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Sample records for human choriocarcinoma cell

  1. Isolation, Culture and Identification of Choriocarcinoma Stem-Like Cells from the Human Choriocarcinoma Cell-Line JEG-3.

    Science.gov (United States)

    Cai, Jingting; Peng, Tianfang; Wang, Jing; Zhang, Jingli; Hu, Hui; Tang, Dihong; Chu, Chaonan; Yang, Ting; Liu, Huining

    2016-01-01

    Cancer stem cells (CSCs) exhibit enhanced proliferative capacity and resistance to chemotherapy; however, choriocarcinoma CSCs have not yet been reported. In this study the human choriocarcinoma cell line JEG-3 was cultured in serum free media, and the characteristics of suspension and parental adherent JEG-3 cells were compared. Cell proliferation, colony-formation, soft agar clonogenicity, and transwell invasion assays were performed in vitro, and tumor xenografts in BALB/c nude mice were used to evaluate stem cell properties. In serum-supplemented medium (SSM), JEG-3 cells were 4.51 ± 1.71% CD44+, 7.67 ± 2.67% CD133+, and 13.85 ± 2.95% ABCG2+. In serum-free medium (SFM), the expression of these markers increased to 53.08 ± 3.15%, 47.40 ± 2.67%, and 78.70 ± 7.16%, respectively. Moreover, suspension JEG-3 cells exhibited enhanced colony-formation capability as well as invasive and proliferative ability in vitro, alongside enhanced tumorigenic properties in vivo. Suspension JEG-3 cells also exhibited resistance to the chemotherapeutic drugs methotrexate, fluorouracil and etoposide. When seeded in serum supplemented medium, suspension JEG-3 cells readopted an adherent phenotype and continued to differentiate with no significant difference in the morphology between suspension and parent cells. In this study, choriocarcinoma stem-like cells (CSLCs) were isolated from the human choriocarcinoma JEG-3 cell line by SFM culture and characterized. © 2016 The Author(s) Published by S. Karger AG, Basel.

  2. In vitro approaches to evaluate placental drug transport by using differentiating JEG-3 human choriocarcinoma cells.

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    Ikeda, Kenji; Utoguchi, Naoki; Tsutsui, Hidenobu; Yamaue, Satoko; Homemoto, Manami; Nakao, Erina; Hukunaga, Yumi; Yamasaki, Kyohei; Myotoku, Michiaki; Hirotani, Yoshihiko

    2011-02-01

    Human choriocarcinoma cells have been used as models for studying transcellular drug transport through placental trophoblasts. However, these models allow the transport of low-molecular-weight drugs through intercellular gap junctions. This study aimed at investigating the differentiation patterns of JEG-3 choriocarcinoma cells under different culture conditions and establishing the appropriate model of in vitro syncytiotrophoblast drug transport. Paracellular permeability was estimated by measuring the transepithelial electrical resistance (TEER) across JEG-3 cell layers. The mRNA expression levels of non-expressed in choriocarcinoma clone 1 (NECC1) and breast cancer resistance protein (BCRP), and those of E-cadherin (ECAD) and cadherin-11 (CDH11), which are adherens junction-associated proteins related to fusogenic ability of syncytiotrophoblasts differentiated from cytotrophoblasts, protein expression levels were considered as the differentiation signals. The highest TEER values were obtained in the JEG-3 cells cultured in the Dulbecco's modified Eagle's medium (DMEM)/Ham's F-12 (1:1) mixed medium (CS-C(®) ; Dainippon Sumitomo Pharma Co. Ltd., Osaka, Japan). By comparing the TEER values and the differentiation signals, the authors identified at least five JEG-3 cell-differentiation patterns. The differentiation pattern of JEG-3 cultured in CS-C resembled the syncytiotrophoblast-like differentiation signal characterizations in vivo. In conclusion, the syncytiotrophoblast-like models of differentiating JEG-3 cells cultured in CS-C might be appropriate for evaluating drug transport across the placental trophoblast. © 2010 The Authors. Basic & Clinical Pharmacology & Toxicology © 2010 Nordic Pharmacological Society.

  3. Coumestrol induces mitochondrial dysfunction by stimulating ROS production and calcium ion influx into mitochondria in human placental choriocarcinoma cells.

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    Lim, Whasun; Yang, Changwon; Jeong, Muhah; Bazer, Fuller W; Song, Gwonhwa

    2017-11-01

    Does coumestrol inhibit proliferation of human placental choriocarcinoma cells? Coumestrol promotes cell death in the choriocarcinoma cells by regulating ERK1/2 MAPK and JNK MAPK signaling pathways and through disruption of Ca2+ and ROS homeostasis. A number of patients who suffer from choriocarcinomas fail to survive due to delayed diagnosis or a recurrent tumor and resistance to traditional chemotherapy using platinum-based agents and methotrexate. To overcome these limitations, it is important to discover novel compounds which have no adverse effects yet can inhibit the expression of a target molecule to develop, as a novel therapeutic for prevention and/or treatment of choriocarcinomas. Effects of coumestrol on human placental choriocarcinoma cell lines, JAR and JEG3, were assessed in diverse assays in a dose- and time-dependent manner. Effects of coumestrol on cell proliferation, apoptosis (annexin V expression, propidium iodide staining, TUNEL and invasion assays), mitochondria-mediated apoptosis, production of reactive oxygen species (ROS), lipid peroxidation, glutathione levels and endoplasmic reticulum (ER) stress proteins in JAR and JEG3 cells were determined. Signal transduction pathways in JAR and JEG3 cells in response to coumestrol were determined by western blot analyses. Results of the present study indicated that coumestrol suppressed proliferation and increased apoptosis in JAR and JEG3 cells by inducing pro-apoptotic proteins, Bax and Bak. In addition, coumestrol increased ROS production, as well as lipid peroxidation and glutathione levels in JAR and JEG3 cells. Moreover, coumestrol-induced depolarization of mitochondrial membrane potential (MMP) and increased cytosolic and mitochondrial Ca2+ levels in JAR and JEG3 cells. Consistent with those results, treatment of JAR and JEG3 cells with a Ca2+ chelator and an inhibitor of IP3 receptor decreased coumestrol-induced depolarization of MMP and increased proliferation in JAR and JEG3 cells. N/A. A

  4. Choriocarcinoma

    Science.gov (United States)

    ... feed the fetus. Choriocarcinoma is a type of gestational trophoblastic disease . ... Goldstein DP, Berkowitz RS. Gestational trophoblastic disease. ... Abeloff's Clinical Oncology . 5th ed. Philadelphia, PA: Elsevier ...

  5. Lysosomal degradation of receptor-bound urokinase-type plasminogen activator is enhanced by its inhibitors in human trophoblastic choriocarcinoma cells

    DEFF Research Database (Denmark)

    Jensen, Poul Henning; Christensen, Erik Ilsø; Ebbesen, P.

    1990-01-01

    We have studied the effect of plasminogen activator inhibitors PAI-1 and PAI-2 on the binding of urokinase-type plasminogen activator (u-PA) to its receptor in the human choriocarcinoma cell line JAR. With 125I-labeled ligands in whole-cell binding assays, both uncomplexed u-PA and u-PA-inhibitor......We have studied the effect of plasminogen activator inhibitors PAI-1 and PAI-2 on the binding of urokinase-type plasminogen activator (u-PA) to its receptor in the human choriocarcinoma cell line JAR. With 125I-labeled ligands in whole-cell binding assays, both uncomplexed u-PA and u...... for the removal of u-PA after its complex formation with a specific inhibitor. The data suggest a novel mechanism by which receptor-mediated endocytosis is initiated by the binding of a secondary ligand. Full text...

  6. The human leukocyte antigen G promotes trophoblast fusion and β-hCG production through the Erk1/2 pathway in human choriocarcinoma cell lines

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    Wang, Ji-meng [School of Medicine, Nankai University, Tianjin 300071 (China); State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China); Zhao, Hong-xi [Department of Obstetrics and Gynecology, Tangdu Hospital, Fourth Military Medical University, Xi’an 710038 (China); Wang, Li [Department of Obstetrics and Gynecology, General Hospital of Chinese People’s Liberation Army, Beijing 100853 (China); Gao, Zhi-ying, E-mail: gaozy301@yahoo.com.cn [Department of Obstetrics and Gynecology, General Hospital of Chinese People’s Liberation Army, Beijing 100853 (China); Yao, Yuan-qing, E-mail: yqyao@126.com [Department of Obstetrics and Gynecology, General Hospital of Chinese People’s Liberation Army, Beijing 100853 (China)

    2013-05-10

    Highlights: •HLA-G expression promotes BeWo cells fusion and fusogenic gene expression. •HLA-G is capable of inducing β-hCG production in human choriocarcinoma cell lines. •Up-regulation of β-hCG production by HLA-G is mediated via the Erk1/2 pathway. -- Abstract: The human leukocyte antigen G (HLA-G) is expressed on the fetal–maternal interface and plays a role in protecting fetal-derived trophoblasts from the maternal immune response, allowing trophoblasts to invade the uterus. However, HLA-G also possesses immune suppressing-independent functions. We found that HLA-G expressing BeWo choriocarcinoma cells increased cell–cell fusion compared to control BeWo cells under forskolin treatment. Regardless of forskolin treatment, the expression of fusogenic gene mRNAs, including syncytin-1, the transcription factor glial cell missing 1 (Gcm1), and beta human chorionic gonadotropin (β-hCG) were elevated. HLA-G up-regulates β-hCG production in human choriocarcinoma cells because HLA-G knockdown in JEG-3 cells induces a dramatic decrease in β-hCG compared with control cells. The defect in β-hCG production in HLA-G knocked-down cells could not be completely overcome by stimulating hCG production through increasing intracellular cAMP levels. HLA-G expressing cells have increased phosphorylation levels for extracellular signal-regulated kinase1/2 (Erk1/2) in BeWo cells. The Erk1/2 pathway is inactivated after the inhibition of HLA-G expression in JEG-3 cells. Finally, Erk1/2 inhibition was able to suppress the increased hCG production induced by HLA-G expression. Together, these data suggest novel roles for HLA-G in regulating β-hCG production via the modulation of the Erk1/2 pathway and by inducing trophoblast cell fusion.

  7. The human leukocyte antigen G promotes trophoblast fusion and β-hCG production through the Erk1/2 pathway in human choriocarcinoma cell lines.

    Science.gov (United States)

    Wang, Ji-meng; Zhao, Hong-xi; Wang, Li; Gao, Zhi-ying; Yao, Yuan-qing

    2013-05-10

    The human leukocyte antigen G (HLA-G) is expressed on the fetal-maternal interface and plays a role in protecting fetal-derived trophoblasts from the maternal immune response, allowing trophoblasts to invade the uterus. However, HLA-G also possesses immune suppressing-independent functions. We found that HLA-G expressing BeWo choriocarcinoma cells increased cell-cell fusion compared to control BeWo cells under forskolin treatment. Regardless of forskolin treatment, the expression of fusogenic gene mRNAs, including syncytin-1, the transcription factor glial cell missing 1 (Gcm1), and beta human chorionic gonadotropin (β-hCG) were elevated. HLA-G up-regulates β-hCG production in human choriocarcinoma cells because HLA-G knockdown in JEG-3 cells induces a dramatic decrease in β-hCG compared with control cells. The defect in β-hCG production in HLA-G knocked-down cells could not be completely overcome by stimulating hCG production through increasing intracellular cAMP levels. HLA-G expressing cells have increased phosphorylation levels for extracellular signal-regulated kinase1/2 (Erk1/2) in BeWo cells. The Erk1/2 pathway is inactivated after the inhibition of HLA-G expression in JEG-3 cells. Finally, Erk1/2 inhibition was able to suppress the increased hCG production induced by HLA-G expression. Together, these data suggest novel roles for HLA-G in regulating β-hCG production via the modulation of the Erk1/2 pathway and by inducing trophoblast cell fusion. Copyright © 2013 Elsevier Inc. All rights reserved.

  8. Treatment of Human Placental Choriocarcinoma Cells with Formaldehyde and Benzene Induced Growth and Epithelial Mesenchymal Transition via Induction of an Antioxidant Effect

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    Hae-Miru Lee

    2017-07-01

    Full Text Available Cigarette smoke (CS causes about 480,000 deaths each year worldwide, and it is well-known to have harmful effects on the human body, leading to heart disease, stroke, lung cancer, and cardiovascular problems. In this study, the effects of formaldehyde (FA and benzene (Bz, the main components of CS, on cell proliferation and epithelial mesenchymal transition (EMT of JEG-3 human choriocarcinoma cells were examined to confirm the relationship between CS components and placenta carcinoma. Upon MTT assay, FA (10−8 M to 10−5 M and Bz (10−11 M to 10−8 M increased JEG-3 cell proliferation. Western blot assay revealed that the protein expression of cyclin D1 and E1 increased, while the levels of p21 and p27 were reduced following treatment. In Scratch assay, FA (10−8 M and 10−5 M and Bz (10−11 M and 10−8 M increased migration of JEG-3 cells at 24 h and 48 h compared with that at 0 h. In addition, the expression of the epithelial marker, E-cadherin, was significantly decreased, while the expression of the mesenchymal marker, N-cadherin, was significantly increased by FA (10−8 M and 10−5 M and Bz (10−11 M and 10−8 M. snail and slug transcriptional factors were associated with EMT, which were also up-regulated by FA and Bz, indicating that FA and Bz lead to an increase in the EMT process in JEG-3 choriocarcinoma cells. We further evaluated reactive oxygen species (ROS and activation of antioxidant effect using dichlorofluorescin diacetate (DCFH-DA and Western blot assay. FA and Bz increased the ROS production and an antioxidant related marker, Nrf2, in JEG-3 cells. However, eIF2α levels were reduced by FA and Bz via activation of the antioxidant reaction. Taken together, these results indicated that FA and Bz induce the growth and migration of human choriocarcinoma cells via regulation of the cell cycle and EMT and activation of ROS and antioxidant related markers.

  9. Combined choriocarcinoma, neuroendocrine cell carcinoma and tubular adenocarcinoma in the stomach

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    Hirano, Yasumitsu; Hara, Takuo; Nozawa, Hiroshi; Oyama, Kaeko; Ohta, Naohiro; Omura, Kenji; Watanabe, Go; Niwa, Hideki

    2008-01-01

    We described a patient with adenocarcinoma of the stomach combined with choriocarcinoma and neuroendocrine cell carcinoma. An 85-year-old man visited our hospital because of appetite loss. Gastric fiberscopy revealed a large tumor occupying the cardial region and anterior wall of the gastric body. The patient underwent total gastrectomy with lymphnode dissection and partial resection of the liver. Choriocarcinoma, small cell carcinoma and tubular adenocarcinoma existed in the gastric tumor. The choriocarcinomatous foci contained cells positive for beta-subunit of human chorionic gonadotropin (B-hCG) and human placental lactogen mainly in syncytiotrophoblastic cells. The small cell carcinomatous foci contained cells positive for synaptophysin, neuron-specific enolase (NSE), and chromogranin A. The prognosis for gastric adenocarcinoma with choriocarcinoma and neuroendocrine cell carcinoma is exceedingly poor. This patient died about 2 mo after the first complaint from hepatic failure. This is the first reported case of gastric cancer with these three pathological features. PMID:18506939

  10. Changes in the expression of pituitary adenylate cyclase-activating polypeptide in the human placenta during pregnancy and its effects on the survival of JAR choriocarcinoma cells.

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    Brubel, R; Boronkai, A; Reglodi, D; Racz, B; Nemeth, J; Kiss, P; Lubics, A; Toth, G; Horvath, G; Varga, T; Szogyi, D; Fonagy, E; Farkas, J; Barakonyi, A; Bellyei, Sz; Szereday, L; Koppan, M; Tamas, A

    2010-11-01

    Pituitary adenylate cyclase-activating polypeptide (PACAP), a neuropeptide with survival-promoting actions, has been observed in endocrine organs and is thought to play a role in reproductive functions, including pregnancy. PACAP occurs in two forms, 27 and 38 amino acid residues, with PACAP38 being the predominant form in human tissues. In the present study, we determined the concentrations of PACAP38 and PACAP27 in first-trimester and full-term human placentas using radioimmunoassay. We found high levels of PACAP38 and lower levels of PACAP27 in different parts of the full-term human placenta. PACAP38 content increased in the placenta during pregnancy, both on the maternal side and on the fetal side. The effects of PACAP on the survival of JAR human choriocarcinoma cells were investigated using flow cytometry and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) cell viability assay in cells exposed to the widely used chemotherapeutic agent methotrexate (MTX). It was found that PACAP neither influenced the survival of JAR cytotrophoblast cells nor affected cellular response to the death-inducing effect of the chemotherapeutic agent MTX. The present observations further support the significance of PACAP in the human placenta. The observation that PACAP did not influence the effects of MTX may have future clinical importance, showing that PACAP does not decrease the effects of certain chemotherapeutic agents.

  11. Differential expression of the HLA class I multigene family by human embryonal carcinoma and choriocarcinoma cell lines

    NARCIS (Netherlands)

    Rinke de Wit, T. F.; Vloemans, S.; van den Elsen, P. J.; Haworth, A.; Stern, P. L.

    1990-01-01

    We have studied the expression of both HLA class and the recently described HLA class I-like genes (HLA-E and HLA-6.0) in two human developmental tumor cell lines, that serologically could not be typed for HLA-A, -B, and -C. Evidence is presented that the teratocarcinoma Tera-2 stem cells express

  12. Downregulation of the gli transcription factors regulator Kif7 facilitates cell survival and migration of choriocarcinoma cells.

    Directory of Open Access Journals (Sweden)

    Joanna Ho

    Full Text Available The kinesin protein Kif7 has been recognized as an integral component of hedgehog signalling. Aberrant activation of hedgehog signalling has been implicated in many human solid tumours. Gestational trophoblastic disease includes frankly malignant choriocarcinoma and potentially malignant hydatidiform mole. Here we investigated the hedgehog signalling components expression profiles in gestational trophoblastic disease. Downregulation of Gli1, Gli2, Gli3 and Kif7 was demonstrated in clinical samples of choriocarcinoma and hydatidiform moles as well as choriocarcinoma cell lines when compared with normal placentas. Ectopic expression of Kif7 in two choriocarcinoma cell lines JAR and JEG-3 led to a decrease in cell growth and increase in apoptosis demonstrated by MTT and TUNEL assays, respectively. Overexpression of Kif7 also led to suppressed cell migration through transwell assay. In contrast, knocking down Kif7 in HTR-8/SVneo, an immortalized trophoblast cell line, increased cell number over time and increased the migratory ability of the cells. Taken together, Kif7 may contribute to pathogenesis of gestational trophoblastic disease through enhancing survival and promoting dissemination of trophoblasts.

  13. Choriocarcinoma: blocking factor and monoclonal antibody iodine 131 imaging

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    Pattillo, R.A.; Khazaeli, M.B.; Ruckert, A.C.; Hussa, R.O.; Collier, B.D.; Beierwaltes, W.; Mattingly, R.F.

    1984-04-01

    Postoperative iodine 131 monoclonal antibody localization in metastatic choriocarcinoma was accomplished in this study. The monoclonal antibody was prepared to male choriocarcinoma which cross reacted with gestational choriocarcinoma. The antibody was raised against whole choriocarcinoma cells and human chorionic gonadotropin (hCG) cross reactivity was excluded. The purified antibody was iodinated with /sup 131/I and successfully imaged BeWo choriocarcinoma transplanted in nude mice; however, imaging of choriocarcinoma in a patient was verified only after resection. It is our belief that failure to sufficiently concentrate the antibody in the tumor before operation was due to blocking factor in the serum of the patient. Blocking factor and hCG dropped postoperatively. Blocking factor activity in 15 patients with metastatic trophoblastic disease was monitored and, like hCG, was found to be a sensitive indicator of the presence of disease. Its efficacy may be in the small number of patients without hCG but with persistent disease.

  14. Gastrointestinal Choriocarcinoma: Case Report

    African Journals Online (AJOL)

    Choriocarcinoma arises from the chorionic epithelium of placentas in association with hydatidiform mole, recent abortion or full term pregnancy (1). It is a rap- idly growing, highly invasive and widely metastasizing tumour that produces human chorionic gonadotropin. (hCG). Choriocarcinomas may also arise from ectopic.

  15. Cellular immune recognition of HLA-G-expressing choriocarcinoma cell line Jeg-3

    NARCIS (Netherlands)

    Burt, D.; Johnston, D.; Rinke de Wit, T.; van den Elsen, P.; Stern, P. L.

    1991-01-01

    Jeg-3 choriocarcinoma cells express class-I MHC HLA-G and low levels of a novel HLA-C product. The functional significance of such novel MHC class-I expression in regard of the cellular immune response has been investigated. Jeg-3 cells are NK-insensitive, but susceptible to LAK cytotoxicity, some

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

    DEFF Research Database (Denmark)

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

    2010-01-01

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

  17. Plasmodium falciparum parasites expressing pregnancy-specific variant surface antigens adhere strongly to the choriocarcinoma cell line BeWo

    DEFF Research Database (Denmark)

    Haase, Rikke N; Megnekou, Rosette; Lundquist, Maja

    2006-01-01

    Placenta-sequestering Plasmodium falciparum parasites causing pregnancy-associated malaria express pregnancy-specific variant surface antigens (VSA(PAM)). We report here that VSA(PAM)-expressing patient isolates adhere strongly to the choriocarcinoma cell line BeWo and that the BeWo line can...

  18. Analysis of a candidate gene associated with growth suppression of choriocarcinoma and differentiation of trophoblasts.

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    Asanoma, Kazuo; Kato, Hidenori; Inoue, Takafumi; Matsuda, Takao; Wake, Norio

    2004-08-01

    To isolate a choriocarcinoma suppressor gene and analyze its structure and function. We constructed a polymerase chain reaction-based subtracted fragmentary cDNA library between normal placental villi and choriocarcinoma cell line CC1. A novel homeobox gene, designated NECC1 (not expressed in differ choriocarcinoma clone 1), is included in this library. We analyzed the structure and function of NECC1 with molecular biology. NECC1 comprises an open reading frame of 219 nucleotides encoding 73 amino acids and contains a homeodomain as a consensus motif. NECC1 is located on human chromosome 4q11-q12, and its expression is ubiquitous in the brain, placenta, lung, smooth muscle, uterus, bladder, kidney and spleen. Normal placental villi abundantly expressed NECC1, but all choriocarcinoma cell lines examined and most surgically removed choriocarcinoma tissue samples failed to express it. We transfected this gene into choriocarcinoma cell lines and observed remarkable alterations in cell morphology and suppression of in vivo tumorigenesis. Induction of chorionic somatomammotropin hormone 1 by NECC1 transfection suggested differentiation of choriocarcinoma cells to syncytiotrophoblast-like cells. Our results suggest that loss of NECC1 expression is involved in malignant conversion of placental trophoblasts.

  19. NECC1, a candidate choriocarcinoma suppressor gene that encodes a homeodomain consensus motif.

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    Asanoma, Kazuo; Matsuda, Takao; Kondo, Haruhiko; Kato, Kiyoko; Kishino, Tatsuya; Niikawa, Norio; Wake, Norio; Kato, Hidenori

    2003-01-01

    We isolated a candidate choriocarcinoma suppressor gene from a PCR-based subtracted fragmentary cDNA library between normal placental villi and the choriocarcinoma cell line CC1. This gene comprises an open reading frame of 219 nt encoding 73 amino acids and contains a homeodomain as a consensus motif. This gene, designated NECC1 (not expressed in choriocarcinoma clone 1), is located on human chromosome 4q11-q12. NECC1 expression is ubiquitous in the brain, placenta, lung, smooth muscle, uterus, bladder, kidney, and spleen. Normal placental villi expressed NECC1, but all choriocarcinoma cell lines examined and most of the surgically removed choriocarcinoma tissue samples failed to express it. We transfected this gene into choriocarcinoma cell lines and observed remarkable alterations in cell morphology and suppression of in vivo tumorigenesis. Induction of CSH1 (chorionic somatomammotropin hormone 1) by NECC1 expression suggested differentiation of choriocarcinoma cells to syncytiotrophoblasts. Our results suggest that loss of NECC1 expression is involved in malignant conversion of placental trophoblasts.

  20. Preparation and characterization of magnetic nanoparticles containing Fe(3)O(4)-dextran-anti-β-human chorionic gonadotropin, a new generation choriocarcinoma-specific gene vector.

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    Jingting, Cai; Huining, Liu; Yi, Zhang

    2011-01-01

    To evaluate the feasibility of using magnetic iron oxide (Fe(3)O(4))-dextran-anti-β-human chorionic gonadotropin (HCG) nanoparticles as a gene vector for cellular transfections. Fe(3)O(4)-dextran-anti-β-HCG nanoparticles were synthesized by chemical coprecipitation. The configuration, diameter, and iron content of the nanoparticles were detected by transmission electron microscopy (TEM), light scatter, and atomic absorption spectrophotometry. A3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetrazoliumromide assay was used to evaluate the cytotoxicity of Fe(3)O(4)-dextran-anti-β-HCG nanoparticles. Enzyme-linked immunosorbent assay and indirect immunofluorescence were used to evaluate immunoreactivity. The efficiency of absorbing DNA and resisting deoxyribonuclease I (DNase I) digestion when bound to Fe(3)O(4)-dextran-anti-β-HCG nanoparticles was examined by agarose gel electrophoresis. The ability of Fe(3)O(4)-dextran-anti-β-HCG nanoparticles to absorb heparanase antisense oligodeoxynucleotides (AS-ODN) nanoparticles in different cell lines was evaluated by flow cytometry. The tissue distribution of heparanase AS-ODN magnetic nanoparticles in choriocarcinoma tumors transplanted in nude mice was detected by atomic absorption spectrophotometry. TEM demonstrated that the shape of nanoparticles is irregular. Light scatter revealed nanoparticles with a mean diameter of 75.5 nm and an iron content of 37.5 μg/mL. No cytotoxicity was observed when the concentration of Fe(3)O(4)-dextran-anti-β-HCG nanoparticles was dextran nanoparticles have a satisfactory potential to combine with β-HCG antibody. Agarose gel electrophoresis analysis of binding experiments showed that after treatment with sodium periodate, Fe(3)O(4)-dextran-anti-β-HCG nanoparticles have a satisfactory potential to absorb DNA, and the protection experiment showed that nanoparticles can effectively protect DNA from DNase I digestion. Aldehyde Fe(3)O(4)-dextran-anti-β-HCG nanoparticles can transfect

  1. Preparation and characterization of magnetic nanoparticles containing Fe3O4-dextran-anti-β-human chorionic gonadotropin, a new generation choriocarcinoma-specific gene vector

    Science.gov (United States)

    Jingting, Cai; Huining, Liu; Yi, Zhang

    2011-01-01

    Objective: To evaluate the feasibility of using magnetic iron oxide (Fe3O4)-dextran-anti-β-human chorionic gonadotropin (HCG) nanoparticles as a gene vector for cellular transfections. Study design: Fe3O4-dextran-anti-β-HCG nanoparticles were synthesized by chemical coprecipitation. The configuration, diameter, and iron content of the nanoparticles were detected by transmission electron microscopy (TEM), light scatter, and atomic absorption spectrophotometry. A3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetrazoliumromide assay was used to evaluate the cytotoxicity of Fe3O4-dextran-anti-β-HCG nanoparticles. Enzyme-linked immunosorbent assay and indirect immunofluorescence were used to evaluate immunoreactivity. The efficiency of absorbing DNA and resisting deoxyribonuclease I (DNase I) digestion when bound to Fe3O4-dextran-anti-β-HCG nanoparticles was examined by agarose gel electrophoresis. The ability of Fe3O4-dextran-anti-β-HCG nanoparticles to absorb heparanase antisense oligodeoxynucleotides (AS-ODN) nanoparticles in different cell lines was evaluated by flow cytometry. The tissue distribution of heparanase AS-ODN magnetic nanoparticles in choriocarcinoma tumors transplanted in nude mice was detected by atomic absorption spectrophotometry. Results: TEM demonstrated that the shape of nanoparticles is irregular. Light scatter revealed nanoparticles with a mean diameter of 75.5 nm and an iron content of 37.5 μg/mL. No cytotoxicity was observed when the concentration of Fe3O4-dextran-anti-β-HCG nanoparticles was dextran nanoparticles have a satisfactory potential to combine with β-HCG antibody. Agarose gel electrophoresis analysis of binding experiments showed that after treatment with sodium periodate, Fe3O4-dextran-anti-β-HCG nanoparticles have a satisfactory potential to absorb DNA, and the protection experiment showed that nanoparticles can effectively protect DNA from DNase I digestion. Aldehyde Fe3O4-dextran-anti-β-HCG nanoparticles can transfect

  2. Preimplantation Factor (PIF Promotes HLA-G, -E, -F, -C Expression in JEG-3 Choriocarcinoma Cells and Endogenous Progesterone Activity

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    Miya Soukaina Hakam

    2017-10-01

    Full Text Available Background/Aims: Pregnancy success requires mandatory maternal tolerance of the semi/ allogeneic embryo involving embryo-derived signals. Expression levels of PreImplantation Factor (PIF, a novel peptide secreted by viable embryos, correlate with embryo development, and its early detection in circulation correlates with a favourable pregnancy outcome. PIF enhances endometrial receptivity to promote embryo implantation. Via the p53 pathway, it increases trophoblast invasion, improving cell survival / immune privilege. PIF also reduces spontaneous and LPS-induced foetal death in immune naïve murine model. We examined PIF effect on gene expression of human leukocyte antigen (HLA-G, -E -F and –C and the influence of PIF on local progesterone activity in JEG-3 choriocarcinoma cells. Methods: PIF and progesterone (P4 effects on JEG-3 cells surface and intracellular HLA molecules was tested using monoclonal antibodies, flow cytometry, and Western blotting. PIF and IL17 effects on P4 and cytokines secretion was determined by ELISA. PIF and P4 effects on JEG-3 cells proteome was examined using 2D gel staining followed by spot analysis, mass spectrometry and bioinformatic analysis. Results: In cytotrophoblastic JEG-3 cells PIF increased intracellular expression of HLA-G, HLA-F, HLA-E and HLA-C and surface expression of HLA-G, HLA-E and HLA-C in dose and time dependent manner. In case of HLA-E, -F results were confirmed also by Western blot. Proteome analysis confirmed an increase in HLA-G, pro-tolerance FOXP3+ regulatory T cells (Tregs, coagulation factors and complement regulator. In contrast, PIF reduced PRDX2 and HSP70s to negate oxidative stress and protein misfolding. PIF enhanced local progesterone activity, increasing steroid secretion and the receptor protein. It also promoted the secretion of the Th1/Th2 cytokines (IL-10, IL-1β, IL-8, GM-CSF and TGF-β1, resulting in improved maternal signalling. Conclusion: PIF can generate a pro

  3. Natural history of postterm choriocarcinoma.

    Science.gov (United States)

    Rodabaugh, K J; Bernstein, M R; Goldstein, D P; Berkowitz, R S

    1998-01-01

    To review the 32-year experience of the New England Trophoblastic Disease Center (NETDC) with choriocarcinoma occurring after a term gestation and to evaluate potential prognostic factors using the World Health Organization (WHO) prognostic score. The charts of 44 women who were treated for postterm choriocarcinoma at the NETDC from August 1964 to January 1996 were retrospectively reviewed. Demographic data and details of the clinical course were determined. Potential risk factors, including disease duration, pretreatment human chorionic gonadotropin (hCG) level, sites of metastases and stage, as well as data regarding the infants and previous and subsequent pregnancies, were evaluated. Five (11%) of the infants suffered significant complications secondary to maternal choriocarcinoma. The time interval from delivery to diagnosis, pretreatment hCG level and sites of metastatic disease were all significant risk factors in predicting outcome. All 31 patients with a WHO score 8 died. Disease duration greater than four months from delivery, pretreatment hCG level > 100,000 mIU/mL, presence of liver or brain metastases, and a WHO score > 8 were all important predictors of outcome in patients with postterm choriocarcinoma.

  4. Incidental placental choriocarcinoma in a term pregnancy: a case report

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    Chung Christopher

    2008-10-01

    Full Text Available Abstract Introduction Gestational choriocarcinoma occurs in 1 in 40,000 pregnancies. Of all forms of gestational choriocarcinoma, placental choriocarcinoma is the most rare. Maternal choriocarcinoma is usually diagnosed in symptomatic patients with metastases. The incidental finding of a choriocarcinoma confined to the placenta with no evidence of dissemination to the mother, or infant is the least common scenario. Case presentation The patient is an 18 year-old Gravida 1 Para 1 African American female who delivered a viable 3641 g female infant at 39 weeks gestation. Her pregnancy course was complicated by gestational hypertension during the third trimester. Her placenta revealed intraplacental choriocarcinoma. She was then followed closely by the Gynecologic Oncology service with a weekly serum beta human chorionic gonadotropin value. Beta human chorionic gonadotropin values dropped from 3070 mIU/ml to less than 2 mIU/ml two months post partum. No chemotherapy was initiated. Metastasis was ruled out by chest x-ray and whole body computed tomography scan. To date, both mother and baby are well. Conclusion Due to the potential fatal outcome of placental choriocarcinoma, careful evaluation of both mother and infant after the diagnosis is made is important. The incidence of placental choriocarcinoma may actually be higher than expected since it is not routine practice to send placentas for pathological evaluation after a normal spontaneous delivery. The obstetrician, pathologist, and pediatrician should have an increased awareness of placental choriocarcinoma and its manifestations.

  5. A Rare Case of Metastatic Choriocarcinoma of Lung Origin

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    Parth Rali

    2017-01-01

    Full Text Available Choriocarcinoma is part of the spectrum of gestational trophoblastic disease that occurs in women of reproductive age. Although the most common metastatic site of choriocarcinoma is the lung, primary pulmonary choriocarcinoma is rare. To diagnose primary pulmonary choriocarcinoma, the patient should have no previous gynecologic malignancy, have elevated human chorionic gonadotropin, and have pathological confirmation of the disease excluding gonadal primary site of the tumor. Due to the paucity of data, there are no guidelines for treatment. Prognosis of this malignancy is extremely poor. We report a rare case of metastatic primary lung choriocarcinoma in a 69-year-old postmenopausal woman who was treated with combination of surgery, chemotherapy, and radiation. The patient had a good outcome and is doing well after 1-year follow-up.

  6. PRIMARY PULMONARY CHORIOCARCINOMA – CASE REPORT

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    Biljana Lazović

    2010-12-01

    Full Text Available Literature has described only twenty-two cases of primary pulmonary choriocarcinoma. Unlike gestational choriocarcinoma which responds well to chemotherapy and has good prognosis, even in cases when detected later, pulmonary choriocarcinoma leads rapidly to fatal outcome. The reason is unknown. This paper presents a case of thirty-four-old young woman with clinical presentation of cough, expectoration, occasionally with hemoptisis, pleural pain, lack of breath and fever, with laboratory findings of pleural effusion on the left, which was treated as a case of tuberculosis for three months. Gynecological finding was regular. Progressive, rapid deterioration of general condition, degree of dispnea, hemoptisis episode led to respiratory failure before having appied the appropriate therapy. In cases of haemoptisis, one should think about this rare malignant entity and check the level of human chorionic gonadotropin in women.

  7. A role for the homeobox protein Distal-less 3 in the activation of the glycoprotein hormone alpha subunit gene in choriocarcinoma cells.

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    Roberson, M S; Meermann, S; Morasso, M I; Mulvaney-Musa, J M; Zhang, T

    2001-03-30

    Synthesis and secretion of chorionic gonadotropin in trophoblast cells of the placenta is required for establishment of early pregnancy in primates. Chorionic gonadotropin is a heterodimeric glycoprotein hormone consisting of alpha and beta subunits. Regulation of the alpha subunit gene within the placenta requires an array of cis elements within the 5'-flanking region of the promoter. Within this array of elements, the junctional regulatory element (JRE) putatively binds a placental-specific transcription factor. The aim of our studies was to determine the identity and role of the transcriptional regulator that binds to the JRE in choriocarcinoma cells (JEG3 cells). Mutations within the JRE resulted in reduction in basal expression of an alpha subunit reporter gene, suggesting that the JRE binding factor was necessary for full basal activity. Using electrophoretic mobility shift assays, we determined that the JRE was capable of serving as a homeobox factor-binding site. The homeobox factor, Distal-less 3 (Dlx 3) was found to be expressed in JEG3 cells and in the trophoblast layer of human chorionic villus but not in a gonadotrope cell line that also expresses the alpha subunit gene. Electrophoretic mobility shift assays revealed that recombinant Dlx 3 could bind specifically to the JRE and endogenous Dlx 3 was present in JRE/JEG3 nuclear protein complexes. Overexpression of Dlx 3 resulted in activation of an alpha subunit reporter gene. A JRE mutation resulted in attenuated activation of the alpha subunit reporter via an adjacent cis element, suggesting that JRE/Dlx 3 interactions may facilitate regulation of the alpha subunit gene at sites immediately upstream of the JRE. Our studies support the conclusion that Dlx 3 is a placental-specific transcriptional regulator that binds to the JRE and contributes to expression of the alpha subunit gene in cells of trophoblast origin.

  8. FBI-1 Is Overexpressed in Gestational Trophoblastic Disease and Promotes Tumor Growth and Cell Aggressiveness of Choriocarcinoma via PI3K/Akt Signaling.

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    Mak, Victor C Y; Wong, Oscar G W; Siu, Michelle K Y; Wong, Esther S Y; Ng, Wai-Yan; Wong, Richard W C; Chan, Ka-Kui; Ngan, Hextan Y S; Cheung, Annie N Y

    2015-07-01

    Human placental trophoblasts can be considered pseudomalignant, with tightly controlled proliferation, apoptosis, and invasiveness. Gestational trophoblastic disease (GTD) represents a family of heterogeneous trophoblastic lesions with aberrant apoptotic and proliferative activities and dysregulation of cell signaling pathways. We characterize the oncogenic effects of factor that binds to the inducer of short transcripts of HIV-1 [FBI-1, alias POZ and Krüppel erythroid myeloid ontogenic factor (POKEMON)/ZBTB7A] in GTD and its role in promoting cell aggressiveness in vitro and tumor growth in vivo. IHC studies showed increased nuclear expression of FBI-1, including hydatidiform moles, choriocarcinoma (CCA), and placental site trophoblastic tumor, in GTD. In JAR and JEG-3 CCA cells, ectopic FBI-1 expression opposed apoptosis through repression of proapoptotic genes (eg, BAK1, FAS, and CASP8). FBI-1 overexpression also promoted Akt activation, as indicated by Akt-pS473 phosphorylation. FBI-1 overexpression promoted mobility and invasiveness of JEG-3 and JAR, but not in the presence of the phosphoinositide 3-kinase inhibitor LY294002. These findings suggest that FBI-1 could promote cell migration and invasion via phosphoinositide 3-kinase/Akt signaling. In vivo, nude mice injected with CCA cells with stable FBI-1 knockdown demonstrated reduced tumor growth compared with that in control groups. These findings suggest that FBI-1 is clinically associated with the progression of, and may be a therapeutic target in, GTD, owing to its diverse oncogenic effects on dysregulated trophoblasts. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  9. Vulva choriocarcinoma | Boufettal | Pan African Medical Journal

    African Journals Online (AJOL)

    Choriocarcinoma vulva is an exceptional location of gestational trophoblastic tumors. We report the case of a female patient of 23 years who had vulvar mass, painful, sitting at the large right lip and measured five centimeters in diameter. Pelvic ultrasound was normal. Beta-h-CG (human chorionic gonadotrophin) ...

  10. Choriocarcinoma in a 73-year-old woman: a case report and review of the literature

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    Said Rabih

    2010-11-01

    Full Text Available Abstract Introduction Choriocarcinoma is a highly malignant tumor of trophoblastic origin. Most cases present within one year of the antecedent pregnancy (molar or non-molar. However, very rarely, choriocarcinoma can develop from germ cells or from dedifferentiation of endometrial carcinoma into choriocarcinoma. This article concerns a case of choriocarcinoma developing 38 years after the patient's last pregnancy and 23 years after menopause. Case presentation A 73-year-old African-American woman presented with a three-week history of vaginal bleeding. A vaginal mass was seen on pelvic examination. Ultrasonography showed a thickened complex endometrial echo. Her β-human chorionic gonadotrophin level was found to be elevated (2,704,040 mIU/mL. Vaginal and uterine biopsies were suggestive of choriocarcinoma. Immunohistochemistry tests were positive for β-human chorionic gonadotrophin as well as cytokeratin and negative for octamer binding transcription factor 3/4 and α-fetoprotein, supporting the diagnosis of choriocarcinoma. A combination of etoposide, methotrexate, and dactinomycin, followed by cyclophosphamide and vincristine (the so-called EMA/CO regimen was initiated. After seven cycles of chemotherapy, her β-human chorionic gonadotrophin level dropped below 5 mIU/mL. Our patient is being followed up at our oncology institute. Conclusions We report an extremely rare case of choriocarcinoma arising 23 years after menopause. A postmenopausal woman presenting with vaginal bleed from a mass and β-human chorionic gonadotrophin elevation should be evaluated by immunohistochemical analysis to rule out the possibilities of a germ cell origin of the tumor or dedifferentiation of an epithelial tumor. Absence of octamer binding transcription factor 3/4, α-fetoprotein and CD-30 staining helps in exclusion of most germ cell tumors. DNA polymorphism studies can be used to differentiate between gestational and non-gestational tumor origin. These

  11. Developing retroperitoneal anaplastic carcinoma with choriocarcinoma focus after ovarian non-gestastional choriocarcinoma: Case report

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    Nikolić Branka

    2012-01-01

    Full Text Available Introduction. Choriocarcinoma is a malignant form of gestational trophoblastic neoplasm (GTN. It is a rare event but also a curable malignancy. In the majority of instancies it developes after any gestational event. In some cases it developes as non-gestational extrauterine malignancy. Prognosis of choriocarcinoma is poor when invasion and metastases appear early and spread fast. This form of choriocarcinoma can lead to incurable and letal outcome. Case report. We presented a 20-year-old patient with abdominal and retroperitoneal malignancy - anaplastic carcinoma combined with choriocarcinoma metastases in. Tumor developed three months after left adnexectomy which had been done because of adnexal tumor. Choriocarcinoma was immunohistochemicaly confirmed in adnexal masses. Two courses of chemotherapy, metotrexate + folic acid (MTX+FA regimen, were administrated. The initial serum beta human chorionic gonadotropin level stayed unknown as well as the last one after the treatment. The patient came from the other country and was hospitalized because of pelvic and abdominal pain and palpable abdominal masses in hypogastrium with progressive anemia. The human chorionic gonadotropin level was 38 mIU/L. Tumor biopsy was done and choriocarcinoma metastases were immunohistochemicaly confirmed with predominant anaplastic carcinoma. Five day course of MTX + cyclophosphamide regimen was administrated and the patient was prepared for operative treatment. Relaparotomy was perforemed and tumor completely exceeded. Tumor mass mostly developed retroperitonely and partialy in abdominal cavity infiltrating intestinal wall with rupture of sigmoid colon. Anaplastic carcinoma, with large fields of necrosis and bleeding, was confirmed after histological examination. Immunohistochemical examination excluded choriocarcinoma in tumor mass. After 20 blood units transfusion, one course of chemotherapy and tumor excision, the patient left hospital on the 9th postoperative day

  12. Concurrent development of testicular seminoma and choriocarcinoma of the superior mediastinum, presented as cervical mass: a case report and implications about pathogenesis of germ-cell tumours

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    Bamias Aristotelis

    2006-11-01

    Full Text Available Abstract Background Synchronous presentation of more than one germ cell tumours of different histology in the same patient is considered to be very rare. In these cases of multiple germ cell tumours, strong theoretical and clinical data suggest an underlying common pathogenetic mechanism concerning genetic instability or abnormalities during the pluripotent embryonic differentiation and maturation of the germ cell. Case presentation A 25 year-old young man presented with an enlarging, slightly painful left cervical mass. Despite the initial disorientation of the diagnosis to a possible thyroid disorder, the patient underwent complete surgical resection of the mass revealing mediastinal choriocarcinoma. Subsequent ultrasound of the scrotum indicated the presence of a small lobular node in the upper pole of the left testicle and the patient underwent radical left inguinal orchiectomy disclosing a typical seminoma. Based on these results, the patient received 4 cycles of Bleomycin, Etoposide and Platinum chemotherapy experiencing only mild toxicity and resulting in complete ongoing clinical and biochemical remission. Conclusion The pathogenesis of concurrent germ cell tumours in the same patient remains an area of controversy. Although the genetic instability of the pluripotent germ cell offers an adequate explanation, the possibility of metastasis from the primary, less differentiated tumour to a distant location as a more mature subtype cannot be excluded. Possible development of a metastatic site of different histology and thus biological behaviour (e.g choriocarcinoma should be anticipated. Furthermore, urologists, pathologists and medical oncologists should be meticulous in the original pathological diagnosis in these patients, since there is a significant frequency of germ cell tumours with mixed or overlapping histological elements with diverse potential of evolution and differentiation.

  13. A pure non-gestational ovarian choriocarcinoma with delayed solitary brain metastases: Case report and review of the literature

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    KVL Narasinga Rao

    2015-01-01

    Full Text Available Choriocarcinoma is the most malignant tumour of gestational trophoblastic origin. Most ovarian choriocarcinomas are gestational in origin and usually metastasize to the ovary from uterine or tubal choriocarcinoma. Non gestational choriocarcinoma (NGOC of the ovary is exceedingly rare and usually seen along with other germ cell tumors. Non gestational choriocarcinoma has been found to be resistant to single-agent chemotherapy and has a worse prognosis than gestational choriocarcinoma. We are reporting long term follow up of published rare case of pure non gestational ovarian choriocarcinoma (NGOC with concurrent metastases to the spleen and adrenal glands, who developed a delayed solitary brain metastases, two years after completion of primary treatment. Surgery along with triple agent chemotherapy and radiotherapy was found to give good remission in this aggressive disease.

