet, no changes in doublecortin expression were apparent, suggesting that the cytokine did not influence the birth of cells destined to become neurons. Essentially, the route of administration and chronicity of cytokine administration had a marked influence upon the nature of hippocampal alterations provoked, suggesting that cytokines may differentially regulate hippocampal plasticity in neuropsychiatric conditions.

  10. Humanism, Administration and Education: The Demand of Documentation and the Production of a New Pedagogical Desire

    ERIC Educational Resources Information Center

    Plum, Maja

    2012-01-01

    Through the example of a Danish reform of educational plans in early childhood education, this paper analyses the emergence of a new pedagogical desire related to administrative educational reforms promoting accountability, visibility and documentation. Two arguments are made: first, it is argued that the changes in administrative practices during…

  11. Cell entry by human pathogenic arenaviruses.

    PubMed

    Rojek, Jillian M; Kunz, Stefan

    2008-04-01

    The arenaviruses Lassa virus (LASV) in Africa and Machupo (MACV), Guanarito (GTOV) and Junin viruses (JUNV) in South America cause severe haemorrhagic fevers in humans with fatality rates of 15-35%. The present review focuses on the first steps of infection with human pathogenic arenaviruses, the interaction with their cellular receptor molecules and subsequent entry into the host cell. While similarities exist in genomic organization, structure and clinical disease caused by pathogenic Old World and New World arenaviruses these pathogens use different primary receptors. The Old World arenaviruses employ alpha-dystroglycan, a cellular receptor for proteins of the extracellular matrix, and the human pathogenic New World arenaviruses use the cellular cargo receptor transferrin receptor 1. While the New World arenavirus JUNV enters cells via clathrin-dependent endocytosis, evidence occurred for clathrin-independent entry of the prototypic Old World arenavirus lymphocytic choriomeningitis virus. Upon internalization, arenaviruses are delivered to the endosome, where pH-dependent membrane fusion is mediated by the envelope glycoprotein (GP). While arenavirus GPs share characteristics with class I fusion GPs of other enveloped viruses, unusual mechanistic features of GP-mediated membrane fusion have recently been discovered for arenaviruses with important implications for viral entry.

  12. Molecular imaging of human embryonic stem cells.

    PubMed

    Narsinh, Kazim H; Cao, Feng; Wu, Joseph C

    2009-01-01

    Human embryonic stem cells (hESCs) are a renewable source of differentiated cell types that may be employed in various tissue regeneration strategies. However, clinical implementation of cell transplantation therapy is hindered by legitimate concerns regarding the in vivo teratoma formation of undifferentiated hESCs and host immune reactions to allogenic cells. Investigating in vivo hESC behaviour and the ultimate feasibility of cell transplantation therapy necessitates the development of novel molecular imaging techniques to longitudinally monitor hESC localization, proliferation, and viability in living subjects. An innovative approach to harness the respective strengths of various imaging platforms is the creation and use of a fusion reporter construct composed of red fluorescent protein (RFP), firefly luciferase (fluc), and herpes simplex virus thymidine kinase (HSV-tk). The imaging modalities made available by use of this construct, including optical fluorescence, bioluminescence, and positron emission tomography (PET), mat be adapted to investigate a variety of physiological phenomena, including the spatio-temporal kinetics of hESC engraftment and proliferation in living subjects. This chapter describes the applications of reporter gene imaging to accelerate basic science research and clinical studies involving hESCs through (1) isolation of a homogenous hESC population, (2) noninvasive, longitudinal tracking of the location and proliferation of hESCs administered to a living subject, and (3) ablation of the hESC graft in the event of cellular misbehavior.

  13. Photodynamic therapy by topical meso-tetraphenylporphinesulfonate tetrasodium salt administration in superficial basal cell carcinomas

    SciTech Connect

    Santoro, O.; Bandieramonte, G.; Melloni, E.; Marchesini, R.; Zunino, F.; Lepera, P.; De Palo, G. )

    1990-08-01

    The efficacy of an originally developed photodynamic approach, using topical administration of tetraphenylporphinesulfonate as the photosensitizer, was evaluated in a series of 292 basal cell carcinoma lesions (less than 2-mm thick) in 50 treated patients. The lack of indication for conventional therapies was the main selection criterion. The photosensitizing agent (2% solution) was topically applied at 0.1 ml/cm2, followed by light irradiation with a dye laser emitting at 645 nm (120 or 150 J/cm2). After initial treatment, all lesions responded, with 273 (93.5%) complete responses. Recurrences were observed in 29 (10.6%). A second application of photoradiation was performed in 15 persistent lesions and 11 relapsed lesions, producing 19/26 complete responses. Our results suggest that this technique can be considered a promising alternative treatment modality in selected cases of superficial basal cell carcinomas.

  14. 96-hour methamphetamine self-administration in male and female rats: a novel model of human methamphetamine addiction.

    PubMed

    Cornett, Elyse M; Goeders, Nicholas E

    2013-10-01

    Methamphetamine (MA) is a highly addictive psychostimulant drug of abuse for which no FDA-approved treatment exists. While high on MA, both male and female MA users report engaging in risky behaviors and are more likely to be involved in violent criminal activities and to engage in domestic and sexual violence. A unique aspect of MA is that it is typically used in binges. However, there is no animal model of MA self-administration that appears to represent a human MA self-administration binge. We recently developed a 96-hour MA self-administration paradigm in rats that more closely resembles how human MA users take the drug. Male and female rats were trained to self-administer MA for 96 consecutive hours for 5 weeks. Responding by female and male rats tended to escalate to binge-like behavior, as the animals responded continuously during their normal periods of activity as well as during their inactive periods for up to 72 h, followed by a crash of 6 or more hours. Thus, this 96-hour model of MA self-administration is a novel way to study MA addition in rats that may contribute to the development of improved treatments for recovering human MA users.

  15. Glycolate kinase activity in human red cells.

    PubMed

    Fujii, S; Beutler, E

    1985-02-01

    Human red cells manifest glycolate kinase activity. This activity copurifies with pyruvate kinase and is decreased in the red cells of subjects with hereditary pyruvate kinase deficiency. Glycolate kinase activity was detected in the presence of FDP or glucose-1,6-P2. In the presence of 1 mmol/L FDP, the Km for adenosine triphosphate (ATP) was 0.28 mmol/L and a half maximum velocity for glycolate was obtained at 40 mmol/L. The pH optimum of the reaction was over 10.5 With 10 mumol/L FDP, 500 mumol/L glucose-1,6-P2, 2 mmol/L ATP, 5 mmol/L MgCl2, and 50 mmol/L glycolate at pH 7.5, glycolate kinase activity was calculated to be approximately 0.0013 U/mL RBC. In view of this low activity even in the presence of massive amounts of glycolate, the glycolate kinase reaction cannot account for the maintenance of the reported phosphoglycolate level in human red cells.

  16. Naive Pluripotent Stem Cells Derived Directly from Isolated Cells of the Human Inner Cell Mass

    PubMed Central

    Guo, Ge; von Meyenn, Ferdinand; Santos, Fatima; Chen, Yaoyao; Reik, Wolf; Bertone, Paul; Smith, Austin; Nichols, Jennifer

    2016-01-01

    Summary Conventional generation of stem cells from human blastocysts produces a developmentally advanced, or primed, stage of pluripotency. In vitro resetting to a more naive phenotype has been reported. However, whether the reset culture conditions of selective kinase inhibition can enable capture of naive epiblast cells directly from the embryo has not been determined. Here, we show that in these specific conditions individual inner cell mass cells grow into colonies that may then be expanded over multiple passages while retaining a diploid karyotype and naive properties. The cells express hallmark naive pluripotency factors and additionally display features of mitochondrial respiration, global gene expression, and genome-wide hypomethylation distinct from primed cells. They transition through primed pluripotency into somatic lineage differentiation. Collectively these attributes suggest classification as human naive embryonic stem cells. Human counterparts of canonical mouse embryonic stem cells would argue for conservation in the phased progression of pluripotency in mammals. PMID:26947977

  17. Adherence of skin bacteria to human epithelial cells.

    PubMed Central

    Romero-Steiner, S; Witek, T; Balish, E

    1990-01-01

    Aerobic and anaerobic bacteria isolated from human axillae were tested for their capacity to adhere to buccal epithelial cells, immortalized human epithelial (HEp-2) cells, and undifferentiated and differentiated human epithelial cells. In general, both aerobic and anaerobic diphtheroids adhered better to differentiated human epithelial cells than to HEp-2 and undifferentiated human epithelial cells (P less than 0.05). Mannose, galactose, fucose, N-acetyl-D-glucosamine, and fibronectin were also assayed for their capacity to inhibit the adherence of diphtheroids to human epithelial cells. A great deal of variability was observed in the capacity of the latter compounds to inhibit the attachment of aerobic diphtheroids to undifferentiated and differentiated epithelial cells. Overall, mannose appeared to be best at inhibiting the adherence of the aerobic diphtheroids to undifferentiated human epithelial cells. Galactose, fucose, N-acetyl-D-glucosamine, and fibronectin showed a greater capacity to inhibit attachment of aerobic diphtheroids to differentiated than to undifferentiated human epithelial cells. The inhibition of adherence to differentiated human epithelial cells varied with the microorganism and the compound tested; however, the highest and most consistent inhibition of adherence (76.1 to 88.6%) was observed with a 5% solution of N-acetyl-D-glucosamine. The in vitro adherence and adherence inhibition assays presented here demonstrate that a number of adhesins and receptors are involved in the adherence of skin bacteria to human epithelial cells and receptors on human epithelial cells are apparently altered during differentiation. PMID:2298877

  18. Methyl jasmonate downregulates expression of proliferating cell nuclear antigen and induces apoptosis in human neuroblastoma cell lines.

    PubMed

    Tong, Qiang-Song; Jiang, Guo-Song; Zheng, Li-Duan; Tang, Shao-Tao; Cai, Jia-Bin; Liu, Yuan; Zeng, Fu-Qing; Dong, Ji-Hua

    2008-07-01

    Recent evidence indicates that methyl jasmonate, a plant stress hormone, exhibits anticancer activity on human cancer cells. Whether methyl jasmonate could inhibit the growth of human neuroblastoma cells still, however, remains largely unknown. In this study, administration of methyl jasmonate to cultured neuroblastoma cell lines, SK-N-SH and BE(2)-C, resulted in a decrease of cell viability in a dose-dependent and time-dependent manner as demonstrated by MTT colorimetry and colony formation assay. The results from RT-PCR indicated that the expression of proliferating cell nuclear antigen, but not of cyclin D1, was downregulated by methyl jasmonate. Accordingly, the cell cycle of methyl jasmonate-treated neuroblastoma cells was arrested at the G0/G1 phase. Moreover, incubation of SK-N-SH and BE(2)-C cells with methyl jasmonate resulted in characteristic changes of apoptosis, as demonstrated by acridine orange-ethidium bromide (AO/EB) staining, Hoechst 33258 staining and flow cytometry. Moreover, methyl jasmonate decreased the expression of the X-linked inhibitor of apoptosis protein and survivin, critical members of the inhibitors of apoptosis protein family, in neuroblastoma cells. These findings indicate that methyl jasmonate suppresses the growth of cultured human neuroblastoma cells associated with downregulation of proliferating cell nuclear antigen, and induces apoptosis accompanied by downregulation of the X-linked inhibitor of apoptosis protein and survivin, which lays the groundwork for further investigation into the mechanisms of methyl jasmonate-mediated anticancer activities.

  19. Human radiation studies: Remembering the early years. Oral history of Donner Lab Administrator Baird G. Whaley, August 15, 1994

    SciTech Connect

    1995-09-01

    Baird G. Whaley, Donner Lab Administrator, was interviewed by representatives of US DOE Office of Human Radiation Experiments (OHRE). The purpose of the interview was to capture the remembrances of Mr. Whaley concerning what he could relate on activities at the Donner Lab that pertain to the OHRE responsibilities. Following a brief biographical sketch, Mr. Whaley relates his experiences in administration at the LAB including funding activities, staffing concerns, intralaboraory politics, and remembrances of John Lawrence, John Gofman, Cornelius Tobias, Jim Born, Alex Margolis, B.V.A. Low- Beer, and Ed Alpen. Further patient care procedures for Donner Clinic Research Programs were discussed.

  20. 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.

  1. Feasibility and Efficacy of Autologous Bone Marrow Aspirate Transplantation Combined with Human Parathyroid Hormone 1-34 Administration to Treat Osteonecrosis in a Rabbit Model

    PubMed Central

    Sugaya, Hisashi; Aoto, Katsuya; Uemura, Kenta; Tanaka, Kenta; Akaogi, Hiroshi; Yamazaki, Masashi; Mishima, Hajime

    2017-01-01

    No studies have examined the transplantation of a bone marrow aspirate (BMA) containing mesenchymal stem cells (MSCs) combined with human parathyroid hormone 1-34 (hPTH1-34) administration. Therefore, we evaluated the feasibility and efficacy of autologous BMA transplantation combined with hPHT1-34 administration in a bone necrosis model. The metatarsal bones of rabbits were necrotized using liquid nitrogen, and the rabbits received a BMA transplantation or saline injection followed by hPTH1-34 (30 μg/kg) or saline administration three times per week (n = 3-4 per group). The rabbits were euthanized at 12 weeks after the initiation of treatment. No systemic adverse effects or local neoplastic lesions were observed. Importantly, the rabbits in the BMA transplantation plus hPTH1-34 group showed the highest bone volumes and histological scores of new bone. These data confirmed the feasibility of BMA transplantation combined with hPTH1-34, at least during the experimental period. The observed efficacy may be explained by a synergistic effect from the stimulation of MSC differentiation to osteoblasts with hPTH1-34-mediated suppression of apoptosis in osteoblasts. These results indicate the promising potential for BMA transplantation combined with hPTH1-34 administration in bone necrosis treatment. Longer term experiments are needed to confirm the safety of this therapeutic strategy. PMID:28386485

  2. Use of human epidermal cells in the study of carcinogenesis

    SciTech Connect

    Kuroki, T.; Chida, K.; Hosomi, J.; Kondo, S. )

    1989-05-01

    Because of the importance of human cells, particularly human epithelial cells, in cancer research, we have studied certain phases or events of carcinogenesis using human epidermal cells in primary culture. (1) We found that human epidermal cells are capable of metabolizing benzo(a)pyrene. Large inter-individual variations are found in the basal and induced arylhydrocarbon-hydroxylase activities. (2) UV-induced unscheduled DNA synthesis was demonstrated in human epidermal cells on autoradiographs. We also found that DNA repair is defective in epidermal cells isolated from xeroderma pigmentosum by a new explant-outgrowth culture. (3) Human epidermal cells are unique in that there is a large number of binding sites to phorbol esters compared with mouse epidermal cells, but there is no down-regulation. Further, human epidermal cells show essentially negative responses to tumor promoters, i.e., no stimulation of DNA synthesis, sugar uptake, and no induction of ornithine decarboxylase activity. (4) Human epidermal cells contain 1.5 x 10(5) binding sites per cell for epidermal growth factor (EGF), whereas squamous cell carcinomas of skin and oral cavity have larger amounts of EGF receptors in the order of 10(6) per cell. (5) Based on the above results, we attempted to transform human epidermal cells by the treatment with chemical carcinogens, but until now no transformation was obtained. 16 references.

  3. Growth requirements of human mammary epithelial cells in culture.

    PubMed

    Taylor-Papadimitriou, J; Shearer, M; Stoker, M G

    1977-12-15

    Colony-forming epithelial cells can be separated from the non-dividing "foam cells" in human milk by differential adhesion to glass and freezing. The growth of such partially purified mammary epithelial cells is stimulated by co-culture with non-dividing feeder cells. Foam cells, mitomycin-treated mouse fibroblast lines and human mammary fibroblasts and calf lens epithelial cells are all effective in promoting mammary epithelial cell growth. Contact between epithelial cells and feeders is not required for the growth-promoting effect. The mitogenic effect of epidermal growth factor on mammary epithelial cells also requires feeder cell activity.

  4. Evaluating the first-in-human clinical trial of a human embryonic stem cell-based therapy.

    PubMed

    Chapman, Audrey R; Scala, Courtney C

    2012-09-01

    Phase I clinical trials generally raise greater ethical and human protection challenges than later stage clinical trials, suggesting a need to proceed cautiously. This is particularly the case for Phase I trials with a novel therapy being tested in humans for the first time, usually termed first-in-human (FIH) trials. In January 2009, the Food and Drug Administration approved the Investigational New Drug application of Geron Corporation, a small California-based biopharmaceutical company, to initiate a clinical trial to assess GRNOPC1, a human embryonic stem cell-derived candidate therapy for severe spinal cord injuries. This article evaluates the ethical and human subject protection issues raised by the Geron FIH trial. It identifies problems with the approval process and with the conduct of the trial, and then recommends ways to improve review of future proposed trials with novel and high-risk therapies.

  5. Characterization of Human Mammary Epithelial Stem Cells

    DTIC Science & Technology

    2008-10-01

    9 Appendix……………………………………………………………………………… 10 Eirew,P., Stingl,J., Raouf,A., Turashvili,G., Aparicio ,S., Emerman,J.T., and Eaves,C.J. A...Peter Eirew, John Stingl, Afshin Raouf, Gulisa Turashvili, Samuel Aparicio , Joanne Emerman and Connie Eaves. A method for quantifying normal human...Eirew, Afshin Raouf, John Stingl, Gulisa Turashvili, Allen Delaney, Joanne Emerman, Marco Marra and Samuel Aparicio . “Stem Cells in the Mammary Gland

  6. Development of translational preclinical models in substance abuse: Effects of cocaine administration on cocaine choice in humans and non-human primates.

    PubMed

    Foltin, Richard W; Haney, Margaret; Rubin, Eric; Reed, Stephanie C; Vadhan, Nehal; Balter, Rebecca; Evans, Suzette M

    2015-07-01

    Human drug use involves repeated choices to take drugs or to engage in alternative behaviors. The purpose of this study was to examine how response cost for cocaine and the value of an alternative reinforcer (opportunity to play a game of chance) and how 'free' doses (with minimal response cost) affected cocaine choice. Two laboratory studies of cocaine self-administration were conducted in a group of humans who were habitual cocaine smokers and in a group of rhesus monkeys that intravenously self-administered cocaine. Nine human cocaine smokers who were not seeking treatment for their cocaine were repeatedly presented with the choice to smoke 25mg cocaine base or play a game of chance for a monetary bonus paid at study completion. The response cost for choosing cocaine varied (up to 4000 responses/dose) and the number of game plays varied (up to 8). In this sample of humans, increasing either the response cost for cocaine or increasing the value of the alternative reinforcer did not significantly affect cocaine choice, while increasing both simultaneously slightly decreased cocaine choice and increased choice of the alternative. In monkeys, the dose-response function for cocaine self-administration (10 choices of 0.0125-0.1mg/kg/infusion vs. candy coated chocolate) was steep and we failed to achieve a 50/50 cocaine/candy choice even after substantially manipulating cost and number of candies available. Providing a large 'free' self-administered cocaine dose to humans did not significantly affect cocaine choice, whereas in monkeys, a large free dose of cocaine decreased cocaine choice when higher doses of cocaine were available for self-administration. The present results demonstrate that in the laboratory, it is difficult to modify on-going cocaine self-administration behavior in both humans and non-human primates.

  7. Targeting eradication of malignant cells derived from human bone marrow mesenchymal stromal cells

    SciTech Connect

    Yang, Yingbin; Cai, Shaoxi; Yang, Li; Yu, Shuhui; Jiang, Jiahuan; Yan, Xiaoqing; Zhang, Haoxing; Liu, Lan; Liu, Qun; Du, Jun; Cai, Shaohui; Sung, K.L. Paul

    2010-12-10

    Human bone marrow mesenchymal stromal cells (hBMSC) have been shown to participate in malignant transformation. However, hampered by the low frequency of malignant transformation of hBMSC, we do not yet know how to prevent malignant transformation of implanted hBMSC. In this study, in order to establish a model for the eradication of hBMSC-derived malignant cells, a gene fusion consisting of a human telomerase (hTERT) promoter modified with both c-Myc and myeloid zinc finger protein2 (MZF-2) binding elements and followed by the E. coli cytosine deaminase (CD) and luciferase genes was stably transferred into hBMSC via lentiviral transduction; n-phosphonacelyl-L-aspartic acid (PALA) selection was used to generate malignant cell colonies derived from transduced hBMSC after treatment with the carcinogenic reagent BPDE. Cells that were amplified after PALA selection were used for transplantation and 5-FC pro-drug cytotoxicity tests. The results showed that PALA-resistant malignant cells could be generated from hBMSC co-induced with lentiviral transduction and treatment with Benzo(a)pyrene Diol Epoxide (BPDE); the modification of c-Myc and MZF-2 binding elements could remarkably enhance the transcriptional activities of the hTERT promoter in malignant cells, whereas transcriptional activity was depressed in normal hBMSC; malignant cells stably expressing CD under the control of the modified hTERT promoter could be eliminated by 5-FC administration. This study has provided a method for targeted eradication of malignant cells derived from hBMSC.

  8. Human embryonic stem cells and respect for life

    PubMed Central

    Meyer, J.

    2000-01-01

    The purpose of this essay is to stimulate academic discussion about the ethical justification of using human primordial stem cells for tissue transplantation, cell replacement, and gene therapy. There are intriguing alternatives to using embryos obtained from elective abortions and in vitro fertilisation to reconstitute damaged or dysfunctional human organs. These include the expansion and transplantation of latent adult progenitor cells. Key Words: Primordial stem cell research • embryonic stem cells • pluripotent stem cells • embryo research PMID:10860206

  9. Restriction of human adenovirus replication in Chinese hamster cell lines and their hybrids with human cells.

    PubMed

    Radna, R L; Foellmer, B; Feldman, L A; Francke, U; Ozer, H L

    1987-11-01

    We have found that the replication of human adenovirus (Ad2) is restricted in multiple Chinese hamster cell lines including CHO and V79. The major site of restriction involves differential accumulation of late viral proteins as demonstrated by immunofluorescence assay and polyacrylamide gel electrophoresis with and without prior immunoprecipitation. Synthesis of fiber and penton base are markedly reduced, whereas others, such as the 100K polypeptide, are synthesized efficiently. This pattern of restriction is similar to that previously reported for Ad2 infection of several monkey cell lines; however, the restriction is more marked in the Chinese hamster cell lines. The restriction is most likely due to a deficient cellular function since stable cell hybrids between V79 or CHO and human cells are permissive for virus replication. By analysis of a series of hybrids with reduced numbers of human chromosomes, fiber synthesis was correlated with the presence of the short arm of human chromosome 3. More hybrids showed restoration of fiber synthesis than production of progeny virus, suggesting that more than one unlinked function is required for the latter.

  10. Lipoprotein binding to cultured human hepatoma cells.

    PubMed Central

    Krempler, F; Kostner, G M; Friedl, W; Paulweber, B; Bauer, H; Sandhofer, F

    1987-01-01

    Binding of various 125I-lipoproteins to hepatic receptors was studied on cultured human hepatoma cells (Hep G2). Chylomicrons, isolated from a chylothorax, chylomicron remnants, hypertriglyceridemic very low-density lipoproteins, normotriglyceridemic very low-density lipoproteins (NTG-VLDL), their remnants, low-density lipoproteins (LDL), and HDL-E (an Apo E-rich high-density lipoprotein isolated from the plasma of a patient with primary biliary cirrhosis) were bound by high-affinity receptors. Chylomicron remnants and HDL-E were bound with the highest affinity. The results, obtained from competitive binding experiments, are consistent with the existence of two distinct receptors on Hep G2 cells: (a) a remnant receptor capable of high-affinity binding of triglyceride-rich lipoproteins and HDL-E, but not of Apo E free LDL, and (b) a LDL receptor capable of high-affinity binding of LDL, NTG-VLDL, and HDL-E. Specific binding of Apo E-free LDL was completely abolished in the presence of 3 mM EDTA, indicating that binding to the LDL receptor is calcium dependent. Specific binding of chylomicron remnants was not inhibited by the presence of even 10 mM EDTA. Preincubation of the Hep G2 cells in lipoprotein-containing medium resulted in complete suppression of LDL receptors but did not affect the remnant receptors. Hep G2 cells seem to be a suitable model for the study of hepatic receptors for lipoprotein in man. Images PMID:3038957

  11. Memory B Cells of Mice and Humans.

    PubMed

    Weisel, Florian; Shlomchik, Mark

    2017-01-30

    Wecomprehensively review memory B cells (MBCs), covering the definition of MBC and their identities and subsets, how MBCs are generated, where they are localized, how they are maintained, and how they are reactivated. Whereas naive B cells adopt multiple fates upon stimulation, MBCs are more restricted in their responses. Evolving work reveals that the MBC compartment in mice and humans consists of distinct subpopulations with differing effector functions. We discuss the various approaches to define subsets and subset-specific roles. A major theme is the need to both deliver faster effector function upon reexposure and readapt to antigenically variant pathogens while avoiding burnout, which would be the result if all MBCs generated only terminal effector function. We discuss cell-intrinsic differences in gene expression and signaling that underlie differences in function between MBCs and naive B cells and among MBC subsets and how this leads to memory responses. Expected final online publication date for the Annual Review of Immunology Volume 35 is April 26, 2017. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

  12. Anti-erythropoietin and anti-thrombopoietin antibodies induced after administration of recombinant human erythropoietin.

    PubMed

    Shin, Sug Kyun; Pack, Seung Pil; Oh, Jin-Gyo; Kang, Nam Kyu; Chang, Myung Hee; Chung, Yoon Hee; Kim, Sung-Jo; Lee, Jong Wook; Heo, Tae-Hwe

    2011-12-01

    Recombinant human erythropoietin (rhEPO) has been successfully used for correcting renal anemia. However, recent studies have raised some concerns about the safety of rhEPO treatment due to its immunogenic side effect - pure red cell aplasia (PRCA). We now report a case of development of anti-EPO neutralizing antibodies (Abs) implicated in thrombocytopenia as well as erythrocytopenia. A 35-year-old man had a history of administering rhEPO (epoetin alfa, epoetin beta and darbepoetin alfa) for 2years to treat renal anemia. The hematological parameters were collected. Anti-EPO, anti-platelet, and anti-thrombopoietin (TPO) Ab assays were performed to test the presence of autoreactive Abs. After performing antibody assays due to severe resistance to rhEPO treatment, a high titer of anti-EPO neutralizing Abs was detected. However, unexpectedly, this patient also showed thrombocytopenia rather than PRCA. We investigated the cause of the marked thrombocytopenia and found anti-TPO Abs in patient serum. To our best knowledge, this is the first report of the development of anti-TPO Abs during rhEPO treatment for anemia.

  13. Systemic Administration of Interleukin 2 Enhances the Therapeutic Efficacy of Dendritic Cell-Based Tumor Vaccines

    NASA Astrophysics Data System (ADS)

    Shimizu, K.; Fields, R. C.; Giedlin, M.; Mule, J. J.

    1999-03-01

    We have reported previously that murine bone marrow-derived dendritic cells (DC) pulsed with whole tumor lysates can mediate potent antitumor immune responses both in vitro and in vivo. Because successful therapy was dependent on host immune T cells, we have now evaluated whether the systemic administration of the T cell stimulatory/growth promoting cytokine interleukin-2 (IL-2) could enhance tumor lysate-pulsed DC-based immunizations to further promote protective immunity toward, and therapeutic rejection of, syngeneic murine tumors. In three separate approaches using a weakly immunogenic sarcoma (MCA-207), the systemic administration of non-toxic doses of recombinant IL-2 (20,000 and 40,000 IU/dose) was capable of mediating significant increases in the potency of DC-based immunizations. IL-2 could augment the efficacy of tumor lysate-pulsed DC to induce protective immunity to lethal tumor challenge as well as enhance splenic cytotoxic T lymphocyte activity and interferon-γ production in these treated mice. Moreover, treatment with the combination of tumor lysate-pulsed DC and IL-2 could also mediate regressions of established pulmonary 3-day micrometastases and 7-day macrometastases as well as established 14- and 28-day s.c. tumors, leading to either significant cure rates or prolongation in overall survival. Collectively, these findings show that nontoxic doses of recombinant IL-2 can potentiate the antitumor effects of tumor lysate-pulsed DC in vivo and provide preclinical rationale for the use of IL-2 in DC-based vaccine strategies in patients with advanced cancer.

  14. Co-transplantation of human hematopoietic stem cells and human breast cancer cells in NSG mice: a novel approach to generate tumor cell specific human antibodies.

    PubMed

    Wege, Anja K; Schmidt, Marcus; Ueberham, Elke; Ponnath, Marvin; Ortmann, Olaf; Brockhoff, Gero; Lehmann, Jörg

    2014-01-01

    Humanized tumor mice (HTM) were generated by the co-transplantation of human hematopoietic stem cells and human breast cancer cells overexpressing HER2 into neonatal NOD-scid IL2Rγ(null) (NSG) mice. These mice are characterized by the development of a human immune system in combination with human breast cancer growth. Due to concurrent transplantation into newborn mice, transfer of MHC-mismatched tumor cells resulted in solid coexistence and immune cell activation (CD4(+) T cells, natural killer cells, and myeloid cells), but without evidence for rejection. Histological staining of the spleen of HTM revealed co-localization of human antigen-presenting cells together with human T and B cells allowing MHC-dependent interaction, and thereby the generation of T cell-dependent antibody production. Here, we investigated the capability of these mice to generate human tumor-specific antibodies and correlated immunoglobulin titers with tumor outgrowth. We found detectable IgM and also IgG amounts in the serum of HTM, which apparently controlled tumor development when IgG serum concentrations were above 10 µg/ml. Western blot analyses revealed that the tumor-specific antibodies generated in HTM did not recognize HER2/neu antigens, but different, possibly relevant antigens for breast cancer therapy. In conclusion, HTM offer a novel approach to generate complete human monoclonal antibodies that do not require further genetic manipulation (e. g., humanization) for a potential application in humans. In addition, efficacy and safety of the generated antibodies can be tested in the same mouse model under human-like conditions. This might be of particular interest for cancer subtypes with no currently available antibody therapy.

  15. Mogoltacin enhances vincristine cytotoxicity in human transitional cell carcinoma (TCC) cell line.

    PubMed

    Behnam Rassouli, F; Matin, M M; Iranshahi, M; Bahrami, A R; Neshati, V; Mollazadeh, S; Neshati, Z

    2009-03-01

    Bladder cancer is the second common cancer of the genitourinary system throughout the world and intravesical chemotherapy is usually used to reduce tumour recurrence and progression. Human transitional cell carcinoma (TCC) is an epithelial-like adherent cell line originally established from primary bladder carcinoma. Here we report the effect of mogoltacin, a sesquiterpene coumarin from Ferula badrakema on TCC cells. Mogoltacin was isolated from the fruits of F. badrakema, using silica gel column chromatography and preparative thin layer chromatography. Mogoltacin did not have any significant cytotoxicity effect on neoplastic TCC cells at 16, 32, 64, 128, 200 and 600 microg ml(-1) concentrations. In order to analyse its combination effect, TCC cells were cultured in the presence of various combining concentrations of mogoltacin and vincristine. Cells were then observed for morphological changes (by light microscopy) and cytotoxicity using MTT assay. The effect of mogoltacin on vincristine toxicity was studied after 24, 48 and 72 h of drug administration. The results of MTT assay showed that mogoltacin can significantly enhance the cytotoxicity of vincristine and confirmed the morphological observations. Results revealed that combination of 40 microg ml(-1) vincristine with 16 microg ml(-1) mogoltacin increased the cytotoxicity of vincristine after 48 h by 32.8%.

  16. T helper cell activation and human retroviral pathogenesis.

    PubMed Central

    Copeland, K F; Heeney, J L

    1996-01-01

    T helper (Th) cells are of central importance in regulating many critical immune effector mechanisms. The profile of cytokines produced by Th cells correlates with the type of effector cells induced during the immune response to foreign antigen. Th1 cells induce the cell-mediated immune response, while Th2 cells drive antibody production. Th cells are the preferential targets of human retroviruses. Infections with human T-cell leukemia virus (HTLV) or human immunodeficiency virus (HIV) result in the expansion of Th cells by the action of HTLV (adult T-cell leukemia) or the progressive loss of T cells by the action of HIV (AIDS). Both retrovirus infections impart a high-level activation state in the host immune cells as well as systemically. However, diverging responses to this activation state have contrasting effects on the Th-cell population. In HIV infection, Th-cell loss has been attributed to several mechanisms, including a selective elimination of cells by apoptosis. The induction of apoptosis in HIV infection is complex, with many different pathways able to induce cell death. In contrast, infection of Th cells with HTLV-1 affords the cell a protective advantage against apoptosis. This advantage may allow the cell to escape immune surveillance, providing the opportunity for the development of Th-cell cancer. In this review, we will discuss the impact of Th-cell activation and general immune activation on human retrovirus expression with a focus upon Th-cell function and the progression to disease. PMID:8987361

  17. Human neural stem cells can target and deliver therapeutic genes to breast cancer brain metastases.

    PubMed

    Joo, Kyeung Min; Park, In H; Shin, Ji Y; Jin, Juyoun; Kang, Bong Gu; Kim, Mi Hyun; Lee, Se Jeong; Jo, Mi-young; Kim, Seung U; Nam, Do-Hyun

    2009-03-01

    The tumor-tropic properties of neural stem cells (NSCs) led to the development of a novel strategy for delivering therapeutic genes to tumors in the brain. To apply this strategy to the treatment of brain metastases, we made a human NSC line expressing cytosine deaminase (F3.CD), which converts 5-fluorocytosine (5-FC) into 5-fluorouracil, an anticancer agent. In vitro, the F3.CD cells significantly inhibited the growth of tumor cell lines in the presence of the prodrug 5-FC. In vivo, MDA-MB-435 human breast cancer cells were implanted into the brain of immune-deficient mouse stereotactically, and F3.CD cells were injected into the contralateral hemisphere followed by systemic 5-FC administration. The F3.CD cells migrated selectively into the brain metastases located in the opposite hemisphere and resulted in significantly reduced volumes. The F3.CD and 5-FC treatment also decreased both tumor volume and number of tumor mass significantly, when immune-deficient mouse had MDA-MB-435 cells injected into the internal carotid artery and F3.CD cells were transplanted into the contralateral brain hemisphere stereotactically. Taken together, brain transplantation of human NSCs, encoding the suicide enzyme CD, combined with systemic administration of the prodrug 5-FC, is an effective treatment regimen for brain metastases of tumors.