  14. Impact of CCN3 (NOV) glycosylation on migration/invasion properties and cell growth of the choriocarcinoma cell line Jeg3.

    Science.gov (United States)

    Yang, Wei; Wagener, Jessica; Wolf, Nadine; Schmidt, Markus; Kimmig, Rainer; Winterhager, Elke; Gellhaus, Alexandra

    2011-10-01

    Recently we have shown that the matricellular CCN3 protein expressed in invasive extravillous trophoblast cells (EVTs) is decreased in early-onset pre-eclampsia and is regulated by oxygen tension. Pathogenesis of pre-eclampsia relies on a shallow invasion of EVTs into the spiral arteries, which leads to hypoxia accompanied by uteroplacental insufficiency. Here we investigated the function of glycosylated and non-glycosylated CCN3 protein on cell growth as well as migration and invasion properties of the malignant trophoblast cell line Jeg3 which is a widely used model for the invasive trophoblast. Stable transfection of Jeg3 choriocarcinoma cells with full length CCN3 resulted in high expression of secreted glycosylated and cellular non-glycosylated CCN3. These cells revealed significantly reduced growth in cell numbers combined with a significantly increased migratory and invasive capacity. Matrix metalloprotease (MMP)-2 and MMP-9 activities were enhanced dependent on CCN3 expression, which could be confirmed by CCN3 knockdown studies. Using recombinant glycosylated and non-glycosylated CCN3, we revealed that CCN3 decreased growth in Jeg3 cell numbers independent of its glycosylation status, whereas only non-glycosylated CCN3 was able to enhance migration and invasion properties. The present results suggest that CCN3 protein regulates the decrease in Jeg3 cell numbers independent of its glycosylation status, whereas migratory and invasive properties are influenced only by non-glycosylated CCN3. An impaired balance in the expression of glycosylated and non-glycosylated CCN3 could contribute to the shallow invasion of EVTs observed in pre-eclampsia.

  15. Mediastinal Choriocarcinoma Masquerading as Relapsed Hodgkin Lymphoma

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    Selay Lam

    2011-10-01

    Full Text Available Primary mediastinal choriocarcinoma is a rare extragonadal germ cell malignancy. We describe the first case of a patient who developed mediastinal choriocarcinoma after treatment for Hodgkin lymphoma (HL. A 25-year-old man with classic HL, nodular sclerosis subtype, underwent treatment with splenectomy followed by radiation therapy. Unfortunately, his disease relapsed with a paraspinal mass, and he was subsequently treated with MOPP (mechlorethamine, Oncovin, procarbazine, and prednisone alternating with ABVD (Adriamycin, bleomycin, vinblastine, and dacarbazine. He achieved a complete remission after 6 cycles. Ten years after treatment, the patient presented with a persistent cough, haemoptysis, right supraclavicular lymphadenopathy, and weight loss. His chest X-ray showed opacification of the lower right hemithorax with a widened mediastinum. Given unresponsiveness to several antibiotics and lack of evidence for lung volume loss, there were concerns over lung infiltration with relapsed lymphoma. Transbronchial fine needle aspiration biopsy suggested recurrence of his HL. MOPP alternating with ABVD was again given. Due to disease progression, brachytherapy as well as a cocktail of dexamethasone, cytarabine, and cisplatin were also tried. However, on a subsequent excisional lymph node biopsy, it turned out that the tumour was in fact choriocarcinoma and not relapsed HL. Unfortunately, despite aggressive therapy, the patient’s disease rapidly progressed, and he died within 2 weeks.

  16. HLA-G regulates the invasive properties of JEG-3 choriocarcinoma cells by controlling STAT3 activation.

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    Liu, X; Gu, W; Li, X

    2013-11-01

    The expression of human leucocyte antigen-G (HLA-G) in trophoblasts plays a crucial role in successful embryonic implantation, and reduced HLA-G expression might contribute to adverse obstetric outcomes. In this study, we silenced HLA-G expression using RNA interference in JEG-3 cells, resulting in a notably attenuated invasion capacity of the cells in a Transwell assay; however, no alterations in cell proliferation or apoptosis were observed. The down-regulation of HLA-G dampened the activation of signal transducer and activator of transcription 3 (STAT3), whereas the up-regulation of HLA-G promoted STAT3 activation and invasion in JEG-3 cells treated with human galectin-1. Most importantly, interleukin-6 (IL-6), but not galectin-1, was shown to rescue invasion deficiency in a dose-dependent manner. Thus, we demonstrate that HLA-G is able to regulate JEG-3 cell invasion by influencing STAT3 activation, which may underlie the implantation defects accompanying HLA-G hypo-expression in pre-eclampsia. Copyright © 2013 Elsevier Ltd. All rights reserved.

  17. Reduced syncytin-1 expression in choriocarcinoma BeWo cells activates the calpain1-AIF-mediated apoptosis, implication for preeclampsia.

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    Huang, Qiang; Chen, Haibin; Wang, Fengchao; Brost, Brian C; Li, Jinping; Gao, Yu; Li, Zongfang; Gao, Ya; Jiang, Shi-Wen

    2014-08-01

    Placentas associated with preeclampsia are characterized by extensive apoptosis in trophoblast lineages. Syncytin-1 (HERVWE1) mediates the fusion of cytotrophoblasts to form syncytiotrophoblasts, which assume the placental barrier, fetal-maternal exchange and endocrine functions. While decreased syncytin-1 expression has been observed in preeclamptic placentas, it is not clear if this alteration is involved in trophoblast apoptosis. In the current study, we found that siRNA-mediated knockdown of syncytin-1 led to apoptosis in choriocarcinoma BeWo, a cell line of trophoblastic origin. Characterization of the apoptotic pathways indicated that this effect does not rely on the activation of caspases. Rather, decreased syncytin-1 levels activated the apoptosis inducing factor (AIF) apoptotic pathway by inducing the expression, cleavage, and nuclear translocation of AIF. Moreover, calpain1, the cysteine protease capable of cleaving AIF, was upregulated by syncytin-1 knockdown. Furthermore, treatment with calpain1 inhibitor MDL28170 effectively reversed AIF cleavage, AIF nuclear translocation, and cell apoptosis triggered by syncytin-1 downregulation, verifying the specific action of calpain1-AIF pathway in trophoblast apoptosis. We confirmed that preeclamptic placentas express lower levels of syncytin-1 than normal placentas, and observed an inverse correlation between syncytin-1 and AIF/calpain1 mRNA levels, a result consistent with the in vitro findings. Immunohistochemistry analyses indicated decreased syncytin-1 and increased AIF and calpain1 protein levels in apoptotic cells of preeclamptic placentas. These findings have for the first time revealed that decreased levels of syncytin-1 can trigger the AIF-mediated apoptosis pathway in BeWo cells. This novel mechanism may contribute to the structural and functional deficiencies of syncytium frequently observed in preeclamptic placentas.

  18. Metastatic Choriocarcinoma Following Live Birth – A Rare Presentation

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    Joya Sree Roy

    2011-09-01

    Full Text Available Choriocarcinoma is the most malignant tumor of gestational trophoblastic neoplasia. Post-partum choriocarcinoma is an infrequent event with poor prognosis. The diagnosis is usually delayed due to failure to recognize the mode of presentation of this disease. Being a rare occurrence, limited data is available regarding its clinical features. We present a 24 years old women with parity one delivered by caesarean section indicated for premature rupture of membrane with fetal distress at 39 wks of pregnancy. Frequent episodes of heavy vaginal bleeding had started 28 days following C/S. For this, she had H/O uterine evacuatin twice within 5 days interval. Biopsy report of second curettage showed choriocarcinoma. On admission to Dhaka Medical College Hospital (DMCH, the pretreatment Human Chorionic Gonadotrophin (βhCG level was >200000.0 IU/L, uterine mass of about 18 wks pregnancy size & X-Ray chest showed segmental consolidation in left mid zone of lung. After consultation with oncologist Etoposide, Methotrexate, Dactinomycin, Cyclophosphamide & Vincristine (EMACO therapy was started but before completion of her proposed cycles, she developed a live threatening condition which was managed very urgently and meticulously. Now the patient is under regular monitoring. Key words: GTD; Choriocarcinoma DOI: http://dx.doi.org/10.3329/bsmmuj.v4i2.8642 BSMMU J 2011; 4(2:116-118

  19. [Computer tomographic findings in metastasizing choriocarcinoma].

    Science.gov (United States)

    Schild, H; Treese, N; Reuss, M; Friese, K

    1983-06-01

    The angiographic picture of metastatic choriocarcinoma is well known since long, computertomographic findings, however, have not yet been described. The article gives a case-report and discusses the computertomographic picture of the disease.

  20. Gestational choriocarcinoma at Ibn Rochd Hospital, Casablanca, 2004-2010.

    Science.gov (United States)

    Boufettal, H; Khalkane, L; Noun, M; Hermas, S; Samouh, N

    2014-01-09

    The aim of this retrospective study was to determine the frequency, risk factors, management and prognosis of gestational choriocarcinoma at the gynaecology-obstetrics C department of the Ibn Rochd Hospital in Casablanca, Morocco over a 7-year period (2004-2010). There were 18 760 pregnancies in the study period, among which 8 cases of gestational choriocarcinoma were recorded, a frequency of 43/100 000 or 1 case of gestational choriocarcinoma in 2345 pregnancies. Maternal age ranged from 20 to 48 years. It occurred after a hydatidiform mole in 4 cases, after spontaneous miscarriage in 2 and after normal delivery in 2 cases. All the patients presented with bleeding. Metastases were found in 3 patients, including 2 with pulmonary localization. Positive diagnosis was made mainly by ultrasound and human chorionic gonadotropin level and histological confirmation was done for all cases. Polychemotherapy was used to treat 4 cases, monochemotherapy to treat 3 and one case received only radiotherapy. Of the 8 cases, 2 patients died and 6 had complete remission.

  1. Colorectal choriocarcinoma in a patient with probable Lynch syndrome

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    Viktor Hendrik Koelzer

    2016-11-01

    Full Text Available Background: Personalized therapy of colorectal cancer (CRC is influenced by morphological, molecular and host-related factors. Here we report the comprehensive clinicopathological and molecular analysis of a pure extra-gestational colorectal choriocarcinoma in a patient with probable Lynch syndrome.Case presentation: A 61 year old female with history of gastric cancer at age 36 presented with a transmurally invasive tumor of the right hemicolon and liver metastasis. A right hemicolectomy was performed. Histopathological analysis showed a mixed trophoblastic and syncytiotrophoblastic differentiation, consistent with choriocarcinoma. Disease progression was rapid under oxaliplatin, capecitabine, irinotecan (XELOXIRI and bevacizumab. Molecular phenotyping identified loss of the mismatch-repair (MMR protein PMS2, microsatellite instability, a lack of MLH1 promoter methylation and lack of of BRAF mutation suggestive of Lynch-Syndrome. Targeted next generation sequencing revealed an Ataxia Telangiectasia Mutated (ATM p.P604S missense mutation. A bleomycin, etoposide and cisplatin (BEP treatment protocol targeting germ-cell neoplasia lead to disease remission and prolonged survival of 34 months.Conclusions: Comprehensive immunohistochemical and genetic testing is essential to identify uncommon cancers possibly related to Lynch syndrome. For rare tumors, personalized therapeutic approaches should take both molecular and morphological information into account.Key words: Colorectal cancer, choriocarcinoma, histopathology, prognostic factors, Lynch syndrome, microsatellite instability, ataxia telangiectasia mutated, molecular pathology, next generation sequencing, personalized medicine

  2. Choriocarcinoma after caesarian section: case report

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    Zohreh Yousefi

    2017-02-01

    Full Text Available Background: The most common symptom of patients with gestational trophoblastic neoplasia is abnormal vaginal bleeding. Despite repeated visits of patients with postpartum choriocarcinoma and abnormal postpartum hemorrhage, delayed diagnosis leads to advanced disease with widespread metastasis. Therefore, occurrence of choriocarcinoma with variable patterns in different diagnosis of late onset postpartum hemorrhage should be considered. Early diagnosis of choriocarcinoma after term pregnancy is important that resulted in decrease of maternal morbidity. Therefor late onset postpartum hemorrhage should have an awareness. The aim of this study was to report a case of choriocarcinoma after caesarian section. Case Presentation: A 33-years-old woman one month after antecedent caesarian section in her second pregnancy admitted with abnormal vaginal bleeding. Based on raised titer concentration of β-hCG was 187000 u, with clinical suspicious of choriocarcinoma she was referred to oncology department of Ghaem Hospital, Mashhad University of Medical Sciences, Iran, in 2016. Vaginal exam revealed an enlarged uterus about 10 weeks of pregnancy. Transvaginal sonography showed an intracavitary heterogeneous mass with irregular surface in fundus without myometrium invasion. Extra pelvic metastasis excluded via vaginal exam, pulmonary X-ray and, ultrasonography and computed tomography scan. Due to early stage of gestational trophoblastic neoplasia and diagnosis of this condition, single agent chemotherapy (methotrexate was recommended, but because of unresponsive disease, subsequently, she was treated with combination chemotherapy (etoposide, methotrexate, and actinomycin, followed by cyclophosphamide and vincristine that led to remarkable response. After three courses of therapy, normal level of β-hCG was observed and now the patient is free of disease and under-serial follow-up visit for choriocarcinoma. Conclusion: Diagnosis of choriocarcinoma should be

  3. Metastatic choriocarcinoma in the small bowel: a case report

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    Zohreh Yousefi

    2014-08-01

    Conclusion: In abnormal postpartum hemorrhage, we should consider the possibility of choriocarcinoma. Although, it is important to note rare manifestations of metastatic choriocarcinoma of small bowel in massive gastrointestinal hemorrhage.

  4. Coexistence of Gastric Adenocarcinoma and Choriocarcinoma: Complete Response to Trastuzumab and Chemotherapy

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    Seyda Gunduz

    2012-07-01

    Full Text Available Gastric choriocarcinoma is a rare neoplasm and usually accompanies gastric adenocarcinoma. The prognosis is poor due to the aggressive course of the disease. A 57-year-old female patient with weight loss and abdominal pain was examined. The patient was operated following the examination, and pathological analysis revealed the presence of a gastric adenocarcinoma associated with choriocarcinoma. Immunohistochemical analysis showed a positive reaction with antibodies to beta-human chorionic gonadotropin and overexpression of the cErbB2 proto-oncogene. Staging revealed multiple metastases in the liver. A complete response was obtained with a combination of trastuzumab and chemotherapy. The diagnosis of gastric choriocarcinomas without pathological examination is difficult due to their rare occurrence. A complete response can be obtained with trastuzumab in the treatment of cases with overexpression of the cErbB2 protein.

  5. Primary ovary choriocarcinoma: individual DNA polymorphic analysis as a strategy to confirm diagnosis and treatment

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    Fernando Nalesso

    2013-04-01

    Full Text Available Primary choriocarcinoma of the ovary is rare. Furthermore, this tumor can arise from gestational tissue or pure germ cells of the ovary, with the latter resulting in non-gestational choriocarcinoma. While the clinical characteristics and histology of both tumor types are identical, differentiation of these tumors is necessary for effective treatment. One strategy for the differentiation of these tumors types is to assay for the presence of paternal DNA. Accordingly, in the present case, a patient with primary choriocarcinoma of the ovary with a non-gestational origin was confirmed by DNA analysis. The patient subsequently exhibited an excellent response to chemotherapy, and following surgery, achieved complete remission. A pathological analysis of surgical specimens further confirmed the absence of tumor.

  6. The choriocarcinoma cell line JEG-3 upregulates regulatory T cell phenotypes and modulates pro-inflammatory cytokines through HLA-G

    DEFF Research Database (Denmark)

    Melsted, Wenna Nascimento; Matzen, Sara Hyldig; Andersen, Mads Hald

    2017-01-01

    -inflammatory cytokines IFN-γ, TNF-α and IL-17A. When JEG-3 cells were stimulated with rhIFN-γ prior to co-culture, CD4+HLA-G+ T cells were significantly increased, and IFN-γ and TNF-α elevated. Taken together, the results indicate that JEG-3 cells upregulate regulatory T cell phenotypes and modulate the level of pro......-inflammatory cytokines, which might be important mechanisms in the tumor microenvironment and at the feto-maternal interface during pregnancy....

  7. Ectopic pregnancy with associated gestational choriocarcinoma in a California sea lion (Zalophus californianus).

    Science.gov (United States)

    Fravel, Vanessa A; Lowenstine, Linda J; Koehne, Amanda

    2016-07-07

    A wild-born, captive-reared, 14 yr old, primiparous female California sea lion Zalophus californianus presented for anorexia of 14 d duration and abdominal distention. Routine complete blood cell count revealed leukocytosis with a neutrophilia, and serum chemistry revealed hypoalbumenemia and hyponatremia. Treatment with broad spectrum antibiotics and non-steroidal anti-inflammatories were started, but the animal continued to decline. Abdominal radiographs revealed a mature mineralized fetal skull and spine in the caudal abdomen and abdominal ultrasound revealed ascites but could not confirm the fetus. The patient was taken to surgery where a full term fetus was found outside of the uterus but within the fetal membranes, representing a secondary ectopic pregnancy. The patient passed away during surgery and was taken to necropsy. Gross necropsy revealed a diffuse peritonitis with yellow deposits over the serosal surfaces of the abdominal organs. The uterus appeared intact grossly and the ovaries appeared abnormal. The mesenteric, renal, and sub-lumbar nodes were enlarged and edematous. Histopathology revealed choriocarcinoma in the right uterine horn with evidence of chronic uterine rupture and protrusion of the placental tissue into the abdomen. The choriocarcinoma had metastasized locally as well as to the liver, spleen and lung. Choriocarcinoma is a highly malignant trophoblastic neoplasm that is rare in domestic animals. This case represents, to the authors' knowledge, the first report of gestational choriocarcinoma causing secondary ectopic pregnancy in a California sea lion and presents questions regarding pregnancy monitoring and management in a population of captive, minimally trained California sea lions.

  8. Active Surveillance, Bleomycin, Carboplatin, Etoposide, or Cisplatin in Treating Pediatric and Adult Patients With Germ Cell Tumors

    Science.gov (United States)

    2017-06-02

    Adult Germ Cell Tumor; Childhood Extracranial Germ Cell Tumor; Childhood Germ Cell Tumor; Extragonadal Embryonal Carcinoma; Grade 2 Immature Ovarian Teratoma; Grade 3 Immature Ovarian Teratoma; Malignant Germ Cell Tumor; Stage I Ovarian Choriocarcinoma; Stage I Ovarian Embryonal Carcinoma; Stage I Ovarian Teratoma; Stage I Ovarian Yolk Sac Tumor; Stage I Testicular Choriocarcinoma; Stage I Testicular Embryonal Carcinoma; Stage I Testicular Yolk Sac Tumor; Stage II Ovarian Choriocarcinoma; Stage II Ovarian Embryonal Carcinoma; Stage II Ovarian Yolk Sac Tumor; Stage II Testicular Choriocarcinoma; Stage II Testicular Embryonal Carcinoma; Stage II Testicular Yolk Sac Tumor; Stage III Ovarian Choriocarcinoma; Stage III Ovarian Embryonal Carcinoma; Stage III Ovarian Yolk Sac Tumor; Stage III Testicular Choriocarcinoma; Stage III Testicular Embryonal Carcinoma; Stage III Testicular Yolk Sac Tumor; Stage IV Ovarian Choriocarcinoma; Stage IV Ovarian Embryonal Carcinoma; Stage IV Ovarian Yolk Sac Tumor; Testicular Mixed Choriocarcinoma and Embryonal Carcinoma; Testicular Mixed Choriocarcinoma and Teratoma; Testicular Mixed Choriocarcinoma and Yolk Sac Tumor

  9. Metastatic choriocarcinoma as initial presentation of small bowel perforation in absence of primary uterine lesion: a case report.

    Science.gov (United States)

    Cho, En Bee; Byun, Jung Mi; Jeong, Dae Hoon; Yoon, Hye Kyoung; Kim, Young Nam; Sung, Moon Su; Lee, Kyung Bok; Kim, Ki Tae

    2016-11-11

    Choriocarcinoma is a highly malignant tumor of gestational trophoblastic neoplasia that characteristically spreads via the bloodstream. Small bowel metastasis is very rare, and a small number of cases of choriocarcinoma metastasis to small bowel have been reported. We report a case of a 40-year-old woman presenting with acute abdominal pain and vaginal bleeding due to small bowel perforation secondary to jejunal metastasis. In our case, metastatic choriocarcinoma was present in the small bowel, lung, and liver, but no primary lesion was apparent. After resection and anastomosis of the perforated small bowel, chemotherapy was performed. The patient began chemotherapy with etoposide, methotrexate, actinomycin D, cyclophosphamide, and vincristine (EMA-CO) for 9 cycles. When the 4th cycle of chemotherapy was finished, β-human chorionic gonadotropin (hCG) level was normalized and follow-up computed tomography scans showed regression of the liver metastasis and small bowel lesion and significantly decreased lung metastasis. After 9 cycles of chemotherapy were completed, the patient showed complete response. If patients have high β-hCG of unknown origin and suspected panperitonitis, which are suspicious of choriocarcinoma metastasis to small bowel, the pathologic findings are important. It is essential to confirm the histopathologic diagnosis postoperatively. Choriocarcinoma can spread to various organs and show aggressive manifestations. Therefore, we should be aware of possible metastatic sites and remain alert to its diagnosis.

  10. Treatment and prognosis of post term choriocarcinoma in The Netherlands

    NARCIS (Netherlands)

    Lok, Christianne A.; Ansink, Anca C.; Grootfaam, Danielle; van der Velden, Jacobus; Verheijen, Rene H.; ten Kate-Booij, Marianne J.

    2006-01-01

    INTRODUCTION: Antecedent term pregnancy is an adverse prognostic factor in Gestational Trophoblastic Disease (GTD). In The Netherlands, patients with post term choriocarcinoma are considered high-risk independent of WHO score. In the present study, we assessed whether post term choriocarcinoma

  11. St. Jude Children's Research Hospital, Memphis, Tennessee: gestational choriocarcinoma.

    Science.gov (United States)

    Weiss, Aaron; Khoury, Joseph D; Kaste, Sue C; Spunt, Sheri L

    2006-10-15

    This paper describes a recent tumor board presentation conducted at our institution involving an adolescent with gestational choriocarcinoma. Despite its rarity in pediatrics, gestational choriocarcinoma offers unique diagnostic, treatment and off-therapy considerations in adolescent patients who have become pregnant. Key features and findings of the case as well as important management issues will be discussed. Copyright (c) 2005 Wiley-Liss, Inc.

  12. A Rare Case of Gestational Choriocarcinoma Presenting as Cornual Ectopic Pregnancy.

    Science.gov (United States)

    Han, Vanessa; Kaye, Stephen

    2017-10-11

    Choriocarcinoma associated with cornual pregnancy is extremely rare. To our knowledge, only three other cases have been reported in the literature. A 38-year-old woman was found to have a left cornual ectopic pregnancy on ultrasound after presenting with abdominal pain, irregular vaginal bleeding, and a positive pregnancy test. Laparoscopy confirmed the diagnosis and she underwent total abdominal hysterectomy. Three weeks later, she presented with vaginal bleeding. A solid ulcerating lesion was found arising from the vaginal wall and biopsy revealed metastatic gestational choriocarcinoma. Careful histopathological examination of the surgical specimen and diligent monitoring of β-human chorionic gonadotropin to zero is crucial to prevent potentially missing this very malignant, but highly curable disease. Early systemic metastases are common and presentation can include bleeding from vaginal metastases. Copyright © 2017 Society of Obstetricians and Gynaecologists of Canada. Published by Elsevier Inc. All rights reserved.

  13. Effect of ATP depletion and temperature on the transferrin-mediated uptake and release of iron by BeWo choriocarcinoma cells

    NARCIS (Netherlands)

    van der Ende, A.; du Maine, A.; Schwartz, A. L.; Strous, G. J.

    1989-01-01

    We have recently described the transferrin-mediated uptake and release of iron by BeWo cells [van der Ende, du Maine, Simmons, Schwartz & Strous (1987) J. Biol. Chem. 262, 8910-8916]. We now extend our studies of the mechanisms responsible for uptake and release of iron by these cells. Following

  14. Brain metastases of choriocarcinoma: A report on two cases

    Directory of Open Access Journals (Sweden)

    Milenković Vera

    2013-01-01

    Full Text Available Introduction. Gestational trophoblastic diseases (GTD are a spectrum of tumors with a various of biological behavior and potential for metastases. It consists of hydatiform mole, invasive mole, choriocarcinoma and placental site trophoblastic tumor. Choriocarcinoma presents a very aggressive tumor with high malignant potential. Case report. We presented the two cases of choriocarcinoma with brain metastases. The first one was manifested by neurological deterioration as the first sign of metastasis, while the second patient had firstly metrorrhagia and in the further couse neurological disturbances that suggested the presence of brain tumor. In both cases we applied a combined treatment of surgery, chemotherapy and radiation therapy. Both patient survived with high quality of life. Conclusion. A successful outcome of brain metastases of choriocarcinoma was obtained by the use of a combined treatment of surgery, chemotherapy and radiation therapy. In cases of young women with brain metastases, gynecological malignancy should be always considered.

  15. Cerebral metastasis prom choriocarcinoma and oncotic aneurysms: case report

    Directory of Open Access Journals (Sweden)

    Pasquale Gallo

    1993-06-01

    Full Text Available Cerebral metastasis occur in 10 to 20% of patients with choriocarcinoma. We describe the twelfth patient with oncotic aneurysms from choriocarcinoma verified by cerebral angiography. The importance to consider this disease in a woman of childbearing age who develop an intracerebral hemorrhage or a lesion with mass effect is emphasized, as well as laboratorial and radiological characteristics. Therapeutic approaches with chemotherapic agents, surgery and irradiation are discussed.

  16. MyD88 and TRIF mediate the cyclic adenosine monophosphate (cAMP induced corticotropin releasing hormone (CRH expression in JEG3 choriocarcinoma cell line

    Directory of Open Access Journals (Sweden)

    Kocak Hande

    2009-07-01

    Full Text Available Abstract Background Classically protein kinase A (PKA and transcription factor activator protein 1 (AP-1 mediate the cyclic AMP (cAMP induced-corticotrophin releasing hormone (CRH expression in the placenta. However enteric Gram (- bacterial cell wall component lipopolysaccharide (LPS may also induce-CRH expression via Toll like receptor (TLR4 and its adaptor molecule Myd88. Here we investigated the role of MyD88, TRIF and IRAK2 on cAMP-induced CRH promoter activation in JEG3 cells in the absence of LPS/TLR4 stimulation. Methods JEG3 cells were transfected with CRH-luciferase and Beta-galactosidase expression vectors and either empty or dominant-negative (DN-MyD88, DN-TRIF or DN-IRAK2 vectors using Fugene6 (Roche. cAMP-induced CRH promoter activation was examined by using a luminometer and luciferase assay. Calorimetric Beta-galactosidase assays were performed to correct for transfection efficiency. Luciferase expression vectors of cAMP-downstream molecules, CRE and AP-1, were used to further examine the signaling cascades. Results cAMP stimulation induced AP-1 and CRE promoter expression and led to dose-dependent CRH promoter activation in JEG3 cells. Inhibition of MyD88 signaling blocked cAMP-induced CRE and CRH promoter activation. Inhibition of TRIF signaling blocked cAMP-induced CRH but not CRE expression, while inhibition of IRAK2 did not have an effect on cAMP-induced CRH expression. Conclusion MyD88 and TRIF exert direct regulatory effect on cAMP-induced CRH promoter activation in JEG3 cells in the absence of infection. MyD88 most likely interacts with molecules upstream of IRAK2 to regulate cAMP-induced CRH expression.

  17. Acquired Pulmonary Arteriovenous Fistula within Metastasis from Choriocarcinoma: A Case Report

    Directory of Open Access Journals (Sweden)

    Tassia Yamanari

    2017-07-01

    Full Text Available Choriocarcinoma is a highly malignant neoplasm arising from the trophoblast of a human pregnancy, which may have distant spread to multiple organs, particularly to the lung. We report a case of a pulmonary arteriovenous fistula that developed after chemotherapy within a choriocarcinoma metastasis. A 24-year-old female with a history of uterine choriocarcinoma presented with multiple hypervascular pulmonary nodules on the initial chest computed tomography (CT scan, consistent with lung metastases. Four months after chemotherapy, follow-up chest CT revealed a decrease in the size and vascularity of the lung metastases, except for 1 nodule that persisted with marked contrast enhancement and developed a communication with a pulmonary artery and vein. These findings were consistent with an acquired arteriovenous fistula. Physicians should be aware of the risk of acquired arteriovenous fistula in pulmonary metastases after chemotherapy because it can lead to a right-to-left shunt, pulmonary hemorrhage, and paradoxical emboli. It is also important for the definition of therapeutic planning, in order to avoid it being misdiagnosed as a site of disease progression.

  18. Iron metabolism in BeWo chorion carcinoma cells. Transferrin-mediated uptake and release of iron

    NARCIS (Netherlands)

    van der Ende, A.; du Maine, A.; Simmons, C. F.; Schwartz, A. L.; Strous, G. J.

    1987-01-01

    Growing human choriocarcinoma BeWo b24 cells contain 1.5 X 10(6) functional cell surface transferrin binding sites and 2.0 X 10(6) intracellular binding sites. These cells rapidly accumulate iron at a rate of 360,000 iron atoms/min/cell. During iron uptake the transferrin and its receptor recycle at

  19. Coexistence of a choriocarcinoma and a gonadoblastoma in the gonad of a 46,XY female: a single nucleotide polymorphism array analysis.

    Science.gov (United States)

    Beaulieu Bergeron, Mélanie; Soglio, Dorothée Bouron-Dal; Maietta, Antonio; Fournet, Jean-Christophe; Blumenkrantz, Miriam; Brochu, Pierre; Lemieux, Nicole

    2010-01-01

    Females with 46,XY complete gonadal dysgenesis are at significant risk of developing germ cell tumors, mostly gonadoblastomas. We present here the case of 2 half-sisters, sharing the same father, diagnosed with 46,XY complete gonadal dysgenesis. The 1st sister developed a gonadoblastoma and an invasive dysgerminoma, whereas the 2nd sister developed a gonadoblastoma and an invasive choriocarcinoma within the same gonad. No SRY mutation, chromosome abnormalities, or mosaicism were detected in blood. Single nucleotide polymorphism (SNP) profiling of the choriocarcinoma revealed a complex hyperdiploid pattern with gains of 1 to 4 copies of material from several autosomes, as well as the loss of the Y chromosome and a homozygous SNP profile without copy number change for the X chromosome. Our results are in agreement with the recurrent chromosome gains and losses previously published in germ cell tumors, and the coexistence of both tumors within the same gonad suggests that choriocarcinomas may derive from gonadoblastomas.

  20. Clinical Outcome of a FIGO Stage IV Gestational Choriocarcinoma

    Directory of Open Access Journals (Sweden)

    Vera Milenković

    2013-09-01

    Full Text Available Choriocarcinoma is the most malignant tumor of gestational trophoblastic disease arising from any gestation. It has a tendency toward relapse as well as metastasis. Here, a case of relapsed high-risk choriocarcinoma (FIGO stage IV, WHO score 12 in a 37-year-old female presenting with vaginal bleedings is described. Relapse developed at the site of the surgical scar from hysterectomy that had been performed 2 years earlier. Although the patient was treated with aggressive chemotherapy, she was in a bad general condition and died from infection and liver insufficiency.

  1. Coriocarcinoma primário do colo uterino Primary choriocarcinoma of the uterine cervix

    Directory of Open Access Journals (Sweden)

    Maria Fernanda Moreira Ferraz

    2003-06-01

    mechanisms for the development of the cervical choriocarcinoma: 1 the patient initially had a cervical pregnancy and this was followed by malignant transformation; 2 the cervical choriocarcinoma may be a metastasis from the primary tumor of the corpus, which had disappeared; 3 the development of cervical choriocarcinoma was due to the transport of chorionic cells from the preceding pregnancy as emboli, which stayed dormant and, later, gave rise to malignant transformation. Total abdominal histerectomy and posterior chemotherapy are recommended. We report a case of a 34-year-old woman, with vaginal bleeding six months after a term pregnancy, which had a hemorrhagic cervical mass and high b-HCG blood levels. The histopathology showed a tumor with a malignant sincicio and citotrophoblastic components involving only the uterine cervix. Our case is one of an ectopic choriocarcinoma and we believe that its development is due to the transport of chorionic cells from the preceding pregnancy and later gave rise to malignant transformation.

  2. Role of chemotherapy and thrombolysis in treatment of choriocarcinoma accompanied with pulmonary embolism: A case report with literature review.

    Science.gov (United States)

    Yang, Mei; Peng, Li

    2017-09-01

    Choriocarcinoma accompanied with pulmonary embolism (PE) is rare and difficult to diagnose and treat. There are about 25 cases reported in literature, which presented variable clinical characteristics and prognosis. We reported a case of choriocarcinoma presenting PE as the first manifestation in a 26-year-old Chinese female. Four months before her admission to our hospital, she presented to a local hospital with respiratory manifestations, and was first diagnosed as bronchitis and treated with antibiotics without improvement, and subsequently suspected of having pulmonary tuberculosis and receive anti-tuberculosis therapy, with no response. Upon admission to our hospital, spiral computed tomography-pulmonary angiography revealed complete occlusion of right pulmonary artery, leading to PE. She received thrombolysis and anticoagulant therapy, without improvement. Further interrogation of the patient revealed a history of irregular vaginal bleeding and multiple pregnancies and abortions. Following the laboratory report of a significantly elevated level of serum beta-human chorionic gonadotropin (β-HCG) combined with other clinical and laboratory findings, a diagnosis of choriocarcinoma accompanied with PE was established based on the criteria formulated by the International Federation of Gynecology and Obstetrics and guideline of European Society for Medical Oncology. With 1 cycle of chemotherapy with etoposide-methotrexateactinomycin D-cyclophosphamide-vincristine (EMA-CO), her dyspnea and other symptoms improved, with a significant decrease in the serum β-HCG level and pulmonary artery pressure. Unfortunately, she showed bone marrow inhibition, could not continue further chemotherapy, and finally died after discharging. We reviewed 25 similar cases in the literature, and found that all 17 cases receiving chemotherapy showed complete recovery while 6 of 8 cases without chemotherapy died during hospitalization. This case report illustrates the challenges in

  3. Standard-Dose Combination Chemotherapy or High-Dose Combination Chemotherapy and Stem Cell Transplant in Treating Patients With Relapsed or Refractory Germ Cell Tumors

    Science.gov (United States)

    2018-02-05

    Germ Cell Tumor; Teratoma; Choriocarcinoma; Germinoma; Mixed Germ Cell Tumor; Yolk Sac Tumor; Childhood Teratoma; Malignant Germ Cell Neoplasm; Extragonadal Seminoma; Non-seminomatous Germ Cell Tumor; Seminoma

  4. Pathogenesis of Testicular Germ Cell Tumors from a Developmental Point of View

    NARCIS (Netherlands)

    K. Biermann (Katharina)

    2010-01-01

    textabstractCurrent classification systems of human germ cell tumors (GCTs) are based on histological composition. In the group of nonseminomas, different variants of teratoma (somatic differentiation), yolk sac tumor and choriocarcinoma (extra-embryonic differentiation), are recognized, as well as

  5. Live cell imaging of in vitro human trophoblast syncytialization.

    Science.gov (United States)

    Wang, Rui; Dang, Yan-Li; Zheng, Ru; Li, Yue; Li, Weiwei; Lu, Xiaoyin; Wang, Li-Juan; Zhu, Cheng; Lin, Hai-Yan; Wang, Hongmei

    2014-06-01

    Human trophoblast syncytialization, a process of cell-cell fusion, is one of the most important yet least understood events during placental development. Investigating the fusion process in a placenta in vivo is very challenging given the complexity of this process. Application of primary cultured cytotrophoblast cells isolated from term placentas and BeWo cells derived from human choriocarcinoma formulates a biphasic strategy to achieve the mechanism of trophoblast cell fusion, as the former can spontaneously fuse to form the multinucleated syncytium and the latter is capable of fusing under the treatment of forskolin (FSK). Live-cell imaging is a powerful tool that is widely used to investigate many physiological or pathological processes in various animal models or humans; however, to our knowledge, the mechanism of trophoblast cell fusion has not been reported using a live- cell imaging manner. In this study, a live-cell imaging system was used to delineate the fusion process of primary term cytotrophoblast cells and BeWo cells. By using live staining with Hoechst 33342 or cytoplasmic dyes or by stably transfecting enhanced green fluorescent protein (EGFP) and DsRed2-Nuc reporter plasmids, we observed finger-like protrusions on the cell membranes of fusion partners before fusion and the exchange of cytoplasmic contents during fusion. In summary, this study provides the first video recording of the process of trophoblast syncytialization. Furthermore, the various live-cell imaging systems used in this study will help to yield molecular insights into the syncytialization process during placental development. © 2014 by the Society for the Study of Reproduction, Inc.

  6. Expression pattern alterations of the serine protease HtrA1 in normal human placental tissues and in gestational trophoblastic diseases.

    Science.gov (United States)

    Marzioni, Daniela; Quaranta, Alexia; Lorenzi, Teresa; Morroni, Manrico; Crescimanno, Caterina; De Nictolis, Michele; Toti, Paolo; Muzzonigro, Giovanni; Baldi, Alfonso; De Luca, Antonio; Castellucci, Mario

    2009-10-01

    HtrA1 is a secreted protein which behaves as a molecular chaperone at low temperatures and as a serine protease at high temperatures. When the placenta escapes the normal growth control mechanisms, which are present during normal pregnancy, it may develop trophoblastic diseases, such as hydatidiform mole and choriocarcinoma. The aim of the study is to investigate the expression of HtrA1 in these gestational trophoblastic diseases and evaluate whether different HtrA1 expression might be associated with increasingly severe forms of disease. We used immunohistochemistry to assess the expression of HtrA1 in normal human placenta, hydatidiform mole (partial and complete) and choriocarcinoma. In addition to that we used the western blotting technique to quantify HtrA1 immunoreaction in normal human placentas. The most striking finding of our investigation is the decrease in immunostaining of this protease with increasing severity of gestational trophoblastic disease. For instance, in partial and complete moles HtrA1 is weakly expressed in the trophoblast. Moreover, absence of immunoreaction for HtrA1 is observable in the choriocarcinoma cells. In conclusion, we suggest that HtrA1 may play an important role in the pathogenesis and progression of hydatidiform moles and choriocarcinomas, and that HtrA1 may play an important role during the normal development of the placenta, as well as in trophoblastic diseases.

  7. The human cell atlas

    DEFF Research Database (Denmark)

    Regev, Aviv; Teichmann, Sarah A.; Lander, Eric S.

    2017-01-01

    The recent advent of methods for high-throughput single-cell molecular profiling has catalyzed a growing sense in the scientific community that the time is ripe to complete the 150-year-old effort to identify all cell types in the human body. The Human Cell Atlas Project is an international...... collaborative effort that aims to define all human cell types in terms of distinctive molecular profiles (such as gene expression profiles) and to connect this information with classical cellular descriptions (such as location and morphology). An open comprehensive reference map of the molecular state of cells...... in healthy human tissues would propel the systematic study of physiological states, developmental trajectories, regulatory circuitry and interactions of cells, and also provide a framework for understanding cellular dysregulation in human disease. Here we describe the idea, its potential utility, early...

  8. Human memory B cells.

    Science.gov (United States)

    Seifert, M; Küppers, R

    2016-12-01

    A key feature of the adaptive immune system is the generation of memory B and T cells and long-lived plasma cells, providing protective immunity against recurring infectious agents. Memory B cells are generated in germinal center (GC) reactions in the course of T cell-dependent immune responses and are distinguished from naive B cells by an increased lifespan, faster and stronger response to stimulation and expression of somatically mutated and affinity matured immunoglobulin (Ig) genes. Approximately 40% of human B cells in adults are memory B cells, and several subsets were identified. Besides IgG(+) and IgA(+) memory B cells, ∼50% of peripheral blood memory B cells express IgM with or without IgD. Further smaller subpopulations have additionally been described. These various subsets share typical memory B cell features, but likely also fulfill distinct functions. IgM memory B cells appear to have the propensity for refined adaptation upon restimulation in additional GC reactions, whereas reactivated IgG B cells rather differentiate directly into plasma cells. The human memory B-cell pool is characterized by (sometimes amazingly large) clonal expansions, often showing extensive intraclonal IgV gene diversity. Moreover, memory B-cell clones are frequently composed of members of various subsets, showing that from a single GC B-cell clone a variety of memory B cells with distinct functions is generated. Thus, the human memory B-cell compartment is highly diverse and flexible. Several B-cell malignancies display features suggesting a derivation from memory B cells. This includes a subset of chronic lymphocytic leukemia, hairy cell leukemia and marginal zone lymphomas. The exposure of memory B cells to oncogenic events during their generation in the GC, the longevity of these B cells and the ease to activate them may be key determinants for their malignant transformation.

  9. Massive fetomaternal hemorrhage caused by an intraplacental choriocarcinoma: a case report

    DEFF Research Database (Denmark)

    Henningsen, Anna-Karina Aaris; Maroun, Lisa Leth; Havsteen, Hanne

    2010-01-01

    Background. Intraplacental choriocarcinoma is a rare but highly malignant trophoblastic neoplasm. When found near term the risk of maternal metastasis is high because of the late diagnosis. Case. We describe a case of an intraplacental choriocarcinoma diagnosed postpartum after a near-term delivery...... of a severely anemic infant. A fetomaternal hemorrhage resulted in a hemoglobin concentration in the infant of only 2,1 g/dL. Neither mother nor child showed signs of metastatic disease. The macroscopic examination showed a hydropic placenta weighing more than 1 kilogram. Microscopy showed an intraplacental...

  10. DNA Analysis in Samples From Younger Patients With Germ Cell Tumors and Their Parents or Siblings

    Science.gov (United States)

    2017-10-05

    Childhood Malignant Ovarian Germ Cell Tumor; Childhood Malignant Testicular Germ Cell Tumor; Ovarian Choriocarcinoma; Ovarian Embryonal Carcinoma; Ovarian Mixed Germ Cell Tumor; Ovarian Teratoma; Ovarian Yolk Sac Tumor; Testicular Choriocarcinoma; Testicular Embryonal Carcinoma; Testicular Seminoma; Testicular Teratoma; Testicular Yolk Sac Tumor

  11. Human mesenchymal stem cells

    DEFF Research Database (Denmark)

    Abdallah, Basem; Kassem, Moustapha

    2008-01-01

    introduced into clinical medicine in variety of applications and through different ways of administration. Here, we discuss approaches for isolation, characterization and directing differentiation of human mesenchymal stem cells (hMSC). An update of the current clinical use of the cells is also provided....

  12. Term pregnancy with choriocarcinoma presenting as severe fetal anemia and postpartum hemorrhage.

    Science.gov (United States)

    Peng, Hsiu-Huei; Ng, Zooi-Ping; Tang, Yun-Hsin; Chua, Angelica Anne A; Huang, Kuan-Gen

    2016-06-01

    Term pregnancy with choriocarcinoma is a rare condition that can be a serious health threat to both the mother and the fetus. We present a rare case of term pregnancy with choriocarcinoma presenting as severe fetal anemia and postpartum hemorrhage. A 34-year-old woman, gravida 3 para 2, was admitted for profuse vaginal bleeding 2 weeks after cesarean delivery of a full-term anemic baby. Transvaginal sonography revealed a 4.7-cm×10.6-cm heterogenous lesion in the endometrial cavity. Dilatation and curettage was done and a pathologic report revealed choriocarcinoma. Metastatic workup showed lung metastasis. The patient achieved remission after eight cycles of chemotherapy in the form of etoposide, methotrexate, actinomycin D, cyclophosphamide, and vincristine. There was no evidence of recurrence in the subsequent 3 years of regular follow up. Although fetomaternal hemorrhage is a rare form of presentation of choriocarcinoma, its presence should alert the physician to investigate the cause further. This chemotherapy regimen was effective in our case and the patient needed to be followed up carefully. Copyright © 2016. Published by Elsevier B.V.

  13. In vitro antiproliferative effect of six Salvia species on human tumor cell lines.

    Science.gov (United States)

    Fiore, Giovina; Nencini, Cristina; Cavallo, Federica; Capasso, Anna; Bader, Ammar; Giorgi, Giorgio; Micheli, Lucia

    2006-08-01

    This study was designed to examine the in vitro antiproliferative activity of the methanol crude extracts of six Salvia species: Salvia dominica L. leaves, Salvia lanigera Desf. aerial parts, Salvia menthaefolia Ten. roots, Salvia palaestina Benth. aerial parts, Salvia sclarea L. roots and Salvia spinosa L. aerial parts. Extracts were screened for their possible antitumoral activity by MTT test on nine human cancer cell lines: glioblastoma (DBTRG-05MG, T98G, U-87MG), colorectal adenocarcinoma (WiDr and HT-29), prostate adenocarcinoma (MDA Pca2b), choriocarcinoma (JEG-3), endometrium adenocarcinoma (HEC-1A) and B lymphoblast (CIR). IC(50) values were determined for only five extracts and ranged from 90 to 400 microg/mL approximately. Salvia menthaefolia extract exhibited marked antiproliferative activity against all tumor cell lines showing lower IC(50) values, while S. spinosa, S. sclarea and S. dominica extracts showed a degree cytotoxic activity dependent on the cell line type. Finally S. palaestina extract revealed a moderate antiproliferative effect only against three cell lines. Salvia lanigera extract displayed toxic activity at all concentrations tested. The results strengthen the evidence that the genus Salvia could be considered a natural resource of potential antitumor agents.