  18. Introduction: characterization and functions of human T regulatory cells.

    PubMed

    Romagnani, Sergio

    2005-06-01

    The field of human T regulatory (Treg) cells is a rapidly progressing, but still confused field of immunology. The effects of dendritic cell (DC) manipulation in Treg generation and the main features of human "natural" Treg cells, as well as of different populations of adaptive Treg subsets, are still partially unclear. However, it is clear that Treg cells play an important role in human diseases, such as autoimmune disorders, allergy, HIV infection, tumors and graft-versus-host disease.

  19. Biokinetics of (13)C in the human body after oral administration of (13)C-labeled glucose as an index for the biokinetics of (14)C.

    PubMed

    Masuda, Tsuyoshi; Tako, Yasuhiro; Matsushita, Kensaku; Takeda, Hiroshi; Endo, Masahiro; Nakamura, Yuji; Hisamatsu, Shun'ichi

    2016-09-01

    The retention of (13)C in the human body after oral administration of (13)C-labeled glucose was studied in three healthy volunteer subjects to estimate the 50 year cumulative body burden for (13)C as an index of the committed dose of the radioisotope (14)C. After administration of (13)C-labeled glucose, the volunteers ingested controlled diets with a fixed number of calories for 112 d. Samples of breath and urine were collected up to 112 d after administration. Samples of feces were collected up to 14 d after administration. Hair samples were obtained at 119 d after administration and analyzed as a representative index of the rate of excretion of organic (13)C via pathways such as skin cell exfoliation and mucus secretion. All samples were analyzed for (13)C/(12)C atomic ratio to determine the rate of excretion via each pathway. We then constructed a metabolic model with a total of four pathways (breath, urine, feces, and other) comprising seven compartments. We determined the values of the biokinetic parameters in the model by using the obtained excretion data. From 74% to 94% of the (13)C administered was excreted in breath, whereas  <2% was excreted in urine and feces. In the other pathway, the excretion rate constant in the compartment with the longest residence time stretched to hundreds of days but the rate constant for each subject was not statistically significant (P value  >  0.1). In addition, the dataset for one of the three subjects was markedly different from those of the other two. When we estimated the 50 year cumulative body burden for (13)C by using our model and we included non-statistically significant parameters, a considerable cumulative body burden was found in the compartments excreting to the other pathway. Although our results on the cumulative body burden of (13)C from orally administered carbon as glucose were inconclusive, we found that the compartments excreting to the other pathway had a markedly long residence time and

  20. Potential therapeutic effect of the secretome from human uterine cervical stem cells against both cancer and stromal cells compared with adipose tissue stem cells

    PubMed Central

    Seoane, Samuel; Bermúdez, María A.; Lamelas, Maria Luz; Garcia-Caballero, Tomás; Schneider, José; Perez-Fernandez, Roman; Vizoso, Francisco J.

    2014-01-01

    Evidences indicate that tumor development and progression towards a malignant phenotype depend not only on cancer cells themselves, but are also deeply influenced by tumor stroma reactivity. The present study uses mesenchymal stem cells from normal human uterine cervix (hUCESCs), isolated by the minimally invasive method of routine Pap cervical smear, to study their effect on the three main cell types in a tumor: cancer cells, fibroblasts and macrophages. Administration of hUCESCs-conditioned medium (CM) to a highly invasive breast cancer MDA-MB-231 cell line and to human breast tumors with high cell proliferation rates had the effect of reducing cell proliferation, modifying the cell cycle, inducing apoptosis, and decreasing invasion. In a xenograft mouse tumor model, hUCESCs-CM reduced tumor growth and increased overall survival. In cancer-associated fibroblasts, administration of hUCESCs-CM resulted in reduced cell proliferation, greater apoptosis and decreased invasion. In addition, hUCESCs-CM inhibited and reverted macrophage differentiation. The analysis of hUCESCs-CM (fresh and lyophilized) suggests that a complex paracrine signaling network could be implicated in the anti-tumor potential of hUCESCs. In light of their anti-tumor potential, the easy cell isolation method, and the fact that lyophilization of their CM conserves original properties make hUCESCs good candidates for experimental or clinical applications in anticancer therapy. PMID:25296979

  1. Potential therapeutic effect of the secretome from human uterine cervical stem cells against both cancer and stromal cells compared with adipose tissue stem cells.

    PubMed

    Eiró, Noemí; Sendon-Lago, Juan; Seoane, Samuel; Bermúdez, María A; Lamelas, Maria Luz; Garcia-Caballero, Tomás; Schneider, José; Perez-Fernandez, Roman; Vizoso, Francisco J

    2014-11-15

    Evidences indicate that tumor development and progression towards a malignant phenotype depend not only on cancer cells themselves, but are also deeply influenced by tumor stroma reactivity. The present study uses mesenchymal stem cells from normal human uterine cervix (hUCESCs), isolated by the minimally invasive method of routine Pap cervical smear, to study their effect on the three main cell types in a tumor: cancer cells, fibroblasts and macrophages. Administration of hUCESCs-conditioned medium (CM) to a highly invasive breast cancer MDA-MB-231 cell line and to human breast tumors with high cell proliferation rates had the effect of reducing cell proliferation, modifying the cell cycle, inducing apoptosis, and decreasing invasion. In a xenograft mouse tumor model, hUCESCs-CM reduced tumor growth and increased overall survival. In cancer-associated fibroblasts, administration of hUCESCs-CM resulted in reduced cell proliferation, greater apoptosis and decreased invasion. In addition, hUCESCs-CM inhibited and reverted macrophage differentiation. The analysis of hUCESCs-CM (fresh and lyophilized) suggests that a complex paracrine signaling network could be implicated in the anti-tumor potential of hUCESCs. In light of their anti-tumor potential, the easy cell isolation method, and the fact that lyophilization of their CM conserves original properties make hUCESCs good candidates for experimental or clinical applications in anticancer therapy.

  2. Implications of the new Food and Drug Administration draft guidance on human factors engineering for diabetes device manufacturers.

    PubMed

    Wilcox, Stephen B; Drucker, Daniel

    2012-03-01

    This article discusses the implications of the new Food and Drug Administration's draft guidance on human factors and usability engineering for the development of diabetes-related devices. Important considerations include the challenge of identifying users, when the user population is so dramatically broad, and the challenge of identifying use environments when the same can be said for use environments. Another important consideration is that diabetes-related devices, unlike many other medical devices, are used constantly as part of the user's lifestyle--adding complexity to the focus on human factors and ease of use emphasized by the draft guidance.

  3. The effect of cell phones on human health

    NASA Astrophysics Data System (ADS)

    Abu-Isbeih, Ibrahim N.; Saad, Dina

    2011-10-01

    The effect of cell phone radiation on human health is the subject of recent interest and study, as a result of the enormous increase in cell phone usage throughout the world. Cell phones use electromagnetic radiation in the microwave range, which some believe may be harmful to human health. Other digital wireless systems, such as data communication networks, produce similar radiation. The objective of this survey is to review the effects of cell phones on human health: A large body of research exists, both epidemiological and experimental, in non-human animals and in humans, of which the majority shows no definite causative relationship between exposure to cell phones and harmful biological effects in humans. This is often paraphrased simply as the balance of evidence showing no harm to humans from cell phones, although a significant number of individual studies do suggest such a relationship, or are inconclusive.

  4. Vascular endothelial growth factor stimulates osteoblastic differentiation of cultured human periosteal-derived cells expressing vascular endothelial growth factor receptors.

    PubMed

    Hah, Young-Sool; Jun, Jin-Su; Lee, Seong-Gyun; Park, Bong-Wook; Kim, Deok Ryong; Kim, Uk-Kyu; Kim, Jong-Ryoul; Byun, June-Ho

    2011-02-01

    Angiogenesis plays an important role in bone development and postnatal bone fracture repair. Vascular endothelial growth factor (VEGF) and vascular endothelial growth factor receptors (VEGFRs) are primarily involved in angiogenesis. This study investigated the expression of VEGF isoforms, VEGFR-1, and VEGFR-2 during the osteoblastic differentiation of cultured human periosteal-derived cells. In addition, the effect of exogenous VEGF on the osteoblastic differentiation of cultured human periosteal-derived cells was also examined. The expression of the VEGF isoforms (VEGF(121), VEGF(165), VEGF(189), and VEGF(206)), VEGFR-1, and VEGFR-2 was observed in the periosteal-derived cells. Administration of KRN633, a VEGFR-1 and VEGFR-2 inhibitor, decreased the alkaline phosphatase (ALP) activity during the osteoblastic differentiation of cultured human periosteal-derived cells. However, the administration of VEGFR2 Kinase Inhibitor IV, a VEGFR-2 inhibitor, did not affect the ALP activity. The addition of recombinant human VEGF(165) elevated the ALP activity and increased the calcium content in the periosteal-derived cells. Treating the periosteal-derived cells with recombinant human VEGF(165) resulted in an increase in Runx2 transactivation in the periosteal-derived cells. These results suggest that exogenous VEGF stimulates the osteoblastic differentiation of cultured human periosteal-derived cells and VEGF might act as an autocrine growth factor for the osteoblastic differentiation of cultured human periosteal-derived cells.

  5. Extracellular vesicle in vivo biodistribution is determined by cell source, route of administration and targeting.

    PubMed

    Wiklander, Oscar P B; Nordin, Joel Z; O'Loughlin, Aisling; Gustafsson, Ylva; Corso, Giulia; Mäger, Imre; Vader, Pieter; Lee, Yi; Sork, Helena; Seow, Yiqi; Heldring, Nina; Alvarez-Erviti, Lydia; Smith, C I Edvard; Le Blanc, Katarina; Macchiarini, Paolo; Jungebluth, Philipp; Wood, Matthew J A; Andaloussi, Samir El

    2015-01-01

    Extracellular vesicles (EVs) have emerged as important mediators of intercellular communication in a diverse range of biological processes. For future therapeutic applications and for EV biology research in general, understanding the in vivo fate of EVs is of utmost importance. Here we studied biodistribution of EVs in mice after systemic delivery. EVs were isolated from 3 different mouse cell sources, including dendritic cells (DCs) derived from bone marrow, and labelled with a near-infrared lipophilic dye. Xenotransplantation of EVs was further carried out for cross-species comparison. The reliability of the labelling technique was confirmed by sucrose gradient fractionation, organ perfusion and further supported by immunohistochemical staining using CD63-EGFP probed vesicles. While vesicles accumulated mainly in liver, spleen, gastrointestinal tract and lungs, differences related to EV cell origin were detected. EVs accumulated in the tumour tissue of tumour-bearing mice and, after introduction of the rabies virus glycoprotein-targeting moiety, they were found more readily in acetylcholine-receptor-rich organs. In addition, the route of administration and the dose of injected EVs influenced the biodistribution pattern. This is the first extensive biodistribution investigation of EVs comparing the impact of several different variables, the results of which have implications for the design and feasibility of therapeutic studies using EVs.

  6. [Changes of biogenic amines in the thymus cell system after T-activin administration].

    PubMed

    Iastrebova, S A; Sergeeva, V E; Spirin, I V

    2004-01-01

    The aim of this work was to study the dynamics of the neurotransmitter (cateholamine, serotonin, histamine) content in the thymic structures, as well as of heparin maturation degree and degranulation in the thymic mast cells population by means of Falck, Cross and Unna luminescent-histochemical methods 1-30 days following daily T-activinum injections. Concentration of the neurotransmitters in all the bioamine-containing thymic structures was found to fall by day 7, but after day 14 it began to increase, reaching a high level by day 30 that significantly exceeded the initial values. The degree of heparin maturation in mast cells was increased at days 21 and 30. Long-term T-activinum administration (for more than 7 days) is not justified, since an excessive increase in bioamine and heparin content suppresses immune reactions. Degranulating mast cells, surrounded by the zones of microenvironment, that were saturated with the bioamines capable of suppressing thymocytes, were found to predominate at the late stages of an experiment with T-activinum injections.

  7. Extracellular vesicle in vivo biodistribution is determined by cell source, route of administration and targeting

    PubMed Central

    Wiklander, Oscar P. B.; Nordin, Joel Z.; O’Loughlin, Aisling; Gustafsson, Ylva; Corso, Giulia; Mäger, Imre; Vader, Pieter; Lee, Yi; Sork, Helena; Seow, Yiqi; Heldring, Nina; Alvarez-Erviti, Lydia; Smith, CI Edvard; Le Blanc, Katarina; Macchiarini, Paolo; Jungebluth, Philipp; Wood, Matthew J. A.; Andaloussi, Samir EL

    2015-01-01

    Extracellular vesicles (EVs) have emerged as important mediators of intercellular communication in a diverse range of biological processes. For future therapeutic applications and for EV biology research in general, understanding the in vivo fate of EVs is of utmost importance. Here we studied biodistribution of EVs in mice after systemic delivery. EVs were isolated from 3 different mouse cell sources, including dendritic cells (DCs) derived from bone marrow, and labelled with a near-infrared lipophilic dye. Xenotransplantation of EVs was further carried out for cross-species comparison. The reliability of the labelling technique was confirmed by sucrose gradient fractionation, organ perfusion and further supported by immunohistochemical staining using CD63-EGFP probed vesicles. While vesicles accumulated mainly in liver, spleen, gastrointestinal tract and lungs, differences related to EV cell origin were detected. EVs accumulated in the tumour tissue of tumour-bearing mice and, after introduction of the rabies virus glycoprotein-targeting moiety, they were found more readily in acetylcholine-receptor-rich organs. In addition, the route of administration and the dose of injected EVs influenced the biodistribution pattern. This is the first extensive biodistribution investigation of EVs comparing the impact of several different variables, the results of which have implications for the design and feasibility of therapeutic studies using EVs. PMID:25899407

  8. Optimal administration routes for adipose-derived stem cells therapy in ischaemic flaps.

    PubMed

    Lee, Dong Won; Jeon, Yeo Reum; Cho, Eul Je; Kang, Jong Hwa; Lew, Dae Hyun

    2014-08-01

    Improvement of flap survival represents an ongoing challenge in reconstructive surgery. The angiogenic potential of adipose-derived stem cells (ASCs) offers a promising approach to improve the viability of random pattern flaps. Recently, to maximize the therapeutic effects of ASCs, increasing focus is being placed on how to deliver the stem cells to target lesions. The purpose of the present study was to compare the effectiveness of different administration routes of ASCs to improve the viability of the random pattern skin flap. ASCs labelled with PKH26 were applied via four methods to the cranially-based random pattern skin flaps of rats: (a) intravenous injection; (b) subcutaneous injection; (c) application with collagen sponge seeding; and (d) application with fibrin glue seeding. ASCs led to a significant increase in flap viability in the subcutaneous injection group and the collagen sponge group. Cutaneous blood flow was increased in the intravenous injection, subcutaneous injection and collagen sponge groups. Capillary density in the intravenous injection group and collagen sponge group was significantly greater than in the control group (no treatment). PKH26-positive cells via the collagen sponge were distributed more densely within the flap than in other groups. This study demonstrated that the collagen sponge method delivered ASCs most effectively within the flap and increased flap vascularity. The clinical therapeutic effects of ASCs can therefore be maximized when the optimal delivery route is chosen.

  9. Intravenous administration of puppy deciduous teeth stem cells in degenerative valve disease

    PubMed Central

    Petchdee, Soontaree; Sompeewong, Sarunya

    2016-01-01

    Aim: The objective of this study is to investigate the improvement of heart function in dogs with chronic valvular heart disease after puppy deciduous teeth stem cells (pDSCs) administration. Materials and Methods: 20 client-owned dogs with degenerative valvular heart disease underwent multiple intravenous injections of allogeneic pDSCs. Dogs were randomly assigned to two groups: (i) Control group (n=10) with standard treatment for heart failure and (ii) group with standard treatment and multiple administrations of pDSCs (n=10). Electrocardiography, complete transthoracic echocardiography, thoracic radiography, and blood pressure were recorded before and after pDSCs injections for 15, 30 and 60 days. Results: Post pDSCs injection showed measurable improvement in left ventricular ejection fraction, American College of Veterinary Internal Medicine (ACVIM) functional class significantly improved and improved quality of life scores were observed. In the control group, there were no significant enhancements in heart function or ACVIM class. Conclusions: This finding suggests that pDSCs could be a supplement for valvular heart disease treatment. PMID:28096616

  10. Trichloroethylene toxicity in a human hepatoma cell line

    SciTech Connect

    Thevenin, E.; McMillian, J.

    1994-12-31

    The experiments conducted in this study were designed to determine the usefullness of hepatocyte cultures and a human hepatoma cell line as model systems for assessing human susceptibility to hepatocellular carcinoma due to exposure to trichloroethylene. The results from these studies will then be analyzed to determine if human cell lines can be used to conduct future experiments of this nature.

  11. Human anti-murine immune response following administration of radiolabeled monoclonal antibodies

    SciTech Connect

    Reynolds, J.C.; Carrasquillo, J.C.; Larson, S.M.

    1985-05-01

    The author's purpose is to measure circulating anti-murine immunoglobulin antibodies (HAMA) in patients who previously received radiolabeled monoclonal antibodies (MoAb) for tumor imaging and therapy. Because the presence of HAMA may negate further use of MoAb in patients, it is important to determine the frequency and rate of HAMA development. Patients received radiolabeled MoAb Fab 96.5 (IgG2a), Fab 48.7 (IgG1), T101 (IgG2a), B72.3 (IgG1), 9.2.27 (IgG2a) and 791T/36 (IgG2b). HAMA was measured by incubating I-125 labeled 96.5, 48.7 or B72.3 with serum and isolating human IgG with Staphyloccocal protein A cells by centrifugation. The assays were capable of detecting HAMA concentrations which bound 20 ng/ml of monoclonal antibody. 12 of 37 patients who received IgG developed HAMA within 4 months of a single injection. For one patient this occurred as early as 1 week post injection. 2 of 18 patients who received Fab developed HAMA. One of these patients received multiple injections of MoAb. 2 of 3 patients who received IgG2B were positive for HAMA. There was no apparent difference in the positive HAMA when antibody or fragment was given SubQ or IV. The authors conclude that the use of IgG MoAb are more likely to lead to the development of antimurine immunoglobulin antibodies.

  12. Drug administration in animal studies of cardiac arrest does not reflect human clinical experience

    PubMed Central

    Reynolds, Joshua C.; Rittenberger, Jon C.; Menegazzi, James J.

    2007-01-01

    Introduction To date, there is no evidence showing a benefit from any advanced cardiac life support (ACLS) medication in out-of-hospital cardiac arrest (OOHCA), despite animal data to the contrary. One explanation may be a difference in the time to first drug administration. Our previous work has shown the mean time to first drug administration in clinical trials is 19.4 minutes. We hypothesized that the average time to drug administration in large animal experiments occurs earlier than in OOHCA clinical trials. Methods We conducted a literature review between 1990 and 2006 in MEDLINE using the following MeSH headings: swine, dogs, resuscitation, heart arrest, EMS, EMT, ambulance, ventricular fibrillation, drug therapy, epinephrine, vasopressin, amiodarone, lidocaine, magnesium, and sodium bicarbonate. We reviewed the abstracts of 331 studies and 197 full manuscripts. Exclusion criteria included: non-peer reviewed, all without primary animal data, and traumatic models. From these, we identified 119 papers that contained unique information on time to medication administration. The data are reported as mean, ranges, and 95% confidence intervals. Mean time to first drug administration in animal laboratory studies and clinical trials was compared with a t-test. Regression analysis was performed to determine if time to drug predicted ROSC. Results Mean time to first drug administration in 2378 animals was 9.5 minutes (range 3.0–28.0; 95% CI around mean 2.78, 16.22). This is less than the time reported in clinical trials (19.4 min, p<0.001). Time to drug predicted ROSC (Odds Ratio 0.844; 95% CI 0.738, 0.966). Conclusion Shorter drug delivery time in animal models of cardiac arrest may be one reason for the failure of animal studies to translate successfully into the clinical arena. PMID:17360097

  13. Adolescent nicotine administration alters serotonin receptors and cell signaling mediated through adenylyl cyclase.

    PubMed

    Xu, Z; Seidler, F J; Cousins, M M; Slikker, W; Slotkin, T A

    2002-10-04

    Nicotine is a neuroteratogen that targets synaptic function during critical developmental stages and recent studies indicate that CNS vulnerability extends into adolescence, the age at which smoking typically commences. We administered nicotine to adolescent rats via continuous minipump infusions from PN30 to PN47.5, using 6 mg/kg/day, a dose rate that replicates the plasma nicotine levels found in smokers, and examined 5HT receptors and related cell signaling during nicotine administration (PN45) and in the post-treatment period (PN50, 60, 75). Adolescent nicotine decreased 5HT(2) receptor binding in brain regions containing 5HT projections (hippocampus and cerebral cortex), with selectivity for females in the cerebral cortex; regions containing 5HT cell bodies showed either an increase (midbrain in males) or no change (brainstem). In contrast, there were no significant changes in 5HT(1A) receptors; however, the ability of the receptors to signal through adenylyl cyclase (AC) showed a switch from stimulatory to inhibitory effects in females during the post-treatment period. There were also transient alterations in AC responses to beta-adrenergic receptor stimulation, as well as pronounced induction of the AC response to the non-receptor-mediated stimulant, forskolin. Our results indicate that adolescent nicotine exposure alters the concentrations and functions of postsynaptic 5HT receptors in a manner commensurate with impaired 5HT synaptic function. The direction of change, emergence of defects after the cessation of nicotine administration, and sex-preference for effects in females, all support a relationship of impaired 5HT function to the higher incidence of depression seen in adolescent smokers.

  14. Microglia activation and cell death in response to diethyl-dithiocarbamate acute administration.

    PubMed

    Zucconi, Gigliola Grassi; Laurenzi, Maria Assunta; Semprevivo, Massimo; Torni, Federica; Lindgren, Jan Ake; Marinucci, Eva

    2002-04-29

    An increasing body of evidence suggests a role for activated microglia in the pathogenesis of neurodegenerative disorders. Hence, it would be useful to have a better understanding of the significance of microglial activation for neuronal damage. Unfortunately, most models of microglial activation use invasive or long-lasting insults, which make it difficult to evaluate the role played by microglia. We have instead developed a model for microglial activation by using brief exposure to the widely available neurotoxin diethyl-dithiocarbamate (DDTC). Despite evidence for the neurotoxic nature of this substance, microglia involvement has not been hitherto investigated. After acute i.p. administration of DDTC at two different doses, microglia were already activated in selected areas of the rat brain (hippocampal dentate gyrus, entorhinal-pyriform cortex and hypothalamus) after 1 hour, reaching a peak at 3-6 hours and subsided within 6-48 hours, depending on the brain region. Microglia activation was associated with interleukin-1 beta immunopositivity between 3 and 6 hours and with up-regulation of major histocompatibility complex class II expression between 24 and 48 hours. No significant changes in astrocyte immunostaining were detected between 6 hours and 6 days. The TUNEL procedure revealed the death of a limited number of cells in the above-mentioned structures that peaked at 6h and then declined rapidly. Cell death was detected in sites with major, minor, or no microglial activation, indicating that these two events can occur concomitantly or independently. The study shows that the administration of DDTC provides a useful model for studying the implications of region-specific reactivity of microglia and its differential interaction with neuronal damage.

  15. Alternative Sources of Adult Stem Cells: Human Amniotic Membrane

    NASA Astrophysics Data System (ADS)

    Wolbank, Susanne; van Griensven, Martijn; Grillari-Voglauer, Regina; Peterbauer-Scherb, Anja

    Human amniotic membrane is a highly promising cell source for tissue engineering. The cells thereof, human amniotic epithelial cells (hAEC) and human amniotic mesenchymal stromal cells (hAMSC), may be immunoprivileged, they represent an early developmental status, and their application is ethically uncontroversial. Cell banking strategies may use freshly isolated cells or involve in vitro expansion to increase cell numbers. Therefore, we have thoroughly characterized the effect of in vitro cultivation on both phenotype and differentiation potential of hAEC. Moreover, we present different strategies to improve expansion including replacement of animal-derived supplements by human platelet products or the introduction of the catalytic subunit of human telomerase to extend the in vitro lifespan of amniotic cells. Characterization of the resulting cultures includes phenotype, growth characteristics, and differentiation potential, as well as immunogenic and immunomodulatory properties.

  16. Prophylactic steroid administration for strictures after endoscopic resection of large superficial esophageal squamous cell carcinoma

    PubMed Central

    Kadota, Tomohiro; Yano, Tomonori; Kato, Tomoji; Imajoh, Maomi; Noguchi, Masaaki; Morimoto, Hiroyuki; Osera, Shozo; Yoda, Yusuke; Oono, Yasuhiro; Ikematsu, Hiroaki; Ohtsu, Atsushi; Kaneko, Kazuhiro

    2016-01-01

    Background and study aims: One of the major complications after endoscopic resection (ER) for large superficial esophageal squamous cell carcinoma (ESCC) is benign esophageal stricture, which can reduce quality of life even if ESCC achieves a cure without organ resection. Recently, steroid administration has been reported as a prophylactic treatment to prevent esophageal strictures. This retrospective study evaluated the stricture rate according to the different width of mucosal defects due to ER and compared it to that seen with prophylactic steroid administration. Patients and methods: Between June 2007 and December 2013, we enrolled patients with ESCC who had 3/4 or larger circumferential mucosal defects due to ER. In December 2009, steroid injections (triamcinolone acetonide 50 mg) into the ulcer bed due to ER were introduced. Beginning in November 2012, we commenced oral steroid administration (prednisolone 30 mg/day, tapered gradually for 8 weeks) in addition to steroid injection. Patients were classified into 3 groups according to the width of mucosal defect after ER (Group A, ≥ 3/4 and < 7/8; Group B, ≥ 7/8 and less than the entire circumference; and Group C, the entire circumference). We retrospectively evaluated the stricture rate by comparing no treatment, steroid injection, or steroid injection followed by oral steroid according to the width of mucosal defect. Results: A total of 115 patients met the selection criteria. In Group B, no treatment had a significantly higher stricture rate (100 %, vs. steroid injection: 56 % P = 0.015; vs steroid injection followed by oral steroid: 20 % P < 0.001). Conversely, in Group C, the stricture rate was high, regardless of treatment (no treatment: 100 %; steroid injection: 100 %; steroid injection followed by oral steroid: 71 %). Conclusions: Although prophylactic steroid administration is effective to prevent strictures for 7/8 circumference or larger mucosal defects, it is

  17. Interleukin 7 independent development of human B cells.

    PubMed Central

    Prieyl, J A; LeBien, T W

    1996-01-01

    Mammalian hematopoietic stem cell (HSC) commitment and differentiation into lymphoid lineage cells proceed through a series of developmentally restricted progenitor compartments. A complete understanding of this process, and how it differs from HSC commitment and differentiation into cells of the myeloid/erythroid lineages, requires the development of model systems that support HSC commitment to the lymphoid lineages. We now describe a human bone marrow stromal cell culture that preferentially supports commitment and differentiation of human HSC to CD19+ B-lineage cells. Fluorescence activated cell sorterpurified CD34++/lineage-cells were isolated from fetal bone marrow and cultured on human fetal bone marrow stromal cells in serum-free conditions containing no exogenous cytokines. Over a period of 3 weeks, CD34++/lineage- cells underwent commitment, differentiation, and expansion into the B lineage. Progressive changes included: loss of CD34, acquisition of and graded increases in the level of cell surface CD19, and appearance of immature B cells expressing mu/kappa or mu/lambda cell surface Ig receptors. The tempo and phenotype of B-cell development was not influenced by the addition of IL-7 (10 ng/ml), or by the addition of goat anti-IL-7 neutralizing antibody. These results indicate a profound difference between mouse and human in the requirement for IL-7 in normal B-cell development, and provide an experimental system to identify and characterize human bone marrow stromal cell-derived molecules crucial for human B lymphopoiesis. PMID:8816803

  18. Polysialic acid in human neuroblastoma cells

    SciTech Connect

    Livingston, B.D.; Jacobs, J.; Shaw, G.W.; Glick, M.C.; Troy, F.A.

    1987-05-01

    Prokaryotic-derived probes that specifically detect ..cap alpha..-2,8-linked polysialic acid (PSA) units on embryonic neural cell adhesion molecules (N-CAM) were used to show that membrane glycoproteins (GPs) from metastatic human neuroblastoma cells (CHP-134) also contain these unique carbohydrate moieties. This conclusion was based on the following evidence: (1) membranes from CHP-134 cells served as an exogenous acceptor of (/sup 14/C)NeuNAc units in an E. coli K1 sialyltransferase (ST) assay. The bacterial ST is specific for the transfer of (/sup 14/C)NeuNAc to exogenous acceptors containing at least 3 sialyl units (DP3); (2) in SDS-PAGE, the (/sup 14/C)NeuNAc-labeled CHP-134 membranes showed a major peak of radioactivity that was polydisperse. N-CAM shows a similar Mr heterogeneity; (3) treatment of the high Mr CHP-134 product with Endo-N-acetylneuraminidase (Endo-N) released the (/sup 14/C)NeuNAc label as a DP4. Endo-N is specific for hydrolysing ..cap alpha..-2,8-linked PSA chains containing a minimum of 5 sialyl residues; (4) treatment of the DP4 with sialidase converted the label to (/sup 14/C)NeuNAc, thus proving the tetramer contained sialic acid; (5) CHP-134 cells were labeled in vivo with (/sup 3/H)GlcN. A glycopeptide fraction representing ca. 1% of the (/sup 3/H)GlcN incorporated was isolated. Based on Endo-N sensitivity, this glycopeptide contained at least 15-20% of the (/sup 3/H)GlcN label as PSA. Endo-N digestion of the (/sup 3/H)-labeled glycopeptide released (/sup 3/H)-DP4. These results suggest that the surface expression of PSA-containing GPs may be important in neuroblastoma metastasis.

  19. Modulation and the Underlying Mechanism of T Cells in Thymus of Mice by Oral Administration of Sodium Fluoride.

    PubMed

    Yin, Songna; Wu, Haibo; Song, Chao; Chen, Xin; Zhang, Yong

    2016-03-01

    The underlying mechanism of thymic T cell regulation has been a hot topic of research in recent years. Fluoride is toxic at high concentrations and fluoride toxicity to thymic T cells was assessed in our study. To explore T cell responses to excess fluoride, different concentrations of fluoride were uptake by mice for 6 weeks. The expression of genes, including Foxn1, Cbx4, DLL4, and IL-7 gene, associated with the development and differentiation of T cells in thymic epithelial cells(TECs) was lower in the experimental groups than that in the control group. The percentages of CD4(+) and CD8(+) T cells that decreased with the fluoride administration were confirmed by flow cytometry. The mRNA levels of immunoregulatory cytokines IL-2 and IL-10, which participate in T cell proliferation, also declined in the experimental groups as compared with the control group. Expression of the T cell function-related genes CD2, PTPRC, CD69, and CD101, which are involved in thymic function in mice, decreased with the fluoride administration. Our findings suggest that the administration of high concentrations of fluoride to mice induces a decrease in CD4(+) and CD8(+) thymus T cells by harming TECs leading to the dysfunction of the thymus by altering the expression of T cell function-related genes and immunoregulatory cytokine production.

  20. Dynamic behaviour of human neuroepithelial cells in the developing forebrain

    PubMed Central

    Subramanian, Lakshmi; Bershteyn, Marina; Paredes, Mercedes F.; Kriegstein, Arnold R.

    2017-01-01

    To understand how diverse progenitor cells contribute to human neocortex development, we examined forebrain progenitor behaviour using timelapse imaging. Here we find that cell cycle dynamics of human neuroepithelial (NE) cells differ from radial glial (RG) cells in both primary tissue and in stem cell-derived organoids. NE cells undergoing proliferative, symmetric divisions retract their basal processes, and both daughter cells regrow a new process following cytokinesis. The mitotic retraction of the basal process is recapitulated by NE cells in cerebral organoids generated from human-induced pluripotent stem cells. In contrast, RG cells undergoing vertical cleavage retain their basal fibres throughout mitosis, both in primary tissue and in older organoids. Our findings highlight developmentally regulated changes in mitotic behaviour that may relate to the role of RG cells to provide a stable scaffold for neuronal migration, and suggest that the transition in mitotic dynamics can be studied in organoid models. PMID:28139695

  1. Remote ischemic postconditioning enhances cell retention in the myocardium after intravenous administration of bone marrow mesenchymal stromal cells.