  14. Extra gonadal non-seminomatous germ cell tumour and PET-T.D.M. with {sup 18}F-F.D.G.: about one case of primitive retroperitoneal choriocarcinoma; Tumeurs germinales non seminomateuses extragonadiques et TEP-TDM au F-18 FDG: a propos d'un cas de choriocarcinome retroperitoneal primitif

    Energy Technology Data Exchange (ETDEWEB)

    Cimarelli, S.; Deshayes, E.; Mognetti, T.; Desuzinges, C. [Service de medecine nucleaire, centre Leon-Berard, Lyon, (France); Biron, P. [departement d' oncologie, centre Leon-Berard, Lyon, (France); Rivoire, M. [departement de chirurgie, centre Leon-Berard, Lyon, (France); Giammarile, F. [service de medecine nucleaire, hopital Lyon-Sud, (France)

    2009-05-15

    The non-seminomatous germinal tumors represent 60% of the germinal tumors, the most frequent cancer for young men.The positron computed tomography/computed tomography (PET/T.D.M.) with {sup 18}F fluorodeoxyglucose seems full of promises for the initial evaluation and the early evaluation of chemotherapy. for this type of tumor. In 1 to 5% of cases these tumors are extra gonadal. We present the case of a twenty three years old man with a retroperitoneal primitive choriocarcinoma with numerous metastases for whom the metabolic imaging was useful. We discuss the interest of this examination in this rare pathology. For the extra gonadal non-seminomatous germinal tumors the PET-F.D.G. seems bring information useful for the determination of the viable character of a post-chemotherapy residual mass, especially when the anatomical imaging show results discordant with the clinico biological data. (N.C.)

  15. Postmolar choriocarcinoma: An independent risk factor for chemotherapy resistance in low-risk gestational trophoblastic neoplasia.

    Science.gov (United States)

    Strohl, Anna E; Lurain, John R

    2016-05-01

    To evaluate the effect of a clinicopathologic diagnosis of choriocarcinoma (CCA) on clinical characteristics, extent of disease, and response to chemotherapy in low-risk gestational trophoblastic neoplasia (GTN). We reviewed the records of 678 patients with low-risk GTN (FIGO stage I and stages IIIII, scoredisease characteristics, treatment course, as well as clinical response and survival were analyzed retrospectively. Patients with a clinicopathologic diagnosis of CCA were compared to those with a clinical diagnosis of postmolar GTN. Of 678 women with low-risk GTN, 129 (19.0%) had a clinicopathologic diagnosis of CCA. Patients with CCA had higher parity (median 1 vs. 0, p=0.003), more pretreatment human chorionic gonadotropin (hCG) levels at >100,000mIU/mL (12.7% vs. 5.9%, p=0.009), longer duration of disease (19.6 vs. 9.9weeks, pdisease (stage I, 83.1% vs 88.2%, p=0.126). Although there was no difference in survival between groups, increased resistance to first-line methotrexate chemotherapy was significantly associated with a diagnosis of postmolar CCA (OR 2.67, p=0.007)), pretreatment hCG level at >10,000mIU/mL (OR 2.62, p=0.002), and higher FIGO score (3-4: OR 2.02, p=0.027; 5-6: OR 5.56, p<0.001) on multivariate analysis. Clinicopathologic diagnosis of postmolar CCA in patients with low-risk GTN is associated with higher pretreatment hCG levels, higher FIGO scores, and increased resistance to first-line single-agent methotrexate chemotherapy. Copyright © 2016 Elsevier Inc. All rights reserved.

  16. Human innate lymphoid cells.

    Science.gov (United States)

    Mjösberg, Jenny; Spits, Hergen

    2016-11-01

    Innate lymphoid cells (ILCs) are increasingly acknowledged as important mediators of immune homeostasis and pathology. ILCs act as early orchestrators of immunity, responding to epithelium-derived signals by expressing an array of cytokines and cell-surface receptors, which shape subsequent immune responses. As such, ILCs make up interesting therapeutic targets for several diseases. In patients with allergy and asthma, group 2 innate lymphoid cells produce high amounts of IL-5 and IL-13, thereby contributing to type 2-mediated inflammation. Group 3 innate lymphoid cells are implicated in intestinal homeostasis and psoriasis pathology through abundant IL-22 production, whereas group 1 innate lymphoid cells are accumulated in chronic inflammation of the gut (inflammatory bowel disease) and lung (chronic obstructive pulmonary disease), where they contribute to IFN-γ-mediated inflammation. Although the ontogeny of mouse ILCs is slowly unraveling, the development of human ILCs is far from understood. In addition, the growing complexity of the human ILC family in terms of previously unrecognized functional heterogeneity and plasticity has generated confusion within the field. Here we provide an updated view on the function and plasticity of human ILCs in tissue homeostasis and disease. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  17. Pegylated Liposomal Doxorubicin Is an Active Agent for Chemotherapy-Resistant Choriocarcinoma: A Report of Two Cases.

    Science.gov (United States)

    Popadiuk, Catherine; Power, Patti

    2016-01-01

    Despite advances in chemotherapy, radiation, surgery, and supportive treatments, a significant proportion of high-risk metastatic gestational trophoblastic disease patients develop resistant disease and die. Of those cured, protracted treatments can lead to long-term morbidity or later toxicity and death. Here we describe 2 patients with brain metastases who failed multiple lines of standard chemotherapy and radiation but had complete response to pegylated liposomal doxorubicin (PLD). A 35-year-old woman presented with choriocarcinoma in the brain, lungs, and subcutaneous tissues 11 months after full-term delivery. Her FIGO risk score was 14. Over 3 years she was treated with EMA-CO, EMA-CE, Taxol, gemcitabine, brain radiation, and excisional craniotomy for recurrent choriocarcinoma. She showed complete response of choriocarcinoma brain metastases following 2 cycles of PLD. She was choriocarcinoma free until her death 9 months later from acute myelogenous leukemia. A 52-year-old multigravid woman presented with choriocarcinoma 3 years following miscarriage. Her FIGO score was 16. Over 18 months she was treated with EMA-CO, TP/TE and IT MTX, and radiation. Her disease proved resistant and midbrain tumor unresectable. She showed complete response to PLD following 3 cycles but ultimately died from neurologic complications. PLD is an active agent in the treatment of high-risk choriocarcinoma.

  18. Choriocarcinoma in Enugu, South east Nigeria: A Need for a Shift ...

    African Journals Online (AJOL)

    while 7 (46.7% were lost to follow-up during chemotherapy 20.0% or after chemotherapy (26.7%). CONCLUSION: The case fatality for choriocarcinoma and loss of patients to follow-up in Enugu, Nigeria were high. To shift from this situation of high mortality to that of survival, an improved follow-up of post-abortal patients ...

  19. Embelin-Induced Apoptosis of Human Prostate Cancer Cells Is Mediated through Modulation of Akt and β-Catenin Signaling.

    Directory of Open Access Journals (Sweden)

    Nahee Park

    Full Text Available There is increasing evidence that embelin, an active component of Embelia ribes, induces apoptosis in human cancer cells, but the detailed mechanisms are still unclear. Here, we have investigated the effect of embelin on the growth of human prostate cancer cells. Embelin strongly inhibited cell growth especially in human prostate cancer cell lines, including PC3, DU145, LNCaP-LN3 and normal prostate epithelial cell, RWPE-1 compared to breast cancer (MDA-MB-231, MCF-7, and T47D, hepatoma (HepG2, Hep3B, and HuH-7, or choriocarcinoma (JEG-3. We observed that embelin induced apoptosis of PC3 cells in a time-dependent manner correlated with decreased expression of Bcl-2, Bcl-xL, and Mcl-1, increased translocation of Bax into mitochondria, and a reduction in the mitochondrial membrane potential. Furthermore, embelin induced voltage-dependent anion channel (VDAC 1 expression and oligomerization, which may promote cytochrome c and AIF release. Because embelin was able to inhibit Akt activation and cyclooxygenase-2 expression, the effects on Wnt/ β-catenin signaling were determined. Embelin activated glycogen synthase kinase (GSK-3β by preventing phosphorylation and suppressed β-catenin expression. Attenuation of β-catenin-mediated TCF transcriptional activity and gene transcription, such as cyclin D1, c-myc, and matrix metalloproteinase (MMP-7, were shown in embelin-treated cells. The changes in β-catenin levels in response to embelin were blocked by lithium chloride, a GSK-3 inhibitor, indicating that embelin may decrease β-catenin expression via GSK-3β activation. Furthermore, exposure of PC3 cells to embelin resulted in a significant decrease in cell migration and invasion. In conclusion, these findings suggest that inhibition of Akt signaling and activation of GSK-3β partially contributes to the pro-apoptotic effect of embelin in prostate cancer cells.

  20. Recurrent hydatidiform mole: A case report of six consecutive molar pregnancies complicated by choriocarcinoma, and review of the literature

    Directory of Open Access Journals (Sweden)

    Ahlam A Al-Ghamdi

    2011-01-01

    Full Text Available Hydatidiform mole (HM is the most common form of gestational trophoblastic neoplasia. Recurrence of HM is extremely rare. Here, we report the case of a patient with six consecutive partial HMs without normal pregnancy. A 42-year-old lady who was referred to us at King Fahad Hospital of the University, Al Khobar, initially as a case of 26-year-old with persistent trophoblastic disease after three recurrent molar pregnancies that were confirmed histologically in the referring hospital. She underwent evacuation and curettage and was followed up by serial β-human chorionic gonadotropin levels, and did not require chemotherapy. She then had three more molar pregnancies in 1995, 1996, and 2004; all molar pregnancies were evacuated by suction curettage at her base hospital, but in the last event, she complained of shortness of breath and abdominal pain. Diagnostic workup in our hospital confirmed choriocarcinoma, for which she received multiple regimen chemotherapy and was cured. Unfortunately, she lately presented with symptoms suggestive of premature menopause.

  1. The first trimester human trophoblast cell line ACH-3P: a novel tool to study autocrine/paracrine regulatory loops of human trophoblast subpopulations--TNF-alpha stimulates MMP15 expression.

    Science.gov (United States)

    Hiden, Ursula; Wadsack, Christian; Prutsch, Nicole; Gauster, Martin; Weiss, Ursula; Frank, Hans-Georg; Schmitz, Ulrike; Fast-Hirsch, Christa; Hengstschläger, Markus; Pötgens, Andy; Rüben, Angela; Knöfler, Martin; Haslinger, Peter; Huppertz, Berthold; Bilban, Martin; Kaufmann, Peter; Desoye, Gernot

    2007-12-19

    The trophoblast compartment of the placenta comprises various subpopulations with distinct functions. They interact among each other by secreted signals thus forming autocrine or paracrine regulatory loops. We established a first trimester trophoblast cell line (ACH-3P) by fusion of primary human first trimester trophoblasts (week 12 of gestation) with a human choriocarcinoma cell line (AC1-1). Expression of trophoblast markers (cytokeratin-7, integrins, matrix metalloproteinases), invasion abilities and transcriptome of ACH-3P closely resembled primary trophoblasts. Morphology, cytogenetics and doubling time was similar to the parental AC1-1 cells. The different subpopulations of trophoblasts e.g., villous and extravillous trophoblasts also exist in ACH-3P cells and can be immuno-separated by HLA-G surface expression. HLA-G positive ACH-3P display pseudopodia and a stronger expression of extravillous trophoblast markers. Higher expression of insulin-like growth factor II receptor and human chorionic gonadotropin represents the basis for the known autocrine stimulation of extravillous trophoblasts. We conclude that ACH-3P represent a tool to investigate interaction of syngeneic trophoblast subpopulations. These cells are particularly suited for studies into autocrine and paracrine regulation of various aspects of trophoblast function. As an example a novel effect of TNF-alpha on matrix metalloproteinase 15 in HLA-G positive ACH-3P and explants was found.

  2. The first trimester human trophoblast cell line ACH-3P: A novel tool to study autocrine/paracrine regulatory loops of human trophoblast subpopulations – TNF-α stimulates MMP15 expression

    Directory of Open Access Journals (Sweden)

    Knöfler Martin

    2007-12-01

    Full Text Available Abstract Background The trophoblast compartment of the placenta comprises various subpopulations with distinct functions. They interact among each other by secreted signals thus forming autocrine or paracrine regulatory loops. We established a first trimester trophoblast cell line (ACH-3P by fusion of primary human first trimester trophoblasts (week 12 of gestation with a human choriocarcinoma cell line (AC1-1. Results Expression of trophoblast markers (cytokeratin-7, integrins, matrix metalloproteinases, invasion abilities and transcriptome of ACH-3P closely resembled primary trophoblasts. Morphology, cytogenetics and doubling time was similar to the parental AC1-1 cells. The different subpopulations of trophoblasts e.g., villous and extravillous trophoblasts also exist in ACH-3P cells and can be immuno-separated by HLA-G surface expression. HLA-G positive ACH-3P display pseudopodia and a stronger expression of extravillous trophoblast markers. Higher expression of insulin-like growth factor II receptor and human chorionic gonadotropin represents the basis for the known autocrine stimulation of extravillous trophoblasts. Conclusion We conclude that ACH-3P represent a tool to investigate interaction of syngeneic trophoblast subpopulations. These cells are particularly suited for studies into autocrine and paracrine regulation of various aspects of trophoblast function. As an example a novel effect of TNF-α on matrix metalloproteinase 15 in HLA-G positive ACH-3P and explants was found.

  3. The first trimester human trophoblast cell line ACH-3P: A novel tool to study autocrine/paracrine regulatory loops of human trophoblast subpopulations – TNF-α stimulates MMP15 expression

    Science.gov (United States)

    Hiden, Ursula; Wadsack, Christian; Prutsch, Nicole; Gauster, Martin; Weiss, Ursula; Frank, Hans-Georg; Schmitz, Ulrike; Fast-Hirsch, Christa; Hengstschläger, Markus; Pötgens, Andy; Rüben, Angela; Knöfler, Martin; Haslinger, Peter; Huppertz, Berthold; Bilban, Martin; Kaufmann, Peter; Desoye, Gernot

    2007-01-01

    Background The trophoblast compartment of the placenta comprises various subpopulations with distinct functions. They interact among each other by secreted signals thus forming autocrine or paracrine regulatory loops. We established a first trimester trophoblast cell line (ACH-3P) by fusion of primary human first trimester trophoblasts (week 12 of gestation) with a human choriocarcinoma cell line (AC1-1). Results Expression of trophoblast markers (cytokeratin-7, integrins, matrix metalloproteinases), invasion abilities and transcriptome of ACH-3P closely resembled primary trophoblasts. Morphology, cytogenetics and doubling time was similar to the parental AC1-1 cells. The different subpopulations of trophoblasts e.g., villous and extravillous trophoblasts also exist in ACH-3P cells and can be immuno-separated by HLA-G surface expression. HLA-G positive ACH-3P display pseudopodia and a stronger expression of extravillous trophoblast markers. Higher expression of insulin-like growth factor II receptor and human chorionic gonadotropin represents the basis for the known autocrine stimulation of extravillous trophoblasts. Conclusion We conclude that ACH-3P represent a tool to investigate interaction of syngeneic trophoblast subpopulations. These cells are particularly suited for studies into autocrine and paracrine regulation of various aspects of trophoblast function. As an example a novel effect of TNF-α on matrix metalloproteinase 15 in HLA-G positive ACH-3P and explants was found. PMID:18093301

  4. Human leucocyte antigen (HLA) expression of primary trophoblast cells and placental cell lines, determined using single antigen beads to characterize allotype specificities of anti-HLA antibodies.

    Science.gov (United States)

    Apps, Richard; Murphy, Shawn P; Fernando, Raymond; Gardner, Lucy; Ahad, Tashmeeta; Moffett, Ashley

    2009-05-01

    Human trophoblast cells express an unusual repertoire of human leucocyte antigen (HLA) molecules which has been difficult to define. Close homology between and extreme polymorphism at the classical HLA class-I (HLA-I) loci has made it difficult to generate locus-specific monoclonal antibodies (mAbs). The problem of defining an antibody's reactivity against the thousands of existing HLA-I allotypes has often made it impossible to determine the HLA bound by a mAb in biological samples from a normal outbred population. Here we have used commercially available beads coated with individual HLA-I to characterize experimentally the reactivity of nine mAb against 96 common HLA-I allotypes. In conjunction with donor HLA-I genotyping, we could then define the specific HLA molecules bound by these antibodies in normal individuals. We used this approach to analyse the HLA expression of primary trophoblast cells from normal pregnancies; the choriocarcinoma cells JEG-3 and JAR; and the placental cell lines HTR-8/SVneo, Swan-71 and TEV-1. We confirm that primary villous trophoblast cells are HLA null whereas extravillous trophoblast cells express HLA-C, HLA-G and HLA-E, but not HLA-A, HLA-B or HLA-DR molecules in normal pregnancy. Tumour-derived JEG-3 and JAR cells reflect extravillous and villous trophoblast HLA phenotypes, respectively, but the HLA repertoire of the in vitro derived placental cell lines is not representative of either in vivo trophoblast phenotype. This study raises questions regarding the validity of using the placental cell lines that are currently available as model systems for immunological interactions between fetal trophoblast and maternal leucocytes bearing receptors for HLA molecules.

  5. Epidermal growth factor-like domain 7 promotes migration and invasion of human trophoblast cells through activation of MAPK, PI3K and NOTCH signaling pathways.

    Science.gov (United States)

    Massimiani, M; Vecchione, L; Piccirilli, D; Spitalieri, P; Amati, F; Salvi, S; Ferrazzani, S; Stuhlmann, H; Campagnolo, L

    2015-05-01

    Epidermal growth factor-like domain 7 (Egfl7) is a gene that encodes a partially secreted protein and whose expression is largely restricted to the endothelia. We recently reported that EGFL7 is also expressed by trophoblast cells in mouse and human placentas. Here, we investigated the molecular pathways that are regulated by EGFL7 in trophoblast cells. Stable EGFL7 overexpression in a Jeg3 human choriocarcinoma cell line resulted in significantly increased cell migration and invasiveness, while cell proliferation was unaffected. Analysis of mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3K) pathways showed that EGFL7 promotes Jeg3 cell motility by activating both pathways. We show that EGFL7 activates the epidermal growth factor receptor (EGFR) in Jeg3 cells, resulting in downstream activation of extracellular regulated kinases (ERKs). In addition, we provide evidence that EGFL7-triggered migration of Jeg3 cells involves activation of NOTCH signaling. EGFL7 and NOTCH1 are co-expressed in Jeg3 cells, and blocking of NOTCH activation abrogates enhanced migration of Jeg3 cells overexpressing EGFL7. We also demonstrate that signaling through EGFR and NOTCH converged to mediate EGFL7 effects. Reduction of endogenous EGFL7 expression in Jeg3 cells significantly decreased cell migration. We further confirmed that EGFL7 stimulates cell migration by using primary human first trimester trophoblast (PTB) cells overexpressing EGFL7. In conclusion, our data suggest that in trophoblast cells, EGFL7 regulates cell migration and invasion by activating multiple signaling pathways. Our results provide a possible explanation for the correlation between reduced expression of EGFL7 and inadequate trophoblast invasion observed in placentopathies. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  6. Changes in clinical presentation of postterm choriocarcinoma at the New England Trophoblastic Disease Center in recent years.

    Science.gov (United States)

    Diver, Elisabeth; May, Taymaa; Vargas, Roberto; Bernstein, Marilyn; Goldstein, Donald; Berkowitz, Ross

    2013-09-01

    To review the last 15 year experience of choriocarcinoma following a term gestation at the New England Trophoblastic Disease Center (NETDC) and compare these results to earlier data to determine any changes in the clinical presentation and outcome of this disease. Women with postterm choriocarcinoma from 1996 through 2011 followed by the NETDC were identified by diagnosis codes. Twenty charts were identified and reviewed. These data were then compared to published results from the NETDC of 44 women from 1964 to 1996. Time from antecedent pregnancy to diagnosis of choriocarcinoma was significantly longer in the current series, 46.1 vs. 19.7 weeks (p = 0.03). Despite this change, patient outcomes remained comparable, with similar overall mortality rates (13% vs. 10%, p = NS). However, patient presentation was notably different. In the early series, five (11%) infants suffered hydrops or stillbirth, while in the recent series there were no adverse infant outcomes (p = 0.08). Six women in the current series presented in the absence of symptoms suspicious for choriocarcinoma (either by an incidental positive pregnancy test without other symptoms or by placental pathology), compared to one woman in the prior series (30% vs. 2%, p = 0.001). In recent years postterm choriocarcinoma is being diagnosed or referred later after the antecedent pregnancy at our regional referral center. Recent patients more commonly have no other symptoms than a question of pregnancy and are less likely diagnosed due to the presence of fetal hydrops or stillbirth. Despite later diagnoses, survival with postterm choriocarcinoma continues to be high. Copyright © 2013 Elsevier Inc. All rights reserved.

  7. Genome engineering in human cells.

    Science.gov (United States)

    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.

  8. Perfluorinated chemicals: Differential toxicity, inhibition of aromatase activity and alteration of cellular lipids in human placental cells

    Energy Technology Data Exchange (ETDEWEB)

    Gorrochategui, Eva; Pérez-Albaladejo, Elisabet [Department of Environmental Chemistry, IDAEA–CSIC, 08034 Barcelona, Catalonia (Spain); Casas, Josefina [Department of Biomedicinal Chemistry, IQAC–CSIC, 08034 Barcelona, Catalonia (Spain); Lacorte, Sílvia, E-mail: slbqam@cid.csic.es [Department of Environmental Chemistry, IDAEA–CSIC, 08034 Barcelona, Catalonia (Spain); Porte, Cinta, E-mail: cinta.porte@cid.csic.es [Department of Environmental Chemistry, IDAEA–CSIC, 08034 Barcelona, Catalonia (Spain)

    2014-06-01

    The cytotoxicity of eight perfluorinated chemicals (PFCs), namely, perfluorobutanoic acid (PFBA), perfluorohexanoic acid (PFHxA), perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), perfluorododecanoic acid (PFDoA), perfluorobutanesulfonate (PFBS), perfluorohexanesulfonate (PFHxS) and perfluorooctanesulfonate (PFOS) was assessed in the human placental choriocarcinoma cell line JEG-3. Only the long chain PFCs – PFOS, PFDoA, PFNA, PFOA – showed significant cytotoxicity in JEG-3 cells with EC50 values in the range of 107 to 647 μM. The observed cytotoxicity was to some extent related to a higher uptake of the longer chain PFCs by cells (PFDoA > PFOS ≫ PFNA > PFOA > PFHxA). Moreover, this work evidences a high potential of PFOS, PFOA and PFBS to act as aromatase inhibitors in placental cells with IC50s in the range of 57–80 μM, the inhibitory effect of PFBS being particularly important despite the rather low uptake of the compound by cells. Finally, exposure of JEG-3 cells to a mixture of the eight PFCs (0.6 μM each) led to a relative increase (up to 3.4-fold) of several lipid classes, including phosphatidylcholines (PCs), plasmalogen PC and lyso plasmalogen PC, which suggests an interference of PFCs with membrane lipids. Overall, this work highlights the ability of the PFC mixture to alter cellular lipid pattern at concentrations well below those that generate toxicity, and the potential of the short chain PFBS, often considered a safe substitute of PFOS, to significantly inhibit aromatase activity in placental cells. - Highlights: • Eight perfluorinated chemicals of different chain lengths have been selected. • Long chain ones – PFOS, PFDoA, PFNA, PFOA – were cytotoxic in placenta cells. • The uptake of long chain perfluorinated chemicals by cells was comparatively higher. • PFOS, PFOA and the short chain PFBS significantly inhibited aromatase activity. • A mixture of perfluorinated chemicals significantly altered placenta cell

  9. Intraplacental Choriocarcinoma: Rare or Underdiagnosed? Report of 2 Cases Diagnosed after an Incomplete Miscarriage and a Preterm Spontaneous Vaginal Delivery

    Science.gov (United States)

    Oliveira, Paula; Scigliano, Horácio; Nogueira, Rosete; Araújo, Célia; Ferreira, Soledade

    2017-01-01

    Intraplacental choriocarcinoma is a rare malignant tumor diagnosed after an abortion, an ectopic pregnancy, or a term or preterm pregnancy or following the diagnosis of a hydatidiform mole. During pregnancy, it may be more common than reported, as most patients are asymptomatic and placental choriocarcinomas are usually inconspicuous macroscopically and are often mistaken for an infarct. Based upon a case study methodology, we describe 2 cases of intraplacental choriocarcinoma: the first case was identified in the product of a uterine curettage following an incomplete miscarriage and the second in one of the placentas of a bichorionic twin pregnancy. Maternal investigation did not reveal evidence of metastatic disease and neither did the infants' one in the second case. The two cases underwent maternal surveillance with serum hCG and remained disease-free until the present. In conclusion, intraplacental choriocarcinoma is easily underdiagnosed but with current treatment, even in the presence of metastasis, the prognosis is excellent. A routine microscopic examination of all the placentas and products of miscarriage can increase the real incidence of this entity and consequently improve its management. PMID:28567059

  10. Clinical characteristics and prognosis of 272 postterm choriocarcinoma patients at Peking Union Medical College Hospital: a retrospective cohort study.

    Science.gov (United States)

    Li, Jie; Yang, Junjun; Liu, Pengfei; Ren, Tong; Zhao, Jun; Feng, Fengzhi; Wan, Xirun; Xiang, Yang

    2016-06-02

    The objective of our study was to investigate the clinical characteristics and prognosis of postterm choriocarcinoma patients at Peking Union Medical College Hospital within the past 30 years. The clinical characteristics and pertinent follow-up data of 272 patients with postterm choriocarcinoma diagnosed from December 1985 through December 2014 in our hospital were reviewed. The clinical characteristics of two cohorts cut off at 2006 were compared using χ (2) tests. Risk factors of prognosis were estimated by multivariate Cox proportional regression analysis. The most common initial symptom was abnormal uterine bleeding. After individualized treatment 239 patients (87.9 %) achieved complete remission, including 140 patients received initial treatment of 5-fluorouracil-based multidrug chemotherapy. There were almost no statistically significant differences in the clinical characteristics and survival rates between the two cohorts. The results of the multivariate analysis showed that history of resistance to multidrug chemotherapy, liver metastasis and FIGO score greater than 12 were independent risk factors of prognosis. Postterm choriocarcinoma patients were usually accompanied by several high-risk factors that should received combined chemotherapy to prevent delay in adequate treatment. 5-fluorouracil-based multidrug chemotherapy, which has been applied at PUMCH for several decades, can be an effective initial treatment for postterm choriocarcinoma patients. More emphasis should be placed on those who have history of resistance to multidrug chemotherapy, liver metastasis or a FIGO score greater than 12.

  11. Quantified colocalization reveals heterotypic histocompatibility class I antigen associations on trophoblast cell membranes: relevance for human pregnancy.

    Science.gov (United States)

    Jabeen, Asma; Miranda-Sayago, José Maria; Obara, Boguslaw; Spencer, Patrick Simon; Dealtry, Gill Barbara; Hayrabedyan, Soren; Shaikly, Valerie; Laissue, Pierre Philippe; Fernández, Nelson

    2013-10-01

    Human placental syncytiotrophoblasts lack expression of most types of human leukocyte antigen (HLA) class I and class II molecules; this is thought to contribute to a successful pregnancy. However, the HLA class Ib antigens HLA-G, -E, and -F and the HLA class Ia antigen HLA-C are selectively expressed on extravillous trophoblast cells, and they are thought to play a major role in controlling feto-maternal tolerance. We have hypothesized that selective expression, coupled with the preferential physical association of pairs of HLA molecules, contribute to the function of HLA at the feto-maternal interface and the maternal recognition of the fetus. We have developed a unique analytical model that allows detection and quantification of the heterotypic physical associations of HLA class I molecules expressed on the membrane of human trophoblast choriocarcinoma cells, ACH-3P and JEG-3. Automated image analysis was used to estimate the degree of overlap of HLA molecules labeled with different fluorochromes. This approach yields an accurate measurement of the degree of colocalization. In both JEG-3 and ACH-3P cells, HLA-C, -E, and -G were detected on the cell membrane, while the expression of HLA-F was restricted to the cytoplasm. Progesterone treatment alone induced a significant increase in the expression level of the HLA-G/HLA-E association, suggesting that this heterotypic association is modulated by this hormone. Our data shows that the cell-surface HLA class I molecules HLA-G, -E, and -C colocalize with each other and have the potential to form preferential heterotypic associations.

  12. Diffusion inside living human cells

    DEFF Research Database (Denmark)

    Leijnse, N.; Jeon, J. -H.; Loft, Steffen

    2012-01-01

    Naturally occurring lipid granules diffuse in the cytoplasm and can be used as tracers to map out the viscoelastic landscape inside living cells. Using optical trapping and single particle tracking we found that lipid granules exhibit anomalous diffusion inside human umbilical vein endothelial...... cells. For these cells the exact diffusional pattern of a particular granule depends on the physiological state of the cell and on the localization of the granule within the cytoplasm. Granules located close to the actin rich periphery of the cell move less than those located towards to the center...... of the cell or within the nucleus. Also, granules in cells which are stressed by intense laser illumination or which have attached to a surface for a long period of time move in a more restricted fashion than those within healthy cells. For granules diffusing in healthy cells, in regions away from the cell...

  13. Human stromal (mesenchymal) stem cells

    DEFF Research Database (Denmark)

    Aldahmash, Abdullah; Zaher, Walid; Al-Nbaheen, May

    2012-01-01

    Human stromal (mesenchymal) stem cells (hMSC) represent a group of non-hematopoietic stem cells present in the bone marrow stroma and the stroma of other organs including subcutaneous adipose tissue, placenta, and muscles. They exhibit the characteristics of somatic stem cells of self......-renewal and multi-lineage differentiation into mesoderm-type of cells, e.g., to osteoblasts, adipocytes, chondrocytes and possibly other cell types including hepatocytes and astrocytes. Due to their ease of culture and multipotentiality, hMSC are increasingly employed as a source for cells suitable for a number...

  14. Human innate lymphoid cells

    NARCIS (Netherlands)

    Hazenberg, Mette D.; Spits, Hergen

    2014-01-01

    Innate lymphoid cells (ILCs) are lymphoid cells that do not express rearranged receptors and have important effector and regulatory functions in innate immunity and tissue remodeling. ILCs are categorized into 3 groups based on their distinct patterns of cytokine production and the requirement of

  15. Human innate lymphoid cells

    NARCIS (Netherlands)

    Mjösberg, Jenny; Spits, Hergen

    2016-01-01

    Innate lymphoid cells (ILCs) are increasingly acknowledged as important mediators of immune homeostasis and pathology. ILCs act as early orchestrators of immunity, responding to epithelium-derived signals by expressing an array of cytokines and cell-surface receptors, which shape subsequent immune

  16. Pulmonary Metastatic Choriocarcinoma in a Patient with Ectopic Pregnancy

    Directory of Open Access Journals (Sweden)

    Fahimeh Nokhostin

    2017-10-01

    Full Text Available Gestational trophoblastic tumor (GTT most commonly follows a molar pregnancy. In fact, it can occur following gestational events such as induced or spontaneous abortion, ectopic pregnancy, and term pregnancy. In this study, we present the case of a patient with ectopic pregnancy who was treated with a single dose of methotrexate (MTX. The maximum titer of beta subunit of human chorionic gonadotropin (BHCG was 402. Work up of occult metastatic GTT was carried out following MTX therapy, due to elevated BHCG titer. In the positron emission tomography (PET scan, a 17-mm lesion was detected in the right lung and treated via thoracotomy resection and adjuvant chemotherapy. Within an 18-month follow up, the patient did not reveal any symptoms and the BHCG titer level did not increase. Findings of this study revealed that after MTX therapy for ectopic pregnancy patients need to be followed up. Moreover, lack of BHCG titer decline, even in low tittering after treatment with MTX, can be related to GTT tumors or inadequate treatment for ectopic pregnancy.

  17. Gestational Choriocarcinoma Presenting with Lacrimal Gland Metastasis: A First Reported Case

    Directory of Open Access Journals (Sweden)

    Naushad A. B. Ahamed

    2015-01-01

    Full Text Available Background. Gestational choriocarcinoma (GC is a recognized clinicopathological subtype of gestational trophoblastic neoplasia that usually metastasizes hematogenously to highly vascular organs like the lung, liver, and brain. However, orbital metastasis to the choroid and lacrimal gland is a rare occurrence. Case Presentation. A 21-year-old female presented with headache and left orbital swelling one year after resection of a complete hydatidiform mole followed by adjuvant methotrexate chemotherapy. A metastatic imaging screening revealed multiple metastases in the lungs, brain, and adrenal gland, in addition to the choroid and lacrimal gland. Based on her modified WHO risk factors scoring she was started on chemotherapy and whole brain radiotherapy, which resulted in a complete response. At two-year follow-up, serum b-HCG level was with normal limits; imaging surveillance was uneventful. Conclusion. We present the first case of lacrimal gland metastasis in a young girl from GC relapse.

  18. "Term delivery following successful treatment of choriocarcinoma with brain metastases, (a case report"

    Directory of Open Access Journals (Sweden)

    F. Behnamfar

    2006-08-01

    Full Text Available Background: Cerebral metastases from choriocarcinoma are poor prognostic indicator of outcome in both the World Health Organization and FIGO classification systems. Although gestational trophoblastic neoplasia has become the most curable gynecological malignancy, failure rate among “high-risk” patients is still high despite the use of aggressive multidrug regimens. case: A 27 year old woman (G4P2Ab1 presented with hemiplegia due to brain metastases of choriocarcinoma one year after spontaneous abortion. She underwent craniotomy and was treated with nine courses of multiple agent etoposide, methotrexate, actinomycin-etoposide and cisplatinum (EMA-EP regimen combined with whole brain irradiation. She delivered a term healthy child two years after termination of treatment. Conclusion: Multiagent EMA-EP chemotherapy and whole brain irradiation with craniotomy in selected patients preserves fertility and may improve a patient overall prognosis. Methods: In a descriptive study from February to April 2005, two hundred sixty six consecutive pregnant women referring to a university hospital were asked to answer a questionnaire containing questions their sexual status and some demographic data. In 122 cases the answers of the spouses was collected also. The answers were compared in divided groups according to age range, duration of marriage, parity and educational status. Results: Fifty five percent of men and fifty eight percent of women had a negative attitude about sexual relations during pregnancy, and 60% of men and 75% of women presented incorrect knowledge about sexuality during pregnancy. Main reasons for decreased sexual relations in pregnancy were mentioned to be dysparaunia, and the fear of trauma to the baby, abortion, membrane rapture, preterm labor and infection. Conclusion: As couples’ knowledge and attitudes about sexuality affect their general sexual behavior during pregnancy it is crucial to provide proper consultation regarding

  19. Gastrointestinal hemorrhage due to metastatic choriocarcinoma with gastric and colonic involvement Hemorragia digestiva secundaria a coriocarcinoma con metástasis gástricas y colónicas

    Directory of Open Access Journals (Sweden)

    J. Molina Infante

    2004-01-01

    Full Text Available Metastatic choriocarcinoma is a rare nonseminomatous germ-cell tumor with a characteristic hemorrhagic tendency due to its trophoblastic origin. Gastrointestinal tube involvement is present in less than 5% of cases, and location or therapy of these lesions can be achieved by endoscopy, angiography or surgery. Despite its being a highly curable malignant disease, the ocurrence of gastrointestinal bleeding worsens prognosis. We report a case of metastatic choriocarcinoma which manifested as melaena and was diagnosed by the presence of metastatic lesions in the stomach and right bowel on endoscopy.El coriocarcinoma metastático es una infrecuente tumoración de células germinales con una marcada tendencia hemorrágica debido a su origen trofloblástico. La invasión del tubo digestivo ocurre en menos del 5% de los casos. A pesar de ser una enfermedad maligna con buena respuesta a la quimioterapia, la hemorragia gastrointestinal ensombrece el pronóstico. Presentamos un caso de un paciente de 37 años con un coriocarcinoma diseminado en el que la presentación clínica fue hemorragia digestiva en forma de melenas, encontrándose por endoscopia lesiones metastáticas en estómago y colon derecho.

  20. The effect of serum from women with preeclampsia on JAR (trophoblast-like) cell line.

    Science.gov (United States)

    Mahameed, Safa; Goldman, Shlomit; Gabarin, Diane; Weiss, Amir; Shalev, Eliezer

    2005-09-01

    Pathologic placentation has been implicated in the pathogenesis of preeclamsia. We sought to assess the effect serum obtained from women with preeclampsia would have on JAR human choriocarcinoma cells regarding growth, invasiveness, and matrix metalloproteinase (MMP) secretion as compared to normotensive pregnant woman. Blood was collected from 11 healthy pregnant women and from10 patients with preeclampsia at 28-33 weeks of gestation. The JAR human choriocarcinoma cell line was cultured in the presence of 10% serum obtained from each group. Cell proliferation, invasiveness, and MMP secretion was measured using a cell proliferation kit, the Matrigel (BD Biosciences, Beit-Ha'Emek, Israel) invasion assay, and gel zymography, respectively. Cell growth increased by 6% when exposed to serum from patients with preeclampsia compared to 30% from controls (P <.01). Trophoblast invasion was significantly (P <.01) reduced in the preeclampsia group (21 +/- 1.9%) compared to controls (27 +/- 2.5%). Valid MMP-2 secretion was reduced by 51% in the preeclampsia group compared to controls (P <.05). Serum obtained from women with preeclampsia contains a factor or factors that exhibit an inhibitory effect on JAR trophoblast cell proliferation, invasiveness, and MMP-2 secretion. These factors may be involved in the pathologic placentation associated with the pathogenesis of preeclampsia.

  1. Interaction of human trophoblast cells with gland-like endometrial spheroids: a model system for trophoblast invasion.

    Science.gov (United States)

    Buck, V U; Gellersen, B; Leube, R E; Classen-Linke, I

    2015-04-01

    Do maternal endometrial epithelial cell (EEC) differentiation and polarity impact the invasive capacity of extravillous trophoblast (EVT) cells during early human implantation? In a three dimensional (3D) confrontation co-culture the invasiveness of the human trophoblast cell line AC-1M88 was inversely correlated with the degree of differentiation and polarization of human endometrial adenocarcinoma cell spheroids. In a previous study desmosomal and adherens junction proteins were shown to spread from a subapically restricted lateral position to the entire lateral membrane in human glandular EECs during the implantation window of the menstrual cycle. Whether this change in EEC junction localization has an impact on the interaction of EVT cells with glandular EECs during early human implantation is not known. A new 3D cell culture system was developed in order to mimic early implantation events in humans. As a model for the invasion of endometrial glands by EVT cells, spheroids of three differently differentiated and polarized endometrial adenocarcinoma cell lines were confronted with an EVT cell line in co-culture experiments. Three human adenocarcinoma EEC lines were chosen for this study because of their differences in differentiation and polarization: HEC-1-A, which is well differentiated and highly polarized, Ishikawa, which is well differentiated and moderately polarized, and RL95-2, which is moderately differentiated and poorly polarized. When the cell lines were grown in reconstituted basement membrane, they formed gland-like, multicellular spheroids. The degree of polarization within the different EEC spheroids was assessed by 3D confocal immunofluorescence microscopy detecting the basal membrane protein integrin α6, the apical tight junction-associated protein ZO-1 and the desmosomal plaque protein desmoplakin 1/2 (Dsp). Cells of the human EVT cell line AC-1M88, which is a fusion cell line of primary EVT cells and choriocarcinoma-derived JEG-3 cells, were

  2. Coriocarcinoma: um estudo de caso Coriocarcinoma: un estudio de caso Choriocarcinoma: a case study

    Directory of Open Access Journals (Sweden)

    Pollyana Alves Silva

    2010-02-01

    Full Text Available O presente estudo objetivou descrever o caso de uma cliente portadora de coriocarcinoma gestacional e a assistência de enfermagem necessária a esta condição. A partir de informações constantes do prontuário médico e de anamnese e exame físico realizados em visita domiciliária, formularam-se diagnósticos e intervenções de enfermagem embasados na literatura. Foram identificados diagnósticos referentes aos períodos perioperatório e de tratamento quimioterápico. Além dos diagnósticos referidos, foi possível identificar outros relativos ao período pós-tratamento da cliente. O processo de enfermagem mostrou-se efetivo para a assistência qualificada à cliente com coriocarcinoma.El presente estudio objetivó describir el caso de una cliente portadora de coriocarcinoma gestacional y la asistencia de enfermería necesaria a esta condición. A partir de informaciones constantes del prontuario médico y de anamnese y examen físico realizados en visita domiciliaria, se formularon diagnósticos e intervenciones de enfermería embasados en la literatura. Fueron identificados diagnósticos referentes a los períodos periquirúrgico y de tratamiento quimioterápico. Además de los diagnósticos referidos, fue posible identificar otros relativos al período pos-tratamiento de la cliente. El proceso de enfermeria se mostro efectivo para la asistencia calificada a la cliente con coriocarcinoma.The present study aimed at describing the case of a client bearer for gestational choriocarcinoma and the nursing assistance needed for this condition. From information contained in the medical history and of anamnesis and physical examination made in a home visit, made up diagnoses and inteventions of nursing based in literature. Diagnoses have been identified related to the periods perioperative and chemotherapy treatment. In addition to these diagnoses, it was possible to identify others on the post-treatment period of client. The process of

  3. Human stem cell ethics: beyond the embryo.

    Science.gov (United States)

    Sugarman, Jeremy

    2008-06-05

    Human embryonic stem cell research has elicited powerful debates about the morality of destroying human embryos. However, there are important ethical issues related to stem cell research that are unrelated to embryo destruction. These include particular issues involving different types of cells used, the procurement of such cells, in vivo use of stem cells, intellectual property, and conflicts of interest.

  4. Failure of somatostatin to affect human chorionic somatomammotropin and human chorionic gonadotropin secretion in vitro.

    Science.gov (United States)

    Macaron, C; Kyncl, M; Rutsky, L; Halpern, B; Brewer, J

    1978-11-01

    The effect of somatostatin (SRIF) on human chorionic somatomammotropin (hCS) secretion was studied in human placental explants cultured in vitro. In the experimental flasks, SRIF was added in a concentration of 10, 100, and 1000 ng/ml media; hCS levels measured by RIA were not different from those found in the control flasks. In separate experiments, we investigated the action of SRIF on hCG secretion by a human malignant choriocarcinoma cell line maintained in tissue culture. SRIF (1000 ng/ml) did not inhibit basal or dibutyryl cAMP-induced stimulation of hCG secretion. These results suggest that somatostatin does not suppress hCS or hCG release in vitro from normal or malignant trophoblast, respectively.

  5. Unusual clinical presentations of choriocarcinoma: A systematic review of case reports

    Directory of Open Access Journals (Sweden)

    Mishu Mangla

    2017-02-01

    Full Text Available Choriocarcinoma (CC is a highly malignant tumor originating in the trophoblastic tissue. The clinical presentation of CC is so much varied that every case may be one of its kinds and thus can be a diagnostic challenge. Numerous case reports have been published in various journals regarding the unusual clinical presentations of this cancer. Therefore, we conducted a systematic review of all case reports in English language on gestational CC published in PubMed-indexed journals from 1998 to 2015. The main aim was to provide a summary and critical analysis of all the data and evidence published regarding the atypical clinical presentations of CC in recent years. In total, 121 case reports pertaining to unusual clinical manifestations of gestational CC were analyzed. The age of patients in whom cases were reported ranged from 17 to 67 years, and the time period between the index pregnancy and development of CC varied from 4 weeks to as long as 25 years. Cardiopulmonary complaints (20.66% followed by gastrointestinal (18.43% and central nervous system manifestations (17.67% were found to be the most common. Through this review, the authors have made an attempt to discuss various manifestations with which a patient with gestational CC can present to clinician so that early diagnosis and timely management can be initiated, thus improving clinical prognosis.

  6. A Fibroid or Cancer? A Rare Case of Mixed Choriocarcinoma and Epithelioid Trophoblastic Tumour

    Directory of Open Access Journals (Sweden)

    Wan Yu Luk

    2013-01-01

    Full Text Available Background. Gestational trophoblastic disease (GTD is a rare complication of pregnancy which is characterised by abnormal growth of the trophoblasts at the placental site. It is categorised into benign and malignant forms, which include hydatidiform moles (HMs and gestational trophoblastic neoplasia (GTN, respectively. A mixed choriocarcinoma (CC and epithelioid trophoblastic tumour (ETT is an extremely rare subgroup of GTN, which is a highly curable but aggressive form of malignancy. Case. We report a case of mixed CC and ETT in a 41-year-old patient who presented with a 2-year history of menorrhagia and fibroid uterus in the absence of previous history of molar pregnancy. She had a 12-year interval between the antecedent pregnancy and presentation. She was treated with intensive regimen of adjuvant chemotherapy, etoposide, methotrexate, and actinomycin-D with etoposide and cisplatin (EMA-EP. She has remained disease free for more than 5 years. Conclusion. This case highlights the importance of considering GTN as one of the differential diagnoses value of β-HCG in patients presented with menorrhagia and growing fibroids.

  7. Pancreatic and Hepatic Metastasis of an Undiagnosed Choriocarcinoma: An Exceptional Cause of Haemoperitoneum in Young Women – Report of a Rare Case

    Directory of Open Access Journals (Sweden)

    Nishat Fatema

    2016-10-01

    Full Text Available Haemoperitoneum in women of reproductive age usually suggests either ruptured ectopic pregnancy or ruptured ovarian cysts. Metastatic choriocarcinoma is considered the least common cause of haemoperitoneum in women of childbearing age. We report a rare case of pancreatic and hepatic metastasis of choriocarcinoma in a young, 30-year-old female who had delivered 10 months prior at term gestation with no previous history of gestational trophoblastic disease or molar pregnancy. She had a short history of fever and pain in the right hypochondrium, with findings of hypovolaemic shock due to intraperitoneal haemorrhage. Unfortunately, the patient expired with massive uncontrolled bleeding from liver metastasis despite 2 emergency laparotomies within 12 h. This case report is an apt reminder to clinicians to include metastatic choriocarcinoma on the list of differential diagnoses for haemoperitoneum with a positive pregnancy test in women of reproductive age to diagnose early and to avoid life-threatening consequences.

  8. Human regulatory B cells control the TFH cell response.

    Science.gov (United States)

    Achour, Achouak; Simon, Quentin; Mohr, Audrey; Séité, Jean-François; Youinou, Pierre; Bendaoud, Boutahar; Ghedira, Ibtissem; Pers, Jacques-Olivier; Jamin, Christophe

    2017-07-01

    Follicular helper T (TFH) cells support terminal B-cell differentiation. Human regulatory B (Breg) cells modulate cellular responses, but their control of TFH cell-dependent humoral immune responses is unknown. We sought to assess the role of Breg cells on TFH cell development and function. Human T cells were polyclonally stimulated in the presence of IL-12 and IL-21 to generate TFH cells. They were cocultured with B cells to induce their terminal differentiation. Breg cells were included in these cultures, and their effects were evaluated by using flow cytometry and ELISA. B-cell lymphoma 6, IL-21, inducible costimulator, CXCR5, and programmed cell death protein 1 (PD-1) expressions increased on stimulated human T cells, characterizing TFH cell maturation. In cocultures they differentiated B cells into CD138(+) plasma and IgD(-)CD27(+) memory cells and triggered immunoglobulin secretions. Breg cells obtained by Toll-like receptor 9 and CD40 activation of B cells prevented TFH cell development. Added to TFH cell and B-cell cocultures, they inhibited B-cell differentiation, impeded immunoglobulin secretions, and expanded Foxp3(+)CXCR5(+)PD-1(+) follicular regulatory T cells. Breg cells modulated IL-21 receptor expressions on TFH cells and B cells, and their suppressive activities involved CD40, CD80, CD86, and intercellular adhesion molecule interactions and required production of IL-10 and TGF-β. Human Breg cells control TFH cell maturation, expand follicular regulatory T cells, and inhibit the TFH cell-mediated antibody secretion. These novel observations demonstrate a role for the Breg cell in germinal center reactions and suggest that deficient activities might impair the TFH cell-dependent control of humoral immunity and might lead to the development of aberrant autoimmune responses. Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.

  9. Endocannabinoids and Human Sperm Cells

    Directory of Open Access Journals (Sweden)

    Giovanna Zolese

    2010-10-01

    Full Text Available N-acylethanolamides (NAEs are naturally occurring signaling lipids consisting of amides and esters of long-chain polyunsaturated fatty acids. Usually they are present in a very small amounts in many mammalian tissues and cells, including human reproductive tracts and fluids. Recently, the presence of N-arachidonoylethanolamide (anandamide, AEA, the most characterised member of endocannabinoids, and its congeners palmitoylethanolamide (PEA and oleylethanolamide (OEA in seminal plasma, oviductal fluid, and follicular fluids was demonstrated. AEA has been shown to bind not only type-1 (CB1 and type-2 (CB2 cannabinoid receptors, but also type-1 vanilloid receptor (TRPV1, while PEA and OEA are inactive with respect to classical cannabinoid CB1 and CB2 but activate TRPV1 or peroxisome proliferator activate receptors (PPARs. This review concerns the most recent experimental data on PEA and OEA, endocannabinoid-like molecules which appear to exert their action exclusively on sperm cells with altered features, such as membrane characteristics and kinematic parameters. Their beneficial effects on these cells could suggest a possible pharmacological use of PEA and OEA on patients affected by some forms of idiopathic infertility.

  10. Stem cells in the human breast

    DEFF Research Database (Denmark)

    Petersen, Ole William; Polyak, Kornelia

    2010-01-01

    The origins of the epithelial cells participating in the development, tissue homeostasis, and cancer of the human breast are poorly understood. However, emerging evidence suggests a role for adult tissue-specific stem cells in these processes. In a hierarchical manner, these generate the two main...... mammary cell lineages, producing an increasing number of cells with distinct properties. Understanding the biological characteristics of human breast stem cells and their progeny is crucial in attempts to compare the features of normal stem cells and cancer precursor cells and distinguish these from...... nonprecursor cells and cells from the bulk of a tumor. A historical overview of research on human breast stem cells in primary tissue and in culture reveals the progress that has been made in this area, whereas a focus on the cell-of-origin and reprogramming that occurs during neoplastic conversion provides...

  11. Satellite cells in human skeletal muscle plasticity.

    Science.gov (United States)

    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. Germ cell cancer presenting as gastrointestinal bleeding and developing brain metastases: case report and review of the literature.

    Science.gov (United States)

    Fu, Shuang; Avezbakiyev, Boris; Zhi, Wanqing; Kodali, Sreenath; Rizvon, Kaleem; Alaverdian, Artur; Freedman, Lester; Mejia, Jose; Shahzad, Ghulamullah; Gotlieb, Vladimir

    2012-11-01

    This paper describes a rare case of germ cell cancer with duodenum, brain and lung metastases. The patient presented with melena and left testicle enlargement. Orchiectomy revealed mixed germ cell cancer, enteroscopy revealed duodenal choriocarcinoma, and chest x-ray and computed tomography (CT) showed bilateral lung metastases. The patient received and tolerated cisplatinum-based chemotherapy, and responded well. However, he developed seizures 3 months later. MRI showed brain metastases and he was treated with whole-brain radiation. One month later, he developed progressive dyspnea. Chest CT showed worsening lung metastases. He received second-line chemotherapy, but died due to multiorgan failure. Germ cell cancer with nonpulmonary metastases has poor prognosis and the management of these patients requires a multimodal approach. Head CT should be considered as routine screening for all germ cell cancer patients on initial diagnosis and brain MRI should be considered for high-risk patients (with an embryo- or choriocarcinoma histology, dramatically elevated β-human chorionic gonadotropin and lung involvement).

  13. Endothelial cells derived from human embryonic stem cells

    Science.gov (United States)

    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.

  14. Induced pluripotent stem cell lines derived from human somatic cells.

    Science.gov (United States)

    Yu, Junying; Vodyanik, Maxim A; Smuga-Otto, Kim; Antosiewicz-Bourget, Jessica; Frane, Jennifer L; Tian, Shulan; Nie, Jeff; Jonsdottir, Gudrun A; Ruotti, Victor; Stewart, Ron; Slukvin, Igor I; Thomson, James A

    2007-12-21

    Somatic cell nuclear transfer allows trans-acting factors present in the mammalian oocyte to reprogram somatic cell nuclei to an undifferentiated state. We show that four factors (OCT4, SOX2, NANOG, and LIN28) are sufficient to reprogram human somatic cells to pluripotent stem cells that exhibit the essential characteristics of embryonic stem (ES) cells. These induced pluripotent human stem cells have normal karyotypes, express telomerase activity, express cell surface markers and genes that characterize human ES cells, and maintain the developmental potential to differentiate into advanced derivatives of all three primary germ layers. Such induced pluripotent human cell lines should be useful in the production of new disease models and in drug development, as well as for applications in transplantation medicine, once technical limitations (for example, mutation through viral integration) are eliminated.

  15. c-Myc-Dependent Cell Competition in Human Cancer Cells.

    Science.gov (United States)

    Patel, Manish S; Shah, Heta S; Shrivastava, Neeta

    2017-07-01

    Cell Competition is an interaction between cells for existence in heterogeneous cell populations of multicellular organisms. This phenomenon is involved in initiation and progression of cancer where heterogeneous cell populations compete directly or indirectly for the survival of the fittest based on differential gene expression. In Drosophila, cells having lower dMyc expression are eliminated by cell competition through apoptosis when present in the milieu of cells having higher dMyc expression. Thus, we designed a study to develop c-Myc (human homolog) dependent in vitro cell competition model of human cancer cells. Cells with higher c-Myc were transfected with c-myc shRNA to prepare cells with lower c-Myc and then co-cultured with the same type of cells having a higher c-Myc in equal ratio. Cells with lower c-Myc showed a significant decrease in numbers when compared with higher c-Myc cells, suggesting "loser" and "winner" status of cells, respectively. During microscopy, engulfment of loser cells by winner cells was observed with higher expression of JNK in loser cells. Furthermore, elimination of loser cells was prevented significantly, when co-cultured cells were treated with the JNK (apoptosis) inhibitor. Above results indicate elimination of loser cells in the presence of winner cells by c-Myc-dependent mechanisms of cell competition in human cancer cells. This could be an important mechanism in human tumors where normal cells are eliminated by c-Myc-overexpressed tumor cells. J. Cell. Biochem. 118: 1782-1791, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

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

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ying, E-mail: ying.chen@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Wang, Kai; Chandramouli, Gadisetti V.R. [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Knott, Jason G. [Developmental Epigenetics Laboratory, Department of Animal Science, Michigan State University (United States); Leach, Richard, E-mail: Richard.leach@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Department of Obstetrics, Gynecology and Women’s Health, Spectrum Health Medical Group (United States)

    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.

  17. Molecular regulation of human hematopoietic stem cells

    NARCIS (Netherlands)

    van Galen, P.L.J.

    2014-01-01

    Peter van Galen focuses on understanding the determinants that maintain the stem cell state. Using human hematopoietic stem cells (HSCs) as a model, processes that govern self-renewal and tissue regeneration were investigated. Specifically, a role for microRNAs in balancing the human HSC

  18. Human hematopoietic cell culture, transduction, and analyses

    DEFF Research Database (Denmark)

    Bonde, Jesper; Wirthlin, Louisa; Kohn, Donald B

    2008-01-01

    This unit provides methods for introducing genes into human hematopoietic progenitor cells. The Basic Protocol describes isolation of CD34(+) cells, transduction of these cells with a retroviral vector on fibronectin-coated plates, assaying the efficiency of transduction, and establishing long-te...

  19. Human cord blood cells can differentiate into retinal nerve cells.

    Science.gov (United States)

    Koike-Kiriyama, Naoko; Adachi, Yasushi; Minamino, Keizo; Iwasaki, Masayoshi; Nakano, Keiji; Koike, Yasushi; Yamada, Haruhiko; Mukaide, Hiromi; Shigematsu, Akio; Mizokami, Tomomi; Matsumura, Miyo; Ikehara, Susumu

    2007-01-01

    Retinal degeneration and dystrophy are the major causes of blindness in the developed world. It has been reported that human cord blood cells (HCBCs) can differentiate into neuron-like cells in vitro. We have recently demonstrated that bone marrow cells (BMCs) of both mice and rats can differentiate into retinal nerve cells (RNCs). In the present study, we show the differentiation capacity of HCBCs into RNCs in vivo. We transplanted lineage-negative HCBCs into the subretinal space of severe combined immunodeficiency (SCID) mice. Two weeks after the transplantation, some of the transplanted cells expressed human nestin, human MAP2, human neuron specific enolase (NSE), beta-III tubulin and also rhodopsin. These results indicate that HCBCs can differentiate into RNCs and suggest that our new strategy could be used for the regeneration of retinal nerve cells in degenerative or dystrophic diseases.

  20. Human Satellite Cell Isolation and Xenotransplantation.

    Science.gov (United States)

    Garcia, Steven M; Tamaki, Stanley; Xu, Xiaoti; Pomerantz, Jason H

    2017-01-01

    Satellite cells are mononucleated cells of the skeletal muscle lineage that exist beneath the basal lamina juxtaposed to the sarcolemma of skeletal muscle fibers. It is widely accepted that satellite cells mediate skeletal muscle regeneration. Within the satellite cell pool of adult muscle are skeletal muscle stem cells (MuSCs), also called satellite stem cells, which fulfill criteria of tissue stem cells: They proliferate and their progeny either occupies the adult MuSC niche during self-renewal or differentiates to regenerate mature muscle fibers. Here, we describe robust methods for the isolation of enriched populations of human satellite cells containing MuSCs from fresh human muscle, utilizing mechanical and enzymatic dissociation and purification by fluorescence-activated cell sorting. We also describe a process for xenotransplantation of human satellite cells into mouse muscle by injection into irradiated, immunodeficient, mouse leg muscle with concurrent notexin or bupivacaine muscle injury to increase engraftment efficiency. The engraftment of human MuSCs and the formation of human muscle can then be analyzed by histological and immunofluorescence staining, or subjected to in vivo experimentation.

  1. Report on Liver Cell Transplantation Using Human Fetal Liver Cells.

    Science.gov (United States)

    Pietrosi, Giada; Chinnici, Cinzia

    2017-01-01

    In an era of organ shortage, human fetuses donated after medically indicated abortion could be considered a potential liver donor for hepatic cell isolation. We investigated transplantation of fetal liver cells as a strategy to support liver functionality in end-stage liver disease. Here, we report our protocol of human fetal liver cells (hFLC) isolation in fetuses from 17 to 22 gestational weeks, and our clinical procedure of hFLC transplantation through the splenic artery.

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

    Science.gov (United States)

    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.

  3. Energy Generation in the Human Body by the Human Cells ...

    African Journals Online (AJOL)

    We adapted the thermodynamics equation for energy generation in a diesel engine in modeling energy generation in human body by the human cells by doing a thorough study on both systems and saw that the process of energy generation is the same in them. We equally saw that the stages involved in energy generation ...

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

    Science.gov (United States)

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

    2017-09-21

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

  5. Mesenchymal origin of multipotent human testis-derived stem cells in human testicular cell cultures

    NARCIS (Netherlands)

    Chikhovskaya, J. V.; van Daalen, S. K. M.; Korver, C. M.; Repping, S.; van Pelt, A. M. M.

    2014-01-01

    In contrast to mouse germ cell-derived pluripotent stem cells, the pluripotent state of human testis-derived embryonic stem cell (ESC)-like that spontaneously arise in primary testicular cell cultures remains controversial. Recent studies have shown that these cells closely resemble multipotent

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

    Energy Technology Data Exchange (ETDEWEB)

    Varga, Nora [Membrane Research Group of the Hungarian Academy of Sciences, Semmelweis University, Budapest (Hungary); Vereb, Zoltan; Rajnavoelgyi, Eva [Department of Immunology, Medical and Health Science Centre, University of Debrecen, Debrecen (Hungary); Nemet, Katalin; Uher, Ferenc; Sarkadi, Balazs [Membrane Research Group of the Hungarian Academy of Sciences, Semmelweis University, Budapest (Hungary); Apati, Agota, E-mail: apati@kkk.org.hu [Membrane Research Group of the Hungarian Academy of Sciences, Semmelweis University, Budapest (Hungary)

    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.

  7. Modeling human infertility with pluripotent stem cells

    Directory of Open Access Journals (Sweden)

    Di Chen

    2017-05-01

    Full Text Available Human fertility is dependent upon the correct establishment and differentiation of the germline. This is because no other cell type in the body is capable of passing a genome and epigenome from parent to child. Terminally differentiated germline cells in the adult testis and ovary are called gametes. However, the initial specification of germline cells occurs in the embryo around the time of gastrulation. Most of our knowledge regarding the cell and molecular events that govern human germline specification involves extrapolating scientific principles from model organisms, most notably the mouse. However, recent work using next generation sequencing, gene editing and differentiation of germline cells from pluripotent stem cells has revealed that the core molecular mechanisms that regulate human germline development are different from rodents. Here, we will discuss the major molecular pathways required for human germline differentiation and how pluripotent stem cells have revolutionized our ability to study the earliest steps in human embryonic lineage specification in order to understand human fertility.

  8. PD-L1 Expression in Human Placentas and Gestational Trophoblastic Diseases.

    Science.gov (United States)

    Veras, Emanuela; Kurman, Robert J; Wang, Tian-Li; Shih, Ie-Ming

    2017-03-01

    One of the major immune checkpoints responsible for immune evasion in cancer cells is the interaction between programmed cell death-1 (PD-1) and its ligand (PD-L1). As human trophoblastic cells display many of the features of malignant cells such as the ability to invade normal tissue including blood vessels and are apparently not eradicated by the host immune system, we undertook the present study to determine whether PD-L1 was upregulated in different types of trophoblastic cells during normal pregnancy and in gestational trophoblastic diseases. Immunohistochemistry using an anti-PD-L1-specific antibody demonstrated that in early and term normal placentas, PD-L1 was highly expressed in syncytiotrophoblast and to a much lower extent in intermediate trophoblastic cells located in the chorion laeve and implantation site. PD-L1 immunoreactivity was undetectable in cytotrophoblastic cells. This staining pattern in normal placenta was recapitulated in various types of gestational trophoblastic disease. PD-L1 was highly expressed by syncytiotrophoblast in complete moles and choriocarcinomas. The intermediate trophoblastic neoplasms, placental site trophoblastic tumors, and epithelioid trophoblastic tumors showed variable PD-L1 immunoreactivity but at a lower intensity than in the neoplastic syncytiotrophoblast in choriocarcinoma. In addition, we observed PD-1-positive lymphocytes located within the implantation site and in trophoblastic tumors. In summary, this study describes a novel mechanism for trophoblastic cells to create a tolerogenic feto-maternal interface by upregulating PD-L1 in syncytiotrophoblast and in intermediate trophoblast. Trophoblastic tumors may also use PD-L1 expression to evade the host immune response thereby promoting their survival.

  9. Cell communication in the basal cells of the human epidermis.

    Science.gov (United States)

    van Heukelom, J S; Slaaf, D W; van der Leun, J C

    1972-10-01

    Electrotonic spread can be measured in the basal cells of the human epidermis. The communication between neighboring cells is high, whereas no leak to the intercellular spaces could be detected. The specific resistance of the membranes between the cells is about 10 Omegacm(2). This finding suggests that for those particles that are able to pass the cell membrane the intracellular path through the epidermis is at least as suitable as the path through the intercellular spaces.

  10. Calcium signaling in human pluripotent stem cells.

    Science.gov (United States)

    Apáti, Ágota; Berecz, Tünde; Sarkadi, Balázs

    2016-03-01

    Human pluripotent stem cells provide new tools for developmental and pharmacological studies as well as for regenerative medicine applications. Calcium homeostasis and ligand-dependent calcium signaling are key components of major cellular responses, including cell proliferation, differentiation or apoptosis. Interestingly, these phenomena have not been characterized in detail as yet in pluripotent human cell sates. Here we review the methods applicable for studying both short- and long-term calcium responses, focusing on the expression of fluorescent calcium indicator proteins and imaging methods as applied in pluripotent human stem cells. We discuss the potential regulatory pathways involving calcium responses in hPS cells and compare these to the implicated pathways in mouse PS cells. A recent development in the stem cell field is the recognition of so called "naïve" states, resembling the earliest potential forms of stem cells during development, as well as the "fuzzy" stem cells, which may be alternative forms of pluripotent cell types, therefore we also discuss the potential role of calcium homeostasis in these PS cell types. Copyright © 2016 Elsevier Ltd. All rights reserved.

  11. Satellite cells in human skeletal muscle plasticity

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    Tim eSnijders

    2015-10-01

    Full Text Available Skeletal muscle satellite cells are considered to play a crucial role in muscle fiber maintenance, repair and remodelling. 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/remodelling 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/remodelling in the wider context of satellite cell biology whose literature is largely based on animal and cell models.

  12. Characterization of human satellite cells

    OpenAIRE

    Gloy, Sina

    2012-01-01

    Satellite cells guarantee the regeneration of skeletal muscle until old age. They are genuine muscle stem cells that are localized in a characteristic anatomic localization between basal lamina and sarkolemma of each muscle fiber. On protein level they are characterized by their expression of the transcription factor Pax7 and different other markers like c-Met and CXCR4. Most of our knowledge is based on studies with mouse models. Due to their availability and remarkable capacity to regen...

  13. Human placental trophoblast invasion and differentiation: a particular focus on Wnt signalling

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    Martin eKnöfler

    2013-09-01

    Full Text Available Wingless ligands, a family of secreted proteins, are critically involved in organ development and tissue homeostasis by ensuring balanced rates of stem cell proliferation, cell death and differentiation. Wnt signalling components also play crucial roles in murine placental development controlling trophoblast lineage determination, chorioallantoic fusion and placental branching morphogenesis. However, the role of the pathway in human placentation, trophoblast development and differentiation is only partly understood. Here, we summarize our present knowledge about Wnt signalling in the human placenta and discuss its potential role in physiological and aberrant trophoblast invasion, gestational diseases and choriocarcinoma formation. Differentiation of proliferative first trimester cytotrophoblasts into invasive extravillous trophoblasts is associated with nuclear recruitment of β-catenin and induction of Wnt-dependent T-cell factor 4 suggesting that canonical Wnt signalling could be important for the formation and function of extravillous trophoblasts. Indeed, activation of the pathway was shown to promote trophoblast invasion in different in vitro trophoblast model systems as well as trophoblast cell fusion. Methylation-mediated silencing of inhibitors of Wnt signalling provided evidence for epigenetic activation of the pathway in placental tissues and choriocarcinoma cells. Similarly, abundant nuclear expression of β-catenin in invasive trophoblasts of complete hydatidiform moles suggested a role for hyper-activated Wnt signalling. In contrast, upregulation of Wnt inhibitors was noticed in placentae of women with preeclampsia, a disease characterized by shallow trophoblast invasion and incomplete spiral artery remodelling. Moreover, changes in Wnt signalling have been observed upon cytomegalovirus infection and in recurrent abortions. In summary, the current literature suggests a critical role of Wnt signalling in physiological and abnormal

  14. Human neutrophils facilitate tumor cell transendothelial migration.

    LENUS (Irish Health Repository)

    Wu, Q D

    2012-02-03

    Tumor cell extravasation plays a key role in tumor metastasis. However, the precise mechanisms by which tumor cells migrate through normal vascular endothelium remain unclear. In this study, using an in vitro transendothelial migration model, we show that human polymorphonuclear neutrophils (PMN) assist the human breast tumor cell line MDA-MB-231 to cross the endothelial barrier. We found that tumor-conditioned medium (TCM) downregulated PMN cytocidal function, delayed PMN apoptosis, and concomitantly upregulated PMN adhesion molecule expression. These PMN treated with TCM attached to tumor cells and facilitated tumor cell migration through different endothelial monolayers. In contrast, MDA-MB-231 cells alone did not transmigrate. FACScan analysis revealed that these tumor cells expressed high levels of intercellular adhesion molecule-1 (ICAM-1) but did not express CD11a, CD11b, or CD18. Blockage of CD11b and CD18 on PMN and of ICAM-1 on MDA-MB-231 cells significantly attenuated TCM-treated, PMN-mediated tumor cell migration. These tumor cells still possessed the ability to proliferate after PMN-assisted transmigration. These results indicate that TCM-treated PMN may serve as a carrier to assist tumor cell transendothelial migration and suggest that tumor cells can exploit PMN and alter their function to facilitate their extravasation.

  15. Human neutrophils facilitate tumor cell transendothelial migration.

    Science.gov (United States)

    Wu, Q D; Wang, J H; Condron, C; Bouchier-Hayes, D; Redmond, H P

    2001-04-01

    Tumor cell extravasation plays a key role in tumor metastasis. However, the precise mechanisms by which tumor cells migrate through normal vascular endothelium remain unclear. In this study, using an in vitro transendothelial migration model, we show that human polymorphonuclear neutrophils (PMN) assist the human breast tumor cell line MDA-MB-231 to cross the endothelial barrier. We found that tumor-conditioned medium (TCM) downregulated PMN cytocidal function, delayed PMN apoptosis, and concomitantly upregulated PMN adhesion molecule expression. These PMN treated with TCM attached to tumor cells and facilitated tumor cell migration through different endothelial monolayers. In contrast, MDA-MB-231 cells alone did not transmigrate. FACScan analysis revealed that these tumor cells expressed high levels of intercellular adhesion molecule-1 (ICAM-1) but did not express CD11a, CD11b, or CD18. Blockage of CD11b and CD18 on PMN and of ICAM-1 on MDA-MB-231 cells significantly attenuated TCM-treated, PMN-mediated tumor cell migration. These tumor cells still possessed the ability to proliferate after PMN-assisted transmigration. These results indicate that TCM-treated PMN may serve as a carrier to assist tumor cell transendothelial migration and suggest that tumor cells can exploit PMN and alter their function to facilitate their extravasation.

  16. Human hair genealogies and stem cell latency

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    Tavaré Simon

    2006-02-01

    Full Text Available Abstract Background Stem cells divide to reproduce themselves and produce differentiated progeny. A fundamental problem in human biology has been the inability to measure how often stem cells divide. Although it is impossible to observe every division directly, one method for counting divisions is to count replication errors; the greater the number of divisions, the greater the numbers of errors. Stem cells with more divisions should produce progeny with more replication errors. Methods To test this approach, epigenetic errors (methylation in CpG-rich molecular clocks were measured from human hairs. Hairs exhibit growth and replacement cycles and "new" hairs physically reappear even on "old" heads. Errors may accumulate in long-lived stem cells, or in their differentiated progeny that are eventually shed. Results Average hair errors increased until two years of age, and then were constant despite decades of replacement, consistent with new hairs arising from infrequently dividing bulge stem cells. Errors were significantly more frequent in longer hairs, consistent with long-lived but eventually shed mitotic follicle cells. Conclusion Constant average hair methylation regardless of age contrasts with the age-related methylation observed in human intestine, suggesting that error accumulation and therefore stem cell latency differs among tissues. Epigenetic molecular clocks imply similar mitotic ages for hairs on young and old human heads, consistent with a restart with each new hair, and with genealogies surreptitiously written within somatic cell genomes.

  17. Poliovirus persistence in human cells in vitro.

    Science.gov (United States)

    Colbère-Garapin, F; Jacques, S; Drillet, A S; Pavio, N; Couderc, T; Blondel, B; Pelletier, I

    2001-01-01

    Poliovirus (PV) can persist in vivo in the intestine of immunocompromised hosts for years. Moreover, immunocompetent individuals who have survived paralytic poliomyelitis sometimes develop the post-poliomyelitis syndrome (PPS), consisting of a variety of symptoms including new muscular atrophies. PPS may be due to PV persistence. We have developed models of PV persistence in neural cells and epidermoid cells. Cell determinants are of crucial importance for the establishment of persistent infections in human neuronal cells, whereas viral determinants play the primary role in human epidermoid HEp-2 cells. The results obtained with these in vitro models show the capacity of PV to persist and reveal a virus and cell co-evolution involving PV-receptor interactions. In addition, they suggest that several mechanisms are used by PV to establish and maintain persistent infections.

  18. Interaction of Staphylococci with Human B cells.

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    Tyler K Nygaard

    Full Text Available 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.

  19. Human ES cells: Starting Culture from Frozen Cells

    OpenAIRE

    Trish, Erin; Dimos, John; Eggan, Kevin

    2006-01-01

    Here we demonstrate how our lab begins a HuES human embryonic stem cell line culture from a frozen stock. First, a one to two day old ten cm plate of approximately one (to two) million irradiated mouse embryonic fibroblast feeder cells is rinsed with HuES media to remove residual serum and cell debris, and then HuES media added and left to equilibrate in the cell culture incubator. A frozen vial of cells from long term liquid nitrogen storage or a -80C freezer is sourced and quickly submer...

  20. Cannabidiol changes P-gp and BCRP expression in trophoblast cell lines

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    Valeria Feinshtein

    2013-09-01

    Full Text Available Objectives. Marijuana is the most commonly used illicit drug during pregnancy. Due to high lipophilicity, cannabinoids can easily penetrate physiological barriers like the human placenta and jeopardize the developing fetus. We evaluated the impact of cannabidiol (CBD, a major non-psychoactive cannabinoid, on P-glycoprotein (P-gp and Breast Cancer Resistance Protein (BCRP expression, and P-gp function in a placental model, BeWo and Jar choriocarcinoma cell lines (using P-gp induced MCF7 cells (MCF7/P-gp for comparison. Study design. Following the establishment of the basal expression of these transporters in the membrane fraction of all three cell lines, P-gp and BCRP protein and mRNA levels were determined following chronic (24–72 h exposure to CBD, by Western Blot and qPCR. CBD impact on P-gp efflux function was examined by uptake of specific P-gp fluorescent substrates (calcein-AM, DiOC2(3 and rhodamine123(rh123. Cyclosporine A (CsA served as a positive control. Results. Chronic exposure to CBD resulted in significant changes in the protein and mRNA levels of both transporters. While P-gp was down-regulated, BCRP levels were up-regulated in the choriocarcinoma cell lines. CBD had a remarkably different influence on P-gp and BCRP expression in MCF7/P-gp cells, demonstrating that these are cell type specific effects. P-gp dependent efflux (of calcein, DiOC2(3 and rh123 was inhibited upon short-term exposure to CBD. Conclusions. Our study shows that CBD might alter P-gp and BCRP expression in the human placenta, and inhibit P-gp efflux function. We conclude that marijuana use during pregnancy may reduce placental protective functions and change its morphological and physiological characteristics.

  1. Autophagy in human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Thien Tra

    Full Text Available Autophagy (macroautophagy is a degradative process that involves the sequestration of cytosolic material including organelles into double membrane vesicles termed autophagosomes for delivery to the lysosome. Autophagy is essential for preimplantation development of mouse embryos and cavitation of embryoid bodies. The precise roles of autophagy during early human embryonic development, remain however largely uncharacterized. Since human embryonic stem cells constitute a unique model system to study early human embryogenesis we investigated the occurrence of autophagy in human embryonic stem cells. We have, using lentiviral transduction, established multiple human embryonic stem cell lines that stably express GFP-LC3, a fluorescent marker for the autophagosome. Each cell line displays both a normal karyotype and pluripotency as indicated by the presence of cell types representative of the three germlayers in derived teratomas. GFP expression and labelling of autophagosomes is retained after differentiation. Baseline levels of autophagy detected in cultured undifferentiated hESC were increased or decreased in the presence of rapamycin and wortmannin, respectively. Interestingly, autophagy was upregulated in hESCs induced to undergo differentiation by treatment with type I TGF-beta receptor inhibitor SB431542 or removal of MEF secreted maintenance factors. In conclusion we have established hESCs capable of reporting macroautophagy and identify a novel link between autophagy and early differentiation events in hESC.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  3. Quercetin Inhibits Cell Migration and Invasion in Human Osteosarcoma Cells.

    Science.gov (United States)

    Lan, Haifeng; Hong, Wei; Fan, Pan; Qian, Dongyang; Zhu, Jianwei; Bai, Bo

    2017-01-01

    Osteosarcoma is a malignant tumor associated with high mortality; however, no effective therapies for the disease have been developed. Several studies have focused on elucidating the pathogenesis of osteosarcoma and have aimed to develop novel therapies for the disease. Quercetin is a vital dietary flavonoid that has been shown to have a variety of anticancer effects, as it induces cell cycle arrest, apoptosis, and differentiation and is involved in cell adhesion, metastasis and angiogenesis. Herein, we aimed to investigate the effects of quercetin on osteosarcoma migration and invasion in vitro and in vivo and to explore the molecular mechanisms underlying its effects on osteosarcoma migration and invasion. Cell viability, cell cycle activity and cell apoptosis were measured using CCK-8 assay and flow cytometry, and cell migration and invasion were evaluated by wound healing and transwell assays, respectively. The mRNA and protein expression levels of several proteins of interest were assessed by real-time quantitative PCR and western blotting, respectively. Moreover, a nude mouse model of human osteosarcoma lung metastasis was established to assess the anti-metastatic effects of quercetin in vivo. We noted no significant differences in cell cycle activity and apoptosis between HOS and MG63 cells and control cells. Treatment with quercetin significantly attenuated cell migration and invasion in HOS and MG63 cells compared with treatment with control medium. Moreover HIF-1α, VEGF, MMP2, and MMP9 mRNA and protein expression levels were significantly downregulated in HOS cells treated with quercetin compared with HOS cells treated with controls. Additionally, treatment with quercetin attenuated metastatic lung tumor formation and growth in the nude mouse model of osteosarcoma compared with treatment with controls. Our findings regarding the inhibitory effects of quercetin on cell migration and invasion suggest that quercetin may have potential as a therapy for human

  4. 3 CFR - Guidelines for Human Stem Cell Research

    Science.gov (United States)

    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 extent...

  5. Merkel cell distribution in the human eyelid

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    C.A. May

    2013-10-01

    Full Text Available Although Merkel cell carcinoma of the eye lid is reported frequently in the literature, only limited information exists about the distribution of Merkel cells in this tissue. Therefore, serial sections of 18 human cadaver eye lids (donors ages ranging between 63 and 97 years were stained for cytokeratin 20 in various planes. The overall appearance of Merkel cells in these samples was low and mainly located in the outer root layer of the cilia hair follicles. Merkel cells were more frequent in the middle, and almost not detectable at the nasal and temporal edges. The localization is in accordance with that of Merkel cell carcinoma, but concerning the scarce appearance within this adulthood group, a specific physiological role of these cells in the eye lid is difficult to establish.

  6. Cell pattern in adult human corneal endothelium.

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    Carlos H Wörner

    Full Text Available A review of the current data on the cell density of normal adult human endothelial cells was carried out in order to establish some common parameters appearing in the different considered populations. From the analysis of cell growth patterns, it is inferred that the cell aging rate is similar for each of the different considered populations. Also, the morphology, the cell distribution and the tendency to hexagonallity are studied. The results are consistent with the hypothesis that this phenomenon is analogous with cell behavior in other structures such as dry foams and grains in polycrystalline materials. Therefore, its driving force may be controlled by the surface tension and the mobility of the boundaries.

  7. Proliferation conditions for human satellite cells

    DEFF Research Database (Denmark)

    Gaster, M; Beck-Nielsen, H; Schrøder, H D

    2001-01-01

    Primary satellite cell cultures have become an important tool as a model system for skeletal muscles. A common problem in human satellite cell culturing is fibroblast overgrowth. We combined N-CAM (Leu19) immunocytochemical staining of satellite cells (Sc) with stereological methods to estimate...... the fraction of Sc in culture. Evaluation of different culture conditions allowed us to find proliferation conditions preferentially for Sc: a) Sc should be cultured on surfaces coated with ECM-gel. b) Primary cell culture should be inoculated in DMEM supplemented with 10% fetal calf serum to increase cell...... adherence. c) Change of media to DMEM supplemented with 2% Ultroser-G and 2% FCS after 24 h.d) Before subcultivation, cells should be preplated for 30 min. The fractional content of Sc in passage four when applying this method of cultivation was 0.82 +/- 0.07 (mean +/- SE, N = 10). Our method enabled us...

  8. Neocortical glial cell numbers in human brains

    DEFF Research Database (Denmark)

    Pelvig, D.P.; Pakkenberg, H.; Stark, A.K.

    2008-01-01

    Stereological cell counting was applied to post-mortem neocortices of human brains from 31 normal individuals, age 18-93 years, 18 females (average age 65 years, range 18-93) and 13 males (average age 57 years, range 19-87). The cells were differentiated in astrocytes, oligodendrocytes, microglia...... while the total astrocyte number is constant through life; finally males have a 28% higher number of neocortical glial cells and a 19% higher neocortical neuron number than females. The overall total number of neocortical neurons and glial cells was 49.3 billion in females and 65.2 billion in males......, a difference of 24% with a high biological variance. These numbers can serve as reference values in quantitative studies of the human neocortex. (C) 2007 Elsevier Inc. All rights reserved Udgivelsesdato: 2008/11...

  9. T-cell response in human leishmaniasis

    DEFF Research Database (Denmark)

    Kharazmi, A; Kemp, K; Ismail, A

    1999-01-01

    In the present communication we provide evidence for the existence of a Th1/Th2 dichotomy in the T-cell response to Leishmania antigens in human leishmaniasis. Our data suggest that the pattern of IL-4 and IFN-gamma response is polarised in these patients. Lymphocytes from individuals recovered...... from cutaneous leishmaniasis (CL) responded by IFN-gamma production following stimulation with Leishmania antigens whereas cells from patients recovered from visceral leishmaniasis (VL) showed a mixed pattern of IFN-gamma and IL-4 responses. The cells producing these cytokines were predominantly CD4......+. Furthermore, IL-10 plays an important role in the development of post kala azar dermal leishmaniasis (PKDL) from VL. The balance between the parasitic-specific T-cell response plays an important regulatory role in determining the outcome of Leishmania infections in humans....

  10. Lobaplatin arrests cell cycle progression in human hepatocellular carcinoma cells

    Directory of Open Access Journals (Sweden)

    Chen Chang-Jie

    2010-10-01

    Full Text Available Abstract Background Hepatocellular carcinoma (HCC still is a big burden for China. In recent years, the third-generation platinum compounds have been proposed as potential active agents for HCC. However, more experimental and clinical data are warranted to support the proposal. In the present study, the effect of lobaplatin was assessed in five HCC cell lines and the underlying molecular mechanisms in terms of cell cycle kinetics were explored. Methods Cytotoxicity of lobaplatin to human HCC cell lines was examined using MTT cell proliferation assay. Cell cycle distribution was determined by flow cytometry. Expression of cell cycle-regulated genes was examined at both the mRNA (RT-PCR and protein (Western blot levels. The phosphorylation status of cyclin-dependent kinases (CDKs and retinoblastoma (Rb protein was also examined using Western blot analysis. Results Lobaplatin inhibited proliferation of human HCC cells in a dose-dependent manner. For the most sensitive SMMC-7721 cells, lobaplatin arrested cell cycle progression in G1 and G2/M phases time-dependently which might be associated with the down-regulation of cyclin B, CDK1, CDC25C, phosphorylated CDK1 (pCDK1, pCDK4, Rb, E2F, and pRb, and the up-regulation of p53, p21, and p27. Conclusion Cytotoxicity of lobaplatin in human HCC cells might be due to its ability to arrest cell cycle progression which would contribute to the potential use of lobaplatin for the management of HCC.

  11. Human ES cells: starting culture from frozen cells.

    Science.gov (United States)

    Trish, Erin; Dimos, John; Eggan, Kevin

    2006-11-09

    Here we demonstrate how our lab begins a HuES human embryonic stem cell line culture from a frozen stock. First, a one to two day old ten cm plate of approximately one (to two) million irradiated mouse embryonic fibroblast feeder cells is rinsed with HuES media to remove residual serum and cell debris, and then HuES media added and left to equilibrate in the cell culture incubator. A frozen vial of cells from long term liquid nitrogen storage or a -80 C freezer is sourced and quickly submerged in a 37 C water bath for quick thawing. Cells in freezing media are then removed from the vial and placed in a large volume of HuES media. The large volume of HuES media facilitates removal of excess serum and DMSO, which can cause HuES human embryonic stem cells to differentiate. Cells are gently spun out of suspension, and then re-suspended in a small volume of fresh HuES media that is then used to seed the MEF plate. It is considered important to seed the MEF plate by gently adding the HuES cells in a drop wise fashion to evenly disperse them throughout the plate. The newly established HuES culture plate is returned to the incubator for 48 hrs before media is replaced, then is fed every 24 hours thereafter.

  12. Sodium Valproate Induces Cell Senescence in Human Hepatocarcinoma Cells

    Directory of Open Access Journals (Sweden)

    Hong-Mei An

    2013-12-01

    Full Text Available Hepatocarcinogenesis is associated with epigenetic changes, including histone deacetylases (HDACs. Epigenetic modulation by HDAC inhibition is a potentially valuable approach for hepatocellular carcinoma treatment. In present study, we evaluated the anticancer effects of sodium valproate (SVP, a known HDAC inhibitor, in human hepatocarcinoma cells. The results showed SVP inhibited the proliferation of Bel-7402 cells in a dose-dependent manner. Low dose SVP treatment caused a large and flat morphology change, positive SA-β-gal staining, and G0/G1 phase cell cycle arrest in human hepatocarcinoma cells. Low dose SVP treatment also increased acetylation of histone H3 and H4 on p21 promoter, accompanied by up-regulation of p21 and down-regulation of RB phosphorylation. These observations suggested that a low dose of SVP could induce cell senescence in hepatocarcinoma cells, which might correlate with hyperacetylation of histone H3 and H4, up-regulation of p21, and inhibition of RB phosphorylation. Since the effective concentration inducing cell senescence in hepatocarcinoma cells is clinically available, whether a clinical dose of SVP could induce cell senescence in clinical hepatocarcinoma is worthy of further study.

  13. Human Neural Cell-Based Biosensor

    Science.gov (United States)

    2013-05-28

    Orlando R, Stice SL. Membrane proteomic signatures of karyotypically normal and abnormal human embryonic stem cell lines and derivatives. Proteomics. 2011...format (96-,384-well) assays, 2) grow as adherent monolayers, and 3) possess a stable karyotype for multiple (>10) passages with a doubling time of ~36...derived neural progenitor cell line working stock has been amplified, characterized for karyotype and evaluated for the expression of neural progenitor

  14. Mast cells in human airways: the culprit?

    Directory of Open Access Journals (Sweden)

    Jonas S. Erjefält

    2014-09-01

    Full Text Available By virtue of their undisputed role in allergy, the study of airway mast cells has focused on nasal and bronchial mast cells and their involvement in allergic rhinitis and asthma. However, recent mechanistic and human studies suggest that peripheral mast cells also have an important role in asthma, as well as chronic obstructive pulmonary disease, respiratory infections and lung fibrosis. Pathogenic roles include immune-modulatory, pro-inflammatory and pro-fibrotic activities. Importantly, mast cells also actively downregulate inflammation and participate in the defence against respiratory infections. Another complicating factor is the notorious mast cell heterogeneity, where each anatomical compartment of the lung harbours site-specific mast cell populations. Alveolar mast cells stand out as they lack the cardinal expression of the high affinity IgE receptor. Supporting the emerging concept of alveolar inflammation in asthma, alveolar mast cells shift to a highly FcϵRI-expressing phenotype in uncontrolled asthma. Site-specific and disease-associated mast cell changes have also recently been described in most other inflammatory conditions of the lung. Thus, in the exploration of new anti-mast cell treatment strategies the search has widened to include the lung periphery and the delicate task of identifying which of the countless potential roles are the critical disease modifying ones in a given clinical situation.

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

    Science.gov (United States)

    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. Copyright © 2014 Elsevier Ltd. All rights reserved.

  16. Human pancreatic islet progenitor cells demonstrate phenotypic ...

    Indian Academy of Sciences (India)

    2009-04-24

    Apr 24, 2009 ... Phenotypic plasticity is a phenomenon that describes the occurrence of 2 or more distinct phenotypes under diverse conditions. This article discusses the work carried out over the past few years in understanding the potential of human pancreatic islet-derived progenitors for cell replacement therapy in ...

  17. Melanopsin expressing human retinal ganglion cells

    DEFF Research Database (Denmark)

    Hannibal, Jens; Christensen, Anders Tolstrup; Heegaard, Steffen

    2017-01-01

    microscopy and 3D reconstruction of melanopsin immunoreactive (-ir) RGCs, we applied the criteria used in mouse on human melanopsin-ir RGCs. We identified M1, displaced M1, M2 and M4 cells. We found two other subtypes of melanopsin-ir RGCs, which were named "gigantic M1 (GM1)" and "gigantic displaced M1 (GDM...

  18. Human CD56bright NK Cells

    DEFF Research Database (Denmark)

    Michel, Tatiana; Poli, Aurélie; Cuapio, Angelica

    2016-01-01

    Human NK cells can be subdivided into various subsets based on the relative expression of CD16 and CD56. In particular, CD56(bright)CD16(-/dim) NK cells are the focus of interest. They are considered efficient cytokine producers endowed with immunoregulatory properties, but they can also become...... cytotoxic upon appropriate activation. These cells were shown to play a role in different disease states, such as cancer, autoimmunity, neuroinflammation, and infection. Although their phenotype and functional properties are well known and have been extensively studied, their lineage relationship with other...... NK cell subsets is not fully defined, nor is their precise hematopoietic origin. In this article, we summarize recent studies about CD56(bright) NK cells in health and disease and briefly discuss the current controversies surrounding them....

  19. Human embryonic stem cells: preclinical perspectives

    Directory of Open Access Journals (Sweden)

    Sarda Kanchan

    2008-01-01

    Full Text Available Abstract Human embryonic stem cells (hESCs have been extensively discussed in public and scientific communities for their potential in treating diseases and injuries. However, not much has been achieved in turning them into safe therapeutic agents. The hurdles in transforming hESCs to therapies start right with the way these cells are derived and maintained in the laboratory, and goes up-to clinical complications related to need for patient specific cell lines, gender specific aspects, age of the cells, and several post transplantation uncertainties. The different types of cells derived through directed differentiation of hESC and used successfully in animal disease and injury models are described briefly. This review gives a brief outlook on the present and the future of hESC based therapies, and talks about the technological advances required for a safe transition from laboratory to clinic.