    PubMed

    Jiang, Qin; Song, Peng; Wang, Enshi; Li, Jun; Hu, Shengshou; Zhang, Hao

    2013-03-01

    Efficacy of intravenous administration of mesenchymal stromal cells (MSCs) for myocardial infarction (MI) is limited by low cell retention in the damaged myocardium. Previous studies indicated that remote ischemic conditioning could protect against ischemia-reperfusion-induced injury by release of various cytokines including stromal cell derived factor-1 alpha (SDF-1α). However, whether remote ischemic postconditioning (RIPostC) can also enhance the retention of infused cells in the myocardium by activating MSC homing is unclear. In this study, RIPostC was induced with 4cycles of 5min occlusion and reperfusion of the abdominal aorta in female Sprague-Dawley (SD) rats which underwent ligation of the coronary artery 1week previously. Cytokine levels in serum and myocardium were evaluated by enzyme-linked immunosorbent assay (ELISA) at 1, 6, 24 and 48h after RIPostC. Then, a total of 4×10(6) male MSCs were infused intravenously at 24h after RIPostC. The number of survived cells in the myocardium was evaluated by real-time polymerase chain reaction analysis for Y chromosome and the heart function was evaluated by echocardiography at 1month after cell infusion. Furthermore, 10μg/kg rabbit anti-rat CXCR4 polyclonal antibody was injected intraperitoneally to prove the role of SDF-1α for RIPostC. RIPostC induced an increase in SDF-1α in serum at 1h and enhanced SDF-1α transcription and protein synthesis in the myocardium at 24h after the procedure. 1month after cell transplantation, RIPostC significantly increased MSC myocardial retention by 79.1±12.3% and thereby contributed to enhanced cardiac function in comparison with cell transplantation without RIPostC. Furthermore, blockade with a CXCR4-specific antibody after RIPostC markedly attenuated the enhancement of therapeutic efficacy. We conclude that RIPostC activated SDF-1α expression and enhanced retention of the infused MSCs in the injured myocardium. Priming of the heart with RIPostC might be a novel

  2. Plasma membrane proteomics of human embryonic stem cells and human embryonal carcinoma cells.

    PubMed

    Dormeyer, Wilma; van Hoof, Dennis; Braam, Stefan R; Heck, Albert J R; Mummery, Christine L; Krijgsveld, Jeroen

    2008-07-01

    Human embryonic stem cells (hESCs) are of immense interest in regenerative medicine as they can self-renew indefinitely and can give rise to any adult cell type. Human embryonal carcinoma cells (hECCs) are the malignant counterparts of hESCs found in testis tumors. hESCs that have acquired chromosomal abnormalities in culture are essentially indistinguishable from hECC. Direct comparison of karyotypically normal hESCs with hECCs could lead to understanding differences between their mechanisms of growth control and contribute to implementing safe therapeutic use of stem cells without the development of germ cell cancer. While several comparisons of hECCs and hESCs have been reported, their cell surface proteomes are largely unknown, partly because plasma membrane proteomics is still a major challenge. Here, we present a strategy for the identification of plasma membrane proteins that has been optimized for application to the relatively small numbers of stem cells normally available, and that does not require tedious cell fractionation. The method led to the identification of 237 and 219 specific plasma membrane proteins in the hESC line HUES-7 and the hECC line NT2/D1, respectively. In addition to known stemness-associated cell surface markers like ALP, CD9, and CTNNB, a large number of receptors, transporters, signal transducers, and cell-cell adhesion proteins were identified. Our study revealed that several Hedgehog and Wnt pathway members are differentially expressed in hESCs and hECCs including NPC1, FZD2, FZD6, FZD7, LRP6, and SEMA4D, which play a pivotal role in stem cell self-renewal and cancer growth. Various proteins encoded on chromosome 12p, duplicated in testicular cancer, were uniquely identified in hECCs. These included GAPDH, LDHB, YARS2, CLSTN3, CSDA, LRP6, NDUFA9, and NOL1, which are known to be upregulated in testicular cancer. Distinct HLA molecules were revealed on the surface of hESCs and hECCs, despite their low abundance. Results were

  3. Human Stem Cell Derived Cardiomyocytes: An Alternative ...

    EPA Pesticide Factsheets

    Chemical spills and associated deaths in the US has increased 2.6-fold and 16-fold from 1983 to 2012, respectfully. In addition, the number of chemicals to which humans are exposed to in the environment has increased almost 10-fold from 2001 to 2013 within the US. Internationally, a WHO report on the global composite impact of chemicals on health reported that 16% of the total burden of cardiovascular disease was attributed to environmental chemical exposure with 2.5 million deaths per year. Clearly, the cardiovascular system, at all its various developmental and life stages, represents a critical target organ system that can be adversely affected by existing and emerging chemicals (e.g., engineered nanomaterials) in a variety of environmental media. The ability to assess chemical cardiac risk and safety is critically needed but extremely challenging due to the number and categories of chemicals in commerce, as indicated. This presentation\\session will evaluate the use of adult human stem cell derived cardiomyocytes, and existing platforms, as an alternative model to evaluate environmental chemical cardiac toxicity as well as provide key information for the development of predictive adverse outcomes pathways associated with environmental chemical exposures. (This abstract does not represent EPA policy) Rapid and translatable chemical safety screening models for cardiotoxicity current status for informing regulatory decisions, a workshop sponsored by the Society

  4. Human cementum tumor cells have different features from human osteoblastic cells in vitro.

    PubMed

    Arzate, H; Alvarez-Pérez, M A; Aguilar-Mendoza, M E; Alvarez-Fregoso, O

    1998-07-01

    Cells obtained from human cementoblastoma and alveolar bone were isolated and cultured. Initial and late stages of mineralization were assessed by using atomic force microscopy, scanning electron microscopy and X-ray microanalysis. In cultures of cementoblastoma-derived cells the initial stages of mineralization showed well-defined spherical-shaped structures, while the osteoblastic cells showed plaque-like deposits. These morphological patterns of mineral deposition could serve as nucleation centers for hydroxyapatite crystals. Late stages of mineralization at 28 and 35 d maintained those morphological differences established in initial cultures. The material deposited by cementoblastoma and osteoblastic cells, analyzed by EDX spectra, revealed similar Ca/P ratios for both cell types. These values were similar to those reported for hydroxyapatite in enamel and bone. Alkaline phosphatase specific activity (AlP), of osteoblastic cells at 3, 7 and 11 d, showed an increase of 27.9, 50.9 and 37.0% (p < 0.001), respectively. However, at 15 and 19 d there was an increase of AlP activity of cementoblastoma cells by 39.4 and 34.5% over osteoblastic cells (p < 0.001). Immunostaining of cementoblastoma and osteoblastic cells using a specific mAb against a cementum-derived attachment protein revealed strong immunostaining of cementoblastoma cells which was localized to the cell membrane and fibril-like structures (96.2 +/- 1.3). A few osteoblastic cells also stained weakly with the anti-CAP mAb (6.4 +/- 0.6). Sections of decalcified paraffin embedded cementoblastoma specimens, when immunostained with anti-CAP mAb, showed strong immunostaining of the cells surrounding the regular and irregularly-shaped calcified masses of the tumor. Putative cementocytes also stained positively. Immunostaining with a polyclonal antibody against osteopontin strongly stained the osteoblastic cells (89.0 +/- 3.6). Cementoblastoma cells showed weaker staining (54.2 +/- 2.4). The results suggest

  5. Effect of vitamin C administration on hydrogen peroxide-induced cytotoxicity in periodontal ligament cells.

    PubMed

    Wu, Wenlei; Yang, Nanfei; Feng, Xiujing; Sun, Tingzhe; Shen, Pingping; Sun, Weibin

    2015-01-01

    Periodontitis is a disease, which is associated with chronic inflammation and leads to significant destruction of periodontal tissues. Periodontal ligament cells (PDLCs) constitute the largest cell population in PDL tissues and a considerable body of evidence has demonstrated an association between oxidative stress and the progression of periodontitis. However, the effects on PDLCs exposed to hydrogen peroxide (H2O2) and the molecular mechanisms by which H2O2 affects periodontitis remain to be elucidated. In the present study, the potential cytotoxic effect of H2O2 and the antioxidative function of vitamin C (Vc) in PDLCs were investigated. The results demonstrated that H2O2 treatment decreased the viability of PDLCs. The decreased PDLC viability was primarily induced by apoptosis, which was evidenced by cleaved caspases-3, caspases-9 and poly (ADP-ribose) polymerase. Following optimal Vc addition, the proapoptotic effects of H2O2 were partially antagonized. Taken together, the present study demonstrated that H2O2 primarily induced the apoptosis of PDLCs and that these adverse effects were partially rescued following treatment with Vc. These results revealed how H2O2 promotes the progression of periodontitis and provide an improved understanding of the reversal effect of antioxidant treatment. Therefore, optimal Vc administration may provide a potentially effective technique in periodontal therapy.

  6. In vivo administration of artificial antigen-presenting cells activates low-avidity T cells for treatment of cancer.

    PubMed

    Ugel, Stefano; Zoso, Alessia; De Santo, Carmela; Li, Yu; Marigo, Ilaria; Zanovello, Paola; Scarselli, Elisa; Cipriani, Barbara; Oelke, Mathias; Schneck, Jonathan P; Bronte, Vincenzo

    2009-12-15

    The development of effective antitumor immune responses is normally constrained by low-avidity, tumor-specific CTLs that are unable to eradicate the tumor. Strategies to rescue antitumor activity of low-avidity melanoma-specific CTLs in vivo may improve immunotherapy efficacy. To boost the in vivo effectiveness of low-avidity CTLs, we immunized mice bearing lung melanoma metastases with artificial antigen-presenting cells (aAPC), made by covalently coupling (pep)MHC-Ig dimers and B7.1-Ig molecules to magnetic beads. aAPC treatment induced significant tumor reduction in a mouse telomerase antigen system, and complete tumor eradication in a mouse TRP-2 antigen system, when low-avidity CTLs specific for these antigens were adoptively transferred. In addition, in an in vivo treatment model of subcutaneous melanoma, aAPC injection also augmented the activity of adoptively transferred CTLs and significantly delayed tumor growth. In vivo tumor clearance due to aAPC administration correlated with in situ proliferation of the transferred CTL. In vitro studies showed that aAPC effectively stimulated cytokine release, enhanced CTL-mediated lysis, and TCR downregulation in low-avidity CTLs. Therefore, in vivo aAPC administration represents a potentially novel approach to improve cancer immunotherapy.

  7. Administration of interleukin-7 increases CD4 T cells in idiopathic CD4 lymphocytopenia

    PubMed Central

    Porter, Brian O.; DerSimonian, Rebecca; Kovacs, Stephen B.; Thompson, William L.; Perez-Diez, Ainhoa; Freeman, Alexandra F.; Roby, Gregg; Mican, JoAnn; Pau, Alice; Rupert, Adam; Adelsberger, Joseph; Higgins, Jeanette; Bourgeois, Jeffrey S.; Jensen, Stig M. R.; Morcock, David R.; Burbelo, Peter D.; Osnos, Leah; Maric, Irina; Natarajan, Ven; Croughs, Therese; Yao, Michael D.; Estes, Jacob D.; Sereti, Irini

    2016-01-01

    Idiopathic CD4 lymphopenia (ICL) is a rare syndrome defined by low CD4 T-cell counts (<300/µL) without evidence of HIV infection or other known cause of immunodeficiency. ICL confers an increased risk of opportunistic infections and has no established treatment. Interleukin-7 (IL-7) is fundamental for thymopoiesis, T-cell homeostasis, and survival of mature T cells, which provides a rationale for its potential use as an immunotherapeutic agent for ICL. We performed an open-label phase 1/2A dose-escalation trial of 3 subcutaneous doses of recombinant human IL-7 (rhIL-7) per week in patients with ICL who were at risk of disease progression. The primary objectives of the study were to assess safety and the immunomodulatory effects of rhIL-7 in ICL patients. Injection site reactions were the most frequently reported adverse events. One patient experienced a hypersensitivity reaction and developed non-neutralizing anti-IL-7 antibodies. Patients with autoimmune diseases that required systemic therapy at screening were excluded from the study; however, 1 participant developed systemic lupus erythematosus while on study and was excluded from further rhIL-7 dosing. Quantitatively, rhIL-7 led to an increase in the number of circulating CD4 and CD8 T cells and tissue-resident CD3 T cells in the gut mucosa and bone marrow. Functionally, these T cells were capable of producing cytokines after mitogenic stimulation. rhIL-7 was well tolerated at biologically active doses and may represent a promising therapeutic intervention in ICL. This trial was registered at www.clinicaltrials.gov as #NCT00839436. PMID:26675348

  8. Reversine Induced Multinucleated Cells, Cell Apoptosis and Autophagy in Human Non-Small Cell Lung Cancer Cells

    PubMed Central

    Lin, Ching-Yen; Chen, Yih-Yuan; Chen, Ping-Tzu; Tseng, Ya-Shih

    2016-01-01

    Reversine, an A3 adenosine receptor antagonist, has been shown to induce differentiated myogenic-lineage committed cells to become multipotent mesenchymal progenitor cells. We and others have reported that reversine has an effect on human tumor suppression. This study revealed anti-tumor effects of reversine on proliferation, apoptosis and autophagy induction in human non-small cell lung cancer cells. Treatment of these cells with reversine suppressed cell growth in a time- and dosage-dependent manner. Moreover, polyploidy occurred after reversine treatment. In addition, caspase-dependent apoptosis and activation of autophagy by reversine in a dosage-dependent manner were also observed. We demonstrated in this study that reversine contributes to growth inhibition, apoptosis and autophagy induction in human lung cancer cells. Therefore, reversine used as a potential therapeutic agent for human lung cancer is worthy of further investigation. PMID:27385117

  9. Folate antagonist, methotrexate induces neuronal differentiation of human embryonic stem cells transplanted into nude mouse retina.

    PubMed

    Hara, Akira; Taguchi, Ayako; Aoki, Hitomi; Hatano, Yuichiro; Niwa, Masayuki; Yamada, Yasuhiro; Kunisada, Takahiro

    2010-06-25

    Transplanted embryonic stem (ES) cells can be integrated into the retinas of adult mice as well-differentiated neuroretinal cells. However, the transplanted ES cells also have a tumorigenic activity as they have the ability for multipotent differentiation to various types of tissues. In the present study, human ES (hES) cells were transplanted into adult nude mouse retinas by intravitreal injections 20 h after intravitreal N-methyl-D-aspartate (NMDA) administration. After the transplantation of hES cells, the folate antagonist, methotrexate (MTX) was administrated in order to control the differentiation of the transplanted hES cells. Neuronal differentiation and teratogenic potential of hES cells were examined immunohistochemically 5 weeks after transplantation. The proliferative activity of transplanted cells was determined by both the mitotic index and the Ki-67 proliferative index. Disappearance of Oct-4-positive hES cells showing undifferentiated morphology was observed after intraperitoneal MTX treatment daily, for 15 days. Decreased mitotic and Ki-67 proliferative indices, and increased neuronal differentiation were detected in the surviving hES cells after the MTX treatment. These results suggest two important effects of intraperitoneal MTX treatment for hES cells transplanted into nude mouse retina: (1) MTX treatment following transplantation induces neuronal differentiation, and (2) MTX decreases proliferative activity and tumorigenic potential.

  10. Molecular aging and rejuvenation of human muscle stem cells

    PubMed Central

    Carlson, Morgan E; Suetta, Charlotte; Conboy, Michael J; Aagaard, Per; Mackey, Abigail; Kjaer, Michael; Conboy, Irina

    2009-01-01

    Very little remains known about the regulation of human organ stem cells (in general, and during the aging process), and most previous data were collected in short-lived rodents. We examined whether stem cell aging in rodents could be extrapolated to genetically and environmentally variable humans. Our findings establish key evolutionarily conserved mechanisms of human stem cell aging. We find that satellite cells are maintained in aged human skeletal muscle, but fail to activate in response to muscle attrition, due to diminished activation of Notch compounded by elevated transforming growth factor beta (TGF-β)/phospho Smad3 (pSmad3). Furthermore, this work reveals that mitogen-activated protein kinase (MAPK)/phosphate extracellular signal-regulated kinase (pERK) signalling declines in human muscle with age, and is important for activating Notch in human muscle stem cells. This molecular understanding, combined with data that human satellite cells remain intrinsically young, introduced novel therapeutic targets. Indeed, activation of MAPK/Notch restored ‘youthful’ myogenic responses to satellite cells from 70-year-old humans, rendering them similar to cells from 20-year-old humans. These findings strongly suggest that aging of human muscle maintenance and repair can be reversed by ‘youthful’ calibration of specific molecular pathways. PMID:20049743

  11. Human embryonic stem cells vs human induced pluripotent stem cells for cardiac repair.

    PubMed

    Barad, Lili; Schick, Revital; Zeevi-Levin, Naama; Itskovitz-Eldor, Joseph; Binah, Ofer

    2014-11-01

    Human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) have the capacity to differentiate into any specialized cell type, including cardiomyocytes. Therefore, hESC-derived and hiPSC-derived cardiomyocytes (hESC-CMs and hiPSC-CMs, respectively) offer great potential for cardiac regenerative medicine. Unlike some organs, the heart has a limited ability to regenerate, and dysfunction resulting from significant cardiomyocyte loss under pathophysiological conditions, such as myocardial infarction (MI), can lead to heart failure. Unfortunately, for patients with end-stage heart failure, heart transplantation remains the main alternative, and it is insufficient, mainly because of the limited availability of donor organs. Although left ventricular assist devices are progressively entering clinical practice as a bridge to transplantation and even as an optional therapy, cell replacement therapy presents a plausible alternative to donor organ transplantation. During the past decade, multiple candidate cells were proposed for cardiac regeneration, and their mechanisms of action in the myocardium have been explored. The purpose of this article is to critically review the comprehensive research involving the use of hESCs and hiPSCs in MI models and to discuss current controversies, unresolved issues, challenges, and future directions.

  12. Oral administration of Enterococcus faecalis FK-23 suppresses Th17 cell development and attenuates allergic airway responses in mice.

    PubMed

    Zhang, Bei; An, Jun; Shimada, Takashi; Liu, Shuang; Maeyama, Kazutaka

    2012-08-01

    Evidence is increasing that oral administration of probiotics can attenuate asthmatic responses both in murine models and clinical trials. T-helper 17 (Th17) cells, a subset of CD4+ T cells have been implicated as having an important role in the development of several allergic disorders, but the relationship between oral administration of probiotics and Th17 development has not been well studied. BALB/c mice were given lysed Enterococcus faecalis FK-23 (LFK) orally for 28 days. After sensitization by subcutaneous injection of ovalbumin (OVA) on Days 14 and 21 and 1% OVA inhalation on Days 25, 26 and 27, they were challenged with a 5% OVA aerosol on Day 28. Twenty-four hours later, airway resistance and accumulation of inflammatory cells in bronchoalveolar lavage fluid (BALF) and lung tissues were determined. Ιnterleukin (IL)-17-expressing CD4+ lymphocytes isolated from lung, spleen and lamina propria of the intestine were detected by flow cytometry. The expression of IL-6 and TGF-β mRNA was assessed by real-time PCR. Increases in airway hyperresponsiveness, and numbers of total leukocytes and mast cells in BALF induced by OVA challenge were significantly suppressed by oral administration of LFK. The increased percentage of IL-17-expressing CD4+ cells from lung, spleen and intestine in OVA-challenged mice was reduced following LFK treatment. We conclude that the oral administration of LFK suppresses the asthmatic response and that this is associated with attenuation of Th17 cell development.

  13. Human Satellite Cell Transplantation and Regeneration from Diverse Skeletal Muscles

    PubMed Central

    Xu, Xiaoti; Wilschut, Karlijn J.; Kouklis, Gayle; Tian, Hua; Hesse, Robert; Garland, Catharine; Sbitany, Hani; Hansen, Scott; Seth, Rahul; Knott, P. Daniel; Hoffman, William Y.; Pomerantz, Jason H.

    2015-01-01

    Summary Identification of human satellite cells that fulfill muscle stem cell criteria is an unmet need in regenerative medicine. This hurdle limits understanding how closely muscle stem cell properties are conserved among mice and humans and hampers translational efforts in muscle regeneration. Here, we report that PAX7 satellite cells exist at a consistent frequency of 2–4 cells/mm of fiber in muscles of the human trunk, limbs, and head. Xenotransplantation into mice of 50–70 fiber-associated, or 1,000–5,000 FACS-enriched CD56+/CD29+ human satellite cells led to stable engraftment and formation of human-derived myofibers. Human cells with characteristic PAX7, CD56, and CD29 expression patterns populated the satellite cell niche beneath the basal lamina on the periphery of regenerated fibers. After additional injury, transplanted satellite cells robustly regenerated to form hundreds of human-derived fibers. Together, these findings conclusively delineate a source of bona-fide endogenous human muscle stem cells that will aid development of clinical applications. PMID:26352798

  14. Human Satellite Cell Transplantation and Regeneration from Diverse Skeletal Muscles.

    PubMed

    Xu, Xiaoti; Wilschut, Karlijn J; Kouklis, Gayle; Tian, Hua; Hesse, Robert; Garland, Catharine; Sbitany, Hani; Hansen, Scott; Seth, Rahul; Knott, P Daniel; Hoffman, William Y; Pomerantz, Jason H

    2015-09-08

    Identification of human satellite cells that fulfill muscle stem cell criteria is an unmet need in regenerative medicine. This hurdle limits understanding how closely muscle stem cell properties are conserved among mice and humans and hampers translational efforts in muscle regeneration. Here, we report that PAX7 satellite cells exist at a consistent frequency of 2-4 cells/mm of fiber in muscles of the human trunk, limbs, and head. Xenotransplantation into mice of 50-70 fiber-associated, or 1,000-5,000 FACS-enriched CD56(+)/CD29(+) human satellite cells led to stable engraftment and formation of human-derived myofibers. Human cells with characteristic PAX7, CD56, and CD29 expression patterns populated the satellite cell niche beneath the basal lamina on the periphery of regenerated fibers. After additional injury, transplanted satellite cells robustly regenerated to form hundreds of human-derived fibers. Together, these findings conclusively delineate a source of bona-fide endogenous human muscle stem cells that will aid development of clinical applications.

  15. Ethanol co-administration moderates 3,4-methylenedioxymethamphetamine effects on human physiology.

    PubMed

    Dumont, G J H; Kramers, C; Sweep, F C G J; Willemsen, J J; Touw, D J; Schoemaker, R C; van Gerven, J M A; Buitelaar, J K; Verkes, R J

    2010-02-01

    Alcohol is frequently used in combination with 3,4-methylenedioxymethamphetamine (MDMA). Both drugs affect cardiovascular function, hydration and temperature regulation, but may have partly opposing effects. The present study aims to assess the acute physiologic effects of (co-) administration of MDMA and ethanol over time. A four-way, double blind, randomized, crossover, placebo-controlled study in 16 healthy volunteers (9 male and 7 female) between the ages of 18 and 29. MDMA (100 mg) was given orally and blood ethanol concentration was maintained at pseudo-steady state levels of 0.6 per thousand by a three-hour 10% intravenous ethanol clamp. Cardiovascular function, temperature and hydration measures were recorded throughout the study days. Ethanol did not significantly affect physiologic function, with the exception of a short lasting increase in heart rate. MDMA potently increased heart rate and blood pressure and induced fluid retention as well as an increase in temperature. Co-administration of ethanol with MDMA did not affect cardiovascular function compared to the MDMA alone condition, but attenuated the effects of MDMA on fluid retention and showed a trend for attenuation of MDMA-induced temperature increase. In conclusion, co-administration of ethanol and MDMA did not exacerbate physiologic effects compared to all other drug conditions, and moderated some effects of MDMA alone.

  16. New frontiers in human cell biology and medicine: can pluripotent stem cells deliver?

    PubMed

    Goldstein, Lawrence S B

    2012-11-12

    Human pluripotent stem cells provide enormous opportunities to treat disease using cell therapy. But human stem cells can also drive biomedical and cell biological discoveries in a human model system, which can be directly linked to understanding disease or developing new therapies. Finally, rigorous scientific studies of these cells can and should inform the many science and medical policy issues that confront the translation of these technologies to medicine. In this paper, I discuss these issues using amyotrophic lateral sclerosis as an example.

  17. Susceptibility of Human Placenta Derived Mesenchymal Stromal/Stem Cells to Human Herpesviruses Infection

    PubMed Central

    Rotola, Antonella; Alviano, Francesco; Solimando, Liliana; Lanzoni, Giacomo; Bonsi, Laura; Di Luca, Dario; Marchionni, Cosetta; Alvisi, Gualtiero; Ripalti, Alessandro

    2013-01-01

    Fetal membranes (FM) derived mesenchymal stromal/stem cells (MSCs) are higher in number, expansion and differentiation abilities compared with those obtained from adult tissues, including bone marrow. Upon systemic administration, ex vivo expanded FM-MSCs preferentially home to damaged tissues promoting regenerative processes through their unique biological properties. These characteristics together with their immune-privileged nature and immune suppressive activity, a low infection rate and young age of placenta compared to other sources of SCs make FM-MSCs an attractive target for cell-based therapy and a valuable tool in regenerative medicine, currently being evaluated in clinical trials. In the present study we investigated the permissivity of FM-MSCs to all members of the human Herpesviridae family, an issue which is relevant to their purification, propagation, conservation and therapeutic use, as well as to their potential role in the vertical transmission of viral agents to the fetus and to their potential viral vector-mediated genetic modification. We present here evidence that FM-MSCs are fully permissive to infection with Herpes simplex virus 1 and 2 (HSV-1 and HSV-2), Varicella zoster virus (VZV), and Human Cytomegalovirus (HCMV), but not with Epstein-Barr virus (EBV), Human Herpesvirus-6, 7 and 8 (HHV-6, 7, 8) although these viruses are capable of entering FM-MSCs and transient, limited viral gene expression occurs. Our findings therefore strongly suggest that FM-MSCs should be screened for the presence of herpesviruses before xenotransplantation. In addition, they suggest that herpesviruses may be indicated as viral vectors for gene expression in MSCs both in gene therapy applications and in the selective induction of differentiation. PMID:23940750

  18. Susceptibility of human placenta derived mesenchymal stromal/stem cells to human herpesviruses infection.

    PubMed

    Avanzi, Simone; Leoni, Valerio; Rotola, Antonella; Alviano, Francesco; Solimando, Liliana; Lanzoni, Giacomo; Bonsi, Laura; Di Luca, Dario; Marchionni, Cosetta; Alvisi, Gualtiero; Ripalti, Alessandro

    2013-01-01

    Fetal membranes (FM) derived mesenchymal stromal/stem cells (MSCs) are higher in number, expansion and differentiation abilities compared with those obtained from adult tissues, including bone marrow. Upon systemic administration, ex vivo expanded FM-MSCs preferentially home to damaged tissues promoting regenerative processes through their unique biological properties. These characteristics together with their immune-privileged nature and immune suppressive activity, a low infection rate and young age of placenta compared to other sources of SCs make FM-MSCs an attractive target for cell-based therapy and a valuable tool in regenerative medicine, currently being evaluated in clinical trials. In the present study we investigated the permissivity of FM-MSCs to all members of the human Herpesviridae family, an issue which is relevant to their purification, propagation, conservation and therapeutic use, as well as to their potential role in the vertical transmission of viral agents to the fetus and to their potential viral vector-mediated genetic modification. We present here evidence that FM-MSCs are fully permissive to infection with Herpes simplex virus 1 and 2 (HSV-1 and HSV-2), Varicella zoster virus (VZV), and Human Cytomegalovirus (HCMV), but not with Epstein-Barr virus (EBV), Human Herpesvirus-6, 7 and 8 (HHV-6, 7, 8) although these viruses are capable of entering FM-MSCs and transient, limited viral gene expression occurs. Our findings therefore strongly suggest that FM-MSCs should be screened for the presence of herpesviruses before xenotransplantation. In addition, they suggest that herpesviruses may be indicated as viral vectors for gene expression in MSCs both in gene therapy applications and in the selective induction of differentiation.

  19. Establishment of human tumoral ependymal cell lines and coculture with tubular-like human endothelial cells.

    PubMed

    Brisson, C; Lelong-Rebel, I; Mottolèse, C; Jouvet, A; Fèvre-Montange, M; Saint Pierre, G; Rebel, G; Belin, M F

    2002-10-01

    Ependymomas, rare neoplasms of the central nervous system, occur predominantly in children. They are highly vascularized, and histological findings show many perivascular rosettes of tumoral cells radially organized around capillaries. Treatment of ependymomas relies on surgery combined with radio- or chemotherapy, but the efficiency of chemotherapy is limited, probably because of their multidrug resistance (MDR) phenotype. Progress in the therapy of these neoplasms is dramatically limited by the absence of cell line models. We established conditions for the long-term culture of human tumoral ependymocytes and their 3D coculture in Matrigel with endothelial cells. Histological, immunological, and ultrastructural studies showed that the morphological features (microvilli, cilia, and caveolae) of these cultured cells were similar to those of the tumor in vivo. The cells expressed potential oncological markers related to the immature state of tumoral cells (nestin and Notch-1), their tumorigenicity [caveolae and epidermal growth factor-receptor (EGF-R)], or the MDR phenotype [P-glycoprotein (P-gp)]. The expression of P-gp, EGF-R, and caveolin-1 by these tumoral ependymocytes could be useful in studies on new drugs. This coculture model might represent a new powerful tool to study new therapeutic delivery strategies in tumoral cells.

  20. Derivation and spontaneous differentiation of human embryonic stem cells*

    PubMed Central

    Amit, Michal; Itskovitz-Eldor, Joseph

    2002-01-01

    Abstract Embryonic stem (ES) cells are unique cells derived from the inner cell mass of the mammalian blastocyst. These cells are immortal and pluripotent, retain their developmental potential after prolonged culture, and can be continuously cultured in an undifferentiated state. Many in vitro differentiation systems have been developed for mouse ES cells, including reproducible methods for mouse ES cell differentiation into haematopoietic and neural precursors, cardiomyocytes, insulin-secreting cells, endothelial cells and various other cell types. The derivation of new human ES cell lines provides the opportunity to develop unique models for developmental research and for cell therapies. In this review we consider the derivation and spontaneous differentiation of human ES cells. PMID:12033726

  1. Propagation of Human Embryonic Stem Cells on Human Amniotic Fluid Cells as Feeder Cells in Xeno-Free Culture Conditions

    PubMed Central

    Jung, Juwon; Baek, Jin Ah; Seol, Hye Won; Choi, Young Min

    2016-01-01

    Human embryonic stem cells (hESCs) have been routinely cultured on mouse embryonic fibroblast feederlayers with a medium containing animal materials. For clinical application of hESCs, animal-derived products from the animal feeder cells, animal substrates such as gelatin or Matrigel and animal serum are strictly to be eliminated in the culture system. In this study, we performed that SNUhES32 and H1 were cultured on human amniotic fluid cells (hAFCs) with KOSR XenoFree and a humanized substrate. All of hESCs were relatively well propagated on hAFCs feeders with xeno-free conditions and they expressed pluripotent stem cell markers, alkaline phosphatase, SSEA-4, TRA1-60, TRA1-81, Oct-4, and Nanog like hESCs cultured on STO or human foreskin fibroblast feeders. In addition, we observed the expression of nonhuman N-glycolylneuraminic acid (Neu5GC) molecules by flow cytometry, which was xenotransplantation components of contamination in hESCs cultured on animal feeder conditions, was not detected in this xeno-free condition. In conclusion, SNUhES32 and H1 could be maintained on hAFCs for humanized culture conditions, therefore, we suggested that new xenofree conditions for clinical grade hESCs culture will be useful data in future clinical studies. PMID:27294211

  2. Effects of single cortisol administrations on human affect reviewed: Coping with stress through adaptive regulation of automatic cognitive processing.

    PubMed

    Putman, Peter; Roelofs, Karin

    2011-05-01

    The human stress hormone cortisol may facilitate effective coping after psychological stress. In apparent agreement, administration of cortisol has been demonstrated to reduce fear in response to stressors. For anxious patients with phobias or posttraumatic stress disorder this has been ascribed to hypothetical inhibition of retrieval of traumatic memories. However, such stress-protective effects may also work via adaptive regulation of early cognitive processing of threatening information from the environment. This paper selectively reviews the available literature on effects of single cortisol administrations on affect and early cognitive processing of affectively significant information. The concluded working hypothesis is that immediate effects of high concentration of cortisol may facilitate stress-coping via inhibition of automatic processing of goal-irrelevant threatening information and through increased automatic approach-avoidance responses in early emotional processing. Limitations in the existing literature and suggestions for future directions are briefly discussed.

  3. Stem Cells: A Renaissance in Human Biology Research.

    PubMed

    Wu, Jun; Izpisua Belmonte, Juan Carlos

    2016-06-16

    The understanding of human biology and how it relates to that of other species represents an ancient quest. Limited access to human material, particularly during early development, has restricted researchers to only scratching the surface of this inherently challenging subject. Recent technological innovations, such as single cell "omics" and human stem cell derivation, have now greatly accelerated our ability to gain insights into uniquely human biology. The opportunities afforded to delve molecularly into scarce material and to model human embryogenesis and pathophysiological processes are leading to new insights of human development and are changing our understanding of disease and choice of therapy options.

  4. Identification of cell lines permissive for human coronavirus NL63.

    PubMed

    Schildgen, Oliver; Jebbink, Maarten F; de Vries, Michel; Pyrc, Krzysztov; Dijkman, Ronald; Simon, Arne; Müller, Andreas; Kupfer, Bernd; van der Hoek, Lia

    2006-12-01

    Six cell lines routinely used in laboratories were tested for permissiveness to the infection with the newly identified human coronavirus NL63. Two monkey epithelial cell lines, LLC-MK2 and Vero-B4, showed a cytopathic effect (CPE) and clear viral replication, whereas no CPE or replication was observed in human lung fibroblasts MRC-5s. In Rhabdomyosarcoma cells, Madin-Darby-Canine-kidney cells and in an undefined monkey kidney cell line some replication was observed but massive exponential rise in virus yield lacked The results will lead to an improved routine diagnostic algorithm for the detection of the human coronavirus NL63.

  5. Mesenchymal stem cell administration at coronary artery reperfusion in the rat by two delivery routes: a quantitative assessment

    PubMed Central

    Hale, Sharon L; Dai, Wangde; Dow, Joan S; Kloner, Robert A

    2008-01-01

    Aims Ideally, mesenchymal stem cells (MSC) home to and/or remain at the site of damaged myocardium when administered after myocardial infarction. However, MSC may not remain in the heart, but instead relocate to other areas. We investigated quantitatively the distribution of labeled rat MSC, given by two routes after coronary artery occlusion/reperfusion in rats. Main Methods Rats were subjected to 45 minutes of coronary artery occlusion and 7 days of reperfusion. Before reperfusion rats received 2×106 MSC, labeled with europium, injected directly into the ischemic region of the heart (n=9) or intravenously (n = 8). After one week tissues were analyzed for label content together with a standard curve of known quantities of labeled MSC. Key Findings In rats receiving cells injected directly into the myocardium, 15% of labeled cells were retained in the heart. When the cells were administered intravenously, no MSC were detected in the heart. The route of administration did not affect distribution to other organs, as the number of MSC in liver, spleen and lung was similar with both routes of delivery. Significance Even with direct intramyocardial injection, only a small proportion of the cells are retained in the heart, instead traveling to other organs. With intravenous injection there was no evidence that cells "homed" to the damaged heart. Although cell delivery to the heart was significantly affected by the route of administration, the distribution of cells to other organs was similar with both routes of administration. PMID:18755200

  6. Comparative mutagenesis of human cells in vivo and in vitro

    SciTech Connect

    Thilly, W.G.

    1992-05-01

    This report discusses measuring methods of point mutations; high density cell cultures for low dose studies; measurement and sequence determination of mutations in DNA; the mutational spectra of styrene oxide and ethlyene oxide in TK-6 cells; mutational spectrum of Cr in human lymphoblast cells; mutational spectra of radon in TK-6 cells; and the mutational spectra of smokeless tobacco. (CBS)

  7. Identification of a candidate stem cell in human gallbladder

    PubMed Central

    Manohar, Rohan; Li, Yaming; Fohrer, Helene; Guzik, Lynda; Stolz, Donna Beer; Chandran, Uma R.; LaFramboise, William A.; Lagasse, Eric

    2015-01-01

    There are currently no reports of the identification of stem cells in the human gallbladder. The differences between human gallbladder and intrahepatic bile duct (IHBD) cells have also not been explored. The goals of this study were to evaluate if human fetal gallbladder contains a candidate stem cell population and if fetal gallbladder cells are distinct from fetal IHBD cells. We found that EpCAM+CD44+CD13+ cells represent the cell population most enriched for clonal self-renewal from primary gallbladder. Primary EpCAM+CD44+CD13+ cells gave rise to EpCAM+CD44+CD13+ and EpCAM+CD44+CD13− cells in vitro, and gallbladder cells expanded in vitro exhibited short-term engraftment in vivo. Last, we found that CD13, CD227, CD66, CD26 and CD49b were differentially expressed between gallbladder and IHBD cells cultured in vitro indicating clear phenotypic differences between the two cell populations. Microarray analyses of expanded cultures confirmed that both cell types have unique transcriptional profiles with predicted functional differences in lipid, carbohydrate, nucleic acid and drug metabolism. In conclusion, we have isolated a distinct clonogenic population of epithelial cells from primary human fetal gallbladder with stem cell characteristics and found it to be unique compared to IHBD cells. PMID:25765520

  8. 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.

  9. Short-term effects of synthetic human parathyroid hormone-(1--34) administration on bone mineral metabolism in osteoporotic patients.

    PubMed Central

    Slovik, D M; Neer, R M; Potts, J T

    1981-01-01

    Since studies in animals and humans have shown that parathyroid hormone can stimulate bone formation and increase trabecular bone, and patients with primary and secondary hyperparathyroidism may exhibit osteosclerosis, we evaluated the effect of short-term administration of human parathyroid hormone, hPTH-(1--34), in patients with osteoporosis. Six patients with osteoporosis underwent detailed studies including blood and urinary measurements of calcium, phosphate, and magnesium; 47Ca kinetic studies; and 18-d balance studies before and during the short-term administration (3--4 wk) of a daily subcutaneous injection of hPTH fragment 1--34 given as 450 or 750 U/dose. The mean fasting plasma calcium values rose slightly after hPTH-(1--34) administration, primarily in the high-dose group. There was no difference in the mean fasting plasma inorganic phosphate levels. The mean daily urinary excretion of calcium and phosphate was significantly increased in patients given the higher dose. In patients given 750 U, net intestinal calcium absorption increased, phosphate absorption increased, calcium balance improved, and phosphate balance improved. In patients given 450 U, calcium balance and phosphate balance worsened. 47Ca kinetic studies showed a minimal increase in bone accretion rate, a decrease in the mean transit time of calcium in the exchangeable pools, and a decrease in the exchangeable-pool size. In all six patients there was an increased renal clearance of 47Ca as a result of hPTH-(1--34) administration. These studies indicate that low doses of parathyroid hormone may promote bone formation, whereas higher doses clearly have an adverse effect on the skeleton. PMID:7298851

  10. Human mast cells costimulate T cells through a CD28-independent interaction.

    PubMed

    Suurmond, Jolien; Dorjée, Annemarie L; Huizinga, Tom W J; Toes, René E M

    2016-05-01

    Mast cells are innate immune cells usually residing in peripheral tissues, where they are likely to activate T-cell responses. Similar to other myeloid immune cells, mast cells can function as antigen-presenting cells. However, little is known about the capacity of human mast cells to costimulate CD4(+) T cells. Here, we studied the T-cell stimulatory potential of human mast cells. Peripheral blood derived mast cells were generated and cocultured with isolated CD4(+) T cells. In the presence of T-cell receptor triggering using anti-CD3, mast cells promoted strong proliferation of T cells, which was two- to fivefold stronger than the "T-cell promoting capacity" of monocytes. The interplay between mast cells and T cells was dependent on cell-cell contact, suggesting that costimulatory molecules on the mast cell surface are responsible for the effect. However, in contrast to monocytes, the T-cell costimulation by mast cells was independent of the classical costimulatory molecule CD28, or that of OX40L, ICOSL, or LIGHT. Our data show that mast cells can costimulate human CD4(+) T cells to induce strong T-cell proliferation, but that therapies aiming at disrupting the interaction of CD28 and B7 molecules do not inhibit mast cell mediated T-cell activation.

  11. The evolution of human cells in terms of protein innovation.

    PubMed

    Sardar, Adam J; Oates, Matt E; Fang, Hai; Forrest, Alistair R R; Kawaji, Hideya; Gough, Julian; Rackham, Owen J L

    2014-06-01

    Humans are composed of hundreds of cell types. As the genomic DNA of each somatic cell is identical, cell type is determined by what is expressed and when. Until recently, little has been reported about the determinants of human cell identity, particularly from the joint perspective of gene evolution and expression. Here, we chart the evolutionary past of all documented human cell types via the collective histories of proteins, the principal product of gene expression. FANTOM5 data provide cell-type-specific digital expression of human protein-coding genes and the SUPERFAMILY resource is used to provide protein domain annotation. The evolutionary epoch in which each protein was created is inferred by comparison with domain annotation of all other completely sequenced genomes. Studying the distribution across epochs of genes expressed in each cell type reveals insights into human cellular evolution in terms of protein innovation. For each cell type, its history of protein innovation is charted based on the genes it expresses. Combining the histories of all cell types enables us to create a timeline of cell evolution. This timeline identifies the possibility that our common ancestor Coelomata (cavity-forming animals) provided the innovation required for the innate immune system, whereas cells which now form the brain of human have followed a trajectory of continually accumulating novel proteins since Opisthokonta (boundary of animals and fungi). We conclude that exaptation of existing domain architectures into new contexts is the dominant source of cell-type-specific domain architectures.

  12. Comparative Proteome Analyses of Human Plasma Following in vivo Lipopolysaccharide Administration Using Multidimensional Separations Coupled with Tandem Mass Spectrometry

    PubMed Central

    Qian, Wei-Jun; Jacobs, Jon M.; Camp, David G.; Monroe, Matthew E.; Moore, Ronald J.; Gritsenko, Marina A.; Calvano, Steve E.; Lowry, Stephen F.; Xiao, Wenzhong; Moldawer, Lyle L.; Davis, Ronald W.; Tompkins, Ronald G.; Smith, Richard D.

    2007-01-01

    There is significant interest in characterization of the human plasma proteome due to its potential for providing biomarkers applicable to clinical diagnosis and treatment and for gaining a better understanding of human diseases. We describe here a strategy for the comparative proteome analyses of human plasma, which is applicable to biomarker identifications for various disease states. Multidimensional liquid chromatography-mass spectrometry has been applied to make comparative proteome analyses of plasma samples from an individual prior to and 9 h after lipopolysaccharide (LPS) administration. Peptide peak areas and the number of peptide identifications for each protein were used to evaluate the reproducibility of LC-MS/MS and to compare relative changes in protein concentration between the samples following LPS treatment. A total of 804 distinct plasma proteins (not including immunoglobulins) were confidently identified with 32 proteins observed to be significantly increased in concentration following LPS administration, including several known inflammatory response or acute-phase mediators such as C-reactive protein, serum amyloid A and A2, LPS-binding protein, LPS-responsive and beige-like anchor protein, hepatocyte growth factor activator, and von Willebrand factor, and thus constituting potential biomarkers for inflammatory response. PMID:15627965

  13. Development of human epithelial cell systems for radiation risk assessment

    NASA Astrophysics Data System (ADS)

    Yang, C. H.; Craise, L. M.

    1994-10-01

    The most important health effect of space radiation for astronauts is cancer induction. For radiation risk assessment, an understanding of carcinogenic effect of heavy ions in human cells is most essential. In our laboratory, we have successfully developed a human mammary epithelial cell system for studying the neoplastic transformation in vitro. Growth variants were obtained from heavy ion irradiated immortal mammary cell line. These cloned growth variants can grow in regular tissue culture media and maintain anchorage dependent growth and density inhibition property. Upon further irradiation with high-LET radiation, transformed foci were found. Experimental results from these studies suggest that multiexposure of radiation is required to induce neoplastic transformation of human epithelial cells. This multihits requirement may be due to high genomic stability of human cells. These growth variants can be useful model systems for space flight experiments to determine the carcinogenic effect of space radiation in human epithelial cells.

  14. Development of human epithelial cell systems for radiation risk assessment

    NASA Technical Reports Server (NTRS)

    Yang, C. H.; Craise, L. M.

    1994-01-01

    The most important health effect of space radiation for astronauts is cancer induction. For radiation risk assessment, an understanding of carcinogenic effect of heavy ions in human cells is most essential. In our laboratory, we have successfully developed a human mammary epithelial cell system for studying the neoplastic transformation in vitro. Growth variants were obtained from heavy ion irradiated immortal mammary cell line. These cloned growth variants can grow in regular tissue culture media and maintain anchorage dependent growth and density inhibition property. Upon further irradiation with high-Linear Energy Transfer (LET) radiation, transformed foci were found. Experimental results from these studies suggest that multiexposure of radiation is required to induce neoplastic tranformation of human epithelial cells. This multihits requirement may be due to high genomic stability of human cells. These growth variants can be useful model systems for space flight experiments to determine the carcinogenic effect of space radiation in human epithelial cells.

  15. Adult human brain cell culture for neuroscience research.

    PubMed

    Gibbons, Hannah M; Dragunow, Mike

    2010-06-01

    Studies of the brain have progressed enormously through the use of in vivo and in vitro non-human models. However, it is unlikely such studies alone will unravel the complexities of the human brain and so far no neuroprotective treatment developed in animals has worked in humans. In this review we discuss the use of adult human brain cell culture methods in brain research to unravel the biology of the normal and diseased human brain. The advantages of using adult human brain cells as tools to study human brain function from both historical and future perspectives are discussed. In particular, studies using dissociated cultures of adult human microglia, astrocytes, oligodendrocytes and neurons are described and the applications of these types of study are evaluated. Alternative sources of human brain cells such as adult neural stem cells, induced pluripotent stem cells and slice cultures of adult human brain tissue are also reviewed. These adult human brain cell culture methods could benefit basic research and more importantly, facilitate the translation of basic neuroscience research to the clinic for the treatment of brain disorders.

  16. In vitro investigation of ultrasound-induced oxidative stress on human lens epithelial cells.

    PubMed

    Rwei, Patrick; Alex Gong, Cihun-Siyong; Luo, Li-Jyuan; Lin, Meng-Bo; Lai, Jui-Yang; Liu, Hao-Li

    2017-01-22

    The effect of ultrasound exposure on human lens epithelial cells (HLE-B3) was investigated in vitro, specifically on the generation of oxidative stress upon ultrasound application using various clinically-relevant settings. In addition to ultrasound-induced heat effects, oxidative stress has been recently proposed as one of the main mechanisms for ultrasound-induced effects on human cells. In this work, the levels of biocompatibility and generation of oxidative stress by exposure of ultrasound to HLE-B3 were evaluated quantitatively and qualitatively by the MTT assay, Live/Dead assay, reactive oxygen species (ROS) and intracellular calcium level. Oxidative stress induction is traditionally achieved through administrations of H2O2 and thus the administration of H2O2 was used as the positive control group for comparison herein. Concerning the administrations of H2O2 are considered invasive and may potentially have side effects, ultrasound as physical stimulation could be a safer and non-invasive method to induce similar oxidative stress environments. The effect of ultrasound on cell viability and induction of oxidative stress increases with ultrasound intensity. The result reveals that the continuous ultrasound has a positive impact on the oxidative stress levels but does negatively on the cell viability, as compared to the pulsed ultrasound. Furthermore, our work demonstrates that the exposure of 58 kPa continuous ultrasound without microbubbles can maintain acceptable cell viability and produce oxidative stress effects similar to the traditional administrations of H2O2. In summary, exposure of ultrasound can generate oxidative stress comparable to traditional administrations of H2O2. The effect of generating oxidative stress is adjustable through ultrasound parameters, including the pulsed or continuous wave, the intensity of ultrasound and addition of microbubbles.

  17. Activation of GPR119 Stimulates Human β-Cell Replication and Neogenesis in Humanized Mice with Functional Human Islets

    PubMed Central

    Ansarullah; Free, Colette; Christopherson, Jenica; Chen, Quanhai; Gao, Jie; Liu, Chengyang; Naji, Ali; Rabinovitch, Alex; Guo, Zhiguang

    2016-01-01

    Using humanized mice with functional human islets, we investigated whether activating GPR119 by PSN632408, a small molecular agonist, can stimulate human β-cell regeneration in vivo. Human islets were transplanted under the left kidney capsule of immunodeficient mice with streptozotocin- (STZ-) induced diabetes. The recipient mice were treated with PSN632408 or vehicle and BrdU daily. Human islet graft function in the mice was evaluated by nonfasting glucose levels, oral glucose tolerance, and removal of the grafts. Immunostaining for insulin, glucagon, and BrdU or Ki67 was performed in islet grafts to evaluate α- and β-cell replication. Insulin and CK19 immunostaining was performed to evaluate β-cell neogenesis. Four weeks after human islet transplantation, 71% of PSN632408-treated mice achieved normoglycaemia compared with 24% of vehicle-treated mice. Also, oral glucose tolerance was significantly improved in the PSN632408-treated mice. PSN632408 treatment significantly increased both human α- and β-cell areas in islet grafts and stimulated α- and β-cell replication. In addition, β-cell neogenesis was induced from pancreatic duct cells in the islet grafts. Our results demonstrated that activation of GPR119 increases β-cell mass by stimulating human β-cell replication and neogenesis. Therefore, GPR119 activators may qualify as therapeutic agents to increase human β-cell mass in patients with diabetes. PMID:27413754

  18. Programming human pluripotent stem cells into white and brown adipocytes

    PubMed Central

    Ahfeldt, Tim; Schinzel, Robert T.; Lee, Youn-Kyoung; Hendrickson, David; Kaplan, Adam; Lum, David H.; Camahort, Raymond; Xia, Fang; Shay, Jennifer; Rhee, Eugene P.; Clish, Clary B.; Deo, Rahul C.; Shen, Tony; Lau, Frank H.; Cowley, Alicia; Mowrer, Greg; Al-Siddiqi, Heba; Nahrendorf, Matthias; Musunuru, Kiran; Gerszten, Robert E.; Rinn, John L.; Cowan, Chad A.

    2012-01-01

    The utility of human pluripotent stem cells is dependent on efficient differentiation protocols that convert these cells into relevant adult cell types. Here we report the robust and efficient differentiation of human pluripotent stem cells into white or brown adipocytes. We found that inducible expression of PPARG2 alone or combined with CEBPB and/or PRDM16 in mesenchymal progenitor cells derived from pluripotent stem cells programmed their development towards a white or brown adipocyte cell fate with efficiencies of 85%–90%. These adipocytes retained their identity independent of transgene expression, could be maintained in culture for several weeks, expressed mature markers and had mature functional properties such as lipid catabolism and insulin-responsiveness. When transplanted into mice, the programmed cells gave rise to ectopic fat pads with the morphological and functional characteristics of white or brown adipose tissue. These results indicate that the cells could be used to faithfully model human disease. PMID:22246346

  19. Excretion and metabolism of milnacipran in humans after oral administration of milnacipran hydrochloride.

    PubMed

    Li, Fanying; Chin, Christina; Wangsa, Julie; Ho, John

    2012-09-01

    The pharmacokinetics, excretion, and metabolism of milnacipran were evaluated after oral administration of a 100-mg dose of [¹⁴C]milnacipran hydrochloride to healthy male subjects. The peak plasma concentration of unchanged milnacipran (∼240 ng/ml) was attained at 3.5 h and was lower than the peak plasma concentration of radioactivity (∼679 ng Eq of milnacipran/ml) observed at 4.3 h, indicating substantial metabolism of milnacipran upon oral administration. Milnacipran has two chiral centers and is a racemic mixture of cis isomers: d-milnacipran (1S, 2R) and l-milnacipran (1R, 2S). After oral administration, the radioactivity of almost the entire dose was excreted rapidly in urine (approximately 93% of the dose). Approximately 55% of the dose was excreted in urine as unchanged milnacipran, which contained a slightly higher proportion of d-milnacipran (∼31% of the dose). In addition to the excretion of milnacipran carbamoyl O-glucuronide metabolite in urine (∼19% of the dose), predominantly as the l-milnacipran carbamoyl O-glucuronide metabolite (∼17% of the dose), approximately 8% of the dose was excreted in urine as the N-desethyl milnacipran metabolite. No additional metabolites of significant quantity were excreted in urine. Similar plasma concentrations of milnacipran and the l-milnacipran carbamoyl O-glucuronide metabolite were observed after dosing, and the maximum plasma concentration of l-milnacipran carbamoyl O-glucuronide metabolite at 4 h after dosing was 234 ng Eq of milnacipran/ml. Lower plasma concentrations (<25 ng Eq of milnacipran/ml) of N-desethyl milnacipran and d-milnacipran carbamoyl O-glucuronide metabolites were observed.

  20. Human Embryonic Stem Cell Derived Vascular Progenitor Cells Capable of Endothelial and Smooth Muscle Cell Function

    PubMed Central

    Hill, Katherine L; Obrtlikova, Petra; Alvarez, Diego F; King, Judy A; Keirstead, Susan A; Allred, Jeremy R; Kaufman, Dan S

    2010-01-01

    OBJECTIVE Previous studies have demonstrated development of endothelial cells (ECs) and smooth muscle cells (SMCs) as separate cell lineages derived from human embryonic stem cells (hESCs). We demonstrate CD34+ cells isolated from differentiated hESCs function as vascular progenitor cells capable of producing both ECs and SMCs. These studies better define the developmental origin and reveal the relationship between these two cell types, as well as provide a more complete biological characterization. MATERIALS AND METHODS hESCs are co-cultured on M2-10B4 stromal cells or Wnt1 expressing M2-10B4 for 13–15 days to generate a CD34+ cell population. These cells are isolated using a magnetic antibody separation kit and cultured on fibronectin coated dishes in EC medium. To induce SMC differentiation, culture medium is changed and a morphological and phenotypic change occurs within 24–48 hours. RESULTS CD34+ vascular progenitor cells give rise to ECs and SMCs. The two populations express respective cell specific transcripts and proteins, exhibit intracellular calcium in response to various agonists, and form robust tube-like structures when co-cultured in Matrigel. Human umbilical vein endothelial cells (HUVEC) cultured under SMC conditions do not exhibit a change in phenotype or genotype. Wnt1 overexpressing stromal cells produced an increased number of progenitor cells. CONCLUSIONS The ability to generate large numbers of ECs and SMCs from a single vascular progenitor cell population is promising for therapeutic use to treat a variety of diseased and ischemic conditions. The step-wise differentiation outlined here is an efficient, reproducible method with potential for large scale cultures suitable for clinical applications. PMID:20067819

  1. Single-cell analysis reveals a stem-cell program in human metastatic breast cancer cells.

    PubMed

    Lawson, Devon A; Bhakta, Nirav R; Kessenbrock, Kai; Prummel, Karin D; Yu, Ying; Takai, Ken; Zhou, Alicia; Eyob, Henok; Balakrishnan, Sanjeev; Wang, Chih-Yang; Yaswen, Paul; Goga, Andrei; Werb, Zena

    2015-10-01

    Despite major advances in understanding the molecular and genetic basis of cancer, metastasis remains the cause of >90% of cancer-related mortality. Understanding metastasis initiation and progression is critical to developing new therapeutic strategies to treat and prevent metastatic disease. Prevailing theories hypothesize that metastases are seeded by rare tumour cells with unique properties, which may function like stem cells in their ability to initiate and propagate metastatic tumours. However, the identity of metastasis-initiating cells in human breast cancer remains elusive, and whether metastases are hierarchically organized is unknown. Here we show at the single-cell level that early stage metastatic cells possess a distinct stem-like gene expression signature. To identify and isolate metastatic cells from patient-derived xenograft models of human breast cancer, we developed a highly sensitive fluorescence-activated cell sorting (FACS)-based assay, which allowed us to enumerate metastatic cells in mouse peripheral tissues. We compared gene signatures in metastatic cells from tissues with low versus high metastatic burden. Metastatic cells from low-burden tissues were distinct owing to their increased expression of stem cell, epithelial-to-mesenchymal transition, pro-survival, and dormancy-associated genes. By contrast, metastatic cells from high-burden tissues were similar to primary tumour cells, which were more heterogeneous and expressed higher levels of luminal differentiation genes. Transplantation of stem-like metastatic cells from low-burden tissues showed that they have considerable tumour-initiating capacity, and can differentiate to produce luminal-like cancer cells. Progression to high metastatic burden was associated with increased proliferation and MYC expression, which could be attenuated by treatment with cyclin-dependent kinase (CDK) inhibitors. These findings support a hierarchical model for metastasis, in which metastases are initiated

  2. CYP2E1-dependent hepatotoxicity and oxidative damage after ethanol administration in human primary hepatocytes

    PubMed Central

    Liu, Lie-Gang; Yan, Hong; Yao, Ping; Zhang, Wen; Zou, Li-Jun; Song, Fang-Fang; Li, Ke; Sun, Xiu-Fa

    2005-01-01

    AIM: To observe the relationship between ethanol-induced oxidative damage in human primary cultured hepatocytes and cytochrome P450 2E1 (CYP2E1) activity, in order to address if inhibition of CYP2E1 could attenuate ethanol-induced cellular damage. METHODS: The dose-dependent (25-100 mmol/L) and time-dependent (0-24 h) exposures of primary human cultured hepatocytes to ethanol were carried out. CYP2E1 activity and protein expression were detected by spectrophotometer and Western blot analysis respectively. Hepatotoxicity was investigated by determination of lactate dehydrogenase (LDH) and aspartate transaminase (AST) level in hepatocyte culture supernatants, as well as the intracellular formation of malondialdehyde (MDA). RESULTS: A dose-and time-dependent response between ethanol exposure and CYP2E1 activity in human hepatocytes was demonstrated. Moreover, there was a time-dependent increase of CYP2E1 protein after 100 mmol/L ethanol exposure. Meanwhile, ethanol exposure of hepatocytes caused a time-dependent increase of cellular MDA level, LDH, and AST activities in supernatants. Furthermore, the inhibitor of CYP2E1, diallyl sulfide (DAS) could partly attenuate the increases of MDA, LDH, and AST in human hepatocytes. CONCLUSION: A positive relationship between ethanol-induced oxidative damage in human primary cultured hepatocytes and CYP2E1 activity was exhibited, and the inhibition of CYP2E1 could partly attenuate ethanol-induced oxidative damage. PMID:16052683

  3. A cell-based approach to the human proteome project.

    PubMed

    Kelleher, Neil L

    2012-10-01

    The general scope of a project to determine the protein molecules that comprise the cells within the human body is framed. By focusing on protein primary structure as expressed in specific cell types, this concept for a cell-based version of the Human Proteome Project (CB-HPP) is crafted in a manner analogous to the Human Genome Project while recognizing that cells provide a primary context in which to define a proteome. Several activities flow from this articulation of the HPP, which enables the definition of clear milestones and deliverables. The CB-HPP highlights major gaps in our knowledge regarding cell heterogeneity and protein isoforms, and calls for development of technology that is capable of defining all human cell types and their proteomes. The main activities will involve mapping and sorting cell types combined with the application of beyond the state-of-the art in protein mass spectrometry.

  4. A Cell-Based Approach to the Human Proteome Project

    NASA Astrophysics Data System (ADS)

    Kelleher, Neil L.

    2012-10-01

    The general scope of a project to determine the protein molecules that comprise the cells within the human body is framed. By focusing on protein primary structure as expressed in specific cell types, this concept for a cell-based version of the Human Proteome Project (CB-HPP) is crafted in a manner analogous to the Human Genome Project while recognizing that cells provide a primary context in which to define a proteome. Several activities flow from this articulation of the HPP, which enables the definition of clear milestones and deliverables. The CB-HPP highlights major gaps in our knowledge regarding cell heterogeneity and protein isoforms, and calls for development of technology that is capable of defining all human cell types and their proteomes. The main activities will involve mapping and sorting cell types combined with the application of beyond the state-of-the art in protein mass spectrometry.

  5. Generation of Corneal Keratocytes from Human Embryonic Stem Cells

    PubMed Central

    Hertsenberg, Andrew J.; Funderburgh, James L.

    2017-01-01

    Human Embryonic Stem Cells (hESC) offer an important resource as a limitless supply of any differentiated cell type of the human body. Keratocytes, cells from the corneal stroma, may have the potential for restoration of vision in cell therapy and biomedical engineering applications, but these specialized cells are not readily expanded in vitro. Here we describe a two-part method to produce keratocytes from the H1 hESC cell line. The hESC cells, maintained and expanded in feeder-free culture medium are first differentiated to neural crest cells using the stromal-derived inducing activity (SDIA) of the PA6 mouse embryonic fibroblast cell line. The resulting neural crest cells are selected by their expression of cell-surface CD271 and subsequently cultured as 3D pellets in a defined differentiation medium to induce a keratocyte phenotype. PMID:26026882

  6. Nucleosome Organization in Human Embryonic Stem Cells.

    PubMed

    Yazdi, Puya G; Pedersen, Brian A; Taylor, Jared F; Khattab, Omar S; Chen, Yu-Han; Chen, Yumay; Jacobsen, Steven E; Wang, Ping H

    2015-01-01

    The fundamental repeating unit of eukaryotic chromatin is the nucleosome. Besides being involved in packaging DNA, nucleosome organization plays an important role in transcriptional regulation and cellular identity. Currently, there is much debate about the major determinants of the nucleosome architecture of a genome and its significance with little being known about its role in stem cells. To address these questions, we performed ultra-deep sequencing of nucleosomal DNA in two human embryonic stem cell lines and integrated our data with numerous epigenomic maps. Our analyses have revealed that the genome is a determinant of nucleosome organization with transcriptionally inactive regions characterized by a "ground state" of nucleosome profiles driven by underlying DNA sequences. DNA sequence preferences are associated with heterogeneous chromatin organization around transcription start sites. Transcription, histone modifications, and DNA methylation alter this "ground state" by having distinct effects on both nucleosome positioning and occupancy. As the transcriptional rate increases, nucleosomes become better positioned. Exons transcribed and included in the final spliced mRNA have distinct nucleosome profiles in comparison to exons not included at exon-exon junctions. Genes marked by the active modification H3K4m3 are characterized by lower nucleosome occupancy before the transcription start site compared to genes marked by the inactive modification H3K27m3, while bivalent domains, genes associated with both marks, lie exactly in the middle. Combinatorial patterns of epigenetic marks (chromatin states) are associated with unique nucleosome profiles. Nucleosome organization varies around transcription factor binding in enhancers versus promoters. DNA methylation is associated with increasing nucleosome occupancy and different types of methylations have distinct location preferences within the nucleosome core particle. Finally, computational analysis of nucleosome

  7. Cognate HLA absence in trans diminishes human NK cell education

    PubMed Central

    Landtwing, Vanessa; Raykova, Ana; Pezzino, Gaetana; Béziat, Vivien; Graf, Claudine; Moretta, Alessandro; Capaul, Riccarda; Zbinden, Andrea; Malmberg, Karl-Johan; Chijioke, Obinna; Münz, Christian

    2016-01-01

    NK cells are innate lymphocytes with protective functions against viral infections and tumor formation. Human NK cells carry inhibitory killer cell Ig-like receptors (KIRs), which recognize distinct HLAs. NK cells with KIRs for self-HLA molecules acquire superior cytotoxicity against HLA– tumor cells during education for improved missing-self recognition. Here, we reconstituted mice with human hematopoietic cells from donors with homozygous KIR ligands or with a mix of hematopoietic cells from these homozygous donors, allowing assessment of the resulting KIR repertoire and NK cell education. We found that co-reconstitution with 2 KIR ligand–mismatched compartments did not alter the frequency of KIR-expressing NK cells. However, NK cell education was diminished in mice reconstituted with parallel HLA compartments due to a lack of cognate HLA molecules on leukocytes for the corresponding KIRs. This change in NK cell education in mixed human donor–reconstituted mice improved NK cell–mediated immune control of EBV infection, indicating that mixed hematopoietic cell populations could be exploited to improve NK cell reactivity against leukotropic pathogens. Taken together, these findings indicate that leukocytes lacking cognate HLA ligands can disarm KIR+ NK cells in a manner that may decrease HLA– tumor cell recognition but allows for improved NK cell–mediated immune control of a human γ-herpesvirus. PMID:27571408

  8. Intravenous vs intraperitoneal mesenchymal stem cells administration: What is the best route for treating experimental colitis?

    PubMed Central

    Gonçalves, Fabiany da Costa; Schneider, Natália; Pinto, Fernanda Otesbelgue; Meyer, Fabíola Schons; Visioli, Fernanda; Pfaffenseller, Bianca; Lopez, Patrícia Luciana da Costa; Passos, Eduardo Pandolfi; Cirne-Lima, Elizabeth Obino; Meurer, Luíse; Paz, Ana Helena

    2014-01-01

    AIM: To investigate the therapeutic effects of mesenchymal stem cells (MSCs) transplanted intraperitoneally and intravenously in a murine model of colitis. METHODS: MSCs were isolated from C57BL/6 mouse adipose tissue. MSC cultures were analyzed according to morphology, cellular differentiation potential, and surface molecular markers. Experimental acute colitis was induced in C57BL/6 mice by oral administration of 2% dextran sulfate sodium (DSS) in drinking water ad libitum from days 0 to 7. Colitis mice were treated with 1 × 106 MSCs via intraperitoneal or intravenous injection on days 2 and 5. The disease activity index was determined daily based on the following parameters: weight loss, stool consistency and presence of blood in the feces and anus. To compare morphological and functional differences in tissue regeneration between different MSC injection modalities, mice were euthanized on day 8, and their colons were examined for length, weight, and histopathological changes. Inflammatory responses were determined by measuring the levels of different serum cytokines using a CBA Th1/Th2/Th17 kit. Apoptotic rates were evaluated by terminal deoxynucleotidyl transferase-mediated dUDP-biotin nick end labeling assay. RESULTS: Intravenous infusion of MSCs was more effective than intraperitoneal treatment (P < 0.001) in reducing the clinical and histopathologic severity of colitis, which includes weight loss, diarrhea and inflammation. An histological evaluation demonstrated decreased colonic inflammation based on reduced crypt loss and reduced infiltration of inflammatory cells. This therapeutic effect was most likely mediated by the down-regulation of pro-inflammatory cytokines [interleukin (IL)-6 and tumor necrosis factor (TNF)]; and by the up-regulation of anti-inflammatory cytokines (IL-10 and IL-4). Intravenous transplantation also induced high levels of IFN that lead to activation of the immunosuppressive activity of the MSCs, which did not occur with

  9. Extracellular protonation modulates cell-cell interaction mechanics and tissue invasion in human melanoma cells

    PubMed Central

    Hofschröer, Verena; Koch, Kevin Alexander; Ludwig, Florian Timo; Friedl, Peter; Oberleithner, Hans; Stock, Christian; Schwab, Albrecht

    2017-01-01

    Detachment of cells from the primary tumour precedes metastatic progression by facilitating cell release into the tissue. Solid tumours exhibit altered pH homeostasis with extracellular acidification. In human melanoma, the Na+/H+ exchanger NHE1 is an important modifier of the tumour nanoenvironment. Here we tested the modulation of cell-cell-adhesion by extracellular pH and NHE1. MV3 tumour spheroids embedded in a collagen matrix unravelled the efficacy of cell-cell contact loosening and 3D emigration into an environment mimicking physiological confinement. Adhesive interaction strength between individual MV3 cells was quantified using atomic force microscopy and validated by multicellular aggregation assays. Extracellular acidification from pHe7.4 to 6.4 decreases cell migration and invasion but increases single cell detachment from the spheroids. Acidification and NHE1 overexpression both reduce cell-cell adhesion strength, indicated by reduced maximum pulling forces and adhesion energies. Multicellular aggregation and spheroid formation are strongly impaired under acidification or NHE1 overexpression. We show a clear dependence of melanoma cell-cell adhesion on pHe and NHE1 as a modulator. These effects are opposite to cell-matrix interactions that are strengthened by protons extruded via NHE1. We conclude that these opposite effects of NHE1 act synergistically during the metastatic cascade. PMID:28205573

  10. Extracellular protonation modulates cell-cell interaction mechanics and tissue invasion in human melanoma cells.

    PubMed

    Hofschröer, Verena; Koch, Kevin Alexander; Ludwig, Florian Timo; Friedl, Peter; Oberleithner, Hans; Stock, Christian; Schwab, Albrecht

    2017-02-13

    Detachment of cells from the primary tumour precedes metastatic progression by facilitating cell release into the tissue. Solid tumours exhibit altered pH homeostasis with extracellular acidification. In human melanoma, the Na(+)/H(+) exchanger NHE1 is an important modifier of the tumour nanoenvironment. Here we tested the modulation of cell-cell-adhesion by extracellular pH and NHE1. MV3 tumour spheroids embedded in a collagen matrix unravelled the efficacy of cell-cell contact loosening and 3D emigration into an environment mimicking physiological confinement. Adhesive interaction strength between individual MV3 cells was quantified using atomic force microscopy and validated by multicellular aggregation assays. Extracellular acidification from pHe7.4 to 6.4 decreases cell migration and invasion but increases single cell detachment from the spheroids. Acidification and NHE1 overexpression both reduce cell-cell adhesion strength, indicated by reduced maximum pulling forces and adhesion energies. Multicellular aggregation and spheroid formation are strongly impaired under acidification or NHE1 overexpression. We show a clear dependence of melanoma cell-cell adhesion on pHe and NHE1 as a modulator. These effects are opposite to cell-matrix interactions that are strengthened by protons extruded via NHE1. We conclude that these opposite effects of NHE1 act synergistically during the metastatic cascade.