  20. Susceptibility of human liver cells to porcine endogenous retrovirus.

    Science.gov (United States)

    Lin, Xinzi; Qi, Lin; Li, Zhiguo; Chi, Hao; Lin, Wanjun; Wang, Yan; Jiang, Zesheng; Pan, Mingxin; Gao, Yi

    2013-12-01

    The risk of porcine endogenous retrovirus infection is a major barrier for pig-to-human xenotransplant. Porcine endogenous retrovirus, present in porcine cells, can infect many human and nonhuman primate cells in vitro, but there is no evidence available about in vitro infection of human liver cells. We investigated the susceptibility of different human liver cells to porcine endogenous retrovirus. The supernatant from a porcine kidney cell line was added to human liver cells, including a normal hepatocyte cell line (HL-7702 cells), primary hepatocytes (Phh cells), and a liver stellate cell line (Lx-2 cells), and to human embryonic kidney cells as a reference control. Expression of the porcine endogenous retrovirus antigen p15E in the human cells was evaluated with polymerase chain reaction, reverse transcription-polymerase chain reaction, and Western blot. The porcine endogenous retrovirus antigen p15E was not expressed in any human liver cells (HL-7702, Phh, or Lx-2 cells) that had been exposed to supernatants from porcine kidney cell lines. Porcine endogenous retrovirus-specific fragments were amplified in human kidney cells. Human liver cells tested were not susceptible to infection by porcine endogenous retrovirus. Therefore, not all human cells are susceptible to porcine endogenous retrovirus.

  1. Dose-dependent and cell type-specific cell death and proliferation following in vitro exposure to radial extracorporeal shock waves

    Science.gov (United States)

    Hochstrasser, Tanja; Frank, Hans-Georg; Schmitz, Christoph

    2016-01-01

    Radial extracorporeal shock wave (rESW) therapy is widely used in musculoskeletal disorders and wound repair. However, the mechanisms of action are still largely unknown. The current study compared the effects of rESWs on two cell types. Human fetal foreskin fibroblasts (HFFF2) and human placental choriocarcinoma cell line JEG-3 were exposed to 0, 100, 200, 500 or 5000 rESWs generated with a Swiss DolorClast device (2.5 bar, 1 Hz). FACS analysis immediately after rESW exposure showed that initially, rESWs rather induced mechanical cell destruction than regulated or programmed cell death. Cell damage was nearly negated by reducing cavitation. Furthermore, cell viability decreased progressively with higher numbers of rESWs. Exposure to rESWs had no impact on growth potential of JEG-3 cells, but dose-dependently increased growth potential of HFFF2 cells. Cultivation of cells that were initially exposed to sham-rESWs in conditioned media increased the growth potential of HFFF2 cells, nevertheless, an even stronger effect was achieved by direct exposure to rESWs. Additionally, cell cycle distribution analysis demonstrated a shift in proportion from G0/G1 to G2/M phase in HFFF2 cells, but not in JEG-3 cells. These data demonstrate that rESWs leads to initial and subsequent dose-dependent and cell type-specific effects in vitro. PMID:27477873

  2. Activation of Human T Cells in Hypertension: Studies of Humanized Mice and Hypertensive Humans.

    Science.gov (United States)

    Itani, Hana A; McMaster, William G; Saleh, Mohamed A; Nazarewicz, Rafal R; Mikolajczyk, Tomasz P; Kaszuba, Anna M; Konior, Anna; Prejbisz, Aleksander; Januszewicz, Andrzej; Norlander, Allison E; Chen, Wei; Bonami, Rachel H; Marshall, Andrew F; Poffenberger, Greg; Weyand, Cornelia M; Madhur, Meena S; Moore, Daniel J; Harrison, David G; Guzik, Tomasz J

    2016-07-01

    Emerging evidence supports an important role for T cells in the genesis of hypertension. Because this work has predominantly been performed in experimental animals, we sought to determine whether human T cells are activated in hypertension. We used a humanized mouse model in which the murine immune system is replaced by the human immune system. Angiotensin II increased systolic pressure to 162 versus 116 mm Hg for sham-treated animals. Flow cytometry of thoracic lymph nodes, thoracic aorta, and kidney revealed increased infiltration of human leukocytes (CD45(+)) and T lymphocytes (CD3(+) and CD4(+)) in response to angiotensin II infusion. Interestingly, there was also an increase in the memory T cells (CD3(+)/CD45RO(+)) in the aortas and lymph nodes. Prevention of hypertension using hydralazine and hydrochlorothiazide prevented the accumulation of T cells in these tissues. Studies of isolated human T cells and monocytes indicated that angiotensin II had no direct effect on cytokine production by T cells or the ability of dendritic cells to drive T-cell proliferation. We also observed an increase in circulating interleukin-17A producing CD4(+) T cells and both CD4(+) and CD8(+) T cells that produce interferon-γ in hypertensive compared with normotensive humans. Thus, human T cells become activated and invade critical end-organ tissues in response to hypertension in a humanized mouse model. This response likely reflects the hypertensive milieu encountered in vivo and is not a direct effect of the hormone angiotensin II. © 2016 American Heart Association, Inc.

  3. How to make a human germ cell

    Directory of Open Access Journals (Sweden)

    Paul S Cooke

    2015-06-01

    Full Text Available How the primordial germ cell (PGC lineage, which eventually gives rise to spermatozoa in males and oocytes in females, is established in the developing mammalian embryo has been a critical topic in both developmental and reproductive biology for many years. There have been significant breakthroughs over the past two decades in establishing both the source of PGCs and the factors that regulate the specification of this lineage in mice, [1] but our understanding of the factors that control PGC development in the human is rudimentary. The SRY-related HMG-box (SOX family of transcription factors consists of 20 genes in humans and mice that are involved in the maintenance of pluripotency, male sexual development, and other processes. A recent paper in Cell has identified one member of this family, SOX17, as an essential factor for inducing the PGC lineage in humans. [2] Surprisingly, this protein does not appear to have a role in PGC specification in mice. This work not only introduces a new and important player to the field of germ cell specification, but also emphasizes the uniqueness of human PGC development compared to more extensively studied mouse models.

  4. 21 CFR 864.2280 - Cultured animal and human cells.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Cultured animal and human cells. 864.2280 Section... (CONTINUED) MEDICAL DEVICES HEMATOLOGY AND PATHOLOGY DEVICES Cell And Tissue Culture Products § 864.2280 Cultured animal and human cells. (a) Identification. Cultured animal and human cells are in vitro...

  5. Human somatic cell nuclear transfer and cloning.

    Science.gov (United States)

    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. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  6. Human induced pluripotent stem cell-derived models to investigate human cytomegalovirus infection in neural cells.

    Directory of Open Access Journals (Sweden)

    Leonardo D'Aiuto

    Full Text Available Human cytomegalovirus (HCMV infection is one of the leading prenatal causes of congenital mental retardation and deformities world-wide. Access to cultured human neuronal lineages, necessary to understand the species specific pathogenic effects of HCMV, has been limited by difficulties in sustaining primary human neuronal cultures. Human induced pluripotent stem (iPS cells now provide an opportunity for such research. We derived iPS cells from human adult fibroblasts and induced neural lineages to investigate their susceptibility to infection with HCMV strain Ad169. Analysis of iPS cells, iPS-derived neural stem cells (NSCs, neural progenitor cells (NPCs and neurons suggests that (i iPS cells are not permissive to HCMV infection, i.e., they do not permit a full viral replication cycle; (ii Neural stem cells have impaired differentiation when infected by HCMV; (iii NPCs are fully permissive for HCMV infection; altered expression of genes related to neural metabolism or neuronal differentiation is also observed; (iv most iPS-derived neurons are not permissive to HCMV infection; and (v infected neurons have impaired calcium influx in response to glutamate.

  7. Colony forming cell (CFC) assay for human hematopoietic cells.

    Science.gov (United States)

    Sarma, Nayan J; Takeda, Akiko; Yaseen, Nabeel R

    2010-12-18

    Human hematopoietic stem/progenitor cells are usually obtained from bone marrow, cord blood, or peripheral blood and are used to study hematopoiesis and leukemogenesis. They have the capacity to differentiate into lymphoid and myeloid lineages. The colony forming cell (CFC) assay is used to study the proliferation and differentiation pattern of hematopoietic progenitors by their ability to form colonies in a semisolid medium. The number and the morphology of the colonies formed by a fixed number of input cells provide preliminary information about the ability of progenitors to differentiate and proliferate. Cells can be harvested from individual colonies or from the whole plate to further assess their numbers and differentiation states using flow cytometry and morphologic evaluation of Giemsa-stained slides. This assay is useful for assessing myeloid but not lymphoid differentiation. The term myeloid in this context is used in its wider sense to encompass granulocytic, monocytic, erythroid, and megakaryocytic lineages. We have used this assay to assess the effects of oncogenes on the differentiation of primary human CD34+ cells derived from peripheral blood. For this purpose cells are transduced with either control retroviral construct or a construct expressing the oncogene of interest, in this case NUP98-HOXA9. We employ a commonly used retroviral vector, MSCV-IRES-GFP, that expresses a bicistronic mRNA that produces the gene of interest and a GFP marker. Cells are pre-activated by growing in the presence of cytokines for two days prior to retroviral transduction. After another two days, GFP+ cells are isolated by fluorescence-activated cell sorting (FACS) and mixed with a methylcellulose-containing semisolid medium supplemented with cytokines and incubated till colonies appear on the surface, typically 14 days. The number and morphology of the colonies are documented. Cells are then removed from the plates, washed, counted, and subjected to flow cytometry and

  8. The core regulatory network in human cells.

    Science.gov (United States)

    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. Copyright © 2017 Elsevier Inc. All rights reserved.

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

    Science.gov (United States)

    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.

  10. Cell Culture Assay for Human Noroviruses [response

    Energy Technology Data Exchange (ETDEWEB)

    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

  11. Characterizing motility dynamics in human RPE cells

    Science.gov (United States)

    Liu, Zhuolin; Kurokawa, Kazuhiro; Zhang, Furu; Miller, Donald T.

    2017-02-01

    Retinal pigment epithelium (RPE) cells are vital to health of the outer retina, however, are often compromised in ageing and ocular diseases that lead to blindness. Early manifestation of RPE disruption occurs at the cellular level, but while in vivo biomarkers at this scale hold considerable promise, RPE cells have proven extremely challenging to image in the living human eye. Recently we addressed this problem by using organelle motility as a novel contrast agent to enhance the RPE cell in conjunction with 3D resolution of adaptive optics-optical coherence tomography (AO-OCT) to section the RPE layer. In this study, we expand on the central novelty of our method - organelle motility - by characterizing the dynamics of the motility in individual RPE cells, important because of its direct link to RPE physiology. To do this, AO-OCT videos of the same retinal patch were acquired at approximately 1 min intervals or less, time stamped, and registered in 3D with sub-cellular accuracy. Motility was quantified by an exponential decay time constant, the time for motility to decorrelate the speckle field across an RPE cell. In two normal subjects, we found the decay time constant to be just 3 seconds, thus indicating rapid motility in normal RPE cells.

  12. Human dendritic cell culture and bacterial infection.

    Science.gov (United States)

    Jones, Hannah E; Klein, Nigel; Dixon, Garth L J

    2012-01-01

    Dendritic cells (DC) play a key role in the development of natural immunity to microbes. The DC form a bridge between the innate and adaptive immune system by providing key instructions particularly to antigen naïve T-cells. The interaction of DC with T lymphocytes involves three signals: (1) antigen processing and presentation in context of MHC Class I and/or II, (2) expression of T cell co-stimulatory molecules, and (3) cytokine production. Studying the interactions of DCs with specific pathogens allows for better understanding of how protective immunity is generated, and may be particularly useful for assessing vaccine components. In this chapter, we describe methods to generate human monocyte-derived DCs and assess their maturation, activation, and function, using interaction with the gram-negative bacterial pathogen Neisseria meningitidis as a model.

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

    Directory of Open Access Journals (Sweden)

    Hayato Fukusumi

    2016-01-01

    Full Text Available 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.

  14. Functional Cardiomyocytes Derived From Human Induced Pluripotent Stem Cells

    National Research Council Canada - National Science Library

    Zhang, Jianhua; Wilson, Gisela F; Soerens, Andrew G; Koonce, Chad H; Yu, Junying; Palecek, Sean P; Thomson, James A; Kamp, Timothy J

    2009-01-01

    Human induced pluripotent stem (iPS) cells hold great promise for cardiovascular research and therapeutic applications, but the ability of human iPS cells to differentiate into functional cardiomyocytes has not yet been demonstrated...

  15. Molecular aging and rejuvenation of human muscle stem cells

    OpenAIRE

    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 ...

  16. Markers of T Cell Senescence in Humans

    Directory of Open Access Journals (Sweden)

    Weili Xu

    2017-08-01

    Full Text Available Many countries are facing the aging of their population, and many more will face a similar obstacle in the near future, which could be a burden to many healthcare systems. Increased susceptibility to infections, cardiovascular and neurodegenerative disease, cancer as well as reduced efficacy of vaccination are important matters for researchers in the field of aging. As older adults show higher prevalence for a variety of diseases, this also implies higher risk of complications, including nosocomial infections, slower recovery and sequels that may reduce the autonomy and overall quality of life of older adults. The age-related effects on the immune system termed as “immunosenescence” can be exemplified by the reported hypo-responsiveness to influenza vaccination of the elderly. T cells, which belong to the adaptive arm of the immune system, have been extensively studied and the knowledge gathered enables a better understanding of how the immune system may be affected after acute/chronic infections and how this matters in the long run. In this review, we will focus on T cells and discuss the surface and molecular markers that are associated with T cell senescence. We will also look at the implications that senescent T cells could have on human health and diseases. Finally, we will discuss the benefits of having these markers for investigators and the future work that is needed to advance the field of T cell senescence markers.

  17. Human Embryonic Stem Cell Research Debates: A Confucian Argument

    National Research Council Canada - National Science Library

    D. F.-C. Tsai

    2005-01-01

    Human embryonic stem cell research can bring about major biomedical breakthroughs and thus contribute enormously to human welfare, yet it raises serious moral problems because it involves using human...

  18. Regulatory networks define phenotypic classes of human stem cell lines

    OpenAIRE

    Müller, Franz-Josef; Laurent, Louise C.; Kostka, Dennis; Ulitsky, Igor; Williams, Roy; Lu, Christina; Park, In-Hyun; Rao, Mahendra?S.; Shamir, Ron; Schwartz, Philip H.; Schmidt, Nils O.; Loring, Jeanne F.

    2008-01-01

    Stem cells are defined as self-renewing cell populations that can differentiate into multiple distinct cell types. However, hundreds of different human cell lines from embryonic, fetal, and adult sources have been called stem cells, even though they range from pluripotent cells, typified by embryonic stem cells, which are capable of virtually unlimited proliferation and differentiation, to adult stem cell lines, which can generate a far more limited repertory of differentiated cell types. The...

  19. Human embryonic stem cells and microenvironment

    Directory of Open Access Journals (Sweden)

    Banu İskender

    2014-09-01

    Full Text Available Human embryonic stem cells (hESCs possess a great potential in the field of regenerative medicine by their virtue of pluripotent potential with indefinite proliferation capabilities. They can self renew themselves and differentiate into three embryonic germ layers. Although they are conventionally grown on mitotically inactivated mouse feeder cells, there are in vitro culture systems utilizing feeder cells of human origin in order to prevent cross-species contamination. Recently established in vitro culture systems suggested that direct interaction with feeder cells is not necessary but rather attachment to a substrate is required to ensure long-term, efficient hESC culture in vitro. This substrate is usually composed of a mixture of extracellular matrix components representing in vivo natural niche. In hESC biology, the mechanism of interaction of hESCs with extracellular matrix molecules remained insufficiently explored area of research due to their transient nature of interaction with the in vivo niche. However, an in vitro culture system established using extracellular matrix molecules may provide a safer alternative to culture systems with feeder cells while paving the way to Good Manufacturing Practice-GMP production of hESCs for therapeutic purposes. Therefore, it is essential to study the interaction of extracellular matrix molecules with hESCs in order to standardize in vitro culture systems for large-scale production of hESCs in a less labor-intensive way. This would not only provide valuable information regarding the mechanisms that control pluripotency but also serve to dissect the molecular signaling pathways of directed differentiation for prospective therapeutic applications in the future. J Clin Exp Invest 2014; 5 (3: 486-495

  20. Arecoline is cytotoxic for human endothelial cells.

    Science.gov (United States)

    Ullah, Mafaz; Cox, Stephen; Kelly, Elizabeth; Boadle, Ross; Zoellner, Hans

    2014-11-01

    Oral submucous fibrosis is a pre-malignant fibrotic condition caused by areca nut use and involves reduced mucosal vascularity. Arecoline is the principal areca nut alkaloid and is cytotoxic for epithelium and fibroblasts. Endothelial cell cycle arrest is reported on exposure to arecoline, as is cytotoxicity for endothelial-lung carcinoma hybrid cells. We here describe cytotoxicity for primary human endothelial cultures from seven separate donors. Human umbilical vein endothelial cells were exposed to increasing concentrations of arecoline and examined by: phase-contrast microscopy, haemocytometer counts, transmission electron microscopy, lactate dehydrogenase release and the methyl-thiazol-tetrazolium assay. Vacuolation and detachment of endothelium were observed at and above arecoline concentrations of 333 μg/ml or more. Ultrastructural features of cellular stress were seen after 24-h treatment with 111 μg/ml arecoline and included reduced ribosomal studding of endoplasmic reticulum, increased autophagolysosomal structures, increased vacuolation and reduced mitochondrial cristae with slight swelling. Similar changes were seen at 4 h with arecoline at 333 μg/ml or above, but with more severe mitochondrial changes including increased electron density of mitochondrial matrix and greater cristal swelling, while by 24 h, these cells were frankly necrotic. Haemocytometer counts were paralleled by both lactate dehydrogenase release and the methyl-thiazol-tetrazolium assays. Arecoline is cytotoxic via necrosis for endothelium, while biochemical assays indicate no appreciable cellular leakage before death and detachment, as well as no clear effect on mitochondrial function in viable cells. Arecoline toxicity may thus contribute to reduced vascularity in oral submucous fibrosis. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  1. ICAM-3 activation modulates cell-cell contacts of human bone marrow endothelial cells

    NARCIS (Netherlands)

    van Buul, J. D.; Mul, F. P. J.; van der Schoot, C. E.; Hordijk, P. L.

    2004-01-01

    The Ig-like cell adhesion molecule ICAM-3 is mainly expressed on human leukocytes and is involved in cell-cell interactions. Its expression on endothelium is observed during disorders such as Crohn's disease and in solid tumors. We found low but detectable expression of ICAM-3 on VE-cadherin-

  2. Human embryonic stem cells and patent protection

    Directory of Open Access Journals (Sweden)

    Radovanović Sanja M.

    2015-01-01

    Full Text Available Given the importance of biotechnological research in modern diagnostics and therapeutics, on the one hand, and stimulative function of a patent, on the other hand, this work deals with the question of the possibility of pa-tent protection of human embryonic stem cells. Taking into account that this is a biotechnological invention, the key question that this paper highlights is the interpretation of the provisions of their patentability. Namely, thanks to the advanced methods of isolation, purification and preparation for implementation, modern patent systems do not exclude a priori living organisms from patent protection. Therefore, the analysis of representative administrative decisions or court rulings sought to define the criteria that would be applied in order to give patent protection to a certain biotechnological invention (stem cells while others do not.

  3. The similarity between human embryonic stem cell-derived epithelial cells and ameloblast-lineage cells.

    Science.gov (United States)

    Zheng, Li-Wei; Linthicum, Logan; DenBesten, Pamela K; Zhang, Yan

    2013-03-01

    This study aimed to compare epithelial cells derived from human embryonic stem cells (hESCs) to human ameloblast-lineage cells (ALCs), as a way to determine their potential use as a cell source for ameloblast regeneration. Induced by various concentrations of bone morphogenetic protein 4 (BMP4), retinoic acid (RA) and lithium chloride (LiCl) for 7 days, hESCs adopted cobble-stone epithelial phenotype (hESC-derived epithelial cells (ES-ECs)) and expressed cytokeratin 14. Compared with ALCs and oral epithelial cells (OE), ES-ECs expressed amelogenesis-associated genes similar to ALCs. ES-ECs were compared with human fetal skin epithelium, human fetal oral buccal mucosal epithelial cells and human ALCs for their expression pattern of cytokeratins as well. ALCs had relatively high expression levels of cytokeratin 76, which was also found to be upregulated in ES-ECs. Based on the present study, with the similarity of gene expression with ALCs, ES-ECs are a promising potential cell source for regeneration, which are not available in erupted human teeth for regeneration of enamel.

  4. The similarity between human embryonic stem cell-derived epithelial cells and ameloblast-lineage cells

    Science.gov (United States)

    Zheng, Li-Wei; Linthicum, Logan; DenBesten, Pamela K; Zhang, Yan

    2013-01-01

    This study aimed to compare epithelial cells derived from human embryonic stem cells (hESCs) to human ameloblast-lineage cells (ALCs), as a way to determine their potential use as a cell source for ameloblast regeneration. Induced by various concentrations of bone morphogenetic protein 4 (BMP4), retinoic acid (RA) and lithium chloride (LiCl) for 7 days, hESCs adopted cobble-stone epithelial phenotype (hESC-derived epithelial cells (ES-ECs)) and expressed cytokeratin 14. Compared with ALCs and oral epithelial cells (OE), ES-ECs expressed amelogenesis-associated genes similar to ALCs. ES-ECs were compared with human fetal skin epithelium, human fetal oral buccal mucosal epithelial cells and human ALCs for their expression pattern of cytokeratins as well. ALCs had relatively high expression levels of cytokeratin 76, which was also found to be upregulated in ES-ECs. Based on the present study, with the similarity of gene expression with ALCs, ES-ECs are a promising potential cell source for regeneration, which are not available in erupted human teeth for regeneration of enamel. PMID:23538640

  5. Eliminating acute lymphoblastic leukemia cells from human testicular cell cultures: a pilot study

    NARCIS (Netherlands)

    Sadri-Ardekani, Hooman; Homburg, Christa H.; van Capel, Toni M. M.; van den Berg, Henk; van der Veen, Fulco; van der Schoot, C. Ellen; van Pelt, Ans M. M.; Repping, Sjoerd

    2014-01-01

    To study whether acute lymphoblastic leukemia (ALL) cells survive in a human testicular cell culture system. Experimental laboratory study. Reproductive biology laboratory, academic medical center. Acute lymphoblastic leukemia cells from three patients and testicular cells from three other patients.

  6. Derivation of human embryonic stem cells in defined conditions.

    Science.gov (United States)

    Ludwig, Tenneille E; Levenstein, Mark E; Jones, Jeffrey M; Berggren, W Travis; Mitchen, Erika R; Frane, Jennifer L; Crandall, Leann J; Daigh, Christine A; Conard, Kevin R; Piekarczyk, Marian S; Llanas, Rachel A; Thomson, James A

    2006-02-01

    We have previously reported that high concentrations of basic fibroblast growth factor (bFGF) support feeder-independent growth of human embryonic stem (ES) cells, but those conditions included poorly defined serum and matrix components. Here we report feeder-independent human ES cell culture that includes protein components solely derived from recombinant sources or purified from human material. We describe the derivation of two new human ES cell lines in these defined culture conditions.

  7. Radiosensitivity of Human Melanoma Cell Lines

    Energy Technology Data Exchange (ETDEWEB)

    Bergoc, R. M.; Medina, V.; Cricco, G.; Mohamed, N.; Martin, G.; Nunez, M.; Croci, M.; Crescenti, E. J.; Rivera, E. S.

    2004-07-01

    Cutaneous melanoma is a skin cancer resulting from the malign transformation of skin-pigment cells, the melanocytes. The radiotherapy, alone or in combination with other treatment, is an important therapy for this disease. the objective of this paper was to determine in vitro the radiosensitivity of two human melanoma cell lines with different metastatic capability: WM35 and MI/15, and to study the effect of drugs on radiobiological parameters. The Survival Curves were adjusted to the mathematical Linear-quadratic model using GrapsPad Prism software. Cells were seeded in RPMI medium (3000-3500 cells/flask), in triplicate and irradiated 24 h later. The irradiation was performed using an IBL 437C H Type equipment (189 TBq, 7.7 Gy/min) calibrated with a TLD 700 dosimeter. The range of Doses covered from 0 to 10 Gy and the colonies formed were counted at day 7th post-irradiation. Results obtained were: for WM35, {alpha}=0.37{+-}0.07 Gy''-1 and {beta}=0.06{+-}0.02 Gy''-2, for M1/15m {alpha}=0.47{+-}0.03 Gy''-1 and {beta}=0.06{+-}0.01 Gy''-2. The {alpha}/{beta} values WM35: {alpha}/{beta} values WM35: {alpha}/{beta}=6.07 Gy and M1/15: {alpha}/{beta}{sub 7}.33 Gy were similar, independently of their metastatic capabillity and indicate that both lines exhibit high radioresistance. Microscopic observation of irradiated cells showed multinuclear cells with few morphologic changes non-compatible with apoptosis. By means of specific fluorescent dyes and flow cytometry analysis we determined the intracellular levels of the radicals superoxide and hydrogen peroxide and their modulation in response to ionizing radiation. The results showed a marked decreased in H{sub 2}O{sub 2} intracellular levels with a simultaneous increase in superoxide that will be part of a mechanism responsible for induction of cell radioresistance. This response triggered by irradiated cells could not be abrogated by different treatments like histamine or the

  8. Interindividual variation in human T regulatory cells

    Science.gov (United States)

    Ferraro, Alessandra; D’Alise, Anna Morena; Raj, Towfique; Asinovski, Natasha; Phillips, Roxanne; Ergun, Ayla; Replogle, Joseph M.; Bernier, Angelina; Laffel, Lori; Stranger, Barbara E.; De Jager, Philip L.; Mathis, Diane; Benoist, Christophe

    2014-01-01

    FOXP3+ regulatory T (Treg) cells enforce immune self-tolerance and homeostasis, and variation in some aspects of Treg function may contribute to human autoimmune diseases. Here, we analyzed population-level Treg variability by performing genome-wide expression profiling of CD4+ Treg and conventional CD4+ T (Tconv) cells from 168 donors, healthy or with established type-1 diabetes (T1D) or type-2 diabetes (T2D), in relation to genetic and immunologic screening. There was a range of variability in Treg signature transcripts, some almost invariant, others more variable, with more extensive variability for genes that control effector function (ENTPD1, FCRL1) than for lineage-specification factors like FOXP3 or IKZF2. Network analysis of Treg signature genes identified coregulated clusters that respond similarly to genetic and environmental variation in Treg and Tconv cells, denoting qualitative differences in otherwise shared regulatory circuits whereas other clusters are coregulated in Treg, but not Tconv, cells, suggesting Treg-specific regulation of genes like CTLA4 or DUSP4. Dense genotyping identified 110 local genetic variants (cis-expression quantitative trait loci), some of which are specifically active in Treg, but not Tconv, cells. The Treg signature became sharper with age and with increasing body-mass index, suggesting a tuning of Treg function with repertoire selection and/or chronic inflammation. Some Treg signature transcripts correlated with FOXP3 mRNA and/or protein, suggesting transcriptional or posttranslational regulatory relationships. Although no single transcript showed significant association to diabetes, overall expression of the Treg signature was subtly perturbed in T1D, but not T2D, patients. PMID:24610777

  9. Studies of innate immune systems against human cells.

    Science.gov (United States)

    Sakai, Rieko; Kitano, Etsuko; Maeda, Akira; Lo, Pei-Chi; Eguchi, Hiroshi; Watanabe, Masahito; Nagashima, Hiroshi; Okuyama, Hiroomi; Miyagawa, Shuji

    2017-02-01

    Pigs are frequently used as animal models for experiments in the surgical field, including allo- and xeno-transplantation. Regeneration studies, including studies dealing with human- and monkey-induced pluripotent stem cells (iPSC), have gradually progressed, with pigs sometimes being used as the scaffold. However, the immunological response of pigs against humans, especially innate immunities, remain unclear. This study reports on a comprehensive study of pig innate immunity against humans. Hemolytic complement activity of pig serum was measured using a microtitration technique. The pig natural anti-human antibody (Ab) was examined using human peripheral blood mononuclear cells (PBMC). The reaction of pig natural killer (NK) cells and monocytes/macrophages against human cells was also assessed. Most of the pig complement titers were measured based on methods used in human complement assays. The alternative pathway for pig complement reacts with human cells, indicating that pig complement can react with human cells. Pig serum contains relatively high levels of natural antibodies, IgM and IgG, to human PBMC. Furthermore, the killing of NK cells- and monocyte/macrophage-mediated human cells was clearly confirmed. The collective findings indicate that the pig innate immunological systems, not only serum but also cellular factors, are able to recognize and injure human cells. Copyright © 2016. Published by Elsevier B.V.

  10. Nonrandom chromosomal changes in human malignant cells

    Energy Technology Data Exchange (ETDEWEB)

    Rowley, J D

    1977-01-01

    The role of chromosomal changes in human malignant cells has been the subject of much debate. The observation of nonrandom chromosomal changes has become well recognized in chronic myelogenous leukemia, and more recently in acute myelogenous leukemia. In the present report, data are presented on the sites of duplication of chromosome No. 1 in hematologic disorders. Trisomy for region lq25 to lq32 was observed in every one of 34 patients whose cells showed duplication of some part of chromosome No. 1. Adjacent regions lq21 to lq25, and lq32 to lqter, also were trisomic in the majority of patients. Two patients had deletions, one of lq32 to qter, and the other, of lp32 to pter. The sites of chromosomal breaks leading to trisomy differ from those involved in balanced reciprocal translocations. Some of these sites are sometimes, but not always, vulnerable in constitutional chromosomal abnormalities. The nature of the proliferative advantage conferred on myeloid cells by these chromosomal changes is unknown.

  11. Human Cytomegalovirus Manipulation of Latently Infected Cells

    Science.gov (United States)

    Sinclair, John H.; Reeves, Matthew B.

    2013-01-01

    Primary infection with human cytomegalovirus (HCMV) results in the establishment of a lifelong infection of the host which is aided by the ability of HCMV to undergo a latent infection. One site of HCMV latency in vivo is in haematopoietic progenitor cells, resident in the bone marrow, with genome carriage and reactivation being restricted to the cells of the myeloid lineage. Until recently, HCMV latency has been considered to be relatively quiescent with the virus being maintained essentially as a “silent partner” until conditions are met that trigger reactivation. However, advances in techniques to study global changes in gene expression have begun to show that HCMV latency is a highly active process which involves expression of specific latency-associated viral gene products which orchestrate major changes in the latently infected cell. These changes are argued to help maintain latent infection and to modulate the cellular environment to the benefit of latent virus. In this review, we will discuss these new findings and how they impact not only on our understanding of the biology of HCMV latency but also how they could provide tantalising glimpses into mechanisms that could become targets for the clearance of latent HCMV. PMID:24284875

  12. Stereological quantification of mast cells in human synovium

    DEFF Research Database (Denmark)

    Damsgaard, T E; Sørensen, Flemming Brandt; Herlin, T

    1999-01-01

    Mast cells participate in both the acute allergic reaction as well as in chronic inflammatory diseases. Earlier studies have revealed divergent results regarding the quantification of mast cells in the human synovium. The aim of the present study was therefore to quantify these cells in the human...... synovium, using stereological techniques. Different methods of staining and quantification have previously been used for mast cell quantification in human synovium. Stereological techniques provide precise and unbiased information on the number of cell profiles in two-dimensional tissue sections of......, in this case, human synovium. In 10 patients suffering from osteoarthritis a median of 3.6 mast cells/mm2 synovial membrane was found. The total number of cells (synoviocytes, fibroblasts, lymphocytes, leukocytes) present was 395.9 cells/mm2 (median). The mast cells constituted 0.8% of all the cell profiles...

  13. Systemic human T cell developmental processes in humanized mice cotransplanted with human fetal thymus/liver tissue and hematopoietic stem cells.

    Science.gov (United States)

    Joo, Sung-Yeon; Chung, Yun Shin; Choi, Bongkum; Kim, Miyoung; Kim, Jong-Hwa; Jun, Tae-Gook; Chang, Jun; Sprent, Jonathan; Surh, Charles D; Joh, Jae-won; Kim, Sung Joo

    2012-12-15

    In many humanized mouse models, there are few T cells in the engrafted human cell, whereas the number of B cells is high. We attempted to overcome this limitation and investigate whether the entire process of human T cell development arose similarly to the process in humans, as previously reported. To produce an advanced humanized mice model, we transplanted human fetal liver/thymus tissue subrenally and injected human CD34(+) stem cells intravenously into NOD/SCID/IL2Rgamma null (NSG) mice. Humanized mice transplanted with fetal thymus/liver tissues and fetal liver-derived CD34(+) stem cells (FLT+FLCD34) showed higher levels of human cells and T cells than mice transplanted with fetal liver-derived CD34(+) stem cells only (FLCD34). In the transplanted thymus tissue of FLT+FLCD34 mice, thymus seeding progenitors (TSPs), early thymic progenitors (ETPs), pre-T cells, and all the other human T cell populations were identified. In the periphery, FLT+FLCD34 mice have high levels of CD45RA(+) T cells; conversely, FLCD34 mice have higher levels of CD45RO(+) T cells. The CD45RO(+) T cells of FLCD34 mice proliferated rapidly after stimulation and exhibited innate T cells properties, expressing PLZF (promyelocytic leukemia zinc finger protein). Human T cells educated by mouse MHC II in mice without a human thymus differ from normal human T cells. On the basis of these findings, numerous T cell-tropic human diseases could be explored in our humanized mice and molecular aspects of human T cell development could be also studied extensively.

  14. Effect of lycopene on cell viability and cell cycle progression in human cancer cell lines

    Directory of Open Access Journals (Sweden)

    Teodoro Anderson

    2012-08-01

    Full Text Available Abstract Background Lycopene, a major carotenoid component of tomato, has a potential anticancer activity in many types of cancer. Epidemiological and clinical trials rarely provide evidence for mechanisms of the compound’s action, and studies on its effect on cancer of different cell origins are now being done. The aim of the present study was to determine the effect of lycopene on cell cycle and cell viability in eight human cancer cell lines. Methods Human cell lines were treated with lycopene (1–5 μM for 48 and 96 h. Cell viability was monitored using the method of MTT. The cell cycle was analyzed by flow cytometry, and apoptotic cells were identified by terminal deoxynucleotidyl transferase-mediated dUTP nick labeling (TUNEL and by DAPI. Results Our data showed a significant decrease in the number of viable cells in three cancer cells lines (HT-29, T84 and MCF-7 after 48 h treatment with lycopene, and changes in the fraction of cells retained in different cell cycle phases. Lycopene promoted also cell cycle arrest followed by decreased cell viability in majority of cell lines after 96 h, as compared to controls. Furthermore, an increase in apoptosis was observed in four cell lines (T-84, HT-29, MCF-7 and DU145 when cells were treated with lycopene. Conclusions Our findings show the capacity of lycopene to inhibit cell proliferation, arrest cell cycle in different phases and increase apoptosis, mainly in breast, colon and prostate lines after 96 h. These observations suggest that lycopene may alter cell cycle regulatory proteins depending on the type of cancer and the dose of lycopene administration. Taken together, these data indicated that the antiproliferative effect of lycopene was cellular type, time and dose-dependent.

  15. The effect of cell phones on human health

    Science.gov (United States)

    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.

  16. New insights into human primordial germ cells and early embryonic development from single-cell analysis.

    Science.gov (United States)

    Otte, Jörg; Wruck, Wasco; Adjaye, James

    2017-08-01

    Human preimplantation developmental studies are difficult to accomplish due to associated ethical and moral issues. Preimplantation cells are rare and exist only in transient cell states. From a single cell, it is very challenging to analyse the origination of the heterogeneity and complexity inherent to the human body. However, recent advances in single-cell technology and data analysis have provided new insights into the process of early human development and germ cell specification. In this Review, we examine the latest single-cell datasets of human preimplantation embryos and germ cell development, compare them to bulk cell analyses, and interpret their biological implications. © 2017 Federation of European Biochemical Societies.

  17. Trichloroethylene toxicity in a human hepatoma cell line

    Energy Technology Data Exchange (ETDEWEB)

    Thevenin, E.; McMillian, J. [Medical Univ. of Charleston South Carolina, SC (United States)

    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.

  18. Human Galectin-3 Promotes Trypanosoma cruzi Adhesion to Human Coronary Artery Smooth Muscle Cells

    OpenAIRE

    Kleshchenko, Yuliya Y.; Moody, Tapria N.; Furtak, Vyacheslav A.; Ochieng, Josiah; Lima, Maria F.; Villalta, Fernando

    2004-01-01

    Human galectin-3 binds to the surface of Trypanosoma cruzi trypomastigotes and human coronary artery smooth muscle (CASM) cells. CASM cells express galectin-3 on their surface and secrete it. Exogenous galectin-3 increased the binding of T. cruzi to CASM cells. Trypanosome binding to CASM cells was enhanced when either T. cruzi or CASM cells were preincubated with galectin-3. Cells stably transfected with galectin-3 antisense show a dramatic decrease in galectin-3 expression and very little T...

  19. Enhanced casein kinase II activity in human tumour cell cultures

    DEFF Research Database (Denmark)

    Prowald, K; Fischer, H; Issinger, O G

    1984-01-01

    Casein kinase II (CKII) activity is enhanced as much as 2-3 fold in established and 4-5-fold in transformed human cell lines when compared to that of fibroblasts and primary human tumour cell cultures where CKII activity never exceeded a basic level. The high activity of CKII in transformed cells...

  20. Isolation and in vitro expansion of human colonic stem cells

    NARCIS (Netherlands)

    Jung, P.; Sato, T.; Merlos-Suarez, A.; Barriga, F.M.; Iglesias, M.; Rossell, D.; Auer, H.; Gallardo, M.; Blasco, M.A.; Sancho, E.; Clevers, H.; Batlle, E.

    2011-01-01

    Here we describe the isolation of stem cells of the human colonic epithelium. Differential cell surface abundance of ephrin type-B receptor 2 (EPHB2) allows the purification of different cell types from human colon mucosa biopsies. The highest EPHB2 surface levels correspond to epithelial colonic

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

    Science.gov (United States)

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

    2009-01-01

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

  2. Simulation of developing human neuronal cell networks.

    Science.gov (United States)

    Lenk, Kerstin; Priwitzer, Barbara; Ylä-Outinen, Laura; Tietz, Lukas H B; Narkilahti, Susanna; Hyttinen, Jari A K

    2016-08-30

    Microelectrode array (MEA) is a widely used technique to study for example the functional properties of neuronal networks derived from human embryonic stem cells (hESC-NN). With hESC-NN, we can investigate the earliest developmental stages of neuronal network formation in the human brain. In this paper, we propose an in silico model of maturating hESC-NNs based on a phenomenological model called INEX. We focus on simulations of the development of bursts in hESC-NNs, which are the main feature of neuronal activation patterns. The model was developed with data from developing hESC-NN recordings on MEAs which showed increase in the neuronal activity during the investigated six measurement time points in the experimental and simulated data. Our simulations suggest that the maturation process of hESC-NN, resulting in the formation of bursts, can be explained by the development of synapses. Moreover, spike and burst rate both decreased at the last measurement time point suggesting a pruning of synapses as the weak ones are removed. To conclude, our model reflects the assumption that the interaction between excitatory and inhibitory neurons during the maturation of a neuronal network and the spontaneous emergence of bursts are due to increased connectivity caused by the forming of new synapses.

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

    Science.gov (United States)

    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. Efficient and Fast Differentiation of Human Neural Stem Cells from Human Embryonic Stem Cells for Cell Therapy

    Directory of Open Access Journals (Sweden)

    Xinxin Han

    2017-01-01

    Full Text Available Stem cell-based therapies have been used for repairing damaged brain tissue and helping functional recovery after brain injury. Aberrance neurogenesis is related with brain injury, and multipotential neural stem cells from human embryonic stem (hES cells provide a great promise for cell replacement therapies. Optimized protocols for neural differentiation are necessary to produce functional human neural stem cells (hNSCs for cell therapy. However, the qualified procedure is scarce and detailed features of hNSCs originated from hES cells are still unclear. In this study, we developed a method to obtain hNSCs from hES cells, by which we could harvest abundant hNSCs in a relatively short time. Then, we examined the expression of pluripotent and multipotent marker genes through immunostaining and confirmed differentiation potential of the differentiated hNSCs. Furthermore, we analyzed the mitotic activity of these hNSCs. In this report, we provided comprehensive features of hNSCs and delivered the knowledge about how to obtain more high-quality hNSCs from hES cells which may help to accelerate the NSC-based therapies in brain injury treatment.

  5. Human pancreatic islet progenitor cells demonstrate phenotypic ...

    Indian Academy of Sciences (India)

    Prakash

    exploring alternative sources of insulin-producing cells for cell based therapy in diabetes. Since in vitro culture of islet β-cells demonstrates loss in insulin (Beattie et al. 1999), several attempts have been made to identify stem / progenitor cells capable of differentiation into insulin-producing cells. Embryonic stem cells, which ...

  6. Human Cell and Tissue Establishment Registration Public Query

    Data.gov (United States)

    U.S. Department of Health & Human Services — This application provides Human Cell and Tissue registration information for registered, inactive, and pre-registered firms. Query options are by Establishment Name,...

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

    Science.gov (United States)

    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.

  8. Effects of Human Umbilical Cord Mesenchymal Stem Cells on Human Trophoblast Cell Functions In Vitro

    Directory of Open Access Journals (Sweden)

    Yajing Huang

    2016-01-01

    Full Text Available Trophoblast cell dysfunction is involved in many disorders during pregnancy such as preeclampsia and intrauterine growth restriction. Few treatments exist, however, that target improving trophoblast cell function. Human umbilical cord mesenchymal stem cells (hUCMSCs are capable of self-renewing, can undergo multilineage differentiation, and have homing abilities; in addition, they have immunomodulatory effects and paracrine properties and thus are a prospective source for cell therapy. To identify whether hUCMSCs can regulate trophoblast cell functions, we treated trophoblast cells with hUCMSC supernatant or cocultured them with hUCMSCs. Both treatments remarkably enhanced the migration and invasion abilities of trophoblast cells and upregulated their proliferation ability. At a certain concentration, hUCMSCs also modulated hCG, PIGF, and sEndoglin levels in the trophoblast culture medium. Thus, hUCMSCs have a positive effect on trophoblast cellular functions, which may provide a new avenue for treatment of placenta-related diseases during pregnancy.

  9. Single-cell analysis reveals a stem-cell program in human metastatic breast cancer cells

    Science.gov (United States)

    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

    2015-01-01

    Despite major advances in understanding the molecular and genetic basis of cancer, metastasis remains the cause of >90% of cancer-related mortality1. 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 tumours2–5. However, the identity of metastasis-initiating cells in human breast cancer remains elusive, and whether metastases are hierarchically organized is unknown2. 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

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

    Directory of Open Access Journals (Sweden)

    Xiaoti Xu

    2015-09-01

    Full Text Available 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.

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

    Science.gov (United States)

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

    2016-03-01

    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.

  12. A Novel Class of Human Cardiac Stem Cells

    OpenAIRE

    Moccetti, Tiziano; Leri, Annarosa; Goichberg, Polina; Rota, Marcello; Anversa, Piero

    2015-01-01

    Following the recognition that hematopoietic stem cells improve the outcome of myocardial infarction in animal models, bone marrow mononuclear cells, CD34-positive cells and mesenchymal stromal cells have been introduced clinically. The intracoronary or intramyocardial injection of these cell classes has been shown to be safe and to produce a modest but significant enhancement in systolic function. However, the identification of resident cardiac stem cells in the human heart (hCSCs) has creat...

  13. Alloimmune Responses of Humanized Mice to Human Pluripotent Stem Cell Therapeutics

    Directory of Open Access Journals (Sweden)

    Nigel G. Kooreman

    2017-08-01

    Full Text Available There is growing interest in using embryonic stem cell (ESC and induced pluripotent stem cell (iPSC derivatives for tissue regeneration. However, an increased understanding of human immune responses to stem cell-derived allografts is necessary for maintaining long-term graft persistence. To model this alloimmunity, humanized mice engrafted with human hematopoietic and immune cells could prove to be useful. In this study, an in-depth analysis of graft-infiltrating human lymphocytes and splenocytes revealed that humanized mice incompletely model human immune responses toward allogeneic stem cells and their derivatives. Furthermore, using an “allogenized” mouse model, we show the feasibility of reconstituting immunodeficient mice with a functional mouse immune system and describe a key role of innate immune cells in the rejection of mouse stem cell allografts.

  14. Osteocalcin Effect on Human β-Cells Mass and Function.

    Science.gov (United States)

    Sabek, Omaima M; Nishimoto, Satoru Ken; Fraga, Daniel; Tejpal, Neelam; Ricordi, Camillo; Gaber, A O

    2015-09-01

    The osteoblast-specific hormone osteocalcin (OC) was found to regulate glucose metabolism, fat mass, and β-cell proliferation in mice. Here, we investigate the effect of decarboxylated OC (D-OC) on human β-cell function and mass in culture and in vivo using a Nonobese diabetic-severe combined immunodeficiency mouse model. We found that D-OC at dose ranges from 1.0 to 15 ng/mL significantly augmented insulin content and enhanced human β-cell proliferation of cultured human islets. This was paralleled by increased expression of sulfonylurea receptor protein; a marker of β-cell differentiation and a component of the insulin-secretory apparatus. Moreover, in a Nonobese diabetic-severe combined immunodeficiency mouse model, systemic administration of D-OC at 4.5-ng/h significantly augmented production of human insulin and C-peptide from the grafted human islets. Finally, histological staining of the human islet grafts showed that the improvement in the β-cell function was attributable to an increase in β-cell mass as a result of β-cell proliferation indicated by MKI67 staining together with the increased β-cell number and decreased α-cell number data obtained using laser scanning cytometry. Our data for the first time show D-OC-enhanced β-cell function in human islets and support future exploitation of D-OC-mediated β-cell regulation for developing useful clinical treatments for patients with diabetes.