  11. Delivery of iron to human cells by bovine transferrin. Implications for the growth of human cells in vitro.

    PubMed Central

    Young, S P; Garner, C

    1990-01-01

    Following suggestions that transferrin present in fetal-bovine serum, a common supplement used in tissue-culture media, may not bind well to human cells, we have isolated the protein and investigated its interaction with both human and bovine cells. Bovine transferrin bound to a human cell line, K562, at 4 degrees C with a kd of 590 nM, whereas human transferrin bound with a kd of 3.57 nM, a 165-fold difference. With a bovine cell line, NBL4, bovine transferrin bound with the higher affinity, kd 9.09 nM, whereas human transferrin bound with a kd of 41.7 nM, only a 5-fold difference. These values were reflected in an 8.6-fold difference in the rate of iron delivery by the two proteins to human cells, whereas delivery to bovine cells was the same. Nevertheless, the bovine transferrin was taken up by the human cells by a specific receptor-mediated process. Human cells cultured in bovine diferric transferrin at 40 micrograms/ml, the concentration expected in the presence of 10% fetal-bovine serum, failed to thrive, whereas cells cultured in the presence of human transferrin proliferated normally. These results suggest that growth of human cells in bovine serum could give rise to a cellular iron deficiency, which may in turn lead to the selection of clones of cells adapted for survival with less iron. This has important consequences for the use of such cells as models, since they may have aberrant iron-dependent pathways and perhaps other unknown alterations in cell function. PMID:2302189

  12. Evidence for G-quadruplex DNA in human cells.

    PubMed

    Xu, Yan; Komiyama, Makoto

    2013-05-27

    Seen in human cells: A fluorochrome-labeled antibody probe selectively and efficiently binds all types of DNA G-quadruplex with similar binding affinities, but hardly binds single- or double-stranded DNA, or RNA hairpins. Thus, this antibody strictly discriminates between G-quadruplex structures and other conformations of DNA and provides evidence for G-quadruplex DNA in human cells.

  13. Effects of Humanism on Education and Educational Administration. Educational Research Report.

    ERIC Educational Resources Information Center

    Lall, Bernard M.; Nicholas, Anthony

    This report summarizes the historical development of humanism from Petrarch and Boccaccio in the Renaissance to the present. The freedom of thought, self-expression, and creative activity that characterized the humanistic schools are considered to be the fundamental bases of education today. A brief description is given of brutalization and…

  14. Canadian Human Rights Act. Office Consolidation = Loi canadienne sur les droits de la personne. Codification administrative.

    ERIC Educational Resources Information Center

    Department of Supply and Services, Ottawa (Ontario).

    The Canadian Human Rights Act extends the laws in Canada that proscribe discrimination by establishing that each individual has the right to make the life for him- or herself that he or she is able and wishes to have, consistent with the duties and obligations of a member of society, without being hindered or prevented from doing so by…

  15. Human white blood cells contain cyclobutyl pyrimidine dimer photolyase

    SciTech Connect

    Sutherland, B.M.; Bennett, P.V.

    1995-10-10

    Although enzymatic photoreactivation of cyclobutyl pyrimidine dimers in DNA is present in almost all organisms, its presence in placental mammals is controversial. We tested human white blood cells for photolyase by using three defined DNAs (suprecoiled pET-2, nonsupercoiled bacteriphage {lambda}, and a defined-sequence 287-bp oligonucleotide), two dimer-specific endonucleases (T4 endonuclease V and UV endonuclease from Micrococcus luteus), and three assay methods. We show that human white blood cells contain photolyase that can photorepair pyrimidine dimers in defined supercoiled and linear DNAs and in a 287-bp oligonucleotide and that human photolyase is active on genomic DNA in intact human cells. 44 refs., 3 figs.

  16. Human dental pulp stem cells: Applications in future regenerative medicine

    PubMed Central

    Potdar, Pravin D; Jethmalani, Yogita D

    2015-01-01

    Stem cells are pluripotent cells, having a property of differentiating into various types of cells of human body. Several studies have developed mesenchymal stem cells (MSCs) from various human tissues, peripheral blood and body fluids. These cells are then characterized by cellular and molecular markers to understand their specific phenotypes. Dental pulp stem cells (DPSCs) are having a MSCs phenotype and they are differentiated into neuron, cardiomyocytes, chondrocytes, osteoblasts, liver cells and β cells of islet of pancreas. Thus, DPSCs have shown great potentiality to use in regenerative medicine for treatment of various human diseases including dental related problems. These cells can also be developed into induced pluripotent stem cells by incorporation of pluripotency markers and use for regenerative therapies of various diseases. The DPSCs are derived from various dental tissues such as human exfoliated deciduous teeth, apical papilla, periodontal ligament and dental follicle tissue. This review will overview the information about isolation, cellular and molecular characterization and differentiation of DPSCs into various types of human cells and thus these cells have important applications in regenerative therapies for various diseases. This review will be most useful for postgraduate dental students as well as scientists working in the field of oral pathology and oral medicine. PMID:26131314

  17. Human dental pulp stem cells: Applications in future regenerative medicine.

    PubMed

    Potdar, Pravin D; Jethmalani, Yogita D

    2015-06-26

    Stem cells are pluripotent cells, having a property of differentiating into various types of cells of human body. Several studies have developed mesenchymal stem cells (MSCs) from various human tissues, peripheral blood and body fluids. These cells are then characterized by cellular and molecular markers to understand their specific phenotypes. Dental pulp stem cells (DPSCs) are having a MSCs phenotype and they are differentiated into neuron, cardiomyocytes, chondrocytes, osteoblasts, liver cells and β cells of islet of pancreas. Thus, DPSCs have shown great potentiality to use in regenerative medicine for treatment of various human diseases including dental related problems. These cells can also be developed into induced pluripotent stem cells by incorporation of pluripotency markers and use for regenerative therapies of various diseases. The DPSCs are derived from various dental tissues such as human exfoliated deciduous teeth, apical papilla, periodontal ligament and dental follicle tissue. This review will overview the information about isolation, cellular and molecular characterization and differentiation of DPSCs into various types of human cells and thus these cells have important applications in regenerative therapies for various diseases. This review will be most useful for postgraduate dental students as well as scientists working in the field of oral pathology and oral medicine.

  18. Asbestos-associated chromosomal changes in human mesothelial cells

    SciTech Connect

    Lechner, J.F.; Tokiwa, T.; LaVeck, M.; Benedict, W.F.; Banks-Schlegel, S.; Yeager, H. Jr.; Banerjee, A.; Harris, C.C.

    1985-06-01

    Replicative cultures of human pleural mesothelial cells were established from noncancerous adult donors. The cells exhibited normal mesothelial cell characteristics including keratin, hyaluronic acid mucin, and long branched microvilli, and they retained the normal human karyotype until senescence. The mesothelial cells were 10 and 100 times more sensitive to the cytotoxic effects of asbestos fibers than normal human bronchial epithelial or fibroblastic cells, respectively. In addition, cultures of mesothelial cells that survived two cytotoxic exposures of amosite fibers were aneuploid with consistent specific chromosomal losses indicative of clonal origin. These aneuploid cells exhibit both altered growth control properties and a population doubling potential of >50 divisions beyond the culture life span (30 doublings) of the control cells.

  19. Effects of mesenchymal stromal cells on human myeloid dendritic cell differentiation and maturation in a humanized mouse model.

    PubMed

    Chen, Ping; Huang, Yanfei; Womer, Karl L

    2015-12-01

    Mesenchymal stromal cells (MSCs) have shown promise as cellular therapy in allogeneic transplantation, although the precise mechanisms underlying their benefit in clinical trials are difficult to study. We previously demonstrated that MSCs exert immunoregulatory effects in mouse bone marrow-derived dendritic cell (DC) culture. Since mouse studies do not reliably reproduce human events, we used a humanized mouse model to study the immunomodulatory effects of human MSCs on human DC immunobiology. Humanized mice were established by injection of cord blood CD34(+) cells into NOD.Cg-Prkdc(scid) Il2rg(tm1Wjl/SzJ) (NOD scid gamma, NSG) mice. Human cells were detected in the mouse bone marrow, blood, and spleen 12weeks after transplantation. Human DCs were differentiated from humanized mouse bone marrow cells during human MSC co-culture. MSCs inhibited DC differentiation and kept DCs in an immature state as demonstrated by phenotype and function. In conclusion, humanized mouse models represent a useful method to study the function of human MSCs on human DC immunobiology.

  20. A proteomic study on a human osteosarcoma cell line Saos-2 treated with diallyl trisulfide.

    PubMed

    Zhang, Yong Kui; Zhang, Xu Hua; Li, Jian Min; Sun, De Sheng; Yang, Qiang; Diao, Dong Mei

    2009-09-01

    Garlic is generally used as a therapeutic reagent against various diseases, and numerous studies have indicated that garlic and its derivatives can reduce the risk of various types of human cancer. Diallyl trisulfide (DATS), a major member of garlic derivatives, could inhibit the cell proliferation by triggering either cell cycle arrest or apoptosis in a variety of cancer cell lines as shown in many studies. However, whether DATS has the same effect on human osteosarcoma cells remains unknown. In this study, we have attempted to analyze the effects of DATS on cell proliferation, cell cycle, induction of apoptosis, global protein expression pattern in a human osteosarcoma cell line Saos-2 cells, and the potential molecular mechanisms of the action of DATS. Saos-2 cells, a human osteosarcoma cell line, were treated with or without 25, 50, and 100 micromol/l DATS for various time intervals. The cell proliferation, cell cycle progression, and apoptosis were examined in this study. Then, after treatment with or without 50 micromol/l DATS for 48 h, protein add pattern in Saos-2 cells were systematically studied using two-dimensional electrophoresis and mass spectrometry. DATS could inhibit the proliferation of Saos-2 cells in a dose-dependent and time-dependent manner. Moreover, the percentage of apoptotic cell and cell arrest in G0/G1 phase was also dose-dependent and time-dependent upon DATS treatment. A total of 27 unique proteins in Saos-2 cells, including 18 downregulated proteins and nine upregulated proteins, were detected with significant changes in their expression levels corresponding to DATS administration. Interestingly, almost half of these proteins (13 of 27) are related to either the cell cycle or apoptosis. DATS has the ability to suppress cell proliferation of Saos-2 cells by blocking cell cycle progression and inducing apoptosis in a dose and time-dependent manner. The proteomic results presented, therefore, provide additional support to the hypothesis

  1. Extracts of centipede Scolopendra subspinipes mutilans induce cell cycle arrest and apoptosis in A375 human melanoma cells.

    PubMed

    Ma, Weina; Liu, Rui; Qi, Junpeng; Zhang, Yanmin

    2014-07-01

    Extracts from the centipede Scolopendra genus, have been used in traditional medicine for the treatment of various diseases and have been found to exhibit anticancer activity in tumor cells. To investigate the potential and associated antitumor mechanism of alcohol extracts of the centipede Scolopendra subspinipes mutilans (AECS), cell viability, cell cycle and cell apoptosis were studied and the results revealed that AECS inhibits A375 cell proliferation in a dose- and time-dependent manner. In addition, AECS was found to arrest the cell cycle of A375 cells at the S phase, which was accompanied by a marked increase in the protein levels of cyclin E and a decrease in the protein levels of cyclin D1. In a cell culture system, AECS markedly induced the apoptosis of A375 cells, which was closely associated with the effects on the Bcl-2 family, whereby decreased Bcl-2 and increased Bak, Bax and Bad expression levels were observed. The underlying mechanism of AECS inhibiting A375 cell proliferation was associated with the induction of cell cycle arrest and apoptosis, indicating that AECS may present as a potential therapeutic agent for administration in human melanoma cancer intervention.

  2. Dendritic cell-mediated immune humanization of mice: implications for allogeneic and xenogeneic stem cell transplantation.

    PubMed

    Salguero, Gustavo; Daenthanasanmak, Anusara; Münz, Christian; Raykova, Ana; Guzmán, Carlos A; Riese, Peggy; Figueiredo, Constanca; Länger, Florian; Schneider, Andreas; Macke, Laura; Sundarasetty, Bala Sai; Witte, Torsten; Ganser, Arnold; Stripecke, Renata

    2014-05-15

    De novo regeneration of immunity is a major problem after allogeneic hematopoietic stem cell transplantation (HCT). HCT modeling in severely compromised immune-deficient animals transplanted with human stem cells is currently limited because of incomplete maturation of lymphocytes and scarce adaptive responses. Dendritic cells (DC) are pivotal for the organization of lymph nodes and activation of naive T and B cells. Human DC function after HCT could be augmented with adoptively transferred donor-derived DC. In this study, we demonstrate that adoptive transfer of long-lived human DC coexpressing high levels of human IFN-α, human GM-CSF, and a clinically relevant Ag (CMV pp65 protein) promoted human lymphatic remodeling in immune-deficient NOD.Rag1(-/-).IL-2rγ(-/-) mice transplanted with human CD34(+) cells. After immunization, draining lymph nodes became replenished with terminally differentiated human follicular Th cells, plasma B cells, and memory helper and cytotoxic T cells. Human Igs against pp65 were detectable in plasma, demonstrating IgG class-switch recombination. Human T cells recovered from mice showed functional reactivity against pp65. Adoptive immunotherapy with engineered DC provides a novel strategy for de novo immune reconstitution after human HCT and a practical and effective tool for studying human lymphatic regeneration in vivo in immune deficient xenograft hosts.

  3. Genetic engineering of human pluripotent cells using TALE nucleases.

    PubMed

    Hockemeyer, Dirk; Wang, Haoyi; Kiani, Samira; Lai, Christine S; Gao, Qing; Cassady, John P; Cost, Gregory J; Zhang, Lei; Santiago, Yolanda; Miller, Jeffrey C; Zeitler, Bryan; Cherone, Jennifer M; Meng, Xiangdong; Hinkley, Sarah J; Rebar, Edward J; Gregory, Philip D; Urnov, Fyodor D; Jaenisch, Rudolf

    2011-07-07

    Targeted genetic engineering of human pluripotent cells is a prerequisite for exploiting their full potential. Such genetic manipulations can be achieved using site-specific nucleases. Here we engineered transcription activator-like effector nucleases (TALENs) for five distinct genomic loci. At all loci tested we obtained human embryonic stem cell (ESC) and induced pluripotent stem cell (iPSC) clones carrying transgenic cassettes solely at the TALEN-specified location. Our data suggest that TALENs employing the specific architectures described here mediate site-specific genome modification in human pluripotent cells with similar efficiency and precision as do zinc-finger nucleases (ZFNs).

  4. A NEW CRYSTAL-CONTAINING CELL IN HUMAN ADRENAL CORTEX

    PubMed Central

    Magalhães, Maria C.

    1972-01-01

    Electron microscope examination of the adrenal cortex from three male human subjects revealed a special type of cell occurring in periendothelial spaces, in all adrenal cortex zones. It is a clear, spindle-shaped cell the principal cytoplasmic features of which are crystalline inclusions with a structure similar to that of the Reinke crystals of human testicular interstitial cells and an abundance of microfilaments. Enzymatic digestions with pronase, pepsin, and ribonuclease were performed, and no digestion of the crystals was obtained. The crystals had no peroxidase or acid phosphatase activities. This cell appears to be exclusive to human males and it may be related to adrenal androgen secretion. PMID:4347248

  5. Purification and cultivation of human pituitary growth hormone secreting cells

    NASA Technical Reports Server (NTRS)

    Hymer, W. C.

    1978-01-01

    The maintainance of actively secreting human pituitary growth hormone cells (somatotrophs) in vitro was studied. The primary approach was the testing of agents which may be expected to increase the release of the human growth hormone (hGH). A procedure for tissue procurement is described along with the methodologies used to dissociate human pituitary tissue (obtained either at autopsy or surgery) into single cell suspensions. The validity of the Biogel cell column perfusion system for studying the dynamics of GH release was developed and documented using a rat pituitary cell system.

  6. Albumin synthesis in humans increases immediately following the administration of endotoxin.

    PubMed

    Barle, Hans; Januszkiewicz, Anna; Hållström, Lars; Essén, Pia; McNurlan, Margaret A; Garlick, Peter J; Wernerman, Jan

    2002-11-01

    In order to investigate the immediate (i.e. within 3 h) response of albumin synthesis to the administration of endotoxin, as a model of a moderate and well controlled catabolic insult, two measurements employing L-[(2)H(5)]phenylalanine were performed in 16 volunteers. One group ( n =8) received an intravenous injection of endotoxin (4 ng/kg; lot EC-6) immediately after the first measurement of albumin synthesis, whereas the other group received saline. A second measurement was initiated 1 h later. In the endotoxin group, the fractional synthesis rate of albumin was 6.9+/-0.6%/day (mean+/-S.D.) in the first measurement. In the second measurement, a significant increase was observed (9.6+/-1.2%/day; P <0.001). The corresponding values in the control group were were 6.6+/-0.6%/day and 7.0+/-0.6%/day respectively (not significant compared with first measurement and P <0.001 compared with the second measurement in the endotoxin group). The absolute synthesis rates of albumin were 148+/-35 and 201+/-49 mg x kg(-1) x day(-1) before and after endotoxin ( P <0.01). In the control group, the corresponding values were 131+/-21 and 132+/-20 mg x kg(-1) x day(-1) (not significant compared with the first measurement and P <0.01 compared with the second measurement in the endotoxin group). In conclusion, these results indicate that albumin synthesis increases in the very early phase after a catabolic insult, as represented by the administration of endotoxin.

  7. Embryonic death and the creation of human embryonic stem cells.

    PubMed

    Landry, Donald W; Zucker, Howard A

    2004-11-01

    The creation of human embryonic stem cells through the destruction of a human embryo pits the value of a potential therapeutic tool against that of an early human life. This contest of values has resulted in a polarized debate that neglects areas of common interest and perspective. We suggest that a common ground for pursuing research on human embryonic stem cells can be found by reconsidering the death of the human embryo and by applying to this research the ethical norms of essential organ donation.

  8. Modeling Aggressive Medulloblastoma Using Human-Induced Pluripotent Stem Cells

    DTIC Science & Technology

    2015-07-01

    AWARD NUMBER: W81XWH-14-1-0176 TITLE: Modeling Aggressive Medulloblastoma Using Human-Induced Pluripotent Stem Cells PRINCIPAL INVESTIGATOR...Prescribed by ANSI Std. Z39.18 July 2015 Annual 01-July 2014 -- 30 Jun 2015 Modeling Aggressive Medulloblastoma Using Human-Induced Pluripotent Stem...induced pluripotent stem cells by Atoh1 induction can be efficiently transformed by MYC oncogene to form aggressive brain tumors that recapitulate human

  9. [Research with human embryo stem cells. Foundations and judicial limits].

    PubMed

    Eser, Albin; Koch, Hans-Georg

    2004-01-01

    Research with human embryos, and particularly, the use for scientific purposes of human embryonic stem cells has given raise to different sort of problems at the international level. One of the most strict regulation in this field, is this lecture Professors Albin Eser and Hans-Georg Koch analyse the german legal framework in relation with the use of embryos and human embryonic stem cells for scientific purposes.

  10. Induction of cell proliferation in the rat liver by the short-term administration of ethyl tertiary-butyl ether.

    PubMed

    Kakehashi, Anna; Hagiwara, Akihiro; Imai, Norio; Wei, Min; Fukushima, Shoji; Wanibuchi, Hideki

    2015-01-01

    In the present study, in continuation of our previous experiment in order to investigate the mode of action (MOA) of ethyl tertiary-butyl ether (ETBE) hepatotumorigenicity in rats, we aimed to examine alterations in cell proliferation, that are induced by short-term administration of ETBE. F344 rats were administered ETBE at doses of 0, and 1,000 mg/kg body weight twice a day by gavage for 3, 10, 17 and 28 days. It was found that the previously observed significant increase of P450 total content and hydroxyl radical levels after 7 days of ETBE administration, and 8-OHdG formation at day 14, accompanied by accumulation of CYP2B1/2B2, CYP3A1/3A2, CYP2C6, CYP2E1 and CYP1A1 and downregulation of DNA oxoguanine glycosylase 1, was preceded by induction of cell proliferation at day 3. Furthermore, we observed an increase in regenerative cell proliferation as a result of ETBE treatment at day 28, followed by induction of cell cycle arrest and apoptosis by day 14. These results indicated that short-term administration of ETBE led to a significant early increase in cell proliferation activity associated with induction of oxidative stress, and to a regenerative cell proliferation as an adaptive response, which could contribute to the hepatotumorigenicity of ETBE in rats.

  11. Histamine release from human buffy coat-derived mast cells.

    PubMed

    Wang, Xian Song; Lau, Hang Yung Alaster

    2007-04-01

    Mast cells are unique immune cells that release a spectrum of chemical mediators contributing to the inflammatory symptoms of allergic disorders. Although mast cell biology has been extensively studied in the rodents, research on human mast cells is hampered by the lack of a convenient preparation source. This problem has now been addressed by culturing human mast cells from CD34(+) progenitors. We have recently discovered that human buffy coat preparations from local blood banks are an abundant and convenient source of progenitors for culturing mature mast cells which express functional high affinity IgE receptors and contain histamine and tryptase in their granules. In the current study, we further characterize these buffy coat-derived mast cells by studying their responses to common mast cell secretagogues and stabilizers. Mature human mast cells were obtained by culturing isolated progenitors in methylcellulose containing stem cell factor (SCF), IL-3 and IL-6 for 6 weeks and subsequently in liquid medium containing SCF and IL-6 for another 6 to 8 weeks. Following sensitisation with human IgE, these cells released histamine dose-dependently upon activation by anti-IgE and calcium ionophores while compound 48/80 and substance P were relatively ineffective. When the effects of anti-asthmatic agents on anti-IgE-induced mediator release from these cells were compared, only the beta(2)-adrenoceptor agonists and phosphodiesterase inhibitors produced dose-dependent inhibition but not cromolyn or nedocromil. In total, mast cells cultured from human buffy coat progenitors shared similar functional properties of MC(T) subtype of mast cells found predominantly in human lung parenchyma and intestinal mucosa.

  12. Regulation of Human Helper T Cell Subset Differentiation by Cytokines

    PubMed Central

    Schmitt, Nathalie; Ueno, Hideki

    2015-01-01

    Since the discovery of Th1 and Th2 cells in the late 80’s, the family of effector CD4+ helper T (Th) cell subsets has expanded. The differentiation of naïve CD4+ T cells is largely determined when they interact with dendritic cells in lymphoid organs, and cytokines play a major role in the regulation of Th differentiation in the early stages. Recent studies show that the developmental mechanism of certain Th subsets is not fully shared between mice and humans. Here we will review recent discoveries on the roles of cytokines in the regulation of Th differentiation in humans, and discuss the differences between mice and humans in the developmental mechanisms of several Th subsets, including Th17 cells and T follicular helper (Tfh) cells. We propose that the differentiation of human Th subsets is largely regulated by the three cytokines, IL-12, IL-23, and TGF-β. PMID:25879814

  13. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2011-06-11

    astrocytes using defined medium conditions, (3) cell-based methods to detect botulinum toxin, and (4) HTS amenable assays for proliferation...progenitors into dopaminergic neurons, motoneurons and astrocytes using defined medium conditions, (3) cell-based methods to detect botulinum toxin...cell line developed for potential commercial distribution. (3) Development of cell based methods to detect botulinum toxin There has been

  14. Downregulation of Rap1GAP in Human Tumor Cells Alters Cell/Matrix and Cell/Cell Adhesion▿ †

    PubMed Central

    Tsygankova, Oxana M.; Ma, Changqing; Tang, Waixing; Korch, Christopher; Feldman, Michael D.; Lv, Yu; Brose, Marcia S.; Meinkoth, Judy L.

    2010-01-01

    Rap1GAP expression is decreased in human tumors. The significance of its downregulation is unknown. We show that Rap1GAP expression is decreased in primary colorectal carcinomas. To elucidate the advantages conferred on tumor cells by loss of Rap1GAP, Rap1GAP expression was silenced in human colon carcinoma cells. Suppressing Rap1GAP induced profound alterations in cell adhesion. Rap1GAP-depleted cells exhibited defects in cell/cell adhesion that included an aberrant distribution of adherens junction proteins. Depletion of Rap1GAP enhanced adhesion and spreading on collagen. Silencing of Rap expression normalized spreading and restored E-cadherin, β-catenin, and p120-catenin to cell/cell contacts, indicating that unrestrained Rap activity underlies the alterations in cell adhesion. The defects in adherens junction protein distribution required integrin signaling as E-cadherin and p120-catenin were restored at cell/cell contacts when cells were plated on poly-l-lysine. Unexpectedly, Src activity was increased in Rap1GAP-depleted cells. Inhibition of Src impaired spreading and restored E-cadherin at cell/cell contacts. These findings provide the first evidence that Rap1GAP contributes to cell/cell adhesion and highlight a role for Rap1GAP in regulating cell/matrix and cell/cell adhesion. The frequent downregulation of Rap1GAP in epithelial tumors where alterations in cell/cell and cell/matrix adhesion are early steps in tumor dissemination supports a role for Rap1GAP depletion in tumor progression. PMID:20439492

  15. Local tetrahydrobiopterin administration augments reflex cutaneous vasodilation through nitric oxide-dependent mechanisms in aged human skin.

    PubMed

    Stanhewicz, Anna E; Bruning, Rebecca S; Smith, Caroline J; Kenney, W Larry; Holowatz, Lacy A

    2012-03-01

    Functional constitutive nitric oxide synthase (NOS) is required for full expression of reflex cutaneous vasodilation that is attenuated in aged skin. Both the essential cofactor tetrahydrobiopterin (BH(4)) and adequate substrate concentrations are necessary for the functional synthesis of nitric oxide (NO) through NOS, both of which are reduced in aged vasculature through increased oxidant stress and upregulated arginase, respectively. We hypothesized that acute local BH(4) administration or arginase inhibition would similarly augment reflex vasodilation in aged skin during passive whole body heat stress. Four intradermal microdialysis fibers were placed in the forearm skin of 11 young (22 ± 1 yr) and 11 older (73 ± 2 yr) men and women for local infusion of 1) lactated Ringer, 2) 10 mM BH(4), 3) 5 mM (S)-(2-boronoethyl)-l-cysteine + 5 mM N(ω)-hydroxy-nor-l-arginine to inhibit arginase, and 4) 20 mM N(G)-nitro-l-arginine methyl ester (l-NAME) to inhibit NOS. Red cell flux was measured at each site by laser-Doppler flowmetry (LDF) as reflex vasodilation was induced. After a 1.0°C rise in oral temperature (T(or)), mean body temperature was clamped and 20 mM l-NAME was perfused at each site. Cutaneous vascular conductance was calculated (CVC = LDF/mean arterial pressure) and expressed as a percentage of maximum (%CVC(max); 28 mM sodium nitroprusside and local heat, 43°C). Vasodilation was attenuated at the control site of the older subjects compared with young beginning at a 0.3°C rise in T(or). BH(4) and arginase inhibition both increased vasodilation in older (BH(4): 55 ± 5%; arginase-inhibited: 47 ± 5% vs. control: 37 ± 3%, both P < 0.01) but not young subjects compared with control (BH(4): 51 ± 4%CVC(max); arginase-inhibited: 55 ± 4%CVC(max) vs. control: 56 ± 6%CVC(max), both P > 0.05) at a 1°C rise in T(or). With a 1°C rise in T(or), local BH(4) increased NO-dependent vasodilation in the older (BH(4): 31.8 ± 2.4%CVC(max) vs. control: 11.7 ± 2.0%CVC

  16. The response of human and rodent cells to hyperthermia

    SciTech Connect

    Roizin-Towle, L.; Pirro, J.P. )

    1991-04-01

    Inherent cellular radiosensitivity in vitro has been shown to be a good predictor of human tumor response in vivo. In contrast, the importance of the intrinsic thermosensitivity of normal and neoplastic human cells as a factor in the responsiveness of human tumors to adjuvant hyperthermia has never been analyzed systematically. A comparison of thermal sensitivity and thermo-radiosensitization in four rodent and eight human-derived cell lines was made in vitro. Arrhenius plots indicated that the rodent cells were more sensitive to heat killing than the human, and the break-point was 0.5 degrees C higher for the human than rodent cells. The relationship between thermal sensitivity and the interaction of heat with X rays at low doses was documented by thermal enhancement ratios (TER's). Cells received either a 1 hr exposure to 43 degrees C or a 20 minute treatment at 45 degrees C before exposure to 300 kVp X rays. Thermal enhancement ratios ranged from 1.0 to 2.7 for human cells heated at 43 degrees C and from 2.1 to 5.3 for heat exposures at 45 degrees C. Thermal enhancement ratios for rodent cells were generally 2 to 3 times higher than for human cells, because of the fact that the greater thermosensitivity of rodent cells results in a greater enhancement of radiation damage. Intrinsic thermosensitivity of human cells has relevance to the concept of thermal dose; intrinsic thermo-radiosensitization of a range of different tumor cells is useful in documenting the interactive effects of radiation combined with heat.

  17. Isolation of inflammatory cells from human tumours.

    PubMed

    Polak, Marta E

    2011-01-01

    Inflammatory cells are present in many tumours, and understanding their function is of increasing importance, particularly to studies of tumour immunology. The tumour-infiltrating leukocytes encompass a variety of cell types, e.g. T lymphocytes, macrophages, dendritic cells, NK cells, and mast cells. Choice of the isolation method greatly depends on the tumour type and the leukocyte subset of interest, but the protocol usually includes tissue disaggregation and cell enrichment. We recommend density centrifugation for initial enrichment, followed by specific magnetic bead negative or positive panning with leukocyte and tumour cell selective antibodies.

  18. Urokinase production by electrophoretically separated cultured human embryonic kidney cells

    NASA Technical Reports Server (NTRS)

    Kunze, M. E.; Plank, L. D.; Giranda, V.; Sedor, K.; Todd, P. W.

    1985-01-01

    Urokinase is a plasminogen activator found in urine. Relatively pure preparations have been tested in Europe, Japan and the United States for the treatment of deep vein thrombosis and other dangerous blood clots. Human embryonic kidney cell cultures have been found to produce urokinase at much higher concentrations, but less than 5% of the cells in typical cultures are producers. Since human diploid cells become senescent in culture the selection of clones derived from single cells will not provide enough material to be useful, so a bulk purification method is needed for the isolation of urokinase producing cell populations. Preparative cell electrophoresis was chosen as the method, since evidence exists that human embryonic cell cultures are richly heterogeneous with respect to electrophoretic mobility, and preliminary electrophoretic separations on the Apollo-Soyuz space flight produced cell populations that were rich in urokinase production. Similarly, erythropoietin is useful in the treatment of certain anemias and is a kidney cell duct, and electrophoretically enriched cell populations producing this product have been reported. Thus, there is a clear need for diploid human cells that produce these products, and there is evidence that such cells should be separable by free-flow cell electrophoresis.

  19. Measurement of naphthoxylactic and naphtoxyacetic acid in human plasma following propranolol administration.

    PubMed

    Pritchard, F; Schneck, D; Hayes, A

    1979-02-01

    A method for the quantitative determination of naphthoxylactic acid and naphthoxyacetic acid, two acidic metabolites of propranolol, in human plasma is described. Preliminary data from hypertensive patients receiving chronic propranolol therapy and from normal volunteers who took a single 80 mg oral propranolol dose revealed that naphthoxylactic acid plasma levels were 4 to 20 times that of propranolol whereas naphthoxyacetic acid levels were equivalent to or less than the plasma concentrations of propranolol.

  20. Immunohistochemical toolkit for tracking and quantifying xenotransplanted human stem cells

    PubMed Central

    Allard, Justine; Li, Ké; Lopez, Xavier Moles; Blanchard, Stéphane; Barbot, Paul; Rorive, Sandrine; Decaestecker, Christine; Pochet, Roland; Bohl, Delphine; Lepore, Angelo C; Salmon, Isabelle; Nicaise, Charles

    2014-01-01

    Aim Biomarker-based tracking of human stem cells xenotransplanted into animal models is crucial for studying their fate in the field of cell therapy or tumor xenografting. Materials & methods Using immunohistochemistry and in situ hybridization, we analyzed the expression of three human-specific biomarkers: Ku80, human mitochondria (hMito) and Alu. Results We showed that Ku80, hMito and Alu biomarkers are broadly expressed in human tissues with no or low cross-reactivity toward rat, mouse or pig tissues. In vitro, we demonstrated that their expression is stable over time and does not change along the differentiation of human-derived induced pluripotent stem cells or human glial-restricted precursors. We tracked in vivo these cell populations after transplantation in rodent spinal cords using aforementioned biomarkers and human-specific antibodies detecting apoptotic, proliferating or neural-committed cells. Conclusion This study assesses the human-species specificity of Ku80, hMito and Alu, and proposes useful biomarkers for characterizing human stem cells in xenotransplantation paradigms. PMID:25159062

  1. Generating trunk neural crest from human pluripotent stem cells.

    PubMed

    Huang, Miller; Miller, Matthew L; McHenry, Lauren K; Zheng, Tina; Zhen, Qiqi; Ilkhanizadeh, Shirin; Conklin, Bruce R; Bronner, Marianne E; Weiss, William A

    2016-01-27

    Neural crest cells (NCC) are stem cells that generate different lineages, including neuroendocrine, melanocytic, cartilage, and bone. The differentiation potential of NCC varies according to the level from which cells emerge along the neural tube. For example, only anterior "cranial" NCC form craniofacial bone, whereas solely posterior "trunk" NCC contribute to sympathoadrenal cells. Importantly, the isolation of human fetal NCC carries ethical and scientific challenges, as NCC induction typically occur before pregnancy is detectable. As a result, current knowledge of NCC biology derives primarily from non-human organisms. Important differences between human and non-human NCC, such as expression of HNK1 in human but not mouse NCC, suggest a need to study human NCC directly. Here, we demonstrate that current protocols to differentiate human pluripotent stem cells (PSC) to NCC are biased toward cranial NCC. Addition of retinoic acid drove trunk-related markers and HOX genes characteristic of a posterior identity. Subsequent treatment with bone morphogenetic proteins (BMPs) enhanced differentiation to sympathoadrenal cells. Our approach provides methodology for detailed studies of human NCC, and clarifies roles for retinoids and BMPs in the differentiation of human PSC to trunk NCC and to sympathoadrenal lineages.

  2. Sex-dimorphic behaviour development in the human: prenatal hormone administration and postnatal socialization.

    PubMed

    Green, R

    An interdisciplinary integrative approach must be utilized in the study of psychosexual differentiation. The approach must capitalize on data derived from non-human models, from experiments of nature, and from experiments of nurture. Studies from non-human primates strongly suggest the influence of prenatal sex hormone levels on postnatal sexually dimorphic behaviours. Starting from this basis we have studied sixty young adult men whose mothers received, during pregnancy, diethylstilboestrol, diethylstilboestrol and natural progesterone, natural progesterone, or synthetic progesterone. They have been compared with matched controls not exposed in utero to exogenous hormones. Studies of socialization patterns must document the differential developmental experiences, if any, of children with atypical and typical sex-typed behaviours. To this end, we are studying 60 boys whose behaviour before puberty was decidedly feminine, and their parents, and contrasting them with masculine boys and their parents. We are also studying 50 girls whose behaviour before puberty was 'masculine', and contrasting them with 'feminine' girls. Additionally, we are studying the sexually dimorphic behaviour of children of sexually atypical parents. The parents have either undergone sex-change surgery (male-to-female or female-to-male) or are homosexual. Data from the three studies are presented. A call is made to researchers working with non-human primates to test and extend these findings.

  3. Intranasal administration of retinal antigens induces transient T cell activation and apoptosis within drainage lymph nodes but not spleen.

    PubMed

    Laliotou, B; Duncan, L; Dick, A D

    1999-05-01

    Mechanisms of mucosal tolerance induction, including anergy/deletion and active suppression are frequently described as mutually exclusive; dependent upon nature, dose and route of antigen administration. We have previously described induction of low-dose tolerance with administration of retinal autoantigens via the nasorespiratory tract which is antigen-specific, suppresses both cell mediated immunity and ultimately tissue destruction in experimental autoimmune uveoretinitis (EAU) and is mediated by splenic-derived regulatory cells. The present data further shows that splenocytes or fractionated splenic T cells, which secrete IL-4 and IL-10 when stimulated with retinal antigen in vitro, and not regional drainage lymph node cells transfer tolerance to naïve animals. Analysis of apparent mechanistic differences shows that during intranasal antigen administration, the proportion of CD4(+)T cells within drainage lymph nodes increases, concurrent with a burst of IFN-gamma. Following subsequent antigen challenge, T cells downregulate alphabetaTCR expression and undergo apoptosis in regional drainage lymph nodes. An increase in functional Th2 cytokine activity was noted in both Con-A and retinal antigen stimulated lymph node cultures in tolerized animals. T cells from tolerized animals secreted IL-4, whereas IL-10 was secreted predominantly by the non-T cell population present equally in control and tolerized animals. Therefore, spleen derived regulatory cells which suppress Th1 responses and T cell deletion/apoptosis in regional drainage lymph nodes are mechanisms which co-exist in tolerant rats. Th2 cytokine production after immunization appears consequential to tolerance-induced Th1 suppression.

  4. E-cadherin-mediated cell-cell adhesion prevents invasiveness of human carcinoma cells

    PubMed Central

    1991-01-01

    The ability of carcinomas to invade and to metastasize largely depends on the degree of epithelial differentiation within the tumors, i.e., poorly differentiated being more invasive than well-differentiated carcinomas. Here we confirmed this correlation by examining various human cell lines derived from bladder, breast, lung, and pancreas carcinomas. We found that carcinoma cell lines with an epithelioid phenotype were noninvasive and expressed the epithelium-specific cell- cell adhesion molecule E-cadherin (also known as Arc-1, uvomorulin, and cell-CAM 120/80), as visualized by immunofluorescence microscopy and by Western and Northern blotting, whereas carcinoma cell lines with a fibroblastoid phenotype were invasive and had lost E-cadherin expression. Invasiveness of these latter cells could be prevented by transfection with E-cadherin cDNA and was again induced by treatment of the transfected cells with anti-E-cadherin mAbs. These findings indicate that the selective loss of E-cadherin expression can generate dedifferentiation and invasiveness of human carcinoma cells, and they suggest further that E-cadherin acts as an invasion suppressor. PMID:2007622

  5. Human mesenchymal stromal cells exert HGF dependent cytoprotective effects in a human relevant pre-clinical model of COPD

    PubMed Central

    Kennelly, Helen; Mahon, Bernard P.; English, Karen

    2016-01-01

    Bone-marrow derived mesenchymal stromal cells (MSCs) have potent immunomodulatory and tissue reparative properties, which may be beneficial in the treatment of inflammatory diseases such as COPD. This study examined the mechanisms by which human MSCs protect against elastase induced emphysema. Using a novel human relevant pre-clinical model of emphysema the efficacy of human MSC therapy and optimal cell dose were investigated. Protective effects were examined in the lung through histological examination. Further in vivo experiments examined the reparative abilities of MSCs after tissue damage was established and the role played by soluble factors secreted by MSCs. The mechanism of MSC action was determined in using shRNA gene knockdown. Human MSC therapy and MSC conditioned media exerted significant cytoprotective effects when administered early at the onset of the disease. These protective effects were due to significant anti-inflammatory, anti-fibrotic and anti-apoptotic mechanisms, mediated in part through MSC production of hepatocyte growth factor (HGF). When MSC administration was delayed, significant protection of the lung architecture was observed but this was less extensive. MSC cell therapy was more effective than MSC conditioned medium in this emphysema model. PMID:27922052

  6. Cell polarity proteins: common targets for tumorigenic human viruses

    PubMed Central

    Javier, RT

    2012-01-01

    Loss of polarity and disruption of cell junctions are common features of epithelial-derived cancer cells, and mounting evidence indicates that such defects have a direct function in the pathology of cancer. Supporting this idea, results with several different human tumor viruses indicate that their oncogenic potential depends in part on a common ability to inactivate key cell polarity proteins. For example, adenovirus (Ad) type 9 is unique among human Ads by causing exclusively estrogen-dependent mammary tumors in experimental animals and in having E4 region-encoded open reading frame 1 (E4-ORF1) as its primary oncogenic determinant. The 125-residue E4-ORF1 protein consists of two separate protein-interaction elements, one of which defines a PDZ domain-binding motif (PBM) required for E4-ORF1 to induce both cellular transformation in vitro and tumorigenesis in vivo. Most notably, the E4-ORF1 PBM mediates interactions with a selected group of cellular PDZ proteins, three of which include the cell polarity proteins Dlg1, PATJ and ZO-2. Data further indicate that these interactions promote disruption of cell junctions and a loss of cell polarity. In addition, one or more of the E4-ORF1-interacting cell polarity proteins, as well as the cell polarity protein Scribble, are common targets for the high-risk human papillomavirus (HPV) E6 or human T-cell leukemia virus type 1 (HTLV-1) Tax oncoproteins. Underscoring the significance of these observations, in humans, high-risk HPV and HTLV-1 are causative agents for cervical cancer and adult T-cell leukemia, respectively. Consequently, human tumor viruses should serve as powerful tools for deciphering mechanisms whereby disruption of cell junctions and loss of cell polarity contribute to the development of many human cancers. This review article discusses evidence supporting this hypothesis, with an emphasis on the human Ad E4-ORF1 oncoprotein. PMID:19029943

  7. Tissuelike 3D Assemblies of Human Broncho-Epithelial Cells

    NASA Technical Reports Server (NTRS)

    Goodwin, Thomas J.

    2010-01-01

    Three-dimensional (3D) tissuelike assemblies (TLAs) of human broncho-epithelial (HBE) cells have been developed for use in in vitro research on infection of humans by respiratory viruses. The 2D monolayer HBE cell cultures heretofore used in such research lack the complex cell structures and interactions characteristic of in vivo tissues and, consequently, do not adequately emulate the infection dynamics of in-vivo microbial adhesion and invasion. In contrast, the 3D HBE TLAs are characterized by more-realistic reproductions of the geometrical and functional complexity, differentiation of cells, cell-to-cell interactions, and cell-to-matrix interactions characteristic of human respiratory epithelia. Hence, the 3D HBE TLAs are expected to make it possible to perform at least some of the research in vitro under more-realistic conditions, without need to infect human subjects. The TLAs are grown on collagen-coated cyclodextran microbeads under controlled conditions in a nutrient liquid in the simulated microgravitational environment of a bioreactor of the rotating- wall-vessel type. Primary human mesenchymal bronchial-tracheal cells are used as a foundation matrix, while adult human bronchial epithelial immortalized cells are used as the overlying component. The beads become coated with cells, and cells on adjacent beads coalesce into 3D masses. The resulting TLAs have been found to share significant characteristics with in vivo human respiratory epithelia including polarization, tight junctions, desmosomes, and microvilli. The differentiation of the cells in these TLAs into tissues functionally similar to in vivo tissues is confirmed by the presence of compounds, including villin, keratins, and specific lung epithelium marker compounds, and by the production of tissue mucin. In a series of initial infection tests, TLA cultures were inoculated with human respiratory syncytial viruses and parainfluenza type 3 viruses. Infection was confirmed by photomicrographs that

  8. Safety of repeated transplantations of neurotrophic factors-secreting human mesenchymal stromal stem cells

    PubMed Central

    2014-01-01

    Background Therapies based on mesenchymal stem cells (MSC) have been shown to have potential benefit in several clinical studies. We have shown that, using a medium-based approach, MSC can be induced to secrete elevated levels of neurotropic factors, which have been shown to have protective effects in animal models of neurodegenerative diseases. These cells, designated MSC-NTF cells (Neurotrophic factor-secreting MSC, also known as NurOwn™) derived from the patient's own bone marrow, have been recently used for Phase I/II and Phase IIa clinical studies in patients with Amyotrophic Lateral Sclerosis (ALS). In these studies, ALS patients were subjected to a single administration of autologous MSC-NTF cells. The data from these studies indicate that the single administration of MSC-NTF cells is safe and well tolerated. In a recently published case report, it was shown that repeated MSC-NTF injections in an ALS patient treated on a compassionate basis were safe and well tolerated [Muscle Nerve 49:455-457, 2014]. Methods In the current study we studied the toxicity and tolerability of three consecutive intramuscular injections (IM) of cryopreserved human MSC-NTF cells in C57BL/B6 mice to investigate the effect of repeated administration of these cells. Results Monitoring of clinical signs and immune reactions showed that repeated injections of the cells did not lead to any serious adverse events. Pathology, histology and blood biochemistry parameters tested were found to be within normal ranges with no sign of tumor formation. Conclusions Based on these results we conclude that repeated injections of human MSC-NTF are well tolerated in mice. The results of this study suggest that if the outcomes of additional clinical studies point to the need for repeated treatments, such option can be considered safe. PMID:25097724

  9. Generation of kidney tubular organoids from human pluripotent stem cells

    PubMed Central

    Yamaguchi, Shintaro; Morizane, Ryuji; Homma, Koichiro; Monkawa, Toshiaki; Suzuki, Sayuri; Fujii, Shizuka; Koda, Muneaki; Hiratsuka, Ken; Yamashita, Maho; Yoshida, Tadashi; Wakino, Shu; Hayashi, Koichi; Sasaki, Junichi; Hori, Shingo; Itoh, Hiroshi

    2016-01-01

    Recent advances in stem cell research have resulted in methods to generate kidney organoids from human pluripotent stem cells (hPSCs), which contain cells of multiple lineages including nephron epithelial cells. Methods to purify specific types of cells from differentiated hPSCs, however, have not been established well. For bioengineering, cell transplantation, and disease modeling, it would be useful to establish those methods to obtain pure populations of specific types of kidney cells. Here, we report a simple two-step differentiation protocol to generate kidney tubular organoids from hPSCs with direct purification of KSP (kidney specific protein)-positive cells using anti-KSP antibody. We first differentiated hPSCs into mesoderm cells using a glycogen synthase kinase-3β inhibitor for 3 days, then cultured cells in renal epithelial growth medium to induce KSP+ cells. We purified KSP+ cells using flow cytometry with anti-KSP antibody, which exhibited characteristics of all segments of kidney tubular cells and cultured KSP+ cells in 3D Matrigel, which formed tubular organoids in vitro. The formation of tubular organoids by KSP+ cells induced the acquisition of functional kidney tubules. KSP+ cells also allowed for the generation of chimeric kidney cultures in which human cells self-assembled into 3D tubular structures in combination with mouse embryonic kidney cells. PMID:27982115

  10. Nanoemulsion-based mucosal adjuvant induces apoptosis in human epithelial cells

    PubMed Central

    Orzechowska, Beata U.; Kukowska-Latallo, Jolanta F.; Coulter, Alexa D.; Szabo, Zsuzsanna; Gamian, Andrzej; Myc, Andrzej

    2015-01-01

    Nanoemulsions (NEs) are adjuvants that enhance antigen penetration of the nasal mucosa, increase cellular uptake of antigens by both epithelial and dendritic cells, and promote the migration of antigen-loaded dendritic cells to regional lymph nodes within 24-hours of vaccine administration. The objective of this study was to elucidate cell death caused by W805EC NE and identify caspases and genes associated with death pathways. Consistent with this aim, we show that exposure of human epithelial cells (EC), both RPMI 2650 and FaDu, to NE results in the activation of caspases (1, 3/7, 6, 8, and 9) and the expression of genes involved in apoptotic as well as authophagy and necrosis pathways. Interestingly, the NE activates caspase 8 which promotes “immunogenic apoptosis”. The rescue assay was employed to investigate the fate of RPMI 2650 cells treated with W805EC NE. After four hour treatment with as little as 0.03% of NE no cells were rescued at 72 hours. Remarkably, immediately after four-hour treatment, the cells morphologically resembled untreated cells and most of the cells were alive. Altogether, these results suggest that NE induces death of human ECs through multiple pathways. Epithelial cell death caused by W805EC may have further implications on antigen uptake, processing, and presentation by DC's. PMID:25817825

  11. L1 Cell Adhesion Molecule-Specific Chimeric Antigen Receptor-Redirected Human T Cells Exhibit Specific and Efficient Antitumor Activity against Human Ovarian Cancer in Mice

    PubMed Central

    Hong, Hao; Brown, Christine E.; Ostberg, Julie R.; Priceman, Saul J.; Chang, Wen-Chung; Weng, Lihong; Lin, Paul; Wakabayashi, Mark T.; Jensen, Michael C.; Forman, Stephen J.

    2016-01-01

    New therapeutic modalities are needed for ovarian cancer, the most lethal gynecologic malignancy. Recent clinical trials have demonstrated the impressive therapeutic potential of adoptive therapy using chimeric antigen receptor (CAR)-redirected T cells to target hematological cancers, and emerging studies suggest a similar impact may be achieved for solid cancers. We sought determine whether genetically-modified T cells targeting the CE7-epitope of L1-CAM, a cell adhesion molecule aberrantly expressed in several cancers, have promise as an immunotherapy for ovarian cancer, first demonstrating that L1-CAM was highly over-expressed on a panel of ovarian cancer cell lines, primary ovarian tumor tissue specimens, and ascites-derived primary cancer cells. Human central memory derived T cells (TCM) were then genetically modified to express an anti-L1-CAM CAR (CE7R), which directed effector function upon tumor antigen stimulation as assessed by in vitro cytokine secretion and cytotoxicity assays. We also found that CE7R+ T cells were able to target primary ovarian cancer cells. Intraperitoneal (i.p.) administration of CE7R+ TCM induced a significant regression of i.p. established SK-OV-3 xenograft tumors in mice, inhibited ascites formation, and conferred a significant survival advantage compared with control-treated animals. Taken together, these studies indicate that adoptive transfer of L1-CAM-specific CE7R+ T cells may offer a novel and effective immunotherapy strategy for advanced ovarian cancer. PMID:26761817

  12. L1 Cell Adhesion Molecule-Specific Chimeric Antigen Receptor-Redirected Human T Cells Exhibit Specific and Efficient Antitumor Activity against Human Ovarian Cancer in Mice.

    PubMed

    Hong, Hao; Brown, Christine E; Ostberg, Julie R; Priceman, Saul J; Chang, Wen-Chung; Weng, Lihong; Lin, Paul; Wakabayashi, Mark T; Jensen, Michael C; Forman, Stephen J

    2016-01-01

    New therapeutic modalities are needed for ovarian cancer, the most lethal gynecologic malignancy. Recent clinical trials have demonstrated the impressive therapeutic potential of adoptive therapy using chimeric antigen receptor (CAR)-redirected T cells to target hematological cancers, and emerging studies suggest a similar impact may be achieved for solid cancers. We sought determine whether genetically-modified T cells targeting the CE7-epitope of L1-CAM, a cell adhesion molecule aberrantly expressed in several cancers, have promise as an immunotherapy for ovarian cancer, first demonstrating that L1-CAM was highly over-expressed on a panel of ovarian cancer cell lines, primary ovarian tumor tissue specimens, and ascites-derived primary cancer cells. Human central memory derived T cells (TCM) were then genetically modified to express an anti-L1-CAM CAR (CE7R), which directed effector function upon tumor antigen stimulation as assessed by in vitro cytokine secretion and cytotoxicity assays. We also found that CE7R+ T cells were able to target primary ovarian cancer cells. Intraperitoneal (i.p.) administration of CE7R+ TCM induced a significant regression of i.p. established SK-OV-3 xenograft tumors in mice, inhibited ascites formation, and conferred a significant survival advantage compared with control-treated animals. Taken together, these studies indicate that adoptive transfer of L1-CAM-specific CE7R+ T cells may offer a novel and effective immunotherapy strategy for advanced ovarian cancer.

  13. Engraftment and long-term expression of human fetal hemopoietic stem cells in sheep following transplantation in utero.

    PubMed Central

    Zanjani, E D; Pallavicini, M G; Ascensao, J L; Flake, A W; Langlois, R G; Reitsma, M; MacKintosh, F R; Stutes, D; Harrison, M R; Tavassoli, M

    1992-01-01

    Hemopoietic stem cells from human fetal liver were transplanted in utero into preimmune fetal sheep (48-54 days of gestation). The fate of donor cells was followed using karyotype analysis, by immunofluorescence labeling with anti-CD antibodies, and by fluorescent in situ hybridization using human-specific DNA probes. Engraftment occurred in 13 of 33 recipients. Of five live born sheep that exhibited chimerism, all expressed human cells in the marrow, whereas three expressed them in blood as well. Engraftment was multilineage (erythroid, myeloid, and lymphoid) and human hemopoietic progenitors (multipotent colony-forming units, colony-forming units-granulocyte, macrophage, and erythroid burst-forming units) capable of forming colonies in vitro were detected in all five lambs for greater than 2 yr. These progenitors responded to human-specific growth factors both in vitro and in vivo. Thus the administration of recombinant human IL-3 and granulocyte macrophage-colony-stimulating factor to chimeric sheep resulted in a 2.1-3.4-fold increase in the relative expression of donor (human) cells. These results demonstrate that the permissive environment of the preimmune fetal sheep provides suitable conditions for the engraftment and long-term multilineage expression of human hemopoietic stem cells in a large animal model. In this model, donor human cells appear to retain certain phenotypic and functional characteristics that can be used to manipulate the size of donor cell pool. PMID:1348253

  14. Limited hair cell induction from human induced pluripotent stem cells using a simple stepwise method.

    PubMed

    Ohnishi, Hiroe; Skerleva, Desislava; Kitajiri, Shin-ichiro; Sakamoto, Tatsunori; Yamamoto, Norio; Ito, Juichi; Nakagawa, Takayuki

    2015-07-10

    Disease-specific induced pluripotent stem cells (iPS) cells are expected to contribute to exploring useful tools for studying the pathophysiology of inner ear diseases and to drug discovery for treating inner ear diseases. For this purpose, stable induction methods for the differentiation of human iPS cells into inner ear hair cells are required. In the present study, we examined the efficacy of a simple induction method for inducing the differentiation of human iPS cells into hair cells. The induction of inner ear hair cell-like cells was performed using a stepwise method mimicking inner ear development. Human iPS cells were sequentially transformed into the preplacodal ectoderm, otic placode, and hair cell-like cells. As a first step, preplacodal ectoderm induction, human iPS cells were seeded on a Matrigel-coated plate and cultured in a serum free N2/B27 medium for 8 days according to a previous study that demonstrated spontaneous differentiation of human ES cells into the preplacodal ectoderm. As the second step, the cells after preplacodal ectoderm induction were treated with basic fibroblast growth factor (bFGF) for induction of differentiation into otic-placode-like cells for 15 days. As the final step, cultured cells were incubated in a serum free medium containing Matrigel for 48 days. After preplacodal ectoderm induction, over 90% of cultured cells expressed the genes that express in preplacodal ectoderm. By culture with bFGF, otic placode marker-positive cells were obtained, although their number was limited. Further 48-day culture in serum free media resulted in the induction of hair cell-like cells, which expressed a hair cell marker and had stereocilia bundle-like constructions on their apical surface. Our results indicate that hair cell-like cells are induced from human iPS cells using a simple stepwise method with only bFGF, without the use of xenogeneic cells.

  15. Growth suppressive efficacy of human lak cells against human lung-cancer implanted into scid mice.

    PubMed

    Teraoka, S; Kyoizumi, S; Suzuki, T; Yamakido, M; Akiyama, M

    1995-06-01

    The purpose of our study was to determine the efficacy of immunotherapy using human lymphokine activated killer (LAK) cells against a human-lung squamous-cell carcinoma cell line (RERF-LC-AI) implanted into severe combined immunodeficient (SCID) mice. A statistically significant growth suppressive effect on RERF-LC-AI implanted into SCID mice was observed when human LAK cells were administered into the caudal vein of the mice treated with a continuous supply (initiated prior to LAK cells injection) of rIL-2. The human LAK cells stained with PKH 2, a fluorescent dye, for later detection using flow cytometry were administered into the caudal vein of RERF-LC-AI bearing SCID mice; the cells persisted for 7 days in the implanted lung cancer tissue and in the mouse peripheral blood, but for 5 days in the mouse spleen. The number of infiltrated human LAK cells in each tissue increased dose-dependently with the number of injected cells. The results indicate that the antitumor effect most likely occurred during the early implantation period of the human LAK cells. These results demonstrate the applicability of this model to the in vivo study of human lung cancer therapy.

  16. Human amniotic epithelial cells as feeder layer to derive and maintain human embryonic stem cells from poor-quality embryos.

    PubMed

    Ávila-González, Daniela; Vega-Hernández, Eva; Regalado-Hernández, Juan Carlos; De la Jara-Díaz, Julio Francisco; García-Castro, Irma Lydia; Molina-Hernández, Anayansi; Moreno-Verduzco, Elsa Romelia; Razo-Aguilera, Guadalupe; Flores-Herrera, Héctor; Portillo, Wendy; Díaz-Martínez, Néstor Emmanuel; García-López, Guadalupe; Díaz, Néstor Fabián

    2015-09-01

    Data from the literature suggest that human embryonic stem cell (hESC) lines used in research do not genetically represent all human populations. The derivation of hESC through conventional methods involve the destruction of viable human embryos, as well the use of mouse embryonic fibroblasts as a feeder layer, which has several drawbacks. We obtained the hESC line (Amicqui-1) from poor-quality (PQ) embryos derived and maintained on human amniotic epithelial cells (hAEC). This line displays a battery of markers of pluripotency and we demonstrated the capacity of these cells to produce derivates of the three germ layers.

  17. Piperlongumine Suppresses Proliferation of Human Oral Squamous Cell Carcinoma through Cell Cycle Arrest, Apoptosis and Senescence.

    PubMed

    Chen, San-Yuan; Liu, Geng-Hung; Chao, Wen-Ying; Shi, Chung-Sheng; Lin, Ching-Yen; Lim, Yun-Ping; Lu, Chieh-Hsiang; Lai, Peng-Yeh; Chen, Hau-Ren; Lee, Ying-Ray

    2016-04-23

    Oral squamous cell carcinoma (OSCC), an aggressive cancer originating in the oral cavity, is one of the leading causes of cancer deaths in males worldwide. This study investigated the antitumor activity and mechanisms of piperlongumine (PL), a natural compound isolated from Piper longum L., in human OSCC cells. The effects of PL on cell proliferation, the cell cycle, apoptosis, senescence and reactive oxygen species (ROS) levels in human OSCC cells were investigated. PL effectively inhibited cell growth, caused cell cycle arrest and induced apoptosis and senescence in OSCC cells. Moreover, PL-mediated anti-human OSCC behavior was inhibited by an ROS scavenger N-acetyl-l-cysteine (NAC) treatment, suggesting that regulation of ROS was involved in the mechanism of the anticancer activity of PL. These findings suggest that PL suppresses tumor growth by regulating the cell cycle and inducing apoptosis and senescence and is a potential chemotherapy agent for human OSCC cells.

  18. [Incidence of the characteristics of induced pluripotent stem cells in vitro in administrative regime of their lines].

    PubMed

    Bobo-Ruiz, Jesús

    2013-01-01

    Recently achieved techniques in the field of stem cell research have permitted the development of human induced pluripotent stem cells (hiPSCs). The characteristics of these cells and their appearance after the passage of Order SCO/393/2006, of 8 February, on Functioning and Organization of a National Bank of Cell Lines and Act 14/2007, of 3 July, on Biomedical Research raise the questions of the legal regime of their deposition at the National Bank of Cell Lines and their control by the Commission of Guarantees for the Donation and Use of Human Cells and Tissues. Drawing on a wider legal frame, the author defends these requirements on the grounds that the legal classification comes from the origin and potentiality of the cells rather than from the techniques employed to obtain them.

  19. Morphine and codeine concentrations in human urine following controlled poppy seeds administration of known opiate content.

    PubMed

    Smith, Michael L; Nichols, Daniel C; Underwood, Paula; Fuller, Zachary; Moser, Matthew A; LoDico, Charles; Gorelick, David A; Newmeyer, Matthew N; Concheiro, Marta; Huestis, Marilyn A

    2014-08-01

    Opiates are an important component for drug testing due to their high abuse potential. Proper urine opiate interpretation includes ruling out poppy seed ingestion; however, detailed elimination studies after controlled poppy seed administration with known morphine and codeine doses are not available. Therefore, we investigated urine opiate pharmacokinetics after controlled oral administration of uncooked poppy seeds with known morphine and codeine content. Participants were administered two 45 g oral poppy seed doses 8 h apart, each containing 15.7 mg morphine and 3mg codeine. Urine was collected ad libitum up to 32 h after the first dose. Specimens were analyzed with the Roche Opiates II immunoassay at 2000 and 300 μg/L cutoffs, and the ThermoFisher CEDIA(®) heroin metabolite (6-acetylmorphine, 6-AM) and Lin-Zhi 6-AM immunoassays with 10 μg/L cutoffs to determine if poppy seed ingestion could produce positive results in these heroin marker assays. In addition, all specimens were quantified for morphine and codeine by GC/MS. Participants (N=22) provided 391 urine specimens over 32 h following dosing; 26.6% and 83.4% were positive for morphine at 2000 and 300 μg/L GC/MS cutoffs, respectively. For the 19 subjects who completed the study, morphine concentrations ranged from <300 to 7522 μg/L with a median peak concentration of 5239 μg/L. The median first morphine-positive urine sample at 2000 μg/L cutoff concentration occurred at 6.6 h (1.2-12.1), with the last positive from 2.6 to 18 h after the second dose. No specimens were positive for codeine at a cutoff concentration of 2000 μg/L, but 20.2% exceeded 300 μg/L, with peak concentrations of 658 μg/L (284-1540). The Roche Opiates II immunoassay had efficiencies greater than 96% for the 2000 and 300 μg/L cutoffs. The CEDIA 6-AM immunoassay had a specificity of 91%, while the Lin-Zhi assay had no false positive results. These data provide valuable information for interpreting urine opiate results.

  20. Morphine and Codeine Concentrations in Human Urine following Controlled Poppy Seeds Administration of Known Opiate Content

    PubMed Central

    Smith, Michael L.; Nichols, Daniel C.; Underwood, Paula; Fuller, Zachary; Moser, Matthew A.; LoDico, Charles; Gorelick, David A.; Newmeyer, Matthew N.; Concheiro, Marta; Huestis, Marilyn A.

    2014-01-01

    Opiates are an important component for drug testing due to their high abuse potential. Proper urine opiate interpretation includes ruling out poppy seed ingestion; however, detailed elimination studies after controlled poppy seed administration with known morphine and codeine doses are not available. Therefore, we investigated urine opiate pharmacokinetics after controlled oral administration of uncooked poppy seeds with known morphine and codeine content. Participants were administered two 45g oral poppy seed doses 8h apart, each containing 15.7mg morphine and 3mg codeine. Urine was collected ad libitum up to 32h after the first dose. Specimens were analyzed with the Roche Opiates II immunoassay at 2,000 and 300μg/L cutoffs, and the ThermoFisher CEDIA® Heroin Metabolite (6-acetylmorphine, 6AM) and Lin-Zhi 6AM immunoassays with 10μg/L cutoffs to determine if poppy seed ingestion could produce positive results in these heroin marker assays. In addition, all specimens were quantified for morphine and codeine by GC/MS. Participants (N=22) provided 391 urine specimens over 32h following dosing; 26.6% and 83.4% were positive for morphine at 2,000 and 300μg/L GC/MS cutoffs, respectively. For the 19 subjects who completed the study, morphine concentrations ranged from <300 to 7,522μg/L with a median peak concentration of 5,239μg/L. The median first morphine-positive urine sample at 2,000μg/L cutoff concentration occurred at 6.6h (1.2-12.1), with the last positive from 2.6 to 18h after the second dose. No specimens were positive for codeine at a cutoff concentration of 2,000μg/L, but 20.2% exceeded 300μg/L, with peak concentrations of 658 μg/L (284-1540). The Roche Opiates II immunoassay had efficiencies greater than 96% for the 2000 and 300μg/L cutoffs. The CEDIA 6AM immunoassay had a specificity of 91%, while the Lin-Zhi assay had no false positive results. These data provide valuable information for interpreting urine opiate results. PMID:24887324

  1. Human embryonic stem cells differentiate into functional renal proximal tubular-like cells.

    PubMed

    Narayanan, Karthikeyan; Schumacher, Karl M; Tasnim, Farah; Kandasamy, Karthikeyan; Schumacher, Annegret; Ni, Ming; Gao, Shujun; Gopalan, Began; Zink, Daniele; Ying, Jackie Y

    2013-04-01

    Renal cells are used in basic research, disease models, tissue engineering, drug screening, and in vitro toxicology. In order to provide a reliable source of human renal cells, we developed a protocol for the differentiation of human embryonic stem cells into renal epithelial cells. The differentiated stem cells expressed markers characteristic of renal proximal tubular cells and their precursors, whereas markers of other renal cell types were not expressed or expressed at low levels. Marker expression patterns of these differentiated stem cells and in vitro cultivated primary human renal proximal tubular cells were comparable. The differentiated stem cells showed morphological and functional characteristics of renal proximal tubular cells, and generated tubular structures in vitro and in vivo. In addition, the differentiated stem cells contributed in organ cultures for the formation of simple epithelia in the kidney cortex. Bioreactor experiments showed that these cells retained their functional characteristics under conditions as applied in bioartificial kidneys. Thus, our results show that human embryonic stem cells can differentiate into renal proximal tubular-like cells. Our approach would provide a source for human renal proximal tubular cells that are not affected by problems associated with immortalized cell lines or primary cells.