  15. Early stages in human and mouse T-cell development

    NARCIS (Netherlands)

    Spits, H.

    1994-01-01

    One important question in lymphopoiesis is where stem cells commit to T-, B- and natural killer (NK)-cell lineages. Recent findings in human and mouse systems suggest that the thymus is seeded by a yet uncommitted progenitor cell. The earliest murine thymic progenitor cells have the capacity to

  16. Characterizing the radioresponse of pluripotent and multipotent human stem cells.

    Directory of Open Access Journals (Sweden)

    Mary L Lan

    Full Text Available The potential capability of stem cells to restore functionality to diseased or aged tissues has prompted a surge of research, but much work remains to elucidate the response of these cells to genotoxic agents. To more fully understand the impact of irradiation on different stem cell types, the present study has analyzed the radioresponse of human pluripotent and multipotent stem cells. Human embryonic stem (ES cells, human induced pluripotent (iPS cells, and iPS-derived human neural stem cells (iPS-hNSCs cells were irradiated and analyzed for cell survival parameters, differentiation, DNA damage and repair and oxidative stress at various times after exposure. While irradiation led to dose-dependent reductions in survival, the fraction of surviving cells exhibited dose-dependent increases in metabolic activity. Irradiation did not preclude germ layer commitment of ES cells, but did promote neuronal differentiation. ES cells subjected to irradiation exhibited early apoptosis and inhibition of cell cycle progression, but otherwise showed normal repair of DNA double-strand breaks. Cells surviving irradiation also showed acute and persistent increases in reactive oxygen and nitrogen species that were significant at nearly all post-irradiation times analyzed. We suggest that stem cells alter their redox homeostasis to adapt to adverse conditions and that radiation-induced oxidative stress plays a role in regulating the function and fate of stem cells within tissues compromised by radiation injury.

  17. Green tea polyphenol induces significant cell death in human lung ...

    African Journals Online (AJOL)

    Green tea polyphenol induces significant cell death in human lung cancer cells. ... Tropical Journal of Pharmaceutical Research ... (8-OHdG), and apoptosis based on 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay were evaluated in non-small cell lung cancer (NSCLC) cell lines, namely, H1155, ...

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

    Energy Technology Data Exchange (ETDEWEB)

    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)

  19. Human memory T cells: generation, compartmentalization and homeostasis.

    Science.gov (United States)

    Farber, Donna L; Yudanin, Naomi A; Restifo, Nicholas P

    2014-01-01

    Memory T cells constitute the most abundant lymphocyte population in the body for the majority of a person's lifetime; however, our understanding of memory T cell generation, function and maintenance mainly derives from mouse studies, which cannot recapitulate the exposure to multiple pathogens that occurs over many decades in humans. In this Review, we discuss studies focused on human memory T cells that reveal key properties of these cells, including subset heterogeneity and diverse tissue residence in multiple mucosal and lymphoid tissue sites. We also review how the function and the adaptability of human memory T cells depend on spatial and temporal compartmentalization.

  20. Nanoscale Mechanical Stimulation of Human Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    H Nikukar

    2014-05-01

    We observed significant responses after 1 and 2-week stimulations in cell number, cell shapes and phenotypical markers. Microarray was performed for all groups. Cell count showed normal cell growth with stimulation. However, cell surface area, cell perimeter, and arboration after 1-week stimulation showed significant increases. Immunofluorescent studies have showed significant increase in osteocalcin production after stimulation. Conclusions: Nanoscale mechanical vibration showed significant changes in human mesenchymal stem cell behaviours. Cell morphology changed to become more polygonal and increased expression of the osteoblast markers were noted. These findings with gene regulation changes suggesting nanoscale mechanostimulation has stimulated osteoblastogenesis.  Keywords:  Mesenchymal, Nanoscale, Stem Cells.

  1. Comparison of the glycosphingolipids of human-induced pluripotent stem cells and human embryonic stem cells.

    Science.gov (United States)

    Säljö, Karin; Barone, Angela; Vizlin-Hodzic, Dzeneta; Johansson, Bengt R; Breimer, Michael E; Funa, Keiko; Teneberg, Susann

    2017-04-01

    High expectations are held for human-induced pluripotent stem cells (hiPSC) since they are established from autologous tissues thus overcoming the risk of allogeneic immune rejection when used in regenerative medicine. However, little is known regarding the cell-surface carbohydrate antigen profile of hiPSC compared with human embryonic stem cells (hESC). Here, glycosphingolipids were isolated from an adipocyte-derived hiPSC line, and hiPSC and hESC glycosphingolipids were compared by concurrent characterization by binding assays with carbohydrate-recognizing ligands and mass spectrometry. A high similarity between the nonacid glycosphingolipids of hiPSC and hESC was found. The nonacid glycosphingolipids P1 pentaosylceramide, x2 pentaosylceramide and H type 1 heptaosylceramide, not previously described in human pluripotent stem cells (hPSC), were characterized in both hiPSC and hESC. The composition of acid glycosphingolipids differed, with increased levels of GM3 ganglioside, and reduced levels of GD1a/GD1b in hiPSC when compared with hESC. In addition, the hESC glycosphingolipids sulf-globopentaosylceramide and sialyl-globotetraosylceramide were lacking in hiPSC. Neural stem cells differentiating from hiPSC had a reduced expression of sialyl-lactotetra, whereas expression of the GD1a ganglioside was significantly increased. Thus, while sialyl-lactotetra is a marker of undifferentiated hPSC, GD1a is a novel marker of neural differentiation. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  2. A DNA Vaccine Protects Human Immune Cells against Zika Virus Infection in Humanized Mice

    Directory of Open Access Journals (Sweden)

    Guohua Yi

    2017-11-01

    Full Text Available A DNA vaccine encoding prM and E protein has been shown to induce protection against Zika virus (ZIKV infection in mice and monkeys. However, its effectiveness in humans remains undefined. Moreover, identification of which immune cell types are specifically infected in humans is unclear. We show that human myeloid cells and B cells are primary targets of ZIKV in humanized mice. We also show that a DNA vaccine encoding full length prM and E protein protects humanized mice from ZIKV infection. Following administration of the DNA vaccine, humanized DRAG mice developed antibodies targeting ZIKV as measured by ELISA and neutralization assays. Moreover, following ZIKV challenge, vaccinated animals presented virtually no detectable virus in human cells and in serum, whereas unvaccinated animals displayed robust infection, as measured by qRT-PCR. Our results utilizing humanized mice show potential efficacy for a targeted DNA vaccine against ZIKV in humans.

  3. The Evolution of Human Cells in Terms of Protein Innovation

    Science.gov (United States)

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

    2014-01-01

    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. PMID:24692656

  4. Human immunodeficiency syndromes affecting human natural killer cell cytolytic activity

    Directory of Open Access Journals (Sweden)

    Daniel Denis Billadeau

    2014-01-01

    Full Text Available NK cells are lymphocytes of the innate immune system that secrete cytokines upon activation and mediate the killing of tumor cells and virus-infected cells, especially those that escape the adaptive T-cell response caused by the down regulation of MHC-I. The induction of cytotoxicity requires that NK cells contact target cells through adhesion receptors, and initiate activation signaling leading to increased adhesion and accumulation of F-actin at the NK cell cytotoxic synapse. Concurrently, lytic granules undergo minus-end directed movement and accumulate at the microtubule-organizing center (MTOC through the interaction with microtubule motor proteins, followed by polarization of the lethal cargo toward the target cell. Ultimately, myosin-dependent movement of the lytic granules toward the NK cell plasma membrane through F-actin channels, along with SNARE-dependent fusion promotes lytic granule release into the cleft between the NK cell and target cell resulting in target cell killing. Herein, we will discuss several disease-causing mutations in primary immunodeficiency syndromes and how they impact NK cell-mediated killing by disrupting distinct steps of this tightly regulated process.

  5. Canthin-6-one induces cell death, cell cycle arrest and differentiation in human myeloid leukemia cells.

    Science.gov (United States)

    Vieira Torquato, Heron F; Ribeiro-Filho, Antonio C; Buri, Marcus V; Araújo Júnior, Roberto T; Pimenta, Renata; de Oliveira, José Salvador R; Filho, Valdir C; Macho, Antonio; Paredes-Gamero, Edgar J; de Oliveira Martins, Domingos T

    2017-04-01

    Canthin-6-one is a natural product isolated from various plant genera and from fungi with potential antitumor activity. In the present study, we evaluate the antitumor effects of canthin-6-one in human myeloid leukemia lineages. Kasumi-1 lineage was used as a model for acute myeloid leukemia. Cells were treated with canthin-6-one and cell death, cell cycle and differentiation were evaluated in both total cells (Lin+) and leukemia stem cell population (CD34+CD38-Lin-/low). Among the human lineages tested, Kasumi-1 was the most sensitive to canthin-6-one. Canthin-6-one induced cell death with apoptotic (caspase activation, decrease of mitochondrial potential) and necrotic (lysosomal permeabilization, double labeling of annexin V/propidium iodide) characteristics. Moreover, canthin-6-one induced cell cycle arrest at G0/G1 (7μM) and G2 (45μM) evidenced by DNA content, BrdU incorporation and cyclin B1/histone 3 quantification. Canthin-6-one also promoted differentiation of Kasumi-1, evidenced by an increase in the expression of myeloid markers (CD11b and CD15) and the transcription factor PU.1. Furthermore, a reduction of the leukemic stem cell population and clonogenic capability of stem cells were observed. These results show that canthin-6-one can affect Kasumi-1 cells by promoting cell death, cell cycle arrest and cell differentiation depending on concentration used. Canthin-6-one presents an interesting cytotoxic activity against leukemic cells and represents a promising scaffold for the development of molecules for anti-leukemic applications, especially by its anti-leukemic stem cell activity. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Molecular aging and rejuvenation of human muscle stem cells

    DEFF Research Database (Denmark)

    Carlson, Morgan E; Suetta, Charlotte; Conboy, Michael J

    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-beta)/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...

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

    Science.gov (United States)

    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. Copyright © 2016 Elsevier Inc. All rights reserved.

  8. In vitro proliferation of adult human beta-cells.

    Directory of Open Access Journals (Sweden)

    Sabine Rutti

    Full Text Available A decrease in functional beta-cell mass is a key feature of type 2 diabetes. Glucagon-like peptide 1 (GLP-1 analogues induce proliferation of rodent beta-cells. However, the proliferative capacity of human beta-cells and its modulation by GLP-1 analogues remain to be fully investigated. We therefore sought to quantify adult human beta-cell proliferation in vitro and whether this is affected by the GLP-1 analogue liraglutide.Human islets from 7 adult cadaveric organ donors were dispersed into single cells. Beta-cells were purified by FACS. Non-sorted cells and the beta-cell enriched ("beta-cells" population were plated on extracellular matrix from rat (804G and human bladder carcinoma cells (HTB9 or bovine corneal endothelial ECM (BCEC. Cells were maintained in culture+/-liraglutide for 4 days in the presence of BrdU.Rare human beta-cell proliferation could be observed either in the purified beta-cell population (0.051±0.020%; 22 beta-cells proliferating out of 84'283 beta-cells counted or in the non-sorted cell population (0.055±0.011%; 104 proliferating beta-cells out of 232'826 beta-cells counted, independently of the matrix or the culture conditions. Liraglutide increased human beta-cell proliferation on BCEC in the non-sorted cell population (0.082±0.034% proliferating beta-cells vs. 0.017±0.008% in control, p<0.05.These results indicate that adult human beta-cell proliferation can occur in vitro but remains an extremely rare event with these donors and particular culture conditions. Liraglutide increases beta-cell proliferation only in the non-sorted cell population and only on BCEC. However, it cannot be excluded that human beta-cells may proliferate to a greater extent in situ in response to natural stimuli.

  9. Nucleosome Organization in Human Embryonic Stem Cells.

    Directory of Open Access Journals (Sweden)

    Puya G Yazdi

    Full Text Available 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

  10. Proteomic Analysis of Human Blastocoel Fluid and Blastocyst Cells

    DEFF Research Database (Denmark)

    Linnert Jensen, Pernille; Beck, Hans Christian; Petersen, Jørgen

    The human blastocyst consists of 100-200 cells that are organized in an outer layer of differentiated trophectoderm (TE) cells lining the blastocyst cavity into which the undifferentiated inner cell mass (ICM) protrudes. The cavity of the blastocyst is filled with blastocoel fluid to which all...... the cells of the blastocyst are exposed. The ICM is the starting point for the development of undifferentiated human embryonic stem cells (hESCs), which posses the potential to develop into any cell type present in the adult human body [1,2]. This ability makes hESCs a potential source of cells...... blastocoel fluid (1-8 nanoliters per blastocyst), has hampered an in-depth study of the human blastocyst proteome. However, recent developments in mass spectrometry-based proteomic techniques allow the identification and characterization of thousands of proteins from low microgram levels of protein extracted...

  11. Studies of lymphocyte reconstitution in a humanized mouse model reveal a requirement of T cells for human B cell maturation.

    Science.gov (United States)

    Lang, Julie; Kelly, Margot; Freed, Brian M; McCarter, Martin D; Kedl, Ross M; Torres, Raul M; Pelanda, Roberta

    2013-03-01

    The hematopoietic humanized mouse (hu-mouse) model is a powerful resource to study and manipulate the human immune system. However, a major and recurrent issue with this model has been the poor maturation of B cells that fail to progress beyond the transitional B cell stage. Of interest, a similar problem has been reported in transplant patients who receive cord blood stem cells. In this study, we characterize the development of human B and T cells in the lymph nodes (LNs) and spleen of BALB/c-Rag2(null)Il2rγ(null) hu-mice. We find a dominant population of immature B cells in the blood and spleen early, followed by a population of human T cells, coincident with the detection of LNs. Notably, in older mice we observe a major population of mature B cells in LNs and in the spleens of mice with higher T cell frequencies. Moreover, we demonstrate that T cells are necessary for B cell maturation, as introduction of autologous human T cells expedites the appearance of mature B cells, whereas in vivo depletion of T cells retards B cell maturation. The presence of the mature B cell population correlates with enhanced IgG and Ag-specific responses to both T cell-dependent and T cell-independent challenges, indicating their functionality. These findings enhance our understanding of human B cell development, provide increased details of the reconstitution dynamics of hu-mice, and validate the use of this animal model to study mechanisms and treatments for the similar delay of functional B cells associated with cord blood transplantations.

  12. Development of human B cells and antibodies following human hematopoietic stem cell transplantation to Rag2(-/-)γc(-/-) mice.

    Science.gov (United States)

    Tanner, Anne; Hallam, Steven J; Nielsen, Stanton J; Cuadra, German I; Berges, Bradford K

    2015-06-01

    Humanized mice represent a valuable model system to study the development and functionality of the human immune system. In the RAG-hu mouse model highly immunodeficient Rag2(-/-)γc(-/-) mice are transplanted with human CD34(+) hematopoietic stem cells, resulting in human hematopoiesis and a predominant production of B and T lymphocytes. Human adaptive immune responses have been detected towards a variety of antigens in humanized mice but both cellular and humoral immune responses tend to be weak and sporadically detected. The underlying mechanisms for inconsistent responses are poorly understood. Here, we analyzed the kinetics of human B cell development and antibody production in RAG-hu mice to better understand the lack of effective antibody responses. We found that T cell levels in blood did not significantly change from 8 to 28 weeks post-engraftment, while B cells reached a peak at 14 weeks. Concentrations of 3 antibody classes (IgM, IgG, IgA) were found to be at levels about 0.1% or less of normal human levels, but human antibodies were still detected up to 32 weeks after engraftment. Human IgM was detected in 92.5% of animals while IgG and IgA were detected in about half of animals. We performed flow cytometric analysis of human B cells in bone marrow, spleen, and blood to examine the presence of precursor B cells, immature B cells, naïve B cells, and plasma B cells. We detected high levels of surface IgM(+) B cells (immature and naïve B cells) and low levels of plasma B cells in these organs, suggesting that B cells do not mature properly in this model. Low levels of human T cells in the spleen were observed, and we suggest that the lack of T cell help may explain poor B cell development and antibody responses. We conclude that human B cells that develop in humanized mice do not receive the signals necessary to undergo class-switching or to secrete antibody effectively, and we discuss strategies to potentially overcome these barriers. Copyright © 2015

  13. Persistent donor cell gene expression among human induced pluripotent stem cells contributes to differences with human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Zhumur Ghosh

    Full Text Available Human induced pluripotent stem cells (hiPSCs generated by de-differentiation of adult somatic cells offer potential solutions for the ethical issues surrounding human embryonic stem cells (hESCs, as well as their immunologic rejection after cellular transplantation. However, although hiPSCs have been described as "embryonic stem cell-like", these cells have a distinct gene expression pattern compared to hESCs, making incomplete reprogramming a potential pitfall. It is unclear to what degree the difference in tissue of origin may contribute to these gene expression differences. To answer these important questions, a careful transcriptional profiling analysis is necessary to investigate the exact reprogramming state of hiPSCs, as well as analysis of the impression, if any, of the tissue of origin on the resulting hiPSCs. In this study, we compare the gene profiles of hiPSCs derived from fetal fibroblasts, neonatal fibroblasts, adipose stem cells, and keratinocytes to their corresponding donor cells and hESCs. Our analysis elucidates the overall degree of reprogramming within each hiPSC line, as well as the "distance" between each hiPSC line and its donor cell. We further identify genes that have a similar mode of regulation in hiPSCs and their corresponding donor cells compared to hESCs, allowing us to specify core sets of donor genes that continue to be expressed in each hiPSC line. We report that residual gene expression of the donor cell type contributes significantly to the differences among hiPSCs and hESCs, and adds to the incompleteness in reprogramming. Specifically, our analysis reveals that fetal fibroblast-derived hiPSCs are closer to hESCs, followed by adipose, neonatal fibroblast, and keratinocyte-derived hiPSCs.

  14. Human dental pulp stem cells: Applications in future regenerative medicine

    Science.gov (United States)

    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

  15. Human dental pulp stem cells: Applications in future regenerative medicine.

    Science.gov (United States)

    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.

  16. Research on human placenta-derived mesenchymal stem cells ...

    African Journals Online (AJOL)

    PCR) technology, amplified hVEGF165 gene fragments from human leukemia cells HL-60. hVEGF165 gene was reconstructed in pIRES2-EGFP and transferred into the human placenta-derived mesenchymal stem cells (HPMSCs) by ...

  17. Novel human multiple myeloma cell line UHKT-893

    Czech Academy of Sciences Publication Activity Database

    Uherková, L.; Vančurová, I.; Vyhlídalová, I.; Pleschnerová, M.; Špička, I.; Mihalová, R.; Březinová, J.; Hodný, Zdeněk; Čermáková, K.; Polanská, V.; Marinov, I.; Jedelský, P.L.; Kuželová, K.; Stöckbauer, P.

    2013-01-01

    Roč. 37, č. 3 (2013), s. 320-326 ISSN 0145-2126 Institutional support: RVO:68378050 Keywords : human myeloma cell line * human multiple myeloma * plasma cell * IL-6 dependence * immunoglobulin * free light chain Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.692, year: 2013

  18. Sphere-forming cell subpopulations with cancer stem cell properties in human hepatoma cell lines

    Directory of Open Access Journals (Sweden)

    Chen Lei

    2011-06-01

    Full Text Available Abstract Background Cancer stem cells (CSCs are regarded as the cause of tumor formation and recurrence. The isolation and identification of CSCs could help to develop novel therapeutic strategies specifically targeting CSCs. Methods Human hepatoma cell lines were plated in stem cell conditioned culture system allowed for sphere forming. To evaluate the stemness characteristics of spheres, the self-renewal, proliferation, chemoresistance, tumorigenicity of the PLC/PRF/5 sphere-forming cells, and the expression levels of stem cell related proteins in the PLC/PRF/5 sphere-forming cells were assessed, comparing with the parental cells. The stem cell RT-PCR array was performed to further explore the biological properties of liver CSCs. Results The PLC/PRF/5, MHCC97H and HepG2 cells could form clonal nonadherent 3-D spheres and be serially passaged. The PLC/PRF/5 sphere-forming cells possessed a key criteria that define CSCs: persistent self-renewal, extensive proliferation, drug resistance, overexpression of liver CSCs related proteins (Oct3/4, OV6, EpCAM, CD133 and CD44. Even 500 sphere-forming cells were able to form tumors in NOD/SCID mice, and the tumor initiating capability was not decreased when spheres were passaged. Besides, downstream proteins DTX1 and Ep300 of the CSL (CBF1 in humans, Suppressor of hairless in Drosophila and LAG1 in C. elegans -independent Notch signaling pathway were highly expressed in the spheres, and a gamma-secretase inhibitor MRK003 could significantly inhibit the sphere formation ability. Conclusions Nonadherent tumor spheres from hepatoma cell lines cultured in stem cell conditioned medium possess liver CSC properties, and the CSL-independent Notch signaling pathway may play a role in liver CSCs.

  19. Human white blood cells contain cyclobutyl pyrimidine dimer photolyase

    Energy Technology Data Exchange (ETDEWEB)

    Sutherland, B.M.; Bennett, P.V. [Brookhaven National Lab., Upton, NY (United States)

    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.

  20. Human serum promotes osteogenic differentiation of human dental pulp stem cells in vitro and in vivo

    National Research Council Canada - National Science Library

    Pisciotta, Alessandra; Riccio, Massimo; Carnevale, Gianluca; Beretti, Francesca; Gibellini, Lara; Maraldi, Tullia; Cavallini, Gian Maria; Ferrari, Adriano; Bruzzesi, Giacomo; De Pol, Anto

    2012-01-01

    Human dental pulp is a promising alternative source of stem cells for cell-based tissue engineering in regenerative medicine, for the easily recruitment with low invasivity for the patient and for the...

  1. Identifying gene expression modules that define human cell fates

    OpenAIRE

    Germanguz, I; Listgarten, J; Cinkornpumin, J.; Solomon, A; Gaeta, X.; Lowry, W. E.

    2016-01-01

    Using a compendium of cell-state-specific gene expression data, we identified genes that uniquely define cell states, including those thought to represent various developmental stages. Our analysis sheds light on human cell fate through the identification of core genes that are altered over several developmental milestones, and across regional specification. Here we present cell-type specific gene expression data for 17 distinct cell states and demonstrate that these modules of genes can in f...

  2. Human organomics: a fresh approach to understanding human development using single-cell transcriptomics.

    Science.gov (United States)

    Camp, J Gray; Treutlein, Barbara

    2017-05-01

    Innovative methods designed to recapitulate human organogenesis from pluripotent stem cells provide a means to explore human developmental biology. New technologies to sequence and analyze single-cell transcriptomes can deconstruct these 'organoids' into constituent parts, and reconstruct lineage trajectories during cell differentiation. In this Spotlight article we summarize the different approaches to performing single-cell transcriptomics on organoids, and discuss the opportunities and challenges of applying these techniques to generate organ-level, mechanistic models of human development and disease. Together, these technologies will move past characterization to the prediction of human developmental and disease-related phenomena. © 2017. Published by The Company of Biologists Ltd.

  3. STELLA facilitates differentiation of germ cell and endodermal lineages of human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Patompon Wongtrakoongate

    Full Text Available Stella is a developmentally regulated gene highly expressed in mouse embryonic stem (ES cells and in primordial germ cells (PGCs. In human, the gene encoding the STELLA homologue lies on chromosome 12p, which is frequently amplified in long-term cultured human ES cells. However, the role played by STELLA in human ES cells has not been reported. In the present study, we show that during retinoic acid (RA-induced differentiation of human ES cells, expression of STELLA follows that of VASA, a marker of germline differentiation. By contrast, human embryonal carcinoma cells express STELLA at a higher level compared with both karyotypically normal and abnormal human ES cell lines. We found that over-expression of STELLA does not interfere with maintenance of the stem cell state of human ES cells, but following retinoic acid induction it leads to up-regulation of germline- and endodermal-associated genes, whereas neural markers PAX6 and NEUROD1 are down-regulated. Further, STELLA over-expression facilitates the differentiation of human ES cells into BE12-positive cells, in which the expression of germline- and endodermal-associated genes is enriched, and suppresses differentiation of the neural lineage. Taken together, this finding suggests a role for STELLA in facilitating germline and endodermal differentiation of human ES cells.

  4. Exposure to Music Alters Cell Viability and Cell Motility of Human Nonauditory Cells in Culture

    Directory of Open Access Journals (Sweden)

    Nathalia R. Lestard

    2016-01-01

    Full Text Available 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.

  5. MR imaging findings of pineal germinoma: focus on differential diagnosis from other germ cell tumors

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Hyun Jin; Lee, Ho Kyu; Kim, Jae Kyun; Shin, Ji Hoon; Choi, Choong Gon; Lee, Myung Jun; Ham, Soo Youn; Lee, Jong Hwa; Suh, Dae Chul [Ulsan Univ. College of Medicine, Seoul (Korea, Republic of)

    1998-10-01

    To determine the characteristic MR imaging findings of pineal germinoma, and differential diagnosis from other germ cell tumors. MR images of patients with histopathologically proven pineal germinoma(n=3D14) and other pineal germ cell tumors(n=3D10) were retrospectively analyzed with regard to size, signal intensity and homogeneity, enhancing features, cyst formation, and multiplicity of lesions. Other pineal germ cell tumors were the mixed germ cell tumors (n=3D4), malignant teratomas (n=3D3), choriocarcinoma(n=3D1), embryonal carcinoma(n=3D1), and endodermal sinus tumor(n=3D1). Tumor markers were evaluated. On T1-weighted images, germinomas showed homogeneous(86%) or iso signal intensity (93%), while other germ cell tumors showed inhomogeneous(70%) or iso signal intensity(70%). On T2-weighted images, germinomas showed homogeneous(64%) or iso signal intensity(57%), while other germ cell tumors showed inhomogeneous(70%) or high signal intensity(80%). On Gd-DTPA enhanced images, germinomas showed homogeneous (93%) or strong enhancement (64%), while other germ cell tumors showed homogeneous(60%) or strong enhancement (70%). Cyst formation was noted in ten Patients (71%) with germinoma and in six (60%) with other germ cell tumors. Invasion on surrounding structures was seen in 11 patients (79%) with germinoma and in five (50%) with other germ cell tumors. Lesions were multiple in three patients(21%) with germinoma. Thirteen of 14 patients with germinoma had normal serum {alpha}-FP(tetoprotein) and {beta}-HCG(human chononic gonafotrophin) levels. Two of four patients with mixed germ cell tumors had elevated serum {beta}-FP and {alpha}-HCG levels; in the ther two, elevated serum {alpha}-FP or {beta}-HCG levels were noted. In the malignant teratoma and embryonal carcinoma patients, serum {alpha}-FP and {beta}-HCG levels were normal. The patient with choriocarcinoma had an elevated serum {beta}-HCG level. On T1W1, the only significant differential point (p<0.01) between

  6. Human embryonic stem cells differentiate into functional renal proximal tubular-like cells.

    Science.gov (United States)

    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.

  7. Rapid induction of senescence in human cervical carcinoma cells

    Science.gov (United States)

    Goodwin, Edward C.; Yang, Eva; Lee, Chan-Jae; Lee, Han-Woong; Dimaio, Daniel; Hwang, Eun-Seong

    2000-09-01

    Expression of the bovine papillomavirus E2 regulatory protein in human cervical carcinoma cell lines repressed expression of the resident human papillomavirus E6 and E7 oncogenes and within a few days caused essentially all of the cells to synchronously display numerous phenotypic markers characteristic of cells undergoing replicative senescence. This process was accompanied by marked but in some cases transient alterations in the expression of cell cycle regulatory proteins and by decreased telomerase activity. We propose that the human papillomavirus E6 and E7 proteins actively prevent senescence from occurring in cervical carcinoma cells, and that once viral oncogene expression is extinguished, the senescence program is rapidly executed. Activation of endogenous senescence pathways in cancer cells may represent an alternative approach to treat human cancers.

  8. Proteomic analysis of human blastocoel fluid and blastocyst cells

    DEFF Research Database (Denmark)

    Jensen, Pernille; Beck, Hans Christian; Petersen, Jørgen

    2013-01-01

    Human embryonic stem cells (hESCs) are derived from the inner cell mass (ICM) of the blastocyst and can differentiate into any cell type in the human body. These cells hold a great potential for regenerative medicine, but to obtain enough cells needed for medical treatment, culture is required......, the blastocoel fluid, which is in contact with all the cells in the blastocyst, including hESCs. Fifty-three surplus human blastocysts were donated after informed consent, and blastocoel fluid was isolated by micromanipulation. Using highly sensitive nano-high-pressure liquid chromatography-tandem mass...... from the ICM of the human blastocyst are exposed to via the blastocoel fluid. These results can be an inspiration for the development of improved culture conditions for hESCs....

  9. Piperlongumine Suppresses Proliferation of Human Oral Squamous Cell Carcinoma through Cell Cycle Arrest, Apoptosis and Senescence

    Directory of Open Access Journals (Sweden)

    San-Yuan Chen

    2016-04-01

    Full Text Available 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.

  10. Advances in culture and manipulation of human pluripotent stem cells.

    Science.gov (United States)

    Qian, X; Villa-Diaz, L G; Krebsbach, P H

    2013-11-01

    Recent advances in the understanding of pluripotent stem cell biology and emerging technologies to reprogram somatic cells to a stem cell-like state are helping bring stem cell therapies for a range of human disorders closer to clinical reality. Human pluripotent stem cells (hPSCs) have become a promising resource for regenerative medicine and research into early development because these cells are able to self-renew indefinitely and are capable of differentiation into specialized cell types of all 3 germ layers and trophoectoderm. Human PSCs include embryonic stem cells (hESCs) derived from the inner cell mass of blastocyst-stage embryos and induced pluripotent stem cells (hiPSCs) generated via the reprogramming of somatic cells by the overexpression of key transcription factors. The application of hiPSCs and the finding that somatic cells can be directly reprogrammed into different cell types will likely have a significant impact on regenerative medicine. However, a major limitation for successful therapeutic application of hPSCs and their derivatives is the potential xenogeneic contamination and instability of current culture conditions. This review summarizes recent advances in hPSC culture and methods to induce controlled lineage differentiation through regulation of cell-signaling pathways and manipulation of gene expression as well as new trends in direct reprogramming of somatic cells.

  11. Abnormal number cell division of human thyroid anaplastic carcinoma cell line, SW 1736

    Directory of Open Access Journals (Sweden)

    Keiichi Ikeda

    2015-12-01

    Full Text Available Cell division, during which a mother cell usually divides into two daughter cells during one cell cycle, is the most important physiological event of cell biology. We observed one-to-four cell division during imaging of live SW1736 human thyroid anaplastic carcinoma cells transfected with a plasmid expressing the hybrid protein of green fluorescent protein and histone 2B (plasmid eGFP-H2B. Analysis of the images revealed a mother cell divided into four daughter cells. And one of the abnormally divided daughter cells subsequently formed a dinucleate cell.

  12. Tissuelike 3D Assemblies of Human Broncho-Epithelial Cells

    Science.gov (United States)

    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

  13. Human mesenchymal stem cell-engineered hepatic cell sheets accelerate liver regeneration in mice

    OpenAIRE

    Noriko Itaba; Yoshiaki Matsumi; Kaori Okinaka; An Afida Ashla; Yohei Kono; Mitsuhiko Osaki; Minoru Morimoto; Naoyuki Sugiyama; Kazuo Ohashi; Teruo Okano; Goshi Shiota

    2015-01-01

    Mesenchymal stem cells (MSCs) are an attractive cell source for cell therapy. Based on our hypothesis that suppression of Wnt/β-catenin signal enhances hepatic differentiation of human MSCs, we developed human mesenchymal stem cell-engineered hepatic cell sheets by a small molecule compound. Screening of 10 small molecule compounds was performed by WST assay, TCF reporter assay, and albumin mRNA expression. Consequently, hexachlorophene suppressed TCF reporter activity in time- and concentrat...

  14. Side Population Cells Derived from Adult Human Liver Generate Hepatocyte-like Cells In Vitro

    OpenAIRE

    HUSSAIN, SUNNY ZAHEED; Strom, Stephen C.; Kirby, Martha R.; Burns, Sean; Langemeijer, Saskia; Ueda, Takahiro; HSIEH, MATTHEW; Tisdale, John F.

    2005-01-01

    We sought to determine whether hepatic side population (SP) cells derived from adult human liver possess the potential of a novel candidate hepatic stem cell. Human cadaveric donor liver was subjected to collagenase perfusion and hepatocytes were separated from nonparenchymal cells by differential centrifugation. SP cells were isolated from the nonparenchymal portion after Hoechst 33342 staining. Since CD45 is a panleukocyte antigen, CD45-negative SP cells were separated from the vast majorit...

  15. Proteins differentially expressed in human beta-cells-enriched pancreatic islet cultures and human insulinomas

    DEFF Research Database (Denmark)

    Terra, Letícia F; Teixeira, Priscila C; Wailemann, Rosangela A M

    2013-01-01

    In view of the great demand for human beta-cells for physiological and medical studies, we generated cell lines derived from human insulinomas which secrete insulin, C-peptide and express neuroendocrine and islet markers. In this study, we set out to characterize their proteomes, comparing them...... to those of primary beta-cells using DIGE followed by MS. The results were validated by Western blotting. An average of 1800 spots was detected with less than 1% exhibiting differential abundance. Proteins more abundant in human islets, such as Caldesmon, are involved in the regulation of cell......, a molecular snapshot of the orchestrated changes in expression of proteins involved in key processes which could be correlated with the altered phenotype of human beta-cells. Collectively our observations prompt research towards the establishment of bioengineered human beta-cells providing a new and needed...

  16. Comparison of human dental follicle cells and human periodontal ligament cells for dentin tissue regeneration.

    Science.gov (United States)

    Tian, Ye; Bai, Ding; Guo, Weihua; Li, Jie; Zeng, Jin; Yang, Longqiang; Jiang, Zongting; Feng, Lian; Yu, Mei; Tian, Weidong

    2015-05-01

    To compare the odontogenic potential of human dental follicle cells (DFCs) and periodontal ligament cells (PDLCs). In vitro and in vivo characterization studies of DFCs and PDLCs were performed comparatively. DFCs and PDLCs were subcutaneously implanted into the dorsum of mice for 8 weeks after combined with treated dentin matrix scaffolds respectively. Proteomic analysis identified 32 differentially expressed proteins in DFCs and PDLCs. Examination of the harvested grafts showed PDLCs could form the dentin-like tissues as DFCs did. However, the structure of dentin tissues generated by DFCs was more complete. PDLCs could contribute to regenerate dentin-like tissues in the inductive microenvironment of treated dentin matrix. DFCs presented more remarkable dentinogenic capability than PDLCs did.

  17. Phenotype and functions of memory Tfh cells in human blood.

    Science.gov (United States)

    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. Copyright © 2014 Elsevier Ltd. All rights reserved.

  18. Genome editing: a robust technology for human stem cells.

    Science.gov (United States)

    Chandrasekaran, Arun Pandian; Song, Minjung; Ramakrishna, Suresh

    2017-09-01

    Human pluripotent stem cells comprise induced pluripotent and embryonic stem cells, which have tremendous potential for biological and therapeutic applications. The development of efficient technologies for the targeted genome alteration of stem cells in disease models is a prerequisite for utilizing stem cells to their full potential. Genome editing of stem cells is possible with the help of synthetic nucleases that facilitate site-specific modification of a gene of interest. Recent advances in genome editing techniques have improved the efficiency and speed of the development of stem cells for human disease models. Zinc finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated system are powerful tools for editing DNA at specific loci. Here, we discuss recent technological advances in genome editing with site-specific nucleases in human stem cells.

  19. Human retinal pigment epithelial cells inhibit proliferation and IL2R expression of activated T cells

    DEFF Research Database (Denmark)

    Kaestel, Charlotte G; Jørgensen, Annette; Nielsen, Mette

    2002-01-01

    The purpose of this study was to characterize the effects of human retinal pigment epithelial (RPE) cells on activated T cells. Activated T cells were cocultured with adult and foetal human RPE cells whereafter apoptosis and proliferation were determined by flow cytometry and (3)H......-Thymidine incorporation assay, respectively. T cells and RPE cells were cultured directly together or in a transwell system for determination of the effect of cell contact. The importance of cell surface molecules was examined by application of a panel of blocking antibodies (CD2, CD18, CD40, CD40L, CD54, CD58...

  20. Human papilloma virus prevalence in laryngeal squamous cell carcinoma.

    Science.gov (United States)

    Gungor, A; Cincik, H; Baloglu, H; Cekin, E; Dogru, S; Dursun, E

    2007-08-01

    To determine the prevalence and type of human papilloma virus deoxyribonucleic acid (DNA) in cases of laryngeal squamous cell carcinoma. We analysed the prevalence of human papilloma virus infection in archived paraffin block specimens taken from 99 cases of laryngeal squamous cell carcinoma between 1990 and 2005, using polymerase chain reaction techniques. Biopsy specimens from five proven verrucous skin lesions were used as positive controls, and peripheral blood samples from five healthy volunteers were used as negative controls. Four test samples were found to have inadequate deoxyribonucleic acid purity and were therefore excluded from the study. Human papilloma virus deoxyribonucleic acid was detected in seven of 95 cases of laryngeal squamous cell carcinoma (7.36 per cent). Human papilloma virus genotyping revealed double human papilloma virus infection in three cases and single human papilloma virus infection in the remaining four cases. The human papilloma virus genotypes detected were 6, 11 and 16 (the latter detected in only one case). In our series, a very low human papilloma virus prevalence was found among laryngeal squamous cell carcinoma cases. The human papilloma virus genotypes detected were mostly 6 and/or 11, and 16 in only one case. To the best of our knowledge, this is the first report of human papilloma virus prevalence in laryngeal squamous cell carcinoma, based on polymerase chain reaction genotyping in a Turkish population.

  1. Human amniotic epithelial cells as feeder layer to derive and maintain human embryonic stem cells from poor-quality embryos

    Directory of Open Access Journals (Sweden)

    Daniela Ávila-González

    2015-09-01

    Full Text Available 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.

  2. Applied Developmental Biology: Making Human Pancreatic Beta Cells for Diabetics.

    Science.gov (United States)

    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. © 2016 Elsevier Inc. All rights reserved.

  3. Enhancement of Bleomycin Sensitivity in Human Lung Cancer Cell ...

    African Journals Online (AJOL)

    Purpose: To demonstrate the effectiveness of Centella asiatica aqueous extract in augmenting the cytotoxic effect of bleomycin in the adenocarcinoma human alveolar basal epithelial A549 cell line. Methods: The inhibitory effect of bleomycin on A549 cells was determined by incubating the cells for 24 h in different ...

  4. Human Liver Stem Cells Suppress T-Cell Proliferation, NK Activity, and Dendritic Cell Differentiation

    Directory of Open Access Journals (Sweden)

    Stefania Bruno

    2016-01-01

    Full Text Available 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.

  5. Transplantation of human embryonic stem cells onto a partially wounded human cornea in vitro.

    Science.gov (United States)

    Hanson, Charles; Hardarson, Thorir; Ellerström, Catharina; Nordberg, Markus; Caisander, Gunilla; Rao, Mahendra; Hyllner, Johan; Stenevi, Ulf

    2013-03-01

    The aim of this study was to investigate whether cells originating from human embryonic stem cells (hESCs) could be successfully transplanted onto a partially wounded human cornea. A second aim was to study the ability of the transplanted cells to differentiate into corneal epithelial-like cells. Spontaneously, differentiated hESCs were transplanted onto a human corneal button (without limbus) with the epithelial layer partially removed. The cells were cultured on Bowman's membrane for up to 9 days, and the culture dynamics documented in a time-lapse system. As the transplanted cells originated from a genetically engineered hESC line, they all expressed green fluorescent protein, which facilitated their identification during the culture experiments, tissue preparation and analysis. To detect any differentiation into human corneal epithelial-like cells, we analysed the transplanted cells by immunohistochemistry using antibodies specific for CK3, CK15 and PAX6. The transplanted cells established and expanded on Bowman's membrane, forming a 1-4 cell layer surrounded by host corneal epithelial cells. Expression of the corneal marker PAX6 appeared 3 days after transplantation, and after 6 days, the cells were expressing both PAX6 and CK3. This shows that it is possible to transplant cells originating from hESCs onto Bowman's membrane with the epithelial layer partially removed and to get these cells to establish, grow and differentiate into corneal epithelial-like cells in vitro. © 2012 The Authors. Acta Ophthalmologica © 2012 Acta Ophthalmologica Scandinavica Foundation.

  6. Male germline stem cells in non-human primates

    Directory of Open Access Journals (Sweden)

    S. Sharma

    2017-09-01

    Full Text Available Over the past few decades, several studies have attempted to decipher the biology of mammalian germline stem cells (GSCs. These studies provide evidence that regulatory mechanisms for germ cell specification and migration are evolutionarily conserved across species. The characteristics and functions of primate GSCs are highly distinct from rodent species; therefore the findings from rodent models cannot be extrapolated to primates. Due to limited availability of human embryonic and testicular samples for research purposes, two non-human primate models (marmoset and macaque monkeys are extensively employed to understand human germline development and differentiation. This review provides a broader introduction to the in vivo and in vitro germline stem cell terminology from primordial to differentiating germ cells. Primordial germ cells (PGCs are the most immature germ cells colonizing the gonad prior to sex differentiation into testes or ovaries. PGC specification and migratory patterns among different primate species are compared in the review. It also reports the distinctions and similarities in expression patterns of pluripotency markers (OCT4A, NANOG, SALL4 and LIN28 during embryonic developmental stages, among marmosets, macaques and humans. This review presents a comparative summary with immunohistochemical and molecular evidence of germ cell marker expression patterns during postnatal developmental stages, among humans and non-human primates. Furthermore, it reports findings from the recent literature investigating the plasticity behavior of germ cells and stem cells in other organs of humans and monkeys. The use of non-human primate models would enable bridging the knowledge gap in primate GSC research and understanding the mechanisms involved in germline development. Reported similarities in regulatory mechanisms and germ cell expression profile in primates demonstrate the preclinical significance of monkey models for development of

  7. Resistin modulates glucose uptake and glucose transporter-1 (GLUT-1) expression in trophoblast cells

    Science.gov (United States)

    Di Simone, Nicoletta; Di Nicuolo, Fiorella; Marzioni, Daniela; Castellucci, Mario; Sanguinetti, Maurizio; D’lppolito, Silvia; Caruso, Alessandro

    2009-01-01

    Abstract The adipocytokine resistin impairs glucose tolerance and insulin sensitivity. Here, we examine the effect of resistin on glucose uptake in human trophoblast cells and we demonstrate that transplacental glucose transport is mediated by glucose transporter (GLUT)-1. Furthermore, we evaluate the type of signal transduction induced by resistin in GLUT-1 regulation. BeWo choriocarcinoma cells and primary cytotrophoblast cells were cultured with increasing resistin concentrations for 24 hrs. The main outcome measures include glucose transport assay using [3H]-2-deoxy glucose, GLUT-1 protein expression by Western blot analysis and GLUT-1 mRNA detection by quantitative real-time RT-PCR. Quantitative determination of phospho(p)-ERK1/2 in cell lysates was performed by an Enzyme Immunometric Assay and Western blot analysis. Our data demonstrate a direct effect of resistin on normal cytotrophoblastic and on BeWo cells: resistin modulates glucose uptake, GLUT-1 messenger ribonucleic acid (mRNA) and protein expression in placental cells. We suggest that ERK1/2 phosphorylation is involved in the GLUT-1 regulation induced by resistin. In conclusion, resistin causes activation of both the ERK1 and 2 pathway in trophoblast cells. ERK1 and 2 activation stimulated GLUT-1 synthesis and resulted in increase of placental glucose uptake. High resistin levels (50–100 ng/ml) seem able to affect glucose-uptake, presumably by decreasing the cell surface glucose transporter. PMID:18410529

  8. Induction of human neuronal cells by defined transcription factors

    Science.gov (United States)

    Pang, Zhiping P.; Yang, Nan; Vierbuchen, Thomas; Ostermeier, Austin; Fuentes, Daniel R.; Yang, Troy Q.; Citri, Ami; Sebastiano, Vittorio; Marro, Samuele; Südhof, Thomas C.; Wernig, Marius

    2011-01-01

    Summary Somatic cell nuclear transfer, cell fusion, or expression of lineage-specific factors have been shown to induce cell-fate changes in diverse somatic cell types1–12. We recently observed that forced expression of a combination of three transcription factors, Brn2 (also known as Pou3f2), Ascl1, and Myt1l can efficiently convert mouse fibroblasts into functional induced neuronal (iN) cells13. Here, we show that the same three factors can generate functional neurons from human pluripotent stem cells as early as 6 days after transgene activation. When combined with the basic helix-loop-helix transcription factor NeuroD1, these factors could also convert fetal and postnatal human fibroblasts into iN cells displaying typical neuronal morphologies and expressing multiple neuronal markers, even after downregulation of the exogenous transcription factors. Importantly, the vast majority of human iN cells were able to generate action potentials and many matured to receive synaptic contacts when co-cultured with primary mouse cortical neurons. Our data demonstrate that non-neural human somatic cells, as well as pluripotent stem cells, can be directly converted into neurons by lineage-determining transcription factors. These methods may facilitate robust generation of patient-specific human neurons for in vitro disease modeling or future applications in regenerative medicine. PMID:21617644

  9. Activation of intracellular angiotensin AT2 receptors induces rapid cell death in human uterine leiomyosarcoma cells

    DEFF Research Database (Denmark)

    Zhao, Yi; Lützen, Ulf; Fritsch, Jürgen

    2015-01-01

    of apoptosis and cell death in cultured human uterine leiomyosarcoma (SK-UT-1) cells and control human uterine smooth muscle cells (HutSMC). The intracellular levels of the AT2 receptor are low in proliferating SK-UT-1 cells but the receptor is substantially up-regulated in quiescent SK-UT-1 cells with high...... densities in mitochondria. Activation of the cell membrane AT2 receptors by a concomitant treatment with angiotensin II and the AT1 receptor antagonist, losartan, induces apoptosis but does not affect the rate of cell death. We demonstrate for the first time that the high-affinity, non-peptide AT2 receptor...... agonist, Compound 21 (C21) penetrates the cell membrane of quiescent SK-UT-1 cells, activates intracellular AT2 receptors and induces rapid cell death; approximately 70% of cells died within 24 h. The cells, which escaped from the cell death, displayed activation of the mitochondrial apoptotic pathway, i...