  2. Patenting human genes and stem cells.

    PubMed

    Martin-Rendon, Enca; Blake, Derek J

    2007-01-01

    Cell lines and genetically modified single cell organisms have been considered patentable subjects for the last two decades. However, despite the technical patentability of genes and stem cell lines, social and legal controversy concerning their 'ownership' has surrounded stem cell research in recent years. Some granted patents on stem cells with extremely broad claims are casting a shadow over the commercialization of these cells as therapeutics. However, in spite of those early patents, the number of patent applications related to stem cells is growing exponentially. Both embryonic and adult stem cells have the ability to differentiate into several cell lineages in an organism as a result of specific genetic programs that direct their commitment and cell fate. Genes that control the pluripotency of stem cells have been recently identified and the genetic manipulation of these cells is becoming more efficient with the advance of new technologies. This review summarizes some of the recent published patents on pluripotency genes, gene transfer into stem cells and genetic reprogramming and takes the hematopoietic and embryonic stem cell as model systems.

  3. Host responses in human skin after conventional intradermal injection or microneedle administration of virus-like-particle influenza vaccine.

    PubMed

    Pearton, Marc; Pirri, Daniela; Kang, Sang-Moo; Compans, Richard W; Birchall, James C

    2013-10-01

    Miniaturized microneedle devices are being developed for painlessly targeting vaccines to the immune cell populations in skin. As skin immunization studies are generally restricted to animal models however, where skin architecture and immunity is greatly different to human, surprisingly little is known about the local human response to intradermal (ID) vaccines. Here surgically excised human skin is used to explore for the first time the complex molecular and cellular host responses to a candidate influenza vaccine comprising nanoparticulate virus-like-particles (VLPs), administered via conventional hypodermic injection or reduced scale microneedles. Responses at the molecular level are determined by microarray analysis (47,296 discrete transcripts) and validated by quantitative PCR (96 genes). Cellular response is probed through monitoring migration of dendritic cells in viable skin tissue. Gene expression mapping, ontological analysis, and qPCR reveal up-regulation of a host of genes responsible for key immunomodulatory processes and host viral response, including cell recruitment, activation, migration, and T cell interaction following both ID and microneedle injection of VLPs; the response from the microneedles being more subtle. Significant morphological and migratory changes to skin dendritic cells are also apparent following microneedle VLP delivery. This is the first study displaying the global, multifaceted immunological events that occur at the site of vaccine deposition in human skin and will subsequently influence the degree and nature of innate and adaptive immune responses. An increased understanding of the detailed similarities and differences in response against antigen administered via different delivery modalities will inform the development of improved vaccines and vaccine delivery systems.

  4. Human induced pluripotent stem cells: A disruptive innovation.

    PubMed

    De Vos, J; Bouckenheimer, J; Sansac, C; Lemaître, J-M; Assou, S

    2016-01-01

    This year (2016) will mark the 10th anniversary of the discovery of induced pluripotent stem cells (iPSCs). The finding that the transient expression of four transcription factors can radically remodel the epigenome, transcriptome and metabolome of differentiated cells and reprogram them into pluripotent stem cells has been a major and groundbreaking technological innovation. In this review, we discuss the major applications of this technology that we have grouped in nine categories: a model to study cell fate control; a model to study pluripotency; a model to study human development; a model to study human tissue and organ physiology; a model to study genetic diseases in a dish; a tool for cell rejuvenation; a source of cells for drug screening; a source of cells for regenerative medicine; a tool for the production of human organs in animals.

  5. Continuous human cell lines and method of making same

    SciTech Connect

    Stampfer, M.R.

    1989-02-28

    Substantially genetically stable continuous human cell lines derived from normal human mammary epithelial cells (HMEC) and processes for making and using the same. In a preferred embodiment, the cell lines are derived by treating normal human mammary epithelial tissue with a chemical carcinogen such as benzo[a]pyrene. The novel cell lines serve as useful substrates for elucidating the potential effects of a number of toxins, carcinogens and mutagens as well as of the addition of exogenous genetic material. The autogenic parent cells from which the cell lines are derived serve as convenient control samples for testing. The cell lines are not neoplastically transformed, although they have acquired several properties which distinguish them from their normal progenitors. No Drawings

  6. Phenotype and functions of memory Tfh cells in human blood.

    PubMed

    Schmitt, Nathalie; Bentebibel, Salah-Eddine; Ueno, Hideki

    2014-09-01

    Our understanding of the origin and functions of human blood CXCR5(+) CD4(+) T cells found in human blood has changed dramatically in the past years. These cells are currently considered to represent a circulating memory compartment of T follicular helper (Tfh) lineage cells. Recent studies have shown that blood memory Tfh cells are composed of phenotypically and functionally distinct subsets. Here, we review the current understanding of human blood memory Tfh cells and the subsets within this compartment. We present a strategy to define these subsets based on cell surface profiles. Finally, we discuss how increased understanding of the biology of blood memory Tfh cells may contribute insight into the pathogenesis of autoimmune diseases and the mode of action of vaccines.

  7. Continuous human cell lines and method of making same

    DOEpatents

    Stampfer, M.R.

    1985-07-01

    Substantially genetically stable continuous human cell lines derived from normal human mammary epithelial cells (HMEC) and processes for making and using the same. In a preferred embodiment, the cell lines are derived by treating normal human mammary epithelial tissue with a chemical carcinogen such as benzo(a)pyrene. The novel cell lines serve as useful substrates for elucidating the potential effects of a number of toxins, carcinogens and mutagens as well as of the addition of exogenous genetic material. The autogenic parent cells from which the cell lines are derived serve as convenient control samples for testing. The cell lines are not neoplastically transformed, although they have acquired several properties which distinguish them from their normal progenitors. 2 tabs.

  8. Continuous human cell lines and method of making same

    DOEpatents

    Stampfer, Martha R.

    1989-01-01

    Substantially genetically stable continuous human cell lines derived from normal human mammary epithelial cells (HMEC) and processes for making and using the same. In a preferred embodiment, the cell lines are derived by treating normal human mammary epithelial tissue with a chemical carcinogen such as benzo[a]pyrene. The novel cell lines serve as useful substrates for elucidating the potential effects of a number of toxins, carcinogens and mutagens as well as of the addition of exogenous genetic material. The autogenic parent cells from which the cell lines are derived serve as convenient control samples for testing. The cell lines are not neoplastically transformed, although they have acquired several properties which distinguish them from their normal progenitors.

  9. Density gradient electrophoresis of cultured human embryonic kidney cells

    NASA Technical Reports Server (NTRS)

    Plank, L. D.; Kunze, M. E.; Giranda, V.; Todd, P. W.

    1985-01-01

    Ground based confirmation of the electrophoretic heterogeneity of human embryonic kidney cell cultures, the general characterization of their electrophoretic migration, and observations on the general properties of cultures derived from electrophoretic subpopulations were studied. Cell migration in a density gradient electrophoresis column and cell electrophoretic mobility was determined. The mobility and heterogeneity of cultured human embryonic kidney cells with those of fixed rat erythrocytes as model test particle was compared. Electrophoretically separated cell subpopulations with respect to size, viability, and culture characteristics were examined.

  10. New Monoclonal Antibodies to Defined Cell Surface Proteins on Human Pluripotent Stem Cells.

    PubMed

    O'Brien, Carmel M; Chy, Hun S; Zhou, Qi; Blumenfeld, Shiri; Lambshead, Jack W; Liu, Xiaodong; Kie, Joshua; Capaldo, Bianca D; Chung, Tung-Liang; Adams, Timothy E; Phan, Tram; Bentley, John D; McKinstry, William J; Oliva, Karen; McMurrick, Paul J; Wang, Yu-Chieh; Rossello, Fernando J; Lindeman, Geoffrey J; Chen, Di; Jarde, Thierry; Clark, Amander T; Abud, Helen E; Visvader, Jane E; Nefzger, Christian M; Polo, Jose M; Loring, Jeanne F; Laslett, Andrew L

    2017-03-01

    The study and application of human pluripotent stem cells (hPSCs) will be enhanced by the availability of well-characterized monoclonal antibodies (mAbs) detecting cell-surface epitopes. Here, we report generation of seven new mAbs that detect cell surface proteins present on live and fixed human ES cells (hESCs) and human iPS cells (hiPSCs), confirming our previous prediction that these proteins were present on the cell surface of hPSCs. The mAbs all show a high correlation with POU5F1 (OCT4) expression and other hPSC surface markers (TRA-160 and SSEA-4) in hPSC cultures and detect rare OCT4 positive cells in differentiated cell cultures. These mAbs are immunoreactive to cell surface protein epitopes on both primed and naive state hPSCs, providing useful research tools to investigate the cellular mechanisms underlying human pluripotency and states of cellular reprogramming. In addition, we report that subsets of the seven new mAbs are also immunoreactive to human bone marrow-derived mesenchymal stem cells (MSCs), normal human breast subsets and both normal and tumorigenic colorectal cell populations. The mAbs reported here should accelerate the investigation of the nature of pluripotency, and enable development of robust cell separation and tracing technologies to enrich or deplete for hPSCs and other human stem and somatic cell types. Stem Cells 2017;35:626-640.

  11. Benzodiazepine self-administration in humans and laboratory animals--implications for problems of long-term use and abuse.

    PubMed

    Griffiths, R R; Weerts, E M

    1997-11-01

    Drug reinforcement may represent the primary behavioral-pharmacological mechanism underlying two types of problematic use of benzodiazepines--recreational abuse by polydrug abusers and inappropriate chronic use by patients. High dose polydrug abuse for the purpose of getting high is readily recognized as a significant social problem. Inappropriate chronic benzodiazepine use is more subtle but relatively common: for anxiolytics, 36% of past-year users (3% of the adult population in the US) report using these drugs for 4 consecutive months or longer. The risks of such long-term use are much better documented than the benefits. This paper provides a current review of various problems that have been identified with the long-term use and the recreational abuse of benzodiazepines, including memory impairment, risk of accidents, falls and hip fractures in the elderly, a withdrawal syndrome, brain damage, overuse in the elderly, overuse by chronic pain patients, overuse by alcoholics and recreational abuse among alcoholics and polydrug abusers. A comprehensive review of the literature on benzodiazepine reinforcing effects in humans and laboratory animals is also provided. Drug self-administration studies in humans and laboratory animals provide models of both types of problematic benzodiazepine use. Recreational abuse of benzodiazepines has been modeled in human research with polydrug abusers and in laboratory animal studies, which show that the reinforcing effect of benzodiazepines is intermediate relative to other sedative compounds and is increased in subjects with histories of previous sedative drug self-administration. The problem of inappropriate long-term use of benzodiazepines by people without histories of drug abuse has been partially modeled in human studies showing that benzodiazepines function as reinforcers in subjects with anxiety, insomnia, and histories of moderate alcohol consumption, and in preclinical studies showing stable, low-rate benzodiazepine self

  12. 78 FR 66366 - Draft Guidance for Industry: Use of Donor Screening Tests To Test Donors of Human Cells, Tissues...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-11-05

    ...The Food and Drug Administration (FDA) is announcing the availability of a draft document entitled ``Guidance for Industry: Use of Donor Screening Tests to Test Donors of Human Cells, Tissues, and Cellular and Tissue-Based Products (HCT/Ps) for Infection with Treponema pallidum (Syphilis),'' dated October 2013. The draft guidance document provides establishments that make donor eligibility......

  13. Cytotoxicity of fig fruit latex against human cancer cells.

    PubMed

    Wang, Jing; Wang, Xiujie; Jiang, Shu; Lin, Ping; Zhang, Jie; Lu, Yanrong; Wang, Qi; Xiong, Zhujuan; Wu, Yaying; Ren, Jingjing; Yang, Hongliang

    2008-03-01

    Fig fruit latex (FFL) contains significant amounts of polyphenolic compounds and can serve as a source of antioxidants after human consumption. The purpose of this study is to confirm anticancer activity of FFL against human cancer cells and to further elucidate its mechanism of activity. Human glioblastoma, hepatocellular carcinoma, and normal liver cells were used for in vitro tests of FFL effects. Those tests included cytotoxicity, colony formation inhibition, Brdu incorporation, acridine orange/ethidium bromide (AO/EB) staining for apoptotic cells, cell cycle distribution through flow cytometry (FCM), and ADP-ribosyltransferase (NAD+; poly(ADP-ribose) polymerase)-like 1 (ADPERL1) mRNA expression through RT-PCR in response to FFL treatment. After FFL treatment, the proliferation, colony formation, and Brdu labeling indices of cancer cells decreased (P<0.05), while the AO/EB stained apoptotic cells increased (P<0.05). By FCM analysis, an increase of G(0)/G(1) phase cell population and decrease of S and G(2)/M phase cells were observed (P<0.01), while both ADPRTL1 mRNA expression and apoptotic indices increased (P<0.01). The findings in these studies suggested that FFL exhibited potent cytotoxicity in some human cancer cells with little effect in normal cells at certain concentration. The mechanism for such effects might be associated with the inhibition of DNA synthesis, induction of apoptosis, and cell cycle arrest of cancer cells.

  14. Applied Developmental Biology: Making Human Pancreatic Beta Cells for Diabetics.

    PubMed

    Melton, Douglas A

    2016-01-01

    Understanding the genes and signaling pathways that determine the differentiation and fate of a cell is a central goal of developmental biology. Using that information to gain mastery over the fates of cells presents new approaches to cell transplantation and drug discovery for human diseases including diabetes.

  15. Cholera toxin stimulation of human mammary epithelial cells in culture

    SciTech Connect

    Stampfer, M.R.

    1982-06-01

    Addition of cholera toxin to human mammary epithelial cultures derived from reduction mammoplasties and primary carcinomas greatly stimulated cell growth and increased the number of times the cells could be successfully subcultured. Other agents known to increase intracellular cAMP levels were also growth stimulatory. The increased growth potential conferred by cholera toxin enhances the usefulness of this cell culture system.

  16. Vulnerability of Normal Human Mammary Epithelial Cells to Oncogenic Transformation

    DTIC Science & Technology

    2012-04-01

    diverse transformed HMEC lines with defined genetic alterations may aid the identification of potential therapeutic treatments , including...human model systems to test potential therapeutics, could facilitate individualized treatment and possibly prevention. The main variables thought to...epithelial cells. Middle, corresponding cell culture models used in this study. Red, treatment or genetic manipulation used. Cell models are described in

  17. Exposure to Music Alters Cell Viability and Cell Motility of Human Nonauditory Cells in Culture

    PubMed Central

    Lestard, Nathalia R.

    2016-01-01

    Although music is part of virtually all cultures in the world, little is known about how it affects us. Since the beginning of this century several studies suggested that the response to music, and to sound in general, is complex and might not be exclusively due to emotion, given that cell types other than auditory hair cells can also directly react to audible sound. The present study was designed to better understand the direct effects of acoustic vibrations, in the form of music, in human cells in culture. Our results suggest that the mechanisms of cell growth arrest and/or cell death induced by acoustic vibrations are similar for auditory and nonauditory cells. PMID:27478480

  18. Xanthosine administration does not affect the proportion of epithelial stem cells in bovine mammary tissue, but has a latent negative effect on cell proliferation

    SciTech Connect

    Rauner, Gat; Barash, Itamar

    2014-10-15

    The challenge in manipulating the proportion of somatic stem cells lies in having to override tissue homeostasis. Xanthosine infusion via the teat canal has been reported to augment the number of label-retaining cells in the mammary gland of 3-month-old bovine calves. To further delineate xanthosine's effect on defined stem cells in the mammary gland of heifers—which are candidates for increased prospective milk production following such manipulation—bovine mammary parenchymal tissue was transplanted and integrated into the cleared mammary fat pad of immunodeficient mice. Xanthosine administration for 14 days did not affect the number of label-retaining cells after 10- and 11-week chases. No change in stem cell proportion, analyzed according to CD49f and CD24 expression, was noted. Clone formation and propagation rate of cultured cells, as well as expression of stem cell markers, were also unaffected. In contrast, a latent 50% decrease in bovine mammary cell proliferation rate was observed 11 weeks after xanthosine administration. Tumor development in mice was also limited by xanthosine administration. These effects may have resulted from an initial decrease in expression of the rate-limiting enzyme in guanine synthesis, IMPDH. The data indicate that caution should be exerted when considering xanthosine for stem cell manipulation. - Highlights: • Novel “bovinized“ mouse model for exogenous effects on bovine mammary gland. • Xanthosine did not affect stem cell number/function in bovine mammary gland. • Xanthosine caused an immediate decrease in IMPDH expression in bovine mammary gland. • Xanthosine had latent negative effect on cell proliferation in bovine mammary gland. • Xanthosine administration limited mammary tumor growth.

  19. Human hepatocyte and kidney cell metabolism of 2-acetylaminofluorene and comparison to the respective rat cells.

    PubMed

    Langenbach, R; Rudo, K

    1988-12-01

    The metabolism and mutagenic activation of 2-acetylaminofluorene by human and rat hepatocytes and kidney cells were measured. High performance liquid chromatography was used to separate the 2-acetylaminofluorene metabolites, and a cell-mediated Salmonella typhimurium mutagenesis assay was used to detect mutagenic intermediates. Rat and human differences were observed with cells from both organs and levels of metabolism and mutagenesis were higher in human cells. Within a species, liver and kidney cell differences were also evident, with levels of hepatocyte-mediated metabolism and mutagenesis being greater than kidney cells. Human inter-individual variation was apparent with cells from both organs, but the variation observed was significantly greater in hepatocytes than kidney cells. A knowledge of such differences, including an understanding that they may vary with the chemical being studied, should be useful in the extrapolation of rodent carcinogenesis data to humans.

  20. Dual role of macrophages in the response of C26 colon carcinoma cells to 5-fluorouracil administration

    PubMed Central

    Patras, Laura; Sesarman, Alina; Licarete, Emilia; Luca, Lavinia; Alupei, Marius Costel; Rakosy-Tican, Elena; Banciu, Manuela

    2016-01-01

    Previous studies have demonstrated that tumor-associated macrophages (TAMs) are pivotal players in tumor progression via modulation of tumor angiogenesis, inflammation, metastasis and oxidative stress, as well as of the response of cancer cells to cytotoxic drugs. Nevertheless, the role of TAMs in the prognosis of colorectal cancer remains controversial. Therefore, the present study aimed to investigate how TAMs mediate the response of C26 colon carcinoma cells to the cytotoxic drug 5-fluorouracil (5-FU), upon TAM co-cultivation with these cancer cells in vitro. In this respect, 5-FU cytotoxicity was assessed in C26 cells in standard culture and in a co-culture with peritoneal macrophages, the production of NF-κB was determined by western blot analysis, and the production of angiogenic/inflammatory proteins in each experimental model was evaluated by protein array analysis. To gain further evidence of the effect of TAMs on oxidative stress, malondialdehyde was measured through high-performance liquid chromatography, and the total nonenzymatic antioxidant levels and the production of nitrites were measured through colorimetric assays. The results demonstrated that TAMs exerted a dual role in the response of C26 cells to 5-FU administration in the co-culture model. Thus, on one side, TAMs sensitized C26 cells to 5-FU administration through inhibition of the production of inflammatory and angiogenic proteins in these cancer cells; however, they also protected cancer cells against 5-FU-induced oxidative stress. Collectively, the present findings suggest that the combined administration of 5-FU with pharmacological agents that prevent TAMs to maintain the physiological range of tumor cell oxidative stress may highly improve the therapeutic potential of this drug. PMID:27446416

  1. Human CD8+ T cells mediate protective immunity induced by a human malaria vaccine in human immune system mice.

    PubMed

    Li, Xiangming; Huang, Jing; Zhang, Min; Funakoshi, Ryota; Sheetij, Dutta; Spaccapelo, Roberta; Crisanti, Andrea; Nussenzweig, Victor; Nussenzweig, Ruth S; Tsuji, Moriya

    2016-08-31

    A number of studies have shown that CD8+ T cells mediate protective anti-malaria immunity in a mouse model. However, whether human CD8+ T cells play a role in protection against malaria remains unknown. We recently established human immune system (HIS) mice harboring functional human CD8+ T cells (HIS-CD8 mice) by transduction with HLA-A∗0201 and certain human cytokines using recombinant adeno-associated virus-based gene transfer technologies. These HIS-CD8 mice mount a potent, antigen-specific HLA-A∗0201-restricted human CD8+ T-cell response upon immunization with a recombinant adenovirus expressing a human malaria antigen, the Plasmodium falciparum circumsporozoite protein (PfCSP), termed AdPfCSP. In the present study, we challenged AdPfCSP-immunized HIS-CD8 mice with transgenic Plasmodium berghei sporozoites expressing full-length PfCSP and found that AdPfCSP-immunized (but not naïve) mice were protected against subsequent malaria challenge. The level of the HLA-A∗0201-restricted, PfCSP-specific human CD8+ T-cell response was closely correlated with the level of malaria protection. Furthermore, depletion of human CD8+ T cells from AdPfCSP-immunized HIS-CD8 mice almost completely abolished the anti-malaria immune response. Taken together, our data show that human CD8+ T cells mediate protective anti-malaria immunity in vivo.

  2. Resveratrol Treatment Inhibits Proliferation of and Induces Apoptosis in Human Colon Cancer Cells

    PubMed Central

    Feng, Miao; Zhong, Lu-Xing; Zhan, Zheng-Yu; Huang, Zhi-Hao; Xiong, Jian-Ping

    2016-01-01

    Background Resveratrol, a natural isolate from plant sources, has a long and important history in traditional Chinese medicine. In the present study we investigated the effect of resveratrol on human colon cancer cell lines. Material/Methods We used the Cell Counting kit-8 (CCK-8) for determination of colon cancer cell viability. Apoptosis induction was analyzed using the DeadEnd™ Colorimetric TUNEL System (Promega, Madison, WI, USA). The siRNA Transfection Reagent kit (Santa Cruz Biotechnology, Inc.) was used for the administration of COX-2 silencer RNA (siRNA) into the colon cancer cells. Primer Express® software for Real-Time PCR ver. 3.0 (Applied Biosystems, Foster City, CA, USA) was used to prepare the primers for RT-PCR. Results The results revealed that exposure of colon cancer cells to resveratrol inhibited cell viability. Resveratrol exhibited a significant inhibitory effect on cell viability at 30 μM concentration after 48 h of exposure. We observed that 30-μM doses of resveratrol for 72 h led to 18, 29, and 34% reduction in the viability of HCA-17, SW480, and HT29 cells, respectively. It also significantly induced apoptosis in both of the tested carcinoma cell lines. The population of apoptotic cells in HCA-17 and SW480 cell lines after 48 h of resveratrol treatment was 59.8±4 and 67.2±4%, respectively, compared to 2.3±1% in the control cells. The colon cancer cells exposed to resveratrol showed significantly lower cyclooxygenase-2 and prostaglandin receptor expression. Treatment of colon cancer cells with the inhibitor of cyclooxygenase-2, indomethacin, and administration of silencer RNA for cyclooxygenase-2 also produced similar results. Conclusions These findings suggest that resveratrol treatment can be a promising strategy for the treatment of colon cancer. PMID:27040803

  3. Human pituitary and placental hormones control human insulin-like growth factor II secretion in human granulosa cells

    SciTech Connect

    Ramasharma, K.; Li, C.H.

    1987-05-01

    Human granulosa cells cultured with calf serum actively proliferated for 18-20 generations and secreted progesterone into the medium; progesterone levels appeared to decline with increase in generation number. Cells cultured under serum-free conditions secreted significant amounts of progesterone and insulin-like growth factor II (IGF-II). The progesterone secretion was enhanced by the addition of human follitropin, lutropin, and chorionic gonadotropin but not by growth hormone. These cells, when challenged to varying concentrations of human growth hormone, human chorionic somatomammotropin, human prolactin, chorionic gonadotropin, follitropin, and lutropin, secreted IGF-II into the medium as measured by specific IGF-II RIA. Among these human hormones, chorionic gonadotropin, follitropin, and lutropin were most effective in inducing IGF-II secretion from these cells. When synthetic lutropin-releasing hormone and ..cap alpha..-inhibin-92 were tested, only lutropin-releasing hormone was effective in releasing IGF-II. The results described suggest that cultured human granulosa cells can proliferate and actively secrete progesterone and IGF-II into the medium. IGF-II production in human granulosa cells was influenced by a multi-hormonal complex including human growth hormone, human chorionic somatomammotropin, and prolactin.

  4. Pre-encoding administration of amphetamine or THC preferentially modulates emotional memory in humans

    PubMed Central

    Ballard, Michael E.; Gallo, David A.; de Wit, Harriet

    2012-01-01

    Rationale Many addictive drugs are known to have effects on learning and memory, and these effects could motivate future drug use. Specifically, addictive drugs may affect memory of emotional events and experiences in ways that are attractive to some users. However, few studies have investigated the effects of addictive drugs on emotional memory in humans. Objectives This study examined the effects of the memory-enhancing drug dextroamphetamine (AMP) and the memory-impairing drug Δ9-tetrahydrocannabinol (THC) on emotional memory in healthy volunteers. Methods Participants completed three experimental sessions across which they received capsules containing placebo and two doses of either AMP (10 and 20 mg; N=25) or THC (7.5 and 15 mg; N=25) before viewing pictures of positive (pleasant), neutral, and negative (unpleasant) scenes. Memory for the pictures was assessed two days later, under drug-free conditions. Results Relative to placebo, memory for emotional pictures was improved by AMP and impaired by THC, but neither drug significantly affected memory for unemotional pictures. Positive memory biases were not observed with either drug, and there was no indication that the drugs’ memory effects were directly related to their subjective or physiological effects alone. Conclusions This study provides the first clear evidence that stimulant drugs can preferentially strengthen, and cannabinoids can preferentially impair, memory for emotional events in humans. Although addictive drugs do not appear to positively bias memory, the possibility remains that these drugs’ effects on emotional memory could influence drug use among certain individuals. PMID:23224510

  5. Effects of chronic lithium administration on renal acid excretion in humans and rats

    PubMed Central

    Weiner, I. David; Leader, John P.; Bedford, Jennifer J.; Verlander, Jill W.; Ellis, Gaye; Kalita, Priyakshi; Vos, Frederiek; de Jong, Sylvia; Walker, Robert J.

    2014-01-01

    Abstract Lithium therapy's most common side effects affecting the kidney are nephrogenic diabetes insipidus (NDI) and chronic kidney disease. Lithium may also induce a distal renal tubular acidosis. This study investigated the effect of chronic lithium exposure on renal acid–base homeostasis, with emphasis on ammonia and citrate excretion. We compared 11 individuals on long‐term lithium therapy with six healthy individuals. Under basal conditions, lithium‐treated individuals excreted significantly more urinary ammonia than did control subjects. Following an acute acid load, urinary ammonia excretion increased approximately twofold above basal rates in both lithium‐treated and control humans. There were no significant differences between lithium‐treated and control subjects in urinary pH or urinary citrate excretion. To elucidate possible mechanisms, rats were randomized to diets containing lithium or regular diet for 6 months. Similar to humans, basal ammonia excretion was significantly higher in lithium‐treated rats; in addition, urinary citrate excretion was also significantly greater. There were no differences in urinary pH. Expression of the critical ammonia transporter, Rhesus C Glycoprotein (Rhcg), was substantially greater in lithium‐treated rats than in control rats. We conclude that chronic lithium exposure increases renal ammonia excretion through mechanisms independent of urinary pH and likely to involve increased collecting duct ammonia secretion via the ammonia transporter, Rhcg. PMID:25501430

  6. Treatment of complete spinal cord injury patients by autologous bone marrow cell transplantation and administration of granulocyte-macrophage colony stimulating factor.

    PubMed

    Park, Hyung Chun; Shim, Yoo Shik; Ha, Yoon; Yoon, Seung Hwan; Park, So Ra; Choi, Byung Hyune; Park, Hyun Seon

    2005-01-01

    Transplantation of bone marrow cells into the injured spinal cord has been found to improve neurologic functions in experimental animal studies. However, it is unclear whether bone marrow cells can similarly improve the neurologic functions of complete spinal cord injury (SCI) in human patients. To address this issue, we evaluated the therapeutic effects of autologous bone marrow cell transplantation (BMT) in conjunction with the administration of granulocyte macrophage-colony stimulating factor (GM-CSF) in six complete SCI patients. BMT in the injury site (1.1 x 10(6) cells/microL in a total of 1.8 mL) and subcutaneous GM-CSF administration were performed on five patients. One patient was treated with GM-CSF only. The follow-up periods were from 6 to 18 months, depending on the patients. Sensory improvements were noted immediately after the operations. Sensory recovery in the sacral segment was noted mainly 3 weeks to 7 months postoperatively. Significant motor improvements were noted 3 to 7 months postoperatively. Four patients showed neurologic improvements in their American Spiral Injury Association Impairment Scale (AIS) grades (from A to C). One patient improved to AIS grade B from A and the last patient remained in AIS grade A. No immediate worsening of neurologic symptoms was found. Side effects of GMCSF treatment such as a fever (>38 degrees C) and myalgia were noted. Serious complications increasing mortality and morbidity were not found. The follow-up study with magnetic resonance imaging 4-6 months after injury showed slight enhancement within the zone of BMT. Syrinx formation was not definitely found. BMT and GM-CSF administration represent a safe protocol to efficiently manage SCI patients, especially those with acute complete injury. To demonstrate the full therapeutic value of this protocol, long-term and more comprehensive case-control clinical studies are required.

  7. Isolation, Culture, and Imaging of Human Fetal Pancreatic Cell Clusters

    PubMed Central

    Lopez, Ana D.; Kayali, Ayse G.; Hayek, Alberto; King, Charles C.

    2014-01-01

    For almost 30 years, scientists have demonstrated that human fetal ICCs transplanted under the kidney capsule of nude mice matured into functioning endocrine cells, as evidenced by a significant increase in circulating human C-peptide following glucose stimulation1-9. However in vitro, genesis of insulin producing cells from human fetal ICCs is low10; results reminiscent of recent experiments performed with human embryonic stem cells (hESC), a renewable source of cells that hold great promise as a potential therapeutic treatment for type 1 diabetes. Like ICCs, transplantation of partially differentiated hESC generate glucose responsive, insulin producing cells, but in vitro genesis of insulin producing cells from hESC is much less robust11-17. A complete understanding of the factors that influence the growth and differentiation of endocrine precursor cells will likely require data generated from both ICCs and hESC. While a number of protocols exist to generate insulin producing cells from hESC in vitro11-22, far fewer exist for ICCs10,23,24. Part of that discrepancy likely comes from the difficulty of working with human fetal pancreas. Towards that end, we have continued to build upon existing methods to isolate fetal islets from human pancreases with gestational ages ranging from 12 to 23 weeks, grow the cells as a monolayer or in suspension, and image for cell proliferation, pancreatic markers and human hormones including glucagon and C-peptide. ICCs generated by the protocol described below result in C-peptide release after transplantation under the kidney capsule of nude mice that are similar to C-peptide levels obtained by transplantation of fresh tissue6. Although the examples presented here focus upon the pancreatic endoderm proliferation and β cell genesis, the protocol can be employed to study other aspects of pancreatic development, including exocrine, ductal, and other hormone producing cells. PMID:24895054

  8. Human Neural Cell-Based Biosensor

    DTIC Science & Technology

    2011-03-11

    neurons, motoneurons and astrocytes using defined medium conditions, (3) cell-based methods to detect botulinum toxin, and (4) fluorescence based assays...medium conditions, (3) cell-based methods to detect botulinum toxin, and (4) fluorescence based assays for proliferation, cell migration, mitochondrial...line will begin shortly. (3) Development of cell based methods to detect botulinum toxin There has been substantial progress in the development

  9. Human cell dedifferentiation in mesenchymal condensates through controlled autophagy

    PubMed Central

    Pennock, Rebecca; Bray, Elen; Pryor, Paul; James, Sally; McKeegan, Paul; Sturmey, Roger; Genever, Paul

    2015-01-01

    Tissue and whole organ regeneration is a dramatic biological response to injury that occurs across different plant and animal phyla. It frequently requires the dedifferentiation of mature cells to a condensed mesenchymal blastema, from which replacement tissues develop. Human somatic cells cannot regenerate in this way and differentiation is considered irreversible under normal developmental conditions. Here, we sought to establish in vitro conditions to mimic blastema formation by generating different three-dimensional (3D) condensates of human mesenchymal stromal cells (MSCs). We identified specific 3D growth environments that were sufficient to dedifferentiate aged human MSCs to an early mesendoderm-like state with reversal of age-associated cell hypertrophy and restoration of organized tissue regenerating capacity in vivo. An optimal auophagic response was required to promote cytoplasmic remodeling, mitochondrial regression, and a bioenergetic shift from oxidative phosphorylation to anaerobic metabolism. Our evidence suggests that human cell dedifferentiation can be achieved through autonomously controlled autophagic flux. PMID:26290392

  10. Virus-Infected Human Mast Cells Enhance Natural Killer Cell Functions.

    PubMed

    Portales-Cervantes, Liliana; Haidl, Ian D; Lee, Patrick W; Marshall, Jean S

    2017-01-01

    Mucosal surfaces are protected from infection by both structural and sentinel cells, such as mast cells. The mast cell's role in antiviral responses is poorly understood; however, they selectively recruit natural killer (NK) cells following infection. Here, the ability of virus-infected mast cells to enhance NK cell functions was examined. Cord blood-derived human mast cells infected with reovirus (Reo-CBMC) and subsequent mast cell products were used for the stimulation of human NK cells. NK cells upregulated the CD69 molecule and cytotoxicity-related genes, and demonstrated increased cytotoxic activity in response to Reo-CBMC soluble products. NK cell interferon (IFN)-γ production was also promoted in the presence of interleukin (IL)-18. In vivo, SCID mice injected with Reo-CBMC in a subcutaneous Matrigel model, could recruit and activate murine NK cells, a property not shared by normal human fibroblasts. Soluble products of Reo-CBMC included IL-10, TNF, type I and type III IFNs. Blockade of the type I IFN receptor abrogated NK cell activation. Furthermore, reovirus-infected mast cells expressed multiple IFN-α subtypes not observed in reovirus-infected fibroblasts or epithelial cells. Our data define an important mast cell IFN response, not shared by structural cells, and a subsequent novel mast cell-NK cell immune axis in human antiviral host defense.

  11. Can intradermal administration of angiotensin II influence human heat loss responses during whole body heat stress?

    PubMed

    Fujii, Naoto; Meade, Robert D; Paull, Gabrielle; McGinn, Ryan; Foudil-bey, Imane; Akbari, Pegah; Kenny, Glen P

    2015-05-01

    It is unclear if angiotensin II, which can increase the production of reactive oxygen species (oxidative stress), modulates heat loss responses of cutaneous blood flow and sweating. We tested the hypothesis that angiotensin II-induced increases in oxidative stress impair cutaneous perfusion and sweating during rest and exercise in the heat. Eleven young (24 ± 4 yr) healthy adults performed two 30-min cycling bouts at a fixed rate of metabolic heat production (400 W) in the heat (35°C). The first and second exercises were followed by a 20- and 40-min recovery. Four microdialysis fibers were placed in the forearm skin for continuous administration of either: 1) lactated Ringer (control), 2) 10 μM angiotensin II, 3) 10 mM ascorbate (an antioxidant), or 4) a combination of 10 μM angiotensin II + 10 mM ascorbate. Cutaneous vascular conductance (CVC; laser-Doppler perfusion units/mean arterial pressure) and sweating (ventilated capsule) were evaluated at each skin site. Compared with control, angiotensin II reduced both CVC and sweating at baseline resting and during each recovery in the heat (all P < 0.05). However, during both exercise bouts, there were no differences in CVC or sweating between the treatment sites (all P > 0.05). When ascorbate was coinfused with angiotensin II, the effect of angiotensin II on sweating was abolished (all P > 0.05); however, its effect on CVC at baseline resting and during each recovery remained intact (all P < 0.05). We show angiotensin II impairs cutaneous perfusion independent of oxidative stress, while it impairs sweating through increasing oxidative stress during exposure to an ambient heat stress before and following exercise.

  12. Methamphetamine and Amphetamine Isomer Concentrations in Human Urine Following Controlled Vicks VapoInhaler Administration

    PubMed Central

    Smith, Michael L.; Nichols, Daniel C.; Underwood, Paula; Fuller, Zachary; Moser, Matthew A.; Flegel, Ron; Gorelick, David A.; Newmeyer, Matthew N.; Concheiro, Marta; Huestis, Marilyn A.

    2014-01-01

    Legitimate use of legal intranasal decongestants containing l-methamphetamine may complicate interpretation of urine drug tests positive for amphetamines. Our study hypotheses were that commonly used immunoassays would produce no false-positive results and a recently developed enantiomer-specific gas chromatography–mass spectrometry (GC–MS) procedure would find no d-amphetamine or d-methamphetamine in urine following controlled Vicks VapoInhaler administration at manufacturer's recommended doses. To evaluate these hypotheses, 22 healthy adults were each administered one dose (two inhalations in each nostril) of a Vicks VapoInhaler every 2 h for 10 h on Day 1 (six doses), followed by a single dose on Day 2. Every urine specimen was collected as an individual void for 32 h after the first dose and assayed for d- and l-amphetamines specific isomers with a GC–MS method with >99% purity of R-(−)-α-methoxy-α-(trifluoromethyl)phenylacetyl derivatives and 10 µg/L lower limits of quantification. No d-methamphetamine or d-amphetamine was detected in any urine specimen by GC–MS. The median l-methamphetamine maximum concentration was 62.8 µg/L (range: 11.0–1,440). Only two subjects had detectable l-amphetamine, with maximum concentrations coinciding with l-methamphetamine peak levels, and always ≤4% of the parent's maximum. Three commercial immunoassays for amphetamines EMIT® II Plus, KIMS® II and DRI® had sensitivities, specificities and efficiencies of 100, 97.8, 97.8; 100, 99.6, 99.6 and 100, 100, 100%, respectively. The immunoassays had high efficiencies, but our first hypothesis was not affirmed. The EMIT® II Plus assay produced 2.2% false-positive results, requiring an enantiomer-specific confirmation. PMID:25217541

  13. Methamphetamine and amphetamine isomer concentrations in human urine following controlled Vicks VapoInhaler administration.

    PubMed

    Smith, Michael L; Nichols, Daniel C; Underwood, Paula; Fuller, Zachary; Moser, Matthew A; Flegel, Ron; Gorelick, David A; Newmeyer, Matthew N; Concheiro, Marta; Huestis, Marilyn A

    2014-10-01

    Legitimate use of legal intranasal decongestants containing l-methamphetamine may complicate interpretation of urine drug tests positive for amphetamines. Our study hypotheses were that commonly used immunoassays would produce no false-positive results and a recently developed enantiomer-specific gas chromatography-mass spectrometry (GC-MS) procedure would find no d-amphetamine or d-methamphetamine in urine following controlled Vicks VapoInhaler administration at manufacturer's recommended doses. To evaluate these hypotheses, 22 healthy adults were each administered one dose (two inhalations in each nostril) of a Vicks VapoInhaler every 2 h for 10 h on Day 1 (six doses), followed by a single dose on Day 2. Every urine specimen was collected as an individual void for 32 h after the first dose and assayed for d- and l-amphetamines specific isomers with a GC-MS method with >99% purity of R-(-)-α-methoxy-α-(trifluoromethyl)phenylacetyl derivatives and 10 µg/L lower limits of quantification. No d-methamphetamine or d-amphetamine was detected in any urine specimen by GC-MS. The median l-methamphetamine maximum concentration was 62.8 µg/L (range: 11.0-1,440). Only two subjects had detectable l-amphetamine, with maximum concentrations coinciding with l-methamphetamine peak levels, and always ≤ 4% of the parent's maximum. Three commercial immunoassays for amphetamines EMIT(®) II Plus, KIMS(®) II and DRI(®) had sensitivities, specificities and efficiencies of 100, 97.8, 97.8; 100, 99.6, 99.6 and 100, 100, 100%, respectively. The immunoassays had high efficiencies, but our first hypothesis was not affirmed. The EMIT(®) II Plus assay produced 2.2% false-positive results, requiring an enantiomer-specific confirmation.

  14. Aripiprazole effects on self-administration and pharmacodynamics of intravenous cocaine and cigarette smoking in humans.

    PubMed

    Lofwall, Michelle R; Nuzzo, Paul A; Campbell, Charles; Walsh, Sharon L

    2014-06-01

    Aripiprazole is a partial agonist at dopamine (D2) and serotonin (5-HT1a) receptors and 5-HT2 antagonist. Because cocaine affects dopamine and serotonin, this study assessed whether aripiprazole could diminish the reinforcing efficacy of cocaine. Secondary aims evaluated aripiprazole on ad lib cigarette smoking and with a novel 40-hr smoking abstinence procedure. Adults with regular cocaine and cigarette use completed this inpatient double blind, randomized, placebo-controlled mixed-design study. A placebo lead-in was followed by randomization to aripiprazole (0, 2 or 10 mg/day/p.o.; n = 7 completed/group). Three sets of test sessions, each consisting of 3 cocaine sample-choice (i.e., self-administration) sessions and 1 dose-response session, were conducted (once during the lead-in and twice after randomization). Sample sessions tested each cocaine dose (0, 20 and 40 mg/70 kg, i.v.) in random order; subjective, observer-rated and physiologic outcomes were collected. Later that day, participants chose between the morning's sample dose or descending amounts of money over 7 trials. In dose response sessions, all doses were given 1 hr apart in ascending order for pharmacodynamic and pharmacokinetic assessment. Two sets of smoking topography sessions were conducted during the lead-in and after randomization; 1 with and 1 without 40 hr of smoking abstinence. Number of ad lib cigarettes smoked during non-session days was collected. Cocaine produced prototypic effects, but aripiprazole did not significantly alter these effects or smoking outcomes. The smoking abstinence procedure reliably produced nicotine withdrawal and craving and increased smoking modestly. These data do not support further investigation of aripiprazole for cocaine or tobacco use disorder treatment.

  15. Can intradermal administration of angiotensin II influence human heat loss responses during whole body heat stress?

    PubMed Central

    Fujii, Naoto; Meade, Robert D.; Paull, Gabrielle; McGinn, Ryan; Foudil-bey, Imane; Akbari, Pegah

    2015-01-01

    It is unclear if angiotensin II, which can increase the production of reactive oxygen species (oxidative stress), modulates heat loss responses of cutaneous blood flow and sweating. We tested the hypothesis that angiotensin II-induced increases in oxidative stress impair cutaneous perfusion and sweating during rest and exercise in the heat. Eleven young (24 ± 4 yr) healthy adults performed two 30-min cycling bouts at a fixed rate of metabolic heat production (400 W) in the heat (35°C). The first and second exercises were followed by a 20- and 40-min recovery. Four microdialysis fibers were placed in the forearm skin for continuous administration of either: 1) lactated Ringer (control), 2) 10 μM angiotensin II, 3) 10 mM ascorbate (an antioxidant), or 4) a combination of 10 μM angiotensin II + 10 mM ascorbate. Cutaneous vascular conductance (CVC; laser-Doppler perfusion units/mean arterial pressure) and sweating (ventilated capsule) were evaluated at each skin site. Compared with control, angiotensin II reduced both CVC and sweating at baseline resting and during each recovery in the heat (all P < 0.05). However, during both exercise bouts, there were no differences in CVC or sweating between the treatment sites (all P > 0.05). When ascorbate was coinfused with angiotensin II, the effect of angiotensin II on sweating was abolished (all P > 0.05); however, its effect on CVC at baseline resting and during each recovery remained intact (all P < 0.05). We show angiotensin II impairs cutaneous perfusion independent of oxidative stress, while it impairs sweating through increasing oxidative stress during exposure to an ambient heat stress before and following exercise. PMID:25767030

  16. Selective binding of human cumulus cell-secreted glycoproteins to human spermatozoa during capacitation in vitro

    SciTech Connect

    Tesarik, J.; Kopecny, V.; Dvorak, M.

    1984-06-01

    The results of this study demonstrate that glycoproteins manufactured by human cumulus cells can be detected bound to human spermatozoa incubated in capacitational medium containing the labeled cumulus-cell secretions. Cumulus-cell-secreted glycoproteins were labeled with a mixture of /sup 3/H-methionine and /sup 3/H-tryptophan or with 3H-fucose, and the binding of the labeled compounds to spermatozoa was evaluated by autoradiography. The binding was highly selective, involving only approximately 1% of the samples of spermatozoa used. The results suggest that the binding of cumulus-cell-secreted glycoproteins to spermatozoa may represent a final and highly selective step in human sperm capacitation.

  17. Derivation of novel human ground state naive pluripotent stem cells.

    PubMed

    Gafni, Ohad; Weinberger, Leehee; Mansour, Abed AlFatah; Manor, Yair S; Chomsky, Elad; Ben-Yosef, Dalit; Kalma, Yael; Viukov, Sergey; Maza, Itay; Zviran, Asaf; Rais, Yoach; Shipony, Zohar; Mukamel, Zohar; Krupalnik, Vladislav; Zerbib, Mirie; Geula, Shay; Caspi, Inbal; Schneir, Dan; Shwartz, Tamar; Gilad, Shlomit; Amann-Zalcenstein, Daniela; Benjamin, Sima; Amit, Ido; Tanay, Amos; Massarwa, Rada; Novershtern, Noa; Hanna, Jacob H

    2013-12-12

    Mouse embryonic stem (ES) cells are isolated from the inner cell mass of blastocysts, and can be preserved in vitro in a naive inner-cell-mass-like configuration by providing exogenous stimulation with leukaemia inhibitory factor (LIF) and small molecule inhibition of ERK1/ERK2 and GSK3β signalling (termed 2i/LIF conditions). Hallmarks of naive pluripotency include driving Oct4 (also known as Pou5f1) transcription by its distal enhancer, retaining a pre-inactivation X chromosome state, and global reduction in DNA methylation and in H3K27me3 repressive chromatin mark deposition on developmental regulatory gene promoters. Upon withdrawal of 2i/LIF, naive mouse ES cells can drift towards a primed pluripotent state resembling that of the post-implantation epiblast. Although human ES cells share several molecular features with naive mouse ES cells, they also share a variety of epigenetic properties with primed murine epiblast stem cells (EpiSCs). These include predominant use of the proximal enhancer element to maintain OCT4 expression, pronounced tendency for X chromosome inactivation in most female human ES cells, increase in DNA methylation and prominent deposition of H3K27me3 and bivalent domain acquisition on lineage regulatory genes. The feasibility of establishing human ground state naive pluripotency in vitro with equivalent molecular and functional features to those characterized in mouse ES cells remains to be defined. Here we establish defined conditions that facilitate the derivation of genetically unmodified human naive pluripotent stem cells from already established primed human ES cells, from somatic cells through induced pluripotent stem (iPS) cell reprogramming or directly from blastocysts. The novel naive pluripotent cells validated herein retain molecular characteristics and functional properties that are highly similar to mouse naive ES cells, and distinct from conventional primed human pluripotent cells. This includes competence in the generation

  18. Galangin inhibits growth of human head and neck squamous carcinoma cells in vitro and in vivo.

    PubMed

    Zhu, Liping; Luo, Qingqiong; Bi, Jianjun; Ding, Jieying; Ge, Shengfang; Chen, Fuxiang

    2014-12-05

    Galangin, an active flavonoid component extracted from the propolis and root of Alpinia officinarum Hance, has anti-tumor activity, but the mechanisms by which galangin affects various cancers, including human head and neck squamous cell carcinoma (HNSCC) remain unclear. In this study, we demonstrated for the first time that galangin suppressed the growth of HNSCC in vivo. With the cell culture system, galangin inhibited the proliferation and colony formation of HNSCC cells in a dose-dependent manner. Galangin induced significant cell cycle arrest of the tumor cells at the G0/G1 phase, which was accompanied by reduced AKT phosphorylation and mammalian target of rapamycin and S6 kinase activation. Decreased expression of cyclin D1, cyclin-dependent kinase (CDK)4, CDK6 and phosphorylation of retinoblastoma protein was observed in galangin-treated HNSCC cells. In addition, galangin induced apoptosis of HNSCC cells, downregulating antiapoptotic protein Bcl-2 and Bcl-xL and upregulating proapoptotic protein Bax and cleaved caspase 3. Immunohistochemical analysis showed a dose-dependent reduction in cyclin-D1-positive cancer cells and an increase in TUNEL-positive cancer cells in galangin-administrated mouse tumor sections. Therefore, galangin may be a novel therapeutic option in human HNSCC treatment.

  19. Natural killer cell distribution and trafficking in human tissues

    PubMed Central

    Carrega, Paolo; Ferlazzo, Guido

    2012-01-01

    Few data are available regarding the recirculation of natural killer (NK) cells among human organs. Earlier studies have been often impaired by the use of markers then proved to be either not sufficiently specific for NK cells (e.g., CD57, CD56) or expressed only by subsets of NK cells (e.g., CD16). At the present, available data confirmed that human NK cells populate blood, lymphoid organs, lung, liver, uterus (during pregnancy), and gut. Several studies showed that NK cell homing appears to be subset-specific, as secondary lymphoid organs and probably several solid tissues are preferentially inhabited by CD56brightCD16neg/dull non-cytotoxic NK cells. Similar studies performed in the mouse model showed that lymph node and bone marrow are preferentially populated by CD11bdull NK cells while blood, spleen, and lung by CD27dull NK cells. Therefore, an important topic to be addressed in the human system is the contribution of factors that regulate NK cell tissue homing and egress, such as chemotactic receptors or homeostatic mechanisms. Here, we review the current knowledge on NK cell distribution in peripheral tissues and, based on recent acquisitions, we propose our view regarding the recirculation of NK cells in the human body. PMID:23230434

  20. Synthetic vs natural scaffolds for human limbal stem cells

    PubMed Central

    Tominac Trcin, Mirna; Dekaris, Iva; Mijović, Budimir; Bujić, Marina; Zdraveva, Emilija; Dolenec, Tamara; Pauk-Gulić, Maja; Primorac, Dragan; Crnjac, Josip; Špoljarić, Branimira; Mršić, Gordan; Kuna, Krunoslav; Špoljarić, Daniel; Popović, Maja

    2015-01-01

    Aim To investigate the impact of synthetic electrospun polyurethane (PU) and polycaprolactone (PCL) nanoscaffolds, before and after hydrolytic surface modification, on viability and differentiation of cultured human eye epithelial cells, in comparison with natural scaffolds: fibrin and human amniotic membrane. Methods Human placenta was taken at elective cesarean delivery. Fibrin scaffolds were prepared from commercial fibrin glue kits. Nanoscaffolds were fabricated by electrospinning. Limbal cells were isolated from surpluses of human cadaveric cornea and seeded on feeder 3T3 cells. The scaffolds used for viability testing and immunofluorescence analysis were amniotic membrane, fibrin, PU, and PCL nanoscaffolds, with or without prior NaOH treatment. Results Scanning electron microscope photographs of all tested scaffolds showed good colony spreading of seeded limbal cells. There was a significant difference in viability performance between cells with highest viability cultured on tissue culture plastic and cells cultured on all other scaffolds. On the other hand, electrospun PU, PCL, and electrospun PCL treated with NaOH had more than 80% of limbal cells positive for stem cell marker p63 compared to only 27%of p63 positive cells on fibrin. Conclusion Natural scaffolds, fibrin and amniotic membrane, showed better cell viability than electrospun scaffolds. On the contrary, high percentages of p63 positive cells obtained on these scaffolds still makes them good candidates for efficient delivery systems for therapeutic purposes. PMID:26088849

  1. Radiogenic transformation of human mammary epithelial cells in vitro

    NASA Technical Reports Server (NTRS)

    Yang, T. C.; Georgy, K. A.; Tavakoli, A.; Craise, L. M.; Durante, M.

    1996-01-01

    Cancer induction by space radiations is a major concern for manned space exploration. Accurate assessment of radiation risk at low doses requires basic understanding of mechanism(s) of radiation carcinogenesis. For determining the oncogenic effects of ionizing radiation in human epithelial cells, we transformed a mammary epithelial cell line (185B5), which was immortalized by benzo(a)pyrene, with energetic heavy ions and obtained several transformed clones. These transformed cells showed growth properties on Matrigel similar to human mammary tumor cells. To better understand the mechanisms of radiogenic transformation of human cells, we systematically examined the alterations in chromosomes and cancer genes. Among 16 autosomes examined for translocations, by using fluorescence in situ hybridization (FISH) technique, chromosomes 3, 12, 13, 15, 16, and 18 appeared to be normal in transformed cells. Chromosomes 1, 4, 6, 8, and 17 in transformed cells, however, showed patterns different from those in nontransformed cells. Southern blot analyses indicated no detectable alterations in myc, ras, Rb, or p53 genes. Further studies of chromosome 17 by using in situ hybridization with unique sequence p53 gene probe and a centromere probe showed no loss of p53 gene in transformed cells. Experimental results from cell fusion studies indicated that the transforming gene(s) is recessive. The role of genomic instability and tumor suppressor gene(s) in radiogenic transformation of human breast cells remains to be identified.

  2. [Human peritoneum in vitro: changes in urate transport after administration of pyrazinoic acid].

    PubMed

    Czyzewska, K; Grzegorzewska, A; Stawny, B; Knapowski, J

    1989-01-01

    The article is an analysis of the dynamics of two-direction transportation of uric acid (UA) through the human peritoneum in vitro, and also changes of the dynamics under the influence of pyrazinoic++ acid. The peritoneum was taken from the anterior abdominal wall of patients undergoing planned abdominal surgery. It was found that the transportation of UA both from the vascular to the mesothelial side of the peritoneal membrane and in the opposite direction remained on a stable level for 120 minutes. The introduction of pyrazinoic++ acid decreased the transportation of UA from the vascular to the mesothelial side of the peritoneum on the average by 50 per cent. The transportation in the opposite direction did not change. The results obtained are consistent with results of clinical examinations. One may suppose that pyrazinoic++ acid induces changes in transportation qualities of the peritoneum.

  3. Endotoxin administration to humans inhibits hepatic cytochrome P450-mediated drug metabolism.

    PubMed Central

    Shedlofsky, S I; Israel, B C; McClain, C J; Hill, D B; Blouin, R A

    1994-01-01

    In experimental animals, injection of gram-negative endotoxin (LPS) decreases hepatic cytochrome P450-mediated drug metabolism. To evaluate this phenomenon in a human model of gram-negative sepsis, LPS was administered on two consecutive days to healthy male volunteers during which time a cocktail of antipyrine (AP-250 mg), hexobarbital (HB-500 mg), and theophylline (TH-150 mg) was ingested and the apparent oral clearance of each drug determined. Each subject had a control drug clearance study with saline injections. In the first experiment, six subjects received the drug cocktail 0.5 h after the first dose of LPS. In the second experiment, another six subjects received the drug cocktail 0.5 h after the second dose of LPS. In both experiments, LPS caused the expected physiologic responses of inflammation including fever with increases in serum concentrations of TNF alpha, IL-1 beta, IL-6, and acute phase reactants. In the first experiment, only minor decreases in clearances of the probe drugs were observed (7-12%). However in the second experiment, marked decreases in the clearances of AP (35, 95% CI 18-48%), HB (27, 95% CI 14-34%), and TH (22, 95% CI 12-32%) were seen. The decreases in AP clearance correlated with initial peak values of TNF alpha (r = 0.82) and IL-6 (r = 0.86). These data show that in humans the inflammatory response to even a very low dose of LPS significantly decreases hepatic cytochrome P450-mediated drug metabolism and this effect evolves over a 24-h period. It is likely that septic patients with much higher exposures to LPS have more profound inhibition of drug metabolism. PMID:7989576

  4. Analysis of free hydroxytyrosol in human plasma following the administration of olive oil.

    PubMed

    Pastor, Antoni; Rodríguez-Morató, Jose; Olesti, Eulàlia; Pujadas, Mitona; Pérez-Mañá, Clara; Khymenets, Olha; Fitó, Montserrat; Covas, María-Isabel; Solá, Rosa; Motilva, María-José; Farré, Magí; de la Torre, Rafael

    2016-03-11

    Hydroxytyrosol (HT) from olive oil, a potent bioactive molecule with health benefits, has a poor bioavailability, its free form (free HT) being undetectable so far. This fact leads to the controversy whether attained HT concentrations after olive oil polyphenol ingestion are too low to explain the observed biological activities. Due to this, an analytical methodology to determine free HT in plasma is crucial for understanding HT biological activity. Plasma HT instability and low concentrations have been major limitations for its quantification in clinical studies. Here, we describe a method to detect and quantify free HT in human plasma by using liquid chromatography coupled to tandem mass spectrometry. The method encompasses different steps of sample preparation including plasma stabilization, protein precipitation, selective derivatization with benzylamine, and purification by solid-phase extraction. A high sensitivity (LOD, 0.3ng/mL), specificity and stability of HT is achieved following these procedures. The method was validated and its applicability was demonstrated by analyzing human plasma samples after olive oil intake. A pharmacokinetic comparison was performed measuring free HT plasma concentrations following the intake of 25mL of ordinary olive oil (nearly undetectable concentrations) versus an extra-virgin olive oil (Cmax=4.40ng/mL). To our knowledge, this is the first time that an analytical procedure for quantifying free HT in plasma after olive oil dietary doses has been reported. The present methodology opens the door to a better understanding of the relationship between HT plasma concentrations and its beneficial health effects.

  5. SU11657 Enhances Radiosensitivity of Human Meningioma Cells

    SciTech Connect

    Milker-Zabel, Stefanie Bois, Angelika Zabel-du; Ranai, Gholamreza; Trinh, Thuy; Unterberg, Andreas; Debus, Juergen; Lipson, Kenneth E.; Abdollahi, Amir; Huber, Peter E.

    2008-03-15

    Purpose: To analyze the effect of the multireceptor tyrosine kinase inhibitor SU11657 (primarily vascular endothelial growth factor, platelet-derived growth factor) in combination with irradiation in freshly isolated primary human meningioma cells. Methods and Materials: Tumor specimens were obtained from meningioma patients undergoing surgery at the Department of Neurosurgery, University of Heidelberg, Germany. For the present study only cells up to passage 6 were used. Benign and atypical meningioma cells and human umbilical vein endothelial cells (HUVEC) were treated with SU11657 alone and in combination with 6-MV photons (0-10 Gy). Clonogenic survival and cell proliferation were determined alone and in coculture assays to determine direct and paracrine effects. Results: Radiation and SU11657 alone reduced cell proliferation in atypical and benign meningioma cells as well as in HUVEC in a dose-dependent manner. SU11657 alone also reduced clonogenic survival of benign and atypical meningioma cells. SU11657 increased radiosensitivity of human meningioma cells in clonogenic survival and cell number/proliferation assays. The anticlonogenic and antiproliferative effects alone and the radiosensitization effects of SU11657 were more pronounced in atypical meningioma cells compared with benign meningioma cells. Conclusion: Small-molecule tyrosine kinase inhibitors like SU11657 are capable of amplifying the growth inhibitory effects of irradiation in meningioma cells. These data provide a rationale for further clinical evaluation of this combination concept, especially in atypical and malignant meningioma patients.

  6. Effects of preset sequential administrations of sunitinib and everolimus on tumour differentiation in Caki-1 renal cell carcinoma

    PubMed Central

    Santos, C D; Tijeras-Raballand, A; Serova, M; Sebbagh, S; Slimane, K; Faivre, S; de Gramont, A; Raymond, E

    2015-01-01

    Background: Sunitinib (VEGFR/PDGFR inhibitor) and everolimus (mTOR inhibitor) are both approved for advanced renal cell carcinoma (RCC) as first-line and second-line therapy, respectively. In the clinics, sunitinib treatment is limited by the emergence of acquired resistance, leading to a switch to second-line treatment at progression, often based on everolimus. No data have been yet generated on programmed alternating sequential strategies combining alternative use of sunitinib and everolimus before progression. Such strategy is expected to delay the emergence of acquired resistance and improve tumour control. The aim of our study was to assess the changes in tumours induced by three different sequences administration of sunitinib and everolimus. Methods: In human Caki-1 RCC xenograft model, sunitinib was alternated with everolimus every week, every 2 weeks, or every 3 weeks. Effects on necrosis, hypoxia, angiogenesis, and EMT status were assessed by immunohisochemistry and immunofluorescence. Results: Sunitinib and everolimus programmed sequential regimens before progression yielded longer median time to tumour progression than sunitinib and everolimus monotherapies. In each group of treatment, tumour growth control was associated with inhibition of mTOR pathway and changes from a mesenchymal towards an epithelial phenotype, with a decrease in vimentin and an increase in E-cadherin expression. The sequential combinations of these two agents in a RCC mouse clinical trial induced antiangiogenic effects, leading to tumour necrosis. Conclusions: In summary, our study showed that alternate sequence of sunitinib and everolimus mitigated the development of mesenchymal phenotype compared with sunitinib as single agent. PMID:25422908

  7. Suppression of Medulloblastoma Lesions by Forced Migration of Preneoplastic Precursor Cells with Intracerebellar Administration of the Chemokine Cxcl3

    PubMed Central

    Ceccarelli, Manuela; Micheli, Laura; Tirone, Felice

    2016-01-01

    Medulloblastoma (MB), tumor of the cerebellum, remains a leading cause of cancer-related mortality in childhood. We previously showed, in a mouse model of spontaneous MB (Ptch1+/-/Tis21-/-), that a defect of the migration of cerebellar granule neuron precursor cells (GCPs) correlates with an increased frequency of MB. This occurs because GCPs, rather than migrating internally and differentiating, remain longer in the proliferative area at the cerebellar surface, becoming targets of transforming insults. Furthermore, we identified the chemokine Cxcl3 as responsible for the inward migration of GCPs. As it is known that preneoplastic GCPs (pGCPs) can still migrate and differentiate like normal GCPs, thus exiting the neoplastic program, in this study we tested the hypothesis that pGCPs within a MB lesion could be induced by Cxcl3 to migrate and differentiate. We observed that the administration of Cxcl3 for 28 days within the cerebellum of 1-month-old Ptch1+/-/Tis21-/- mice, i.e., when MB lesions are already formed, leads to complete disappearance of the lesions. However, a shorter treatment with Cxcl3 (2 weeks) was ineffective, suggesting that the suppression of MB lesions is dependent on the duration of Cxcl3 application. We verified that the treatment with Cxcl3 causes a massive migration of pGCPs from the lesion to the internal granular layer, where they differentiate. Thus, the induction of migration of pGCPs in MB lesions may open new ways to treat MB that exploit the plasticity of the pGCPs, forcing their differentiation. It remains to be tested whether this plasticity continues at advanced stages of MB. If so, these findings would set a potential use of the chemokine Cxcl3 as therapeutic agent against MB development in human preclinical studies. PMID:28018222

  8. Isolation and Culture of Embryonic Stem Cells, Mesenchymal Stem Cells, and Dendritic Cells from Humans and Mice.

    PubMed

    Kar, Srabani; Mitra, Shinjini; Banerjee, Ena Ray

    2016-01-01

    Stem cells are cells capable of proliferation, self-renewal, and differentiation into specific phenotypes. They are an essential part of tissue engineering, which is used in regenerative medicine in case of degenerative diseases. In this chapter, we describe the methods of isolating and culturing various types of stem cells, like human embryonic stem cells (hESCs), human umbilical cord derived mesenchymal stem cells (hUC-MSCs), murine bone marrow derived mesenchymal stem cells (mBM-MSCs), murine adipose tissue derived mesenchymal stem cells (mAD-MSCs), and murine bone marrow derived dendritic cells (mBMDCs). All these cell types can be used in tissue engineering techniques.

  9. 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.

  10. Comparative mutagenesis of human cells in vivo and in vitro

    SciTech Connect

    Thilly, W.G.

    1990-01-01

    Our goal is to develop the tools of mutational spectrometry in order to discover the cause(s) of genetic change in somatic and germinal cells in humans. Our study of the spectrum of point mutations in human mitochrondrial DNA sequences has revealed that there are multiple point mutation hotspots in each of four separate sequences in the mitochrondrial genome. These spectra were revealed by a combination of high fidelity PCR (modified T{sub 7} polymerase) and denaturing gradient gel electrophoresis which has a limit of detection of about 10{sup {minus}3}. There appear to be identical hotspot mutations in both cultured B cell and fresh human blood T cell samples.

  11. Oral adenosine-5’-triphosphate (ATP) administration increases blood flow following exercise in animals and humans

    PubMed Central

    2014-01-01

    Introduction Extracellular adenosine triphosphate (ATP) stimulates vasodilation by binding to endothelial ATP-selective P2Y2 receptors; a phenomenon, which is posited to be accelerated during exercise. Herein, we used a rat model to examine how different dosages of acute oral ATP administration affected the femoral blood flow response prior to, during, and after an exercise bout. In addition, we performed a single dose chronic administration pilot study in resistance trained athletes. Methods Animal study: Male Wistar rats were gavage-fed the body surface area, species adjusted human equivalent dose (HED) of either 100 mg (n=4), 400 mg (n=4), 1,000 mg (n=5) or 1,600 mg (n=5) of oral ATP as a disodium salt (Peak ATP®, TSI, Missoula, MT). Rats that were not gavage-fed were used as controls (CTL, n=5). Blood flow was monitored continuously: a) 60 min prior to, b) during and c) 90 min following an electrically-evoked leg-kicking exercise. Human Study: In a pilot study, 12 college-aged resistance-trained subjects were given 400 mg of ATP (Peak ATP®, TSI, Missoula, MT) daily for 12 weeks, and prior to an acute arm exercise bout at weeks 1, 4, 8, and 12. Ultrasonography-determined volumetric blood flow and vessel dilation in the brachial artery was measured at rest, at rest 30 minutes after supplementation, and then at 0, 3, and 6 minutes after the exercise. Results Animal Study: Rats fed 1,000 mg HED demonstrated significantly greater recovery blood flow (p < 0.01) and total blood flow AUC values (p < 0.05) compared to CTL rats. Specifically, blood flow was elevated in rats fed 1,000 mg HED versus CTL rats at 20 to 90 min post exercise when examining 10-min blood flow intervals (p < 0.05). When examining within-group differences relative to baseline values, rats fed the 1,000 mg and 1,600 mg HED exhibited the most robust increases in blood flow during exercise and into the recovery period. Human study: At weeks 1, 8, and 12, ATP supplementation significantly increased

  12. Human Liver Stem Cells Suppress T-Cell Proliferation, NK Activity, and Dendritic Cell Differentiation

    PubMed Central

    Bruno, Stefania; Grange, Cristina; Tapparo, Marta; Pasquino, Chiara; Romagnoli, Renato; Dametto, Ennia; Amoroso, Antonio; Tetta, Ciro; Camussi, Giovanni

    2016-01-01

    Human liver stem cells (HLSCs) are a mesenchymal stromal cell-like population resident in the adult liver. Preclinical studies indicate that HLSCs could be a good candidate for cell therapy. The aim of the present study was to evaluate the immunogenicity and the immunomodulatory properties of HLSCs on T-lymphocytes, natural killer cells (NKs), and dendritic cells (DCs) in allogeneic experimental settings. We found that HLSCs inhibited T-cell proliferation by a mechanism independent of cell contact and dependent on the release of prostaglandin E2 (PGE2) and on indoleamine 2,3-dioxygenase activity. When compared with mesenchymal stromal cells (MSCs), HLSCs were more efficient in inhibiting T-cell proliferation. At variance with MSCs, HLSCs did not elicit NK degranulation. Moreover, HLSCs inhibited NK degranulation against K562, a NK-sensitive target, by a mechanism dependent on HLA-G release. When tested on DC generation from monocytes, HLSCs were found to impair DC differentiation and DCs ability to induce T-cell proliferation through PGE2. This study shows that HLSCs have immunomodulatory properties similar to MSCs, but, at variance with MSCs, they do not elicit a NK response. PMID:27127520