  10. Human normal bronchial epithelial cells: a novel in vitro cell model for toxicity evaluation.

    Directory of Open Access Journals (Sweden)

    Wenqiang Feng

    Full Text Available Human normal cell-based systems are needed for drug discovery and toxicity evaluation. hTERT or viral genes transduced human cells are currently widely used for these studies, while these cells exhibited abnormal differentiation potential or response to biological and chemical signals. In this study, we established human normal bronchial epithelial cells (HNBEC using a defined primary epithelial cell culture medium without transduction of exogenous genes. This system may involve decreased IL-1 signaling and enhanced Wnt signaling in cells. Our data demonstrated that HNBEC exhibited a normal diploid karyotype. They formed well-defined spheres in matrigel 3D culture while cancer cells (HeLa formed disorganized aggregates. HNBEC cells possessed a normal cellular response to DNA damage and did not induce tumor formation in vivo by xenograft assays. Importantly, we assessed the potential of these cells in toxicity evaluation of the common occupational toxicants that may affect human respiratory system. Our results demonstrated that HNBEC cells are more sensitive to exposure of 10~20 nm-sized SiO2, Cr(VI and B(aP compared to 16HBE cells (a SV40-immortalized human bronchial epithelial cells. This study provides a novel in vitro human cells-based model for toxicity evaluation, may also be facilitating studies in basic cell biology, cancer biology and drug discovery.

  11. Human normal bronchial epithelial cells: a novel in vitro cell model for toxicity evaluation.

    Science.gov (United States)

    Feng, Wenqiang; Guo, Juanjuan; Huang, Haiyan; Xia, Bo; Liu, Hongya; Li, Jie; Lin, Shaolin; Li, Tiyuan; Liu, Jianjun; Li, Hui

    2015-01-01

    Human normal cell-based systems are needed for drug discovery and toxicity evaluation. hTERT or viral genes transduced human cells are currently widely used for these studies, while these cells exhibited abnormal differentiation potential or response to biological and chemical signals. In this study, we established human normal bronchial epithelial cells (HNBEC) using a defined primary epithelial cell culture medium without transduction of exogenous genes. This system may involve decreased IL-1 signaling and enhanced Wnt signaling in cells. Our data demonstrated that HNBEC exhibited a normal diploid karyotype. They formed well-defined spheres in matrigel 3D culture while cancer cells (HeLa) formed disorganized aggregates. HNBEC cells possessed a normal cellular response to DNA damage and did not induce tumor formation in vivo by xenograft assays. Importantly, we assessed the potential of these cells in toxicity evaluation of the common occupational toxicants that may affect human respiratory system. Our results demonstrated that HNBEC cells are more sensitive to exposure of 10~20 nm-sized SiO2, Cr(VI) and B(a)P compared to 16HBE cells (a SV40-immortalized human bronchial epithelial cells). This study provides a novel in vitro human cells-based model for toxicity evaluation, may also be facilitating studies in basic cell biology, cancer biology and drug discovery.

  12. Surface differentiation antigens of human mammary epithelial cells carried on the human milk fat globule.

    Science.gov (United States)

    Ceriani, R L; Thompson, K; Peterson, J A; Abraham, S

    1977-02-01

    Rabbit antibodies against components of the human milk fat globule bind specifically to normal human breast epithelial cells and cell lines derived from breast carcinomas, as well as to the outer surface of the human milk fat globule. Variation in indirect immunofluorescence staining in both intensity per cell and percentage of cells stained is observed for the different brest cell lines. Cells derived from other epithelial and other ectodermal tissues, fetal fibroblasts, cells of the blood buffy coat, and even fibroblasts of the breast itself do not bind the antibodies. This suggests that these antibodies are detecting cell-type-specific antigens. These normal breast epithelial cell antigens are on the cell surface and their expression is stable in long-term cultured cell lines, even after much chromosomal variation in a given line. By affinity chromatography, three distinct antigenic components can be isolated from the milk fat globule, one of which contains carbohydrate. These differentiation antigens of the human breast epithelial cell are not only useful as specific cell-type markers, but also can provide a tool to study the role of the cell surface in normal and neoplastic mammary development.

  13. Cell therapy with human renal cell cultures containing erythropoietin-positive cells improves chronic kidney injury.

    Science.gov (United States)

    Yamaleyeva, Liliya M; Guimaraes-Souza, Nadia K; Krane, Louis S; Agcaoili, Sigrid; Gyabaah, Kenneth; Atala, Anthony; Aboushwareb, Tamer; Yoo, James J

    2012-05-01

    New therapeutic strategies for chronic kidney disease (CKD) are necessary to offset the rising incidence of CKD and donor shortage. Erythropoietin (EPO), a cytokine produced by fibroblast-like cells in the kidney, has recently emerged as a renoprotective factor with anti-inflammatory, antioxidant properties. This study (a) determined whether human renal cultures (human primary kidney cells [hPKC]) can be enriched in EPO-positive cells (hPKC(F+)) by using magnetic-bead sorting; (b) characterized hPKC(F+) following cell separation; and (c) established that intrarenal delivery of enriched hPKC(F+) cells would be more beneficial in treatment of renal injury, inflammation, and oxidative stress than unsorted hPKC cultures in a chronic kidney injury model. Fluorescence-activated cell sorting analysis revealed higher expression of EPO (36%) and CD73 (27%) in hPKC(F+) as compared with hPKC. After induction of renal injury, intrarenal delivery of hPKC(F+) or hPKC significantly reduced serum creatinine, interstitial fibrosis in the medulla, and abundance of CD68-positive cells in the cortex and medulla (p renal cortex and decreased urinary albumin (3.5-fold) and urinary tubular injury marker kidney injury molecule 1 (16-fold). hPKC(F+) also significantly reduced levels of renal cortical monocyte chemotactic protein 1 (1.8-fold) and oxidative DNA marker 8-hydroxy-deoxyguanosine (8-OHdG) (2.4-fold). After 12 weeks, we detected few injected cells, which were localized mostly to the cortical interstitium. Although cell therapy with either hPKC(F+) or hPKC improved renal function, the hPKC(F+) subpopulation provides greater renoprotection, perhaps through attenuation of inflammation and oxidative stress. We conclude that hPKC(F+) may be used as components of cell-based therapies for degenerative kidney diseases.

  14. Mesenchymal stem cell isolation and characterization from human spinal ligaments.

    Science.gov (United States)

    Asari, Toru; Furukawa, Ken-Ichi; Tanaka, Sunao; Kudo, Hitoshi; Mizukami, Hiroki; Ono, Atsushi; Numasawa, Takuya; Kumagai, Gentaro; Motomura, Shigeru; Yagihashi, Soroku; Toh, Satoshi

    2012-01-27

    Mesenchymal stem cells (MSCs) have a fibroblast-like morphology, multilineage potential, long-term viability and capacity for self-renewal. While several articles describe isolating MSCs from various human tissues, there are no reports of isolating MSCs from human spinal ligaments, and their localization in situ. If MSCs are found in human spinal ligaments, they could be used to investigate hypertrophy or ossification of spinal ligaments. To isolate and characterize MSCs from human spinal ligaments, spinal ligaments were harvested aseptically from eight patients during surgery for lumbar spinal canal stenosis and ossification of the posterior longitudinal ligament. After collagenase digestion, nucleated cells were seeded at an appropriate density to avoid colony-to-colony contact. Cells were cultured in osteogenic, adipogenic or chondrogenic media to evaluate their multilineage differentiation potential. Immunophenotypic analysis of cell surface markers was performed by flow cytometry. Spinal ligaments were processed for immunostaining using MSC-related antibodies. Cells from human spinal ligaments could be extensively expanded with limited senescence. They were able to differentiate into osteogenic, adipogenic or chondrogenic cells. Flow cytometry revealed that their phenotypic characteristics met the minimum criteria of MSCs. Immunohistochemistry revealed the localization of CD90-positive cells in the collagenous matrix of the ligament, and in adjacent small blood vessels. We isolated and expanded MSCs from human spinal ligaments and demonstrated localization of MSCs in spinal ligaments. These cells may play an indispensable role in elucidating the pathogenesis of numerous spinal diseases. Copyright © 2011 Elsevier Inc. All rights reserved.

  15. Chemical Conversion of Human Fibroblasts into Functional Schwann Cells

    Directory of Open Access Journals (Sweden)

    Eva C. Thoma

    2014-10-01

    Full Text Available Direct transdifferentiation of somatic cells is a promising approach to obtain patient-specific cells for numerous applications. However, conversion across germ-layer borders often requires ectopic gene expression with unpredictable side effects. Here, we present a gene-free approach that allows efficient conversion of human fibroblasts via a transient progenitor stage into Schwann cells, the major glial cell type of peripheral nerves. Using a multikinase inhibitor, we transdifferentiated fibroblasts into transient neural precursors that were subsequently further differentiated into Schwann cells. The resulting induced Schwann cells (iSCs expressed numerous Schwann cell-specific proteins and displayed neurosupportive and myelination capacity in vitro. Thus, we established a strategy to obtain mature Schwann cells from human postnatal fibroblasts under chemically defined conditions without the introduction of ectopic genes.

  16. Calorimetric signatures of human cancer cells and their nuclei

    Energy Technology Data Exchange (ETDEWEB)

    Todinova, S. [Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, Sofia 1113 (Bulgaria); Stoyanova, E. [Department of Molecular Immunology, Institute of Biology and Immunology of Reproduction, Bulgarian Academy of Sciences, Tzarigradsko shose Blvd. 73, Sofia 1113 (Bulgaria); Krumova, S., E-mail: sakrumo@gmail.com [Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, Sofia 1113 (Bulgaria); Iliev, I. [Institute of Experimental Morphology, Pathology and Anthropology with Museum, Acad. G. Bonchev Str., Bl. 25, Sofia 1113 (Bulgaria); Taneva, S.G. [Institute of Biophysics and Biomedical Engineering, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. 21, Sofia 1113 (Bulgaria)

    2016-01-10

    Graphical abstract: - Highlights: • Two temperature ranges are distinguished in the thermograms of cells/nuclei. • Different thermodynamic properties of cancer and normal human cells/nuclei. • Dramatic reduction of the enthalpy of the low-temperature range in cancer cells. • Oxaliplatin and 5-FU affect the nuclear matrix proteins and the DNA stability. - Abstract: The human cancer cell lines HeLa, JEG-3, Hep G2, SSC-9, PC-3, HT-29, MCF7 and their isolated nuclei were characterized by differential scanning calorimetry. The calorimetric profiles differed from normal human fibroblast (BJ) cells in the two well distinguished temperature ranges—the high-temperature range (H{sub T}, due to DNA-containing structures) and the low-temperature range (L{sub T}, assigned to the nuclear matrix and cellular proteins). The enthalpy of the L{sub T} range, and, respectively the ratio of the enthalpies of the L{sub T}- vs. H{sub T}-range, ΔH{sub L}/ΔH{sub H}, is strongly reduced for all cancer cells compared to normal fibroblasts. On the contrary, for most of the cancer nuclei this ratio is higher compared to normal nuclei. The HT-29 human colorectal cancer cells/nuclei differed most drastically from normal human fibroblast cells/nuclei. Our data also reveal that the treatment of HT-29 cancer cells with cytostatic drugs affects not only the DNA replication but also the cellular proteome.

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

    Science.gov (United States)

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

    2016-02-09

    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.

  18. Autoreactivity to Sulfatide by Human Invariant NKT Cells.

    Science.gov (United States)

    Stax, Annelein M; Tuengel, Jessica; Girardi, Enrico; Kitano, Naoki; Allan, Lenka L; Liu, Victor; Zheng, Dongjun; Panenka, William J; Guillaume, Joren; Wong, Chi-Huey; van Calenbergh, Serge; Zajonc, Dirk M; van den Elzen, Peter

    2017-07-01

    Invariant NKT (iNKT) cells are innate-like lymphocytes that recognize lipid Ags presented by CD1d. The prototypical Ag, α-galactosylceramide, strongly activates human and mouse iNKT cells, leading to the assumption that iNKT cell physiology in human and mouse is similar. In this article, we report the surprising finding that human, but not mouse, iNKT cells directly recognize myelin-derived sulfatide presented by CD1d. We propose that sulfatide is recognized only by human iNKT cells because of the unique positioning of the 3-O-sulfated β-galactose headgroup. Surface plasmon resonance shows that the affinity of human CD1d-sulfatide for the iNKT cell receptor is relatively low compared with CD1d-α-galactosylceramide (KD of 19-26 μM versus 1 μM). Apolipoprotein E isolated from human cerebrospinal fluid carries sulfatide that can be captured by APCs and presented by CD1d to iNKT cells. APCs from patients with metachromatic leukodystrophy, who accumulate sulfatides due to a deficiency in arylsulfatase-A, directly activate iNKT cells. Thus, we have identified sulfatide as a self-lipid recognized by human iNKT cells and propose that sulfatide recognition by innate T cells may be an important pathologic feature of neuroinflammatory disease and that sulfatide in APCs may contribute to the endogenous pathway of iNKT cell activation. Copyright © 2017 by The American Association of Immunologists, Inc.

  19. Rhein Induces Apoptosis in Human Breast Cancer Cells

    Directory of Open Access Journals (Sweden)

    Ching-Yao Chang

    2012-01-01

    Full Text Available Human breast cancers cells overexpressing HER2/neu are more aggressive tumors with poor prognosis, and resistance to chemotherapy. This study investigates antiproliferation effects of anthraquinone derivatives of rhubarb root on human breast cancer cells. Of 7 anthraquinone derivatives, only rhein showed antiproliferative and apoptotic effects on both HER2-overexpressing MCF-7 (MCF-7/HER2 and control vector MCF-7 (MCF-7/VEC cells. Rhein induced dose- and time-dependent manners increase in caspase-9-mediated apoptosis correlating with activation of ROS-mediated activation of NF-κB- and p53-signaling pathways in both cell types. Therefore, this study highlighted rhein as processing anti-proliferative activity against HER2 overexpression or HER2-basal expression in breast cancer cells and playing important roles in apoptotic induction of human breast cancer cells.

  20. Cardiac Progenitor Cell Extraction from Human Auricles

    KAUST Repository

    Di Nardo, Paolo

    2017-02-22

    For many years, myocardial tissue has been considered terminally differentiated and, thus, incapable of regenerating. Recent studies have shown, instead, that cardiomyocytes, at least in part, are slowly substituted by new cells originating by precursor cells mostly embedded into the heart apex and in the atria. We have shown that an elective region of progenitor cell embedding is represented by the auricles, non-contractile atria appendages that can be easily sampled without harming the patient. The protocol here reported describes how from auricles a population of multipotent, cardiogenic cells can be isolated, cultured, and differentiated. Further studies are needed to fully exploit this cell population, but, sampling auricles, it could be possible to treat cardiac patients using their own cells circumventing rejection or organ shortage limitations.

  1. Epithelial cells as alternative human biomatrices for comet assay

    OpenAIRE

    Emilio eRojas; Yolanda eLorenzo; Kristiane eHuag-Berg; Bjørn eNicolaissen; Mahara eValverde

    2014-01-01

    The comet assay is a valuable experimental tool aimed at mapping DNA damage in human cells in vivo for environmental and occupational monitoring, as well as for therapeutic purposes, such as storage prior to transplant, during tissue engineering, and in experimental ex vivo assays. Furthermore, due to its great versatility, the comet assay allows to explore the use of alternative cell types to assess DNA damage, such as epithelial cells. Epithelial cells, as specialized components of many org...

  2. 3D map of the human corneal endothelial cell

    OpenAIRE

    Zhiguo He; Fabien Forest; Philippe Gain; Damien Rageade; Aurélien Bernard; Sophie Acquart; Michel Peoc’h; Dennis M. Defoe; Gilles Thuret

    2016-01-01

    Corneal endothelial cells (CECs) are terminally differentiated cells, specialized in regulating corneal hydration and transparency. They are highly polarized flat cells that separate the cornea from the aqueous humor. Their apical surface, in contact with aqueous humor is hexagonal, whereas their basal surface is irregular. We characterized the structure of human CECs in 3D using confocal microscopy of immunostained whole corneas in which cells and their interrelationships remain intact. Hexa...

  3. Cell sources for in vitro human liver cell culture models.

    Science.gov (United States)

    Zeilinger, Katrin; Freyer, Nora; Damm, Georg; Seehofer, Daniel; Knöspel, Fanny

    2016-09-01

    In vitro liver cell culture models are gaining increasing importance in pharmacological and toxicological research. The source of cells used is critical for the relevance and the predictive value of such models. Primary human hepatocytes (PHH) are currently considered to be the gold standard for hepatic in vitro culture models, since they directly reflect the specific metabolism and functionality of the human liver; however, the scarcity and difficult logistics of PHH have driven researchers to explore alternative cell sources, including liver cell lines and pluripotent stem cells. Liver cell lines generated from hepatomas or by genetic manipulation are widely used due to their good availability, but they are generally altered in certain metabolic functions. For the past few years, adult and pluripotent stem cells have been attracting increasing attention, due their ability to proliferate and to differentiate into hepatocyte-like cells in vitro However, controlling the differentiation of these cells is still a challenge. This review gives an overview of the major human cell sources under investigation for in vitro liver cell culture models, including primary human liver cells, liver cell lines, and stem cells. The promises and challenges of different cell types are discussed with a focus on the complex 2D and 3D culture approaches under investigation for improving liver cell functionality in vitro Finally, the specific application options of individual cell sources in pharmacological research or disease modeling are described. © 2016 by the Society for Experimental Biology and Medicine.

  4. Dark cells in human oral leukoplakias

    Energy Technology Data Exchange (ETDEWEB)

    Klein-Szanto, A.J.P. (Oak Ridge National Lab., TN); Sega, M.; Banoczy, J.; Albrecht, M.

    1982-01-01

    Dark basal keratinocytes, characterized by a strong affinity for basic dyes and by electron density of cytoplasm and nucleus, could be recognized in eleven oral leukoplakias. The percentage of dark cells was higher in the group comprising leukoplakias verrucosa, and erosiva (28% of the basal cells) than in the leukoplakia simplex group (10%). The presence of these cells is a good indicator of the degree of histological dysplasia and correlates well with the preneoplastic potential of these lesions.

  5. The contribution of human/non-human animal chimeras to stem cell research

    Directory of Open Access Journals (Sweden)

    Sonya Levine

    2017-10-01

    Full Text Available Chimeric animals are made up of cells from two separate zygotes. Human/non-human animal chimeras have been used for a number of research purposes, including human disease modeling. Pluripotent stem cell (PSC research has relied upon the chimera approach to examine the developmental potential of stem cells, to determine the efficacy of cell replacement therapies, and to establish a means of producing human organs. Based on ethical issues, this work has faced pushback from various sources including funding agencies. We discuss here the essential role these studies have played, from gaining a better understanding of human biology to providing a stepping stone to human disease treatments. We also consider the major ethical issues, as well as the current status of support for this work in the United States.

  6. PPAR-γ Regulates Trophoblast Differentiation in the BeWo Cell Model

    Science.gov (United States)

    Levytska, Khrystyna; Drewlo, Sascha; Baczyk, Dora; Kingdom, John

    2014-01-01

    Common pregnancy complications, such as severe preeclampsia and intrauterine growth restriction, disrupt pregnancy progression and impair maternal and fetal wellbeing. Placentas from such pregnancies exhibit lesions principally within the syncytiotrophoblast (SCT), a layer in direct contact with maternal blood. In humans and mice, glial cell missing-1 (GCM-1) promotes differentiation of underlying cytotrophoblast cells into the outer SCT layer. GCM-1 may be regulated by the transcription factor peroxisome proliferator-activated receptor-gamma (PPAR-γ); in mice, PPAR-γ promotes labyrinthine trophoblast differentiation via Gcm-1, and, as we previously demonstrated, PPAR-γ activation ameliorates disease features in rat model of preeclampsia. Here, we aimed to characterize the baseline activity of PPAR-γ in the human choriocarcinoma BeWo cell line that mimics SCT formation in vitro and modulate PPAR-γ activity to study its effects on cell proliferation versus differentiation. We report a novel negative autoregulatory mechanism between PPAR-γ activity and expression and show that blocking PPAR-γ activity induces cell proliferation at the expense of differentiation, while these remain unaltered following treatment with the agonist rosiglitazone. Gaining a deeper understanding of the role and activity of PPAR-γ in placental physiology will offer new avenues for the development of secondary prevention and/or treatment options for placentally-mediated pregnancy complications. PMID:24711815

  7. PPAR-γ Regulates Trophoblast Differentiation in the BeWo Cell Model

    Directory of Open Access Journals (Sweden)

    Khrystyna Levytska

    2014-01-01

    Full Text Available Common pregnancy complications, such as severe preeclampsia and intrauterine growth restriction, disrupt pregnancy progression and impair maternal and fetal wellbeing. Placentas from such pregnancies exhibit lesions principally within the syncytiotrophoblast (SCT, a layer in direct contact with maternal blood. In humans and mice, glial cell missing-1 (GCM-1 promotes differentiation of underlying cytotrophoblast cells into the outer SCT layer. GCM-1 may be regulated by the transcription factor peroxisome proliferator-activated receptor-gamma (PPAR-γ; in mice, PPAR-γ promotes labyrinthine trophoblast differentiation via Gcm-1, and, as we previously demonstrated, PPAR-γ activation ameliorates disease features in rat model of preeclampsia. Here, we aimed to characterize the baseline activity of PPAR-γ in the human choriocarcinoma BeWo cell line that mimics SCT formation in vitro and modulate PPAR-γ activity to study its effects on cell proliferation versus differentiation. We report a novel negative autoregulatory mechanism between PPAR-γ activity and expression and show that blocking PPAR-γ activity induces cell proliferation at the expense of differentiation, while these remain unaltered following treatment with the agonist rosiglitazone. Gaining a deeper understanding of the role and activity of PPAR-γ in placental physiology will offer new avenues for the development of secondary prevention and/or treatment options for placentally-mediated pregnancy complications.

  8. Immune and Inflammatory Cell Composition of Human Lung Cancer Stroma.

    Directory of Open Access Journals (Sweden)

    G-Andre Banat

    Full Text Available Recent studies indicate that the abnormal microenvironment of tumors may play a critical role in carcinogenesis, including lung cancer. We comprehensively assessed the number of stromal cells, especially immune/inflammatory cells, in lung cancer and evaluated their infiltration in cancers of different stages, types and metastatic characteristics potential. Immunohistochemical analysis of lung cancer tissue arrays containing normal and lung cancer sections was performed. This analysis was combined with cyto-/histomorphological assessment and quantification of cells to classify/subclassify tumors accurately and to perform a high throughput analysis of stromal cell composition in different types of lung cancer. In human lung cancer sections we observed a significant elevation/infiltration of total-T lymphocytes (CD3+, cytotoxic-T cells (CD8+, T-helper cells (CD4+, B cells (CD20+, macrophages (CD68+, mast cells (CD117+, mononuclear cells (CD11c+, plasma cells, activated-T cells (MUM1+, B cells, myeloid cells (PD1+ and neutrophilic granulocytes (myeloperoxidase+ compared with healthy donor specimens. We observed all of these immune cell markers in different types of lung cancers including squamous cell carcinoma, adenocarcinoma, adenosquamous cell carcinoma, small cell carcinoma, papillary adenocarcinoma, metastatic adenocarcinoma, and bronchioloalveolar carcinoma. The numbers of all tumor-associated immune cells (except MUM1+ cells in stage III cancer specimens was significantly greater than those in stage I samples. We observed substantial stage-dependent immune cell infiltration in human lung tumors suggesting that the tumor microenvironment plays a critical role during lung carcinogenesis. Strategies for therapeutic interference with lung cancer microenvironment should consider the complexity of its immune cell composition.

  9. Antiproliferative action of metformin in human lung cancer cell lines.

    Science.gov (United States)

    Ashinuma, Hironori; Takiguchi, Yuichi; Kitazono, Satoru; Kitazono-Saitoh, Miyako; Kitamura, Atsushi; Chiba, Tetsuhiro; Tada, Yuji; Kurosu, Katsushi; Sakaida, Emiko; Sekine, Ikuo; Tanabe, Nobuhiro; Iwama, Atsushi; Yokosuka, Osamu; Tatsumi, Koichiro

    2012-07-01

    The oral antidiabetic agent metformin has anticancer properties, probably via adenosine monophosphate-activated protein kinase activation. In the present study, growth inhibition was assessed by a clonogenic and by a cell survival assay, apoptosis induction was assessed by Hoechst staining and caspase activities and cell cycle alteration after exposure to metformin, and the interaction of metformin with cisplatin in vitro were elucidated in four human lung cancer cell lines representing squamous, adeno-, large cell and small cell carcinoma. Clonogenicity and cell proliferation were inhibited by metformin in all the cell lines examined. This inhibitory effect was not specific to cancer cells because it was also observed in a non-transformed human mesothelial cell line and in mouse fibroblast cell lines. Inhibition of clonogenicity was observed only when the cells were exposed to metformin for a long period, (10 days) and the surviving fraction, obtained after inhibiting proliferation by increasing the dose, reached a plateau at approximately 0.1-0.3, indicating the cytostatic characteristics of metformin. Metformin induced significant apoptosis only in the small cell carcinoma cell line. A tendency of cell cycle accumulation at the G0/G1 phase was observed in all four cell lines. Cisplatin, in a dose-dependent manner, severely antagonized the growth inhibitory effect of metformin, and even reversed the effect in three cell lines but not in the adenocarcinoma cell line. The present data obtained using various histological types of human lung cancer cell lines in vitro illustrate the cytostatic nature of metformin and its cytoprotective properties against cisplatin.

  10. Human retinal pigment epithelial cell-induced apoptosis in activated T cells

    DEFF Research Database (Denmark)

    Jørgensen, A; Wiencke, A K; la Cour, M

    1998-01-01

    human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction...... of apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...... block alloactivation, had no inhibitory effect on RPE-mediated T-cell apoptotic responses in MHC class II-specific CD4+ T-cell lines. CONCLUSIONS: Retinal pigment epithelial cells express FasL and induce TCR-independent apoptosis in activated human T cells through Fas-FasL interaction. Retinal pigment...

  11. Human retinal pigment epithelial cell-induced apoptosis in activated T cells

    DEFF Research Database (Denmark)

    Jørgensen, A; Wiencke, A K; la Cour, M

    1998-01-01

    block alloactivation, had no inhibitory effect on RPE-mediated T-cell apoptotic responses in MHC class II-specific CD4+ T-cell lines. CONCLUSIONS: Retinal pigment epithelial cells express FasL and induce TCR-independent apoptosis in activated human T cells through Fas-FasL interaction. Retinal pigment...... human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction...... of apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...

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

    Science.gov (United States)

    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. © The Author 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  13. Hydroxytyrosol Protects against Oxidative DNA Damage in Human Breast Cells

    Directory of Open Access Journals (Sweden)

    José J. Gaforio

    2011-10-01

    Full Text Available Over recent years, several studies have related olive oil ingestion to a low incidence of several diseases, including breast cancer. Hydroxytyrosol and tyrosol are two of the major phenols present in virgin olive oils. Despite the fact that they have been linked to cancer prevention, there is no evidence that clarifies their effect in human breast tumor and non-tumor cells. In the present work, we present hydroxytyrosol and tyrosol’s effects in human breast cell lines. Our results show that hydroxytyrosol acts as a more efficient free radical scavenger than tyrosol, but both fail to affect cell proliferation rates, cell cycle profile or cell apoptosis in human mammary epithelial cells (MCF10A or breast cancer cells (MDA-MB-231 and MCF7. We found that hydroxytyrosol decreases the intracellular reactive oxygen species (ROS level in MCF10A cells but not in MCF7 or MDA-MB-231 cells while very high amounts of tyrosol is needed to decrease the ROS level in MCF10A cells. Interestingly, hydroxytyrosol prevents oxidative DNA damage in the three breast cell lines. Therefore, our data suggest that simple phenol hydroxytyrosol could contribute to a lower incidence of breast cancer in populations that consume virgin olive oil due to its antioxidant activity and its protection against oxidative DNA damage in mammary cells.

  14. Myxoma virus is oncolytic for human pancreatic adenocarcinoma cells.

    Science.gov (United States)

    Woo, Yanghee; Kelly, Kaitlyn J; Stanford, Marianne M; Galanis, Charles; Chun, Yun Shin; Fong, Yuman; McFadden, Grant

    2008-08-01

    Viral oncolytic therapy, which seeks to exploit the use of live viruses to treat cancer, has shown promise in the treatment of cancers resistant to conventional anticancer therapies. Among the most difficult to treat cancers is advanced pancreatic adenocarcinoma. Our study investigates the ability of a novel oncolytic agent, myxoma virus, to infect, productively replicate in, and kill human pancreatic cancer cells in vitro. The myxoma virus vMyxgfp was tested against a panel of human pancreatic adenocarcinoma cell lines. Infectivity, viral proliferation, and tumor cell kill were assessed. Infection of tumor cells was assessed by expression of the marker gene enhanced green fluorescent protein (e-GFP). vMyxgfp had the ability to infect all pancreatic cancer cell lines tested. Killing of tumor cells varied among the 6 cell lines tested, ranging from >90% cell kill at 7 days for the most sensitive Panc-1 cells, to 39% in the most resistant cell line Capan-2. Sensitivity correlated to replication of virus, and was found to maximally exhibit a four-log increase in foci-forming units for the most sensitive Panc-1 cells within 72 h. Our study demonstrates for the first time the ability of the myxoma virus to productively infect, replicate in, and lyse human pancreatic adenocarcinoma cells in vitro. These data encourage further investigation of this virus, which is pathogenic only in rabbits, for treatment of this nearly uniformly fatal cancer.

  15. Variations in Humanized and Defined Culture Conditions Supporting Derivation of New Human Embryonic Stem Cell Lines

    DEFF Research Database (Denmark)

    Fletcher, Judy M; Ferrier, Patricia M; Gardner, John O

    2006-01-01

    The evolution of "humanized" (i.e., free of animal sourced reagents) and ultimately chemically defined culture systems for human embryo stem cell (hESC) isolation and culture is of importance to improving their efficacy and safety in research and therapeutic applications. This can be achieved...... serum-free medium (SFM) containing only human sourced and recombinant protein. Further, outgrowth of embryonic cells from whole blastocysts in both media could be achieved for up to 1 week without reliance on feeder cells. All variant conditions sustained undifferentiated cell status, a stable karyotype......, with a transitional requirement for human feeder cells. This represents another sequential step in the generation of therapeutic grade stem cells with reduced risk of zoonotic pathogen transmission....

  16. On the Sulfation and Methylation of Catecholestrogens in Human Mammary Epithelial Cells and Breast Cancer Cells

    National Research Council Canada - National Science Library

    Hui, Ying; Yasuda, Shin; Liu, Ming-Yih; Wu, Yi-yong; Liu, Ming-Cheh

    2008-01-01

    .... The present study was designed to examine the role of sulfation in the metabolism of CEs. MCF-7 breast cancer cells and MCF 10A human mammary epithelial cells were metabolically labeled with [35S...

  17. Human normal bronchial epithelial cells: a novel in vitro cell model for toxicity evaluation

    National Research Council Canada - National Science Library

    Feng, Wenqiang; Guo, Juanjuan; Huang, Haiyan; Xia, Bo; Liu, Hongya; Li, Jie; Lin, Shaolin; Li, Tiyuan; Liu, Jianjun; Li, Hui

    2015-01-01

    .... hTERT or viral genes transduced human cells are currently widely used for these studies, while these cells exhibited abnormal differentiation potential or response to biological and chemical signals...

  18. [Isolation and identification of side population cells in human lung adenocarcinoma cell line A549].

    Science.gov (United States)

    Xie, Tong; Li, Li; Li, Dan-rong; Mao, Nai-quan; Liu, De-seng; Zuo, Chuan-tian; Zhang, Wei; Huang, Ding-ming

    2011-02-01

    To isolate and characterize the side population cells (SP cells) in the lung adenocarcinomas cell line A549. The protein expression of ABCG2 in human lung adenocarcinoma cell line A549 was detected by immunohistochemistry. SP and NSP cells in the cell line A549 were isolated by FACS, and their differentiation was analysed. ABCG2 expression in the two cell subsets was detected by RT-PCR. The cell growth curves, cell division indexes, cell cycles, plate clone formation tests, migration and invasion assays, chemotherapeutic susceptibility tests, tests of the intracellular drug levels, and the tumor cell implantation experiments on nude mice were applied to study the biological properties of the two cell subsets. The expression of ABCG2 in the transplanted tumor in nude mice was detected by immunohistochemistry and RT-PCR. The positive rate of ABCG2 expression in the A549 cells by immunohistochemistry was 2.13%. SP and NSP cells were isolated by FACS. The SP cells could produce both SP and NSP cells, while NSP cells only produced NSP cells. SP cells expressed ABCG2, but NSP cells did not. The proliferation and migration abilities of the two cell subsets were similar, but the invasion and tumorigenic ability of SP cells was significantly higher than that of NSP cells. The susceptibilities to DDP and its intracellular levels of the two cell subsets were similar, but the susceptibilities to 5-FU, VP16, NVB and GEM and their intracellular levels of NSP cells were significantly higher than those of the SP cells. SP cells in the human lung adenocarcinomas cell line A549 is enriched with tumor stem cells. An effective way to get lung adenocarcinomas stem cells is to isolate SP cells by FACS.

  19. The association between human papillomavirus and oropharyngeal squamous cell Carcinoma

    DEFF Research Database (Denmark)

    Walvik, Lena; Svensson, Amanda Björk; Friborg, Jeppe

    2016-01-01

    There is emerging evidence of the association between human papillomavirus and a subset of head and neck cancers. However, the role of human papillomavirus as a causal factor is still debated. This review addresses the association between human papillomavirus and oropharyngeal squamous cell...... carcinoma using the Bradford Hill criteria. The strength of the association is supported by, detection of human papillomavirus infection and antibodies prior to oropharyngeal squamous cell carcinoma. This is furthermore reinforced by the absence of human papillomavirus DNA in healthy tonsils....... The association is geographically consistent throughout the economically developed world. The presence and integration of high-risk human papillomavirus genome in tonsillar tumours, and expression of viral oncogenes, are specific and plausible. Analogous to human papillomavirus in cervical cancer, the rising...

  20. Phenotypic and functional heterogeneity of human memory B cells.

    Science.gov (United States)

    Sanz, Iñaki; Wei, Chungwen; Lee, F Eun-Hyung; Anolik, Jennifer

    2008-02-01

    Memory B cells are more heterogeneous than previously thought. Given that B cells play powerful antibody-independent effector functions, it seems reasonable to assume division of labor between distinct memory B cells subpopulations in both protective and pathogenic immune responses. Here we review the information emerging regarding the heterogeneity of human memory B cells. A better understanding of this topic should greatly improve our ability to target specific B cell subsets either in vaccine responses or in autoimmune diseases and organ rejection among other pathological conditions where B cells play central pathogenic roles.

  1. Identifying gene expression modules that define human cell fates.

    Science.gov (United States)

    Germanguz, I; Listgarten, J; Cinkornpumin, J; Solomon, A; Gaeta, X; Lowry, W E

    2016-05-01

    Using a compendium of cell-state-specific gene expression data, we identified genes that uniquely define cell states, including those thought to represent various developmental stages. Our analysis sheds light on human cell fate through the identification of core genes that are altered over several developmental milestones, and across regional specification. Here we present cell-type specific gene expression data for 17 distinct cell states and demonstrate that these modules of genes can in fact define cell fate. Lastly, we introduce a web-based database to disseminate the results. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  2. The synthetic cannabinoid WIN-55,212 induced-apoptosis in cytotrophoblasts cells by a mechanism dependent on CB1 receptor.

    Science.gov (United States)

    Almada, Marta; Costa, Lia; Fonseca, Bruno Miguel; Amaral, Cristina; Teixeira, Natércia; Correia-da-Silva, Georgina

    2017-06-15

    The endocannabinoid system has evolved as a key regulator in several pathological and physiological processes, including placentation, decidualization and implantation. In addition, it is known that Cannabis and cannabinoids negatively affect female reproduction. Although, the biological action of synthetic cannabinoids, such as WIN-55,212, in human fertility and pregnancy outcome remain to be unveiled. A tight balance between proliferation, differentiation and apoptosis of trophoblast cells is required for placental development and pregnancy outcome. Therefore, in this work, the effects of the synthetic cannabinoid WIN-55,212 in placental cytotrophoblast cells were explored. For that, it was used a human choriocarcinoma cell line, BeWo cells, and primary cultures of human cytotrophoblasts isolated from term placentas. Results demonstrate that this synthetic cannabinoid induces cell cycle arrest. We also observed that cell viability loss was associated with a disruption of mitochondrial membrane potential and activation of caspases -9 and -3/-7 independently of reactive oxygen species (ROS) production or recruitment of the endoplasmic reticulum stress marker CHOP. Moreover, these effects were prevented by pre-incubation with a selective cannabinoid receptor 1 (CBR1) antagonist (AM281). Thus, our results provide strong evidences of the apoptotic process induced by WIN-55,212 through the activation of the CBR1, which may reveal the impact of cannabinoids consumption during placental development. Copyright © 2017 Elsevier B.V. All rights reserved.

  3. Using Human Induced Pluripotent Stem Cells to Model Skeletal Diseases.

    Science.gov (United States)

    Barruet, Emilie; Hsiao, Edward C

    2016-01-01

    Musculoskeletal disorders affecting the bones and joints are major health problems among children and adults. Major challenges such as the genetic origins or poor diagnostics of severe skeletal disease hinder our understanding of human skeletal diseases. The recent advent of human induced pluripotent stem cells (human iPS cells) provides an unparalleled opportunity to create human-specific models of human skeletal diseases. iPS cells have the ability to self-renew, allowing us to obtain large amounts of starting material, and have the potential to differentiate into any cell types in the body. In addition, they can carry one or more mutations responsible for the disease of interest or be genetically corrected to create isogenic controls. Our work has focused on modeling rare musculoskeletal disorders including fibrodysplasia ossificans progressive (FOP), a congenital disease of increased heterotopic ossification. In this review, we will discuss our experiences and protocols differentiating human iPS cells toward the osteogenic lineage and their application to model skeletal diseases. A number of critical challenges and exciting new approaches are also discussed, which will allow the skeletal biology field to harness the potential of human iPS cells as a critical model system for understanding diseases of abnormal skeletal formation and bone regeneration.

  4. The effect of human papillomavirus infection on sperm cell motility.

    Science.gov (United States)

    Lai, Y M; Lee, J F; Huang, H Y; Soong, Y K; Yang, F P; Pao, C C

    1997-06-01

    To investigate the presence of human papillomavirus (HPV) in human sperm cells and to evaluate potential effects of HPV on the sperm functions. A descriptive clinical study. Specimens of semen were collected from 24 randomly selected patients who attended the fertility clinics at Chang Gung Memorial Hospital. The presence of HPV DNA and RNA were examined by polymerase chain reaction. Semen quality and sperm cell function were analyzed by computer-aided autoanalyzer. Human papillomavirus type 16 DNA and RNA were found in 6 (25%) and 2 (8%) of the sperm cells specimens, respectively. Human papillomavirus type 18 DNA and RNA were present in 11 (46%) and 5 (21%) of the same sperm cells specimens, respectively. Incidence of asthenozoospermia among patients infected with either HPV was significantly higher than in those without HPV in their sperm cells (75% versus 8%). Although performance of curvilinear velocity, straight-line velocity, and mean amplitude of lateral head displacement was significantly lower in HPV-infected specimens, the differences of linearity, beat cross frequency, and straightness were not statistically significant. These results suggest that human papillomavirus can be found in human sperm cells and that certain HPV-specific genes are actively transcribed. Sperm motility parameters seem to be affected by the presence of HPV in the sperm cells, and also the incidence of asthenozoospermia may be associated with HPV infection.

  5. [Progress in the research of germ cell from human embryonic stem cells].

    Science.gov (United States)

    Guo, Xin; Cai, Zhi-Ming; Gui, Yao-Ting

    2006-01-01

    As the development of spontaneous differentiation of germ cells and gametogenesis from mouse embryonic stem cells (mES) in vitro, hES (human embryonic stem cells) also have potential to differentiated into germ cells in theory. This review focuses on the stem cells niches and genes regulating the hES differentiation toward germ cells, as well as the recent advance and application on the reproductive medicine and therapy of infertility.

  6. Human Red Cells With Paroxysmal Nocturnal Haemoglobinuria ...

    African Journals Online (AJOL)

    Paroxysmal nocturnal haemoglobinuria (PNH) is an acquired blood disorder characterised by acute intravascular haemolysis, due to increased susceptibility of red cells to complement -mediated lysis. Any red cell abnormality that would affect the invasion and growth of Plasmodium falciparum would serve as a useful tool ...

  7. Controlled Delivery of Human Cells by Temperature Responsive Microcapsules

    Science.gov (United States)

    Mak, W.C.; Olesen, K.; Sivlér, P.; Lee, C.J.; Moreno-Jimenez, I.; Edin, J.; Courtman, D.; Skog, M.; Griffith, M.

    2015-01-01

    Cell therapy is one of the most promising areas within regenerative medicine. However, its full potential is limited by the rapid loss of introduced therapeutic cells before their full effects can be exploited, due in part to anoikis, and in part to the adverse environments often found within the pathologic tissues that the cells have been grafted into. Encapsulation of individual cells has been proposed as a means of increasing cell viability. In this study, we developed a facile, high throughput method for creating temperature responsive microcapsules comprising agarose, gelatin and fibrinogen for delivery and subsequent controlled release of cells. We verified the hypothesis that composite capsules combining agarose and gelatin, which possess different phase transition temperatures from solid to liquid, facilitated the destabilization of the capsules for cell release. Cell encapsulation and controlled release was demonstrated using human fibroblasts as model cells, as well as a therapeutically relevant cell line—human umbilical vein endothelial cells (HUVECs). While such temperature responsive cell microcapsules promise effective, controlled release of potential therapeutic cells at physiological temperatures, further work will be needed to augment the composition of the microcapsules and optimize the numbers of cells per capsule prior to clinical evaluation. PMID:26096147

  8. Controlled Delivery of Human Cells by Temperature Responsive Microcapsules

    Directory of Open Access Journals (Sweden)

    W.C. Mak

    2015-06-01

    Full Text Available Cell therapy is one of the most promising areas within regenerative medicine. However, its full potential is limited by the rapid loss of introduced therapeutic cells before their full effects can be exploited, due in part to anoikis, and in part to the adverse environments often found within the pathologic tissues that the cells have been grafted into. Encapsulation of individual cells has been proposed as a means of increasing cell viability. In this study, we developed a facile, high throughput method for creating temperature responsive microcapsules comprising agarose, gelatin and fibrinogen for delivery and subsequent controlled release of cells. We verified the hypothesis that composite capsules combining agarose and gelatin, which possess different phase transition temperatures from solid to liquid, facilitated the destabilization of the capsules for cell release. Cell encapsulation and controlled release was demonstrated using human fibroblasts as model cells, as well as a therapeutically relevant cell line—human umbilical vein endothelial cells (HUVECs. While such temperature responsive cell microcapsules promise effective, controlled release of potential therapeutic cells at physiological temperatures, further work will be needed to augment the composition of the microcapsules and optimize the numbers of cells per capsule prior to clinical evaluation.

  9. Inhibition of fatty acid metabolism reduces human myeloma cells proliferation.

    Directory of Open Access Journals (Sweden)

    José Manuel Tirado-Vélez

    Full Text Available Multiple myeloma is a haematological malignancy characterized by the clonal proliferation of plasma cells. It has been proposed that targeting cancer cell metabolism would provide a new selective anticancer therapeutic strategy. In this work, we tested the hypothesis that inhibition of β-oxidation and de novo fatty acid synthesis would reduce cell proliferation in human myeloma cells. We evaluated the effect of etomoxir and orlistat on fatty acid metabolism, glucose metabolism, cell cycle distribution, proliferation, cell death and expression of G1/S phase regulatory proteins in myeloma cells. Etomoxir and orlistat inhibited β-oxidation and de novo fatty acid synthesis respectively in myeloma cells, without altering significantly glucose metabolism. These effects were associated with reduced cell viability and cell cycle arrest in G0/G1. Specifically, etomoxir and orlistat reduced by 40-70% myeloma cells proliferation. The combination of etomoxir and orlistat resulted in an additive inhibitory effect on cell proliferation. Orlistat induced apoptosis and sensitized RPMI-8226 cells to apoptosis induction by bortezomib, whereas apoptosis was not altered by etomoxir. Finally, the inhibitory effect of both drugs on cell proliferation was associated with reduced p21 protein levels and phosphorylation levels of retinoblastoma protein. In conclusion, inhibition of fatty acid metabolism represents a potential therapeutic approach to treat human multiple myeloma.

  10. Cannabinoids induce incomplete maturation of cultured human leukemia cells

    Energy Technology Data Exchange (ETDEWEB)

    Murison, G.; Chubb, C.B.H.; Maeda, S.; Gemmell, M.A.; Huberman, E.

    1987-08-01

    Monocyte maturation markers were induced in cultured human myeloblastic ML-2 leukemia cells after treatment for 1-6 days with 0.03-30 ..mu..M ..delta../sup 9/-tetrahydrocannabinol (THC), the major psychoactive component of marijuana. After a 2-day or longer treatment, 2- to 5-fold increases were found in the percentages of cells exhibiting reactivity with either the murine OKM1 monoclonal antibody of the Leu-M5 monoclonal antibody, staining positively for nonspecific esterase activity, and displaying a promonocyte morphology. The increases in these differentiation markers after treatment with 0.03-1 ..mu..M THC were dose dependent. At this dose range, THC did not cause an inhibition of cell growth. The THC-induced cell maturation was also characterized by specific changes in the patterns of newly synthesized proteins. The THC-induced differentiation did not, however, result in cells with a highly developed mature monocyte phenotype. However, treatment of these incompletely matured cells with either phorbol 12-myristate 13-acetate of 1..cap alpha..,25-dihydroxycholecalciferol, which are inducers of differentiation in myeloid leukemia cells (including ML-2 cells), produced cells with a mature monocyte morphology. The ML-2 cell system described here may be a useful tool for deciphering critical biochemical events that lead to the cannabinoid-induced incomplete cell differentiation of ML-2 cells and other related cell types. Findings obtained from this system may have important implications for studies of cannabinoid effects on normal human bone-marrow progenitor cells.

  11. Human B cells produce chemokine CXCL10 in the presence of Mycobacterium tuberculosis specific T cells.

    Science.gov (United States)

    Hoff, Soren T; Salman, Ahmed M; Ruhwald, Morten; Ravn, Pernille; Brock, Inger; Elsheikh, Nabila; Andersen, Peter; Agger, Else Marie

    2015-01-01

    The role of B cells in human host response to Mycobacterium tuberculosis (Mtb) infection is still controversial, but recent evidence suggest that B cell follicle like structures within the lung may influence host responses through regulation of the local cytokine environment. A candidate for such regulation could be the chemokine CXCL10. CXCL10 is mainly produced by human monocytes, but a few reports have also found CXCL10 production by human B cells. The objective of this study was to investigate CXCL10 production by human B cells in response to in vitro stimulation with Mtb antigens. We analyzed human blood samples from 30 volunteer donors using multiparameter flow cytometry, and identified a subgroup of B cells producing CXCL10 in response to in vitro stimulation with antigens. T cells did not produce CXCL10, but CXCL10 production by B cells appeared to be mediated via IFN-γ and dependent on contact with antigen-specific T cells recognizing the antigen. Human B cells are able to produce CXCL10 in an IFN-γ and T cell contact-dependent manner. The present findings suggest a possible mechanism through which B cells in part may influence granuloma formation in human tuberculosis (TB) and participate in infection control. Copyright © 2014 Elsevier Ltd. All rights reserved.

  12. Chlamydia pneumoniae induces aponecrosis in human aortic smooth muscle cells

    Directory of Open Access Journals (Sweden)

    Walch Michael

    2005-01-01

    Full Text Available Abstract Background The intracellular bacterium Chlamydia pneumoniae is suspected to play a role in formation and progression of atherosclerosis. Many studies investigated cell death initiation versus inhibition by Chlamydia pneumoniae in established cell lines but nothing is known in primary human aortic smooth muscle cells, a cell type among others known to be involved in the formation of the atherosclerotic plaque. Type of cell death was analyzed by various methods in primary aortic smooth muscle cells after infection with Chlamydia pneumoniae to investigate a possible pathogenic link in atherosclerosis. Results Chlamydiae were found to be localized up to 72 h post infection in aortic smooth muscle cells either as single bacteria or inside of large inclusions. Quantification of host cell death by lactate dehydrogenase release assay revealed strictly dose and time dependent lysis for all tested isolates of Chlamydia pneumoniae. Phosphatidylserine exposure was detected by flow cytometry in Chlamydia pneumoniae infected cells. Ultrastructure of Chlamydia pneumoniae infected human aortic smooth muscle cells showed extensive membrane- and organelle damage, chromatin condensation but no nuclear fragmentation. DNA fragmentation as well as cell membrane permeability was analyzed by TUNEL and NHS-biotin staining and occurred exclusively in cells carrying Chlamydia pneumoniae spots but not in smooth muscle cells with inclusions. These morphological features of cell death were not accompanied by an activation of caspase-3 as revealed by analysis of enzyme activity but involved mitochondrial membrane depolarization as shown by TMRE uptake and release of cytochrome c from mitochondria. Conclusion This study provides evidence that Chlamydia pneumoniae induce a spot like infection in human aortic smooth muscle cells, which results in a chimeric cell death with both apoptotic and necrotic characteristics. This aponecrotic cell death may assist chronic

  13. Rho GTPases and regulation of cell migration and polarization in human corneal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Aihua Hou

    Full Text Available PURPOSE: Epithelial cell migration is required for regeneration of tissues and can be defective in a number of ocular surface diseases. This study aimed to determine the expression pattern of Rho family small G-proteins in human corneal epithelial cells to test their requirement in directional cell migration. METHODS: Rho family small G-protein expression was assessed by reverse transcription-polymerase chain reaction. Dominant-inhibitory constructs encoding Rho proteins or Rho protein targeting small interfering RNA were transfected into human corneal epithelial large T antigen cells, and wound closure rate were evaluated by scratch wounding assay, and a complementary non-traumatic cell migration assay. Immunofluorescence staining was performed to study cell polarization and to assess Cdc42 downstream effector. RESULTS: Cdc42, Chp, Rac1, RhoA, TC10 and TCL were expressed in human corneal epithelial cells. Among them, Cdc42 and TCL were found to significantly affect cell migration in monolayer scratch assays. These results were confirmed through the use of validated siRNAs directed to Cdc42 and TCL. Scramble siRNA transfected cells had high percentage of polarized cells than Cdc42 or TCL siRNA transfected cells at the wound edge. We showed that the Cdc42-specific effector p21-activated kinase 4 localized predominantly to cell-cell junctions in cell monolayers, but failed to translocate to the leading edge in Cdc42 siRNA transfected cells after monolayer wounding. CONCLUSION: Rho proteins expressed in cultured human corneal epithelial cells, and Cdc42, TCL facilitate two-dimensional cell migration in-vitro. Although silencing of Cdc42 and TCL did not noticeably affect the appearance of cell adhesions at the leading edge, the slower migration of these cells indicates both GTP-binding proteins play important roles in promoting cell movement of human corneal epithelial cells.

  14. Toona Sinensis Extracts Induced Cell Cycle Arrest and Apoptosis in the Human Lung Large Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Cheng-Yuan Wang

    2010-02-01

    Full Text Available Toona sinensis extracts have been shown to exhibit anti-cancer effects in human ovarian cancer cell lines, human promyelocytic leukemia cells and human lung adenocarcinoma. Its safety has also been confirmed in animal studies. However, its anti-cancer properties in human lung large cell carcinoma have not been studied. Here, we used a powder obtained by freeze-drying the super-natant of centrifuged crude extract from Toona sinensis leaves (TSL-1 to treat the human lung carcinoma cell line H661. Cell viability was evaluated by the 3-(4-,5-dimethylthiazol-2-yl-2,5-diphenyl tetrazolium bromide assay. Flow cytometry analysis revealed that TSL-1 blocked H661 cell cycle progression. Western blot analysis showed decreased expression of cell cycle proteins that promote cell cycle progression, including cyclin-dependent kinase 4 and cyclin D1, and increased the expression of proteins that inhibit cell cycle progression, including p27. Furthermore, flow cytometry analysis showed that TSL-1 induced H661 cell apoptosis. Western blot analysis showed that TSL-1 reduced the expression of the anti-apoptotic protein B-cell lymphoma 2, and degraded the DNA repair protein, poly(ADP-ribose polymerase. TSL-1 shows potential as a novel therapeutic agent or for use as an adjuvant for treating human lung large cell carcinoma.

  15. Side Population Cells Derived from Adult Human Liver Generate Hepatocyte-like Cells In Vitro

    Science.gov (United States)

    HUSSAIN, SUNNY ZAHEED; STROM, STEPHEN C.; KIRBY, MARTHA R.; BURNS, SEAN; LANGEMEIJER, SASKIA; UEDA, TAKAHIRO; HSIEH, MATTHEW; TISDALE, JOHN F.

    2009-01-01

    We sought to determine whether hepatic side population (SP) cells derived from adult human liver possess the potential of a novel candidate hepatic stem cell. Human cadaveric donor liver was subjected to collagenase perfusion and hepatocytes were separated from nonparenchymal cells by differential centrifugation. SP cells were isolated from the nonparenchymal portion after Hoechst 33342 staining. Since CD45 is a panleukocyte antigen, CD45-negative SP cells were separated from the vast majority of CD45-positive SP cells (90%), and hepatic growth medium was used to culture both groups. Both CD45-negative and CD45-positive hepatic SP cells generated colonies in the hepatic growth medium in 2–3 weeks. The colonies yielded large cells morphologically consistent with human hepatocytes, demonstrating granule-rich cytoplasm, dense, often double nuclei, and intracellular lipofuscin pigment. The cultured cells from both sources were positive for markers of human hepatocytes: HepPar, cytokeratin 8 (CK8), and human albumin. Reverse transcriptase–polymerase chain reaction (RT-PCR) performed on both groups demonstrated positivity for additional liver markers including human albumin, CK18, α-1 anti-trypsin, and the human cytochrome P450 enzyme CYP2B6. Double immunostaining (CD45 and HepPar) and RT-PCR confirmed that the hepatocyte-like cells derived from the CD45-negative SP cells acquired HepPar positivity but had no detectable CD45 antigen expression. In contrast, the cultured cells derived from the CD45-positive SP cells also acquired HepPar positivity, but only a minimal fraction expressed the CD45 antigen. We conclude that hepatic SP cells derived from the nonparenchymal portion of human liver are a potential source of human hepatocytes irrespective of their CD45 status, and further animal studies will be required to assess their regenerative potential. PMID:16187169

  16. Cell shape regulates global histone acetylation in human mammaryepithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Le Beyec, Johanne; Xu, Ren; Lee, Sun-Young; Nelson, Celeste M.; Rizki, Aylin; Alcaraz, Jordi; Bissell, Mina J.

    2007-02-28

    Extracellular matrix (ECM) regulates cell morphology and gene expression in vivo; these relationships are maintained in three-dimensional (3D) cultures of mammary epithelial cells. In the presence of laminin-rich ECM (lrECM), mammary epithelial cells round up and undergo global histone deacetylation, a process critical for their functional differentiation. However, it remains unclear whether lrECM-dependent cell rounding and global histone deacetylation are indeed part of a common physical-biochemical pathway. Using 3D cultures as well as nonadhesive and micropatterned substrata, here we showed that the cell 'rounding' caused by lrECM was sufficient to induce deacetylation of histones H3 and H4 in the absence of biochemical cues. Microarray and confocal analysis demonstrated that this deacetylation in 3D culture is associated with a global increase in chromatin condensation and a reduction in gene expression. Whereas cells cultured on plastic substrata formed prominent stress fibers, cells grown in 3D lrECM or on micropatterns lacked these structures. Disruption of the actin cytoskeleton with cytochalasin D phenocopied the lrECM-induced cell rounding and histone deacetylation. These results reveal a novel link between ECM-controlled cell shape and chromatin structure, and suggest that this link is mediated by changes in the actin cytoskeleton.

  17. Human fetal liver stromal cells that overexpress bFGF support growth and maintenance of human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Jiafei Xi

    Full Text Available In guiding hES cell technology toward the clinic, one key issue to be addressed is to culture and maintain hES cells much more safely and economically in large scale. In order to avoid using mouse embryonic fibroblasts (MEFs we isolated human fetal liver stromal cells (hFLSCs from 14 weeks human fetal liver as new human feeder cells. hFLSCs feeders could maintain hES cells for 15 passages (about 100 days. Basic fibroblast growth factor (bFGF is known to play an important role in promoting self-renewal of human embryonic stem (hES cells. So, we established transgenic hFLSCs that stably express bFGF by lentiviral vectors. These transgenic human feeder cells--bFGF-hFLSCs maintained the properties of H9 hES cells without supplementing with any exogenous growth factors. H9 hES cells culturing under these conditions maintained all hES cell features after prolonged culture, including the developmental potential to differentiate into representative tissues of all three embryonic germ layers, unlimited and undifferentiated proliferative ability, and maintenance of normal karyotype. Our results demonstrated that bFGF-hFLSCs feeder cells were central to establishing the signaling network among bFGF, insulin-like growth factor 2 (IGF-2, and transforming growth factor β (TGF-β, thereby providing the framework in which hES cells were instructed to self-renew or to differentiate. We also found that the conditioned medium of bFGF-hFLSCs could maintain the H9 hES cells under feeder-free conditions without supplementing with bFGF. Taken together, bFGF-hFLSCs had great potential as feeders for maintaining pluripotent hES cell lines more safely and economically.

  18. Analysis of allelic differential expression in human white blood cells

    National Research Council Canada - National Science Library

    Pant, P V Krishna; Tao, Heng; Beilharz, Erica J; Ballinger, Dennis G; Cox, David R; Frazer, Kelly A

    2006-01-01

    .... To identify human genes with allelic expression differences, we genotype DNA and examine mRNA isolated from the white blood cells of 12 unrelated individuals using oligonucleotide arrays containing 8406 exonic SNPs...

  19. Recent developments on human cell lines for the bioartificial liver

    NARCIS (Netherlands)

    Hoekstra, R.; Chamuleau, R. A. F. M.

    2002-01-01

    Most bioartificial liver (BAL) devices contain porcine primary hepatocytes as their biological component. However, alternatives are needed due to xenotransplantation associated risks. Human liver cell lines have excellent growth characteristics and are therefore candidates for application in BAL

  20. The development of human mast cells. An historical reappraisal

    Energy Technology Data Exchange (ETDEWEB)

    Ribatti, Domenico, E-mail: domenico.ribatti@uniba.it

    2016-03-15

    The understanding of mast cell (MC) differentiation is derived mainly from in vitro studies of different stages of stem and progenitor cells. The hematopoietic lineage development of human MCs is unique compared to other myeloid-derived cells. Human MCs originate from CD34{sup +}/CD117{sup +}/CD13{sup +}multipotent hematopoietic progenitors, which undergo transendothelial recruitment into peripheral tissues, where they complete differentiation. Stem cell factor (SCF) is a major chemotactic factor for MCs and their progenitors. SCF also elicits cell-cell and cell-substratum adhesion, facilitates the proliferation, and sustains the survival, differentiation, and maturation, of MCs. Because MC maturation is influenced by local microenvironmental factors, different MC phenotypes can develop in different tissues and organs. - Highlights: • Human mast cells originate from CD34/CD117/CD13 positive multipotent hematopoietic progenitors. • Stem cell factor is a major chemotactic factor for mast cells and their progenitors. • Different mast cell phenotypes can develop in different tissues and organs.

  1. β-CELL SPECIFIC CYTOPROTECTION BY PROLACTIN ON HUMAN ISLETS

    Science.gov (United States)

    Yamamoto, Toshiyuki; Ricordi, Camillo; Mita, Atsuyoshi; Miki, Atsushi; Sakuma, Yasunaru; Damaris Molano, R.; Fornoni, Alessia; Pileggi, Antonello; Inverardi, Luca; Ichii, Hirohito

    2008-01-01

    Many cytoprotective agents have been reported to improve islet isolation and transplantation outcomes. However, several cytoprotective agents may improve all cell subsets within an islet preparation, and selected non- β-cells components may have a negative effect on β-cell function and survival (e.g., acinar cells). In this study, we examined the effect of prolactin (PRL) supplementation to the culture medium, to determine whether it could exert β-cell-selective cytoprotection (islet viability and function) towards a possible use of PRL during pre transplant islet culture. Materials and Methods Human islets pre-cultured or not with recombinant human PRL (500 μg/L) for 48 hours. Non-β-cells and β-cell-specific fractional viability and cellular composition were assessed by FACS and Laser Scanning Cytometry (LSC). Islet potency was assessed in vivo by transplantation into chemically-induced diabetic immunodeficient mice. Results The relative viable β-cell mass and the relative islet β-cell content in PRL group were 28% higher (p=0.018) and 19% higher (p=0.029) than control group, respectively. All transplanted mice achieved normoglycemia in both groups, indicating that PRL treatment did not alter islet function. Conclusions PRL treatment improves β-cell-specific viability and survival in human islets in vitro. The development of novel β-cell-specific cytoprotective strategies will be of assistance in improving islet transplantation. PMID:18374075

  2. Nuclear DNA Content Varies with Cell Size across Human Cell Types

    Science.gov (United States)

    Gillooly, James F.; Hein, Andrew; Damiani, Rachel

    2015-01-01

    Variation in the size of cells, and the DNA they contain, is a basic feature of multicellular organisms that affects countless aspects of their structure and function. Within humans, cell size is known to vary by several orders of magnitude, and differences in nuclear DNA content among cells have been frequently observed. Using published data, here we describe how the quantity of nuclear DNA across 19 different human cell types increases with cell volume. This observed increase is similar to intraspecific relationships between DNA content and cell volume in other species, and interspecific relationships between diploid genome size and cell volume. Thus, we speculate that the quantity of nuclear DNA content in somatic cells of humans is perhaps best viewed as a distribution of values that reflects cell size distributions, rather than as a single, immutable quantity. PMID:26134319

  3. Nuclear DNA Content Varies with Cell Size across Human Cell Types.

    Science.gov (United States)

    Gillooly, James F; Hein, Andrew; Damiani, Rachel

    2015-07-01

    Variation in the size of cells, and the DNA they contain, is a basic feature of multicellular organisms that affects countless aspects of their structure and function. Within humans, cell size is known to vary by several orders of magnitude, and differences in nuclear DNA content among cells have been frequently observed. Using published data, here we describe how the quantity of nuclear DNA across 19 different human cell types increases with cell volume. This observed increase is similar to intraspecific relationships between DNA content and cell volume in other species, and interspecific relationships between diploid genome size and cell volume. Thus, we speculate that the quantity of nuclear DNA content in somatic cells of humans is perhaps best viewed as a distribution of values that reflects cell size distributions, rather than as a single, immutable quantity. Copyright © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.

  4. T CELL REPLICATIVE SENESCENCE IN HUMAN AGING

    Science.gov (United States)

    Chou, Jennifer P.; Effros, Rita B.

    2013-01-01

    The decline of the immune system appears to be an intractable consequence of aging, leading to increased susceptibility to infections, reduced effectiveness of vaccination and higher incidences of many diseases including osteoporosis and cancer in the elderly. These outcomes can be attributed, at least in part, to a phenomenon known as T cell replicative senescence, a terminal state characterized by dysregulated immune function, loss of the CD28 costimulatory molecule, shortened telomeres and elevated production of pro-inflammatory cytokines. Senescent CD8 T cells, which accumulate in the elderly, have been shown to frequently bear antigen specificity against cytomegalovirus (CMV), suggesting that this common and persistent infection may drive immune senescence and result in functional and phenotypic changes to the T cell repertoire. Senescent T cells have also been identified in patients with certain cancers, autoimmune diseases and chronic infections, such as HIV. This review discusses the in vivo and in vitro evidence for the contribution of CD8 T cell replicative senescence to a plethora of age-related pathologies and a few possible therapeutic avenues to delay or prevent this differentiative end-state in T cells. The age-associated remodeling of the immune system, through accumulation of senescent T cells has far-reaching consequences on the individual and society alike, for the current healthcare system needs to meet the urgent demands of the increasing proportions of the elderly in the US and abroad. PMID:23061726

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

    Science.gov (United States)

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

    2016-01-01

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

  6. Human B cells produce chemokine CXCL10 in the presence of Mycobacterium tuberculosis specific T cells

    DEFF Research Database (Denmark)

    Hoff, Soren T; Salman, Ahmed M; Ruhwald, Morten

    2015-01-01

    BACKGROUND: The role of B cells in human host response to Mycobacterium tuberculosis (Mtb) infection is still controversial, but recent evidence suggest that B cell follicle like structures within the lung may influence host responses through regulation of the local cytokine environment. A candid......BACKGROUND: The role of B cells in human host response to Mycobacterium tuberculosis (Mtb) infection is still controversial, but recent evidence suggest that B cell follicle like structures within the lung may influence host responses through regulation of the local cytokine environment....... A candidate for such regulation could be the chemokine CXCL10. CXCL10 is mainly produced by human monocytes, but a few reports have also found CXCL10 production by human B cells. The objective of this study was to investigate CXCL10 production by human B cells in response to in vitro stimulation with Mtb...

  7. Human placenta-derived adherent cells induce tolerogenic immune responses

    Science.gov (United States)

    Liu, Wei; Morschauser, Andrew; Zhang, Xin; Lu, Xiaohua; Gleason, Joseph; He, Shuyang; Chen, Hong-Jung; Jankovic, Vladimir; Ye, Qian; Labazzo, Kristen; Herzberg, Uri; Albert, Vivian R; Abbot, Stewart E; Liang, Bitao; Hariri, Robert

    2014-01-01

    Human placenta-derived adherent cells (PDAC cells) are a culture expanded, undifferentiated mesenchymal-like population derived from full-term placental tissue, with immunomodulatory and anti-inflammatory properties. PDA-001 (cenplacel-L), an intravenous formulation of PDAC cells, is in clinical development for the treatment of autoimmune and inflammatory diseases. To elucidate the mechanisms underlying the immunoregulatory properties of PDAC cells, we investigated their effects on immune cell populations, including T cells and dendritic cells (DC) in vitro and in vivo. PDAC cells suppressed T-cell proliferation in an OT-II T-cell adoptive transfer model, reduced the severity of myelin oligodendrocyte glycoprotein peptide-induced experimental autoimmune encephalomyelitis and ameliorated inflammation in a delayed type hypersensitivity response model. In vitro, PDAC cells suppressed T-cell proliferation and inhibited Th1 and Th17 differentiation. Analysis of tissues derived from PDAC cell-treated animals revealed diminished CD86 expression on splenic DC, suggesting that they can also modulate DC populations. Furthermore, PDAC cells modulate the differentiation and maturation of mouse bone marrow-derived DC. Similarly, human DC differentiated from CD14+ monocytes in the presence of PDAC cells acquired a tolerogenic phenotype. These tolerogenic DC failed to induce allogeneic T-cell proliferation and differentiation toward Th1, but skewed T-cell differentiation toward Th2. Inhibition of cyclo-oxygenase-2 activity resulted in a significant, but not complete, abrogation of PDAC cells' effects on DC phenotype and function, implying a role for prostaglandin E2 in PDAC-mediated immunomodulation. This study identifies modulation of DC differentiation toward immune tolerance as a key mechanism underlying the immunomodulatory activities of PDAC cells. PMID:25505962

  8. Chimeric animal models in human stem cell biology.

    Science.gov (United States)

    Glover, Joel C; Boulland, Jean-Luc; Halasi, Gabor; Kasumacic, Nedim

    2009-01-01

    The clinical use of stem cells for regenerative medicine is critically dependent on preclinical studies in animal models. In this review we examine some of the key issues and challenges in the use of animal models to study human stem cell biology-experimental standardization, body size, immunological barriers, cell survival factors, fusion of host and donor cells, and in vivo imaging and tracking. We focus particular attention on the various imaging modalities that can be used to track cells in living animals, comparing their strengths and weaknesses and describing technical developments that are likely to lead to new opportunities for the dynamic assessment of stem cell behavior in vivo. We then provide an overview of some of the most commonly used animal models, their advantages and disadvantages, and examples of their use for xenotypic transplantation of human stem cells, with separate reviews of models involving rodents, ungulates, nonhuman primates, and the chicken embryo. As the use of human somatic, embryonic, and induced pluripotent stem cells increases, so too will the range of applications for these animal models. It is likely that increasingly sophisticated uses of human/animal chimeric models will be developed through advances in genetic manipulation, cell delivery, and in vivo imaging.

  9. Nanoparticle labeling identifies slow cycling human endometrial stromal cells

    Science.gov (United States)

    2014-01-01

    Introduction Evidence suggests that the human endometrium contains stem or progenitor cells that are responsible for its remarkable regenerative capability. A common property of somatic stem cells is their quiescent state. It remains unclear whether slow-cycling cells exist in the human endometrium. We hypothesized that the human endometrium contains a subset of slow-cycling cells with somatic stem cell properties. Here, we established an in vitro stem cell assay to isolate human endometrial-derived mesenchymal stem-like cells (eMSC). Methods Single-cell stromal cultures were initially labeled with fluorescent nanoparticles and a small population of fluorescent persistent cells (FPC) remained after culture of 21 days. Two populations of stromal cells, namely FPC and non-FPC were sorted. Results Quantitative analysis of functional assays demonstrated that the FPC had higher colony forming ability, underwent more rounds of self-renewal and had greater enrichment of phenotypically defined prospective eMSC markers: CD146+/CD140b+ and W5C5+ than the non-FPC. They also differentiate into multiple mesenchymal lineages and the expression of lineage specific markers was lower than that of non-FPC. The FPC exhibit low proliferation activities. A proliferation dynamics study revealed that more FPC had a prolonged G1 phase. Conclusions With this study we present an efficient method to label and isolate slow-proliferating cells obtained from human endometrial stromal cultures without genetic modifications. The FPC population could be easily maintained in vitro and are of interest for tissue-repair and engineering perspectives. In summary, nanoparticle labeling is a promising tool for the identification of putative somatic stem or progenitor cells when their surface markers are undefined. PMID:24996487

  10. Human retinal pigment epithelial cell-induced apoptosis in activated T cells

    DEFF Research Database (Denmark)

    Jørgensen, A; Wiencke, A K; la Cour, M

    1998-01-01

    human retinal pigment epithelial (RPE) cells can induce apoptosis in activated T cells. METHODS: Fas ligand (FasL) expression was detected by flow cytometry and immunohistochemistry. Cultured RPE cells were cocultured with T-cell lines and peripheral blood lymphocytes for 6 hours to 2 days. Induction...... of apoptosis was detected by 7-amino-actinomycin D and annexin V staining. RESULTS: Retinal pigment epithelial cells expressed FasL and induced apoptosis in activated Fas+ T cells. Blocking of Fas-FasL interaction with antibody strongly inhibited RPE-mediated T-cell apoptosis. Retinal pigment epithelial cells...... induced apoptosis in several activated T-cell populations and T-cell lines, including T-cell antigen receptor (TCR)-CD3-negative T-cell lines. In contrast, RPE cells induced little or no apoptosis in resting peripheral T cells. Major histocompatibility complex (MHC) class II monoclonal antibodies, which...

  11. [Isolation and characterization of side population cells in human gastric cancer cell line BGC-823].

    Science.gov (United States)

    Wu, Ting; Li, Jin-yi; Lu, Shi-xin

    2012-04-01

    Isolate and characterize the side population (SP) cells with potency of stem cells from human gastric carcinoma cell line BGC-823. SP and non-SP cells were sorted from BGC-823 cells by fluorescence-activated cell sorting (FACS) using Hoechst33342 staining. The tumorigenic ability of the SP cells was assessed by in vivo transplantation into non-obese diabetic/severe combined immunodeficiency mice. SP cells were isolated from BGC-823 cells in a proportion of 0.9% to 2.1% with respect to the whole cell population. The colony formation assay showed that the colony formation rate of the SP cells was significantly higher than that of the non-SP cells (72.56% vs. 49.00%, P line BGC-823 cells. Further characterization of this SP cell population may provide new insights for diagnosis and treatment of gastric cancer.

  12. Cell diversity and network dynamics in photosensitive human brain organoids

    Science.gov (United States)

    Quadrato, Giorgia; Nguyen, Tuan; Macosko, Evan Z.; Sherwood, John L.; Yang, Sung Min; Berger, Daniel; Maria, Natalie; Scholvin, Jorg; Goldman, Melissa; Kinney, Justin; Boyden, Edward S.; Lichtman, Jeff; Williams, Ziv M.; McCarroll, Steven A.; Arlotta, Paola

    2017-01-01

    In vitro models of the developing brain such as 3D brain organoids offer an unprecedented opportunity to study aspects of human brain development and disease. However, it remains undefined what cells are generated within organoids and to what extent they recapitulate the regional complexity, cellular diversity, and circuit functionality of the brain. Here, we analyzed gene expression in over 80,000 individual cells isolated from 31 human brain organoids. We find that organoids can generate a broad diversity of cells, which are related to endogenous classes, including cells from the cerebral cortex and the retina. Organoids could be developed over extended periods (over 9 months) enabling unprecedented levels of maturity including the formation of dendritic spines and of spontaneously-active neuronal networks. Finally, neuronal activity within organoids could be controlled using light stimulation of photoreceptor-like cells, which may offer ways to probe the functionality of human neuronal circuits using physiological sensory stimuli. PMID:28445462

  13. Salidroside induces cell-cycle arrest and apoptosis in human breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Hu, Xiaolan, E-mail: huxiaolan1998@yahoo.com.cn [Department of Pathology and Pathophysiology, Zhejiang University School of Medicine, Hangzhou (China); Zhang, Xianqi [The 2nd Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou (China); Qiu, Shuifeng [Department of Pathology and Pathophysiology, Zhejiang University School of Medicine, Hangzhou (China); Yu, Daihua; Lin, Shuxin [Fourth Military Medical University, Xi' an (China)

    2010-07-16

    Research highlights: {yields} Salidroside inhibits the growth of human breast cancer cells. {yields} Salidroside induces cell-cycle arrest of human breast cancer cells. {yields} Salidroside induces apoptosis of human breast cancer cell lines. -- Abstract: Recently, salidroside (p-hydroxyphenethyl-{beta}-D-glucoside) has been identified as one of the most potent compounds isolated from plants of the Rhodiola genus used widely in traditional Chinese medicine, but pharmacokinetic data on the compound are unavailable. We were the first to report the cytotoxic effects of salidroside on cancer cell lines derived from different tissues, and we found that human breast cancer MDA-MB-231 cells (estrogen receptor negative) were sensitive to the inhibitory action of low-concentration salidroside. To further investigate the cytotoxic effects of salidroside on breast cancer cells and reveal possible ER-related differences in response to salidroside, we used MDA-MB-231 cells and MCF-7 cells (estrogen receptor-positive) as models to study possible molecular mechanisms; we evaluated the effects of salidroside on cell growth characteristics, such as proliferation, cell cycle duration, and apoptosis, and on the expression of apoptosis-related molecules. Our results demonstrated for the first time that salidroside induces cell-cycle arrest and apoptosis in human breast cancer cells and may be a promising candidate for breast cancer treatment.

  14. Interactions of human microglia cells with Japanese encephalitis virus.

    Science.gov (United States)

    Lannes, Nils; Neuhaus, Viviane; Scolari, Brigitte; Kharoubi-Hess, Solange; Walch, Michael; Summerfield, Artur; Filgueira, Luis

    2017-01-14

    Japanese encephalitis virus (JEV) is a neurotropic flavivirus causing mortality and morbidity in humans. Severe Japanese encephalitis cases display strong inflammatory responses in the central nervous system and an accumulation of viral particles in specific brain regions. Microglia cells are the unique brain-resident immune cell population with potent migratory functions and have been proposed to act as a viral reservoir for JEV. Animal models suggest that the targeting of microglia by JEV is partially responsible for inflammatory reactions in the brain. Nevertheless, the interactions between human microglia and JEV are poorly documented. Using human primary microglia and a new model of human blood monocyte-derived microglia, the present study explores the interaction between human microglia and JEV as well as the role of these cells in viral transmission to susceptible cells. To achieve this work, vaccine-containing inactivated JEV and two live JEV strains were applied on human microglia. Live JEV was non-cytopathogenic to human microglia but increased levels of CCL2, CXCL9 and CXCL10 in such cultures. Furthermore, human microglia up-regulated the expression of the fraktalkine receptor CX3CR1 upon exposure to both JEV vaccine and live JEV. Although JEV vaccine enhanced MHC class II on all microglia, live JEV enhanced MHC class II mainly on CX3CR1+ microglia cells. Importantly, human microglia supported JEV replication, but infectivity was only transmitted to neighbouring cells in a contact-dependent manner. Our findings suggest that human microglia may be a source of neuronal infection and sustain JEV brain pathogenesis.

  15. Human T cell receptor gamma delta + T cells

    NARCIS (Netherlands)

    Spits, H.

    1991-01-01

    TCR gamma delta + T cells represent a minority of CD3+ T cells in many species including man. The molecular structure of the TCR gamma and delta loci in man is well understood. The gamma and delta loci contain V, D, J and C gene segments. These segments do not rearrange randomly but in a

  16. Generation of human induced pluripotent stem cells from osteoarthritis patient-derived synovial cells.

    Science.gov (United States)

    Kim, Min-Jeong; Son, Myung Jin; Son, Mi-Young; Seol, Binna; Kim, Janghwan; Park, Jongjin; Kim, Jung Hwa; Kim, Yong-Hoon; Park, Su A; Lee, Chul-Ho; Lee, Kang-Sik; Han, Yong-Mahn; Chang, Jae-Suk; Cho, Yee Sook

    2011-10-01

    This study was undertaken to generate and characterize human induced pluripotent stem cells (PSCs) from patients with osteoarthritis (OA) and to examine whether these cells can be developed into disease-relevant cell types for use in disease modeling and drug discovery. Human synovial cells isolated from two 71-year-old women with advanced OA were characterized and reprogrammed into induced PSCs by ectopic expression of 4 transcription factors (Oct-4, SOX2, Klf4, and c-Myc). The pluripotency status of each induced PSC line was validated by comparison with human embryonic stem cells (ESCs). We found that OA patient-derived human synovial cells had human mesenchymal stem cell (MSC)-like characteristics, as indicated by the expression of specific markers, including CD14-, CD19-, CD34-, CD45-, CD44+, CD51+, CD90+, CD105+, and CD147+. Microarray analysis of human MSCs and human synovial cells further determined their unique and overlapping gene expression patterns. The pluripotency of established human induced PSCs was confirmed by their human ESC-like morphology, expression of pluripotency markers, gene expression profiles, epigenetic status, normal karyotype, and in vitro and in vivo differentiation potential. The potential of human induced PSCs to differentiate into distinct mesenchymal cell lineages, such as osteoblasts, adipocytes, and chondrocytes, was further confirmed by positive expression of markers for respective cell types and positive staining with alizarin red S (osteoblasts), oil red O (adipocytes), or Alcian blue (chondrocytes). Functional chondrocyte differentiation of induced PSCs in pellet culture and 3-dimensional polycaprolactone scaffold culture was assessed by chondrocyte self-assembly and histology. Our findings indicate that patient-derived synovial cells are an attractive source of MSCs as well as induced PSCs and have the potential to advance cartilage tissue engineering and cell-based models of cartilage defects. Copyright © 2011 by the

  17. Engineering antigen-specific T cells from genetically modified human hematopoietic stem cells in immunodeficient mice.

    Directory of Open Access Journals (Sweden)

    Scott G Kitchen

    Full Text Available There is a desperate need for effective therapies to fight chronic viral infections. The immune response is normally fastidious at controlling the majority of viral infections and a therapeutic strategy aimed at reestablishing immune control represents a potentially powerful approach towards treating persistent viral infections. We examined the potential of genetically programming human hematopoietic stem cells to generate mature CD8+ cytotoxic T lymphocytes that express a molecularly cloned, "transgenic" human anti-HIV T cell receptor (TCR. Anti-HIV TCR transduction of human hematopoietic stem cells directed the maturation of a large population of polyfunctional, HIV-specific CD8+ cells capable of recognizing and killing viral antigen-presenting cells. Thus, through this proof-of-concept we propose that genetic engineering of human hematopoietic stem cells will allow the tailoring of effector T cell responses to fight HIV infection or other diseases that are characterized by the loss of immune control.

  18. Freeze-Dried Human Red Blood Cells

    Science.gov (United States)

    1992-04-15

    after reinfusion. Blood 1982; 56: 1141-1147. 11. Danon D, Marikovsky Y. Determination of density distribution of red cell population. J Lab Clin Med...back to the study site. The lyophilized red cells will be stored refrigerated until use. Fourteen days after phlebotomy, the subject will return to the...The subject will remain in confinement at the study site for 24 hours following the transfusion and return on the following four days (Study Days 2

  19. Isolating human DNA repair genes using rodent-cell mutants

    Energy Technology Data Exchange (ETDEWEB)

    Thompson, L.H.; Weber, C.A.; Brookman, K.W.; Salazar, E.P.; Stewart, S.A.; Mitchell, D.L.

    1987-03-23

    The DNA repair systems of rodent and human cells appear to be at least as complex genetically as those in lower eukaryotes and bacteria. The use of mutant lines of rodent cells as a means of identifying human repair genes by functional complementation offers a new approach toward studying the role of repair in mutagenesis and carcinogenesis. In each of six cases examined using hybrid cells, specific human chromosomes have been identified that correct CHO cell mutations affecting repair of damage from uv or ionizing radiations. This finding suggests that both the repair genes and proteins may be virtually interchangeable between rodent and human cells. Using cosmid vectors, human repair genes that map to chromosome 19 have cloned as functional sequences: ERCC2 and XRCC1. ERCC1 was found to have homology with the yeast excision repair gene RAD10. Transformants of repair-deficient cell lines carrying the corresponding human gene show efficient correction of repair capacity by all criteria examined. 39 refs., 1 fig., 1 tab.

  20. Human muscle satellite cells as targets of Chikungunya virus infection.

    Directory of Open Access Journals (Sweden)

    Simona Ozden

    Full Text Available BACKGROUND: Chikungunya (CHIK virus is a mosquito-transmitted alphavirus that causes in humans an acute infection characterised by fever, polyarthralgia, head-ache, and myalgia. Since 2005, the emergence of CHIK virus was associated with an unprecedented magnitude outbreak of CHIK disease in the Indian Ocean. Clinically, this outbreak was characterized by invalidating poly-arthralgia, with myalgia being reported in 97.7% of cases. Since the cellular targets of CHIK virus in humans are unknown, we studied the pathogenic events and targets of CHIK infection in skeletal muscle. METHODOLOGY/PRINCIPAL FINDINGS: Immunohistology on muscle biopsies from two CHIK virus-infected patients with myositic syndrome showed that viral antigens were found exclusively inside skeletal muscle progenitor cells (designed as satelllite cells, and not in muscle fibers. To evaluate the ability of CHIK virus to replicate in human satellite cells, we assessed virus infection on primary human muscle cells; viral growth was observed in CHIK virus-infected satellite cells with a cytopathic effect, whereas myotubes were essentially refractory to infection. CONCLUSIONS/SIGNIFICANCE: This report provides new insights into CHIK virus pathogenesis, since it is the first to identify a cellular target of CHIK virus in humans and to report a selective infection of muscle satellite cells by a viral agent in humans.

  1. Establishment of human cell lines showing circadian rhythms of bioluminescence.

    Science.gov (United States)

    Yoshikawa, Aki; Shimada, Hiroko; Numazawa, Kahori; Sasaki, Tsukasa; Ikeda, Masaaki; Kawashima, Minae; Kato, Nobumasa; Tokunaga, Katsushi; Ebisawa, Takashi

    2008-11-28

    We have established human retinal pigment epithelial cell lines stably expressing the luciferase gene, driven by the human Bmal1 promoter, to obtain human-derived cells that show circadian rhythms of bioluminescence after dexamethasone treatment. The average circadian period of bioluminescence for the obtained clones was 24.07+/-0.48 h. Lithium (10 mM) in the medium significantly lengthened the circadian period of bioluminescence, which is consistent with previous reports, while 2 mM or 5 mM lithium had no effect. This is the first report on the establishment of human-derived cell lines that proliferate infinitely and show circadian rhythms of bioluminescence, and also the first to investigate the effects of low-dose lithium on the circadian rhythms of human-derived cells in vitro. The established cells will be useful for various in vitro studies of human circadian rhythms and for the development of new therapies for human disorders related to circadian rhythm disturbances.

  2. Carrageenan delays cell cycle progression in human cancer cells in vitro demonstrated by FUCCI imaging.

    Science.gov (United States)

    Prasedya, Eka Sunarwidhi; Miyake, Masao; Kobayashi, Daisuke; Hazama, Akihiro

    2016-08-04

    Carrageenan is a sulfated polysaccharide that exists in red seaweeds recently shown to have anticancer properties. Previous findings show various effects of carrageenan suppressing tumor cell growth. One of the hallmarks of cancer is uncontrolled proliferation, a consequence of loss of normal cell-cycle control, that underlies tumor growth. Recently there is an increasing interest in potential anticancer agents that affect cell cycle in cancer cells. Thus, in this study we investigated the effects of carrageenan on the tumor cell cycle. Using human cervical carcinoma cells (HeLa) cells as and human umbilical vein endothelial cells (HUVEC), the cytotoxic effects of kappa carrageenan (k-CO) and lambda carrageenan (λ-CO) at the concentrations of 250-2500 μg/mL were observed. Cell viability was determined using the MTT assay while cell death rates were determined using staining with calcein-AM/propidium iodide. Cell-cycle profile and progression were demonstrated with HeLa cells expressing FUCCI (fluorescence ubiquitination-based cell-cycle indicator) probes (HeLa-FUCCI). Carrageenan had no significant effect on HUVEC (normal cells). In contrast both forms of carrageenan were cytotoxic towards HeLa cells (cancer cells). Furthermore, according to cell-cycle analysis with FUCCI cells, the cell cycle of HeLa cells was delayed in specific phases due to different carrageenan treatments. Considering these results, it could be suggested that carrageenan affects the cell-cycle of HeLa cells not only by arresting the cell cycle in specific phases but also by delaying the time needed for the cell to progress through the cell cycle. Additionally, different types of carrageenans have different effects on cell cycle progression. This effect of carrageenan towards cancer cells could possibly be developed into a tumor cell-specific anticancer agent.

  3. The endocannabinoid anandamide impairs in vitro decidualization of human cells.

    Science.gov (United States)

    Almada, M; Amaral, C; Diniz-da-Costa, M; Correia-da-Silva, G; Teixeira, N A; Fonseca, B M

    2016-10-01

    Endocannabinoids (eCBs) are endogenous mediators that along with the cannabinoid receptors (CB1 and CB2), a membrane transporter and metabolic enzymes form the endocannabinoid system (ECS). Several eCBs have been discovered with emphasis on anandamide (AEA). They are involved in several biological processes such as energy balance, immune response and reproduction. Decidualization occurs during the secretory phase of human menstrual cycle, which involves proliferation and differentiation of endometrial stromal cells into decidual cells and is crucial for the establishment and progression of pregnancy. In this study, a telomerase-immortalized human endometrial stromal cell line (St-T1b) and non-differentiated primary cultures of human decidual fibroblasts from term placenta were used to characterize the ECS using immunoblotting and qRT-PCR techniques. It was shown that St-T1b cells express CB1, but not CB2, and that both receptors are expressed in HdF cells. Furthermore, the expression of fatty acid amide hydrolase (FAAH), the main degrading enzyme of AEA, increased during stromal cell differentiation. AEA inhibited cell proliferation, through deregulation of cell cycle progression and induced polyploidy. Moreover, through CB1 binding receptor, AEA also impaired cell differentiation. Therefore, AEA is proposed as a modulator of human decidualization. Our findings may provide wider implications, as deregulated levels of AEA, due to Cannabis sativa consumption or altered expression of the metabolic enzymes, may negatively regulate human endometrial stromal cell decidualization with an impact on human (in)fertility.Free Portuguese abstract: A Portuguese translation of this abstract is freely available at http://www.reproduction-online.org/content/152/4/351/suppl/DC1. © 2016 Society for Reproduction and Fertility.

  4. Autophagy in the human placenta throughout gestation.

    Science.gov (United States)

    Hung, Tai-Ho; Hsieh, T'sang-T'ang; Chen, Szu-Fu; Li, Meng-Jen; Yeh, Yi-Lin

    2013-01-01

    Autophagy has been reported to be essential for pre-implantation development and embryo survival. However, its role in placental development and regulation of autophagy during pregnancy remain unclear. The aims of this study were to (1) study autophagy by characterizing changes in levels of beclin-1, DRAM, and LC3B in human placenta throughout gestation; (2) determine whether autophagy is involved in regulation of trophoblast invasion in JEG-3 cells (a choriocarcinoma cell line); (3) examine the effects of reduced oxygen and glucose on the autophagic changes; and (4) investigate the effect of reoxygenation and supplementation of glucose after oxygen-glucose deprivation (OGD) on the autophagic changes in primary cytotrophoblasts obtained from normal term pregnancy. An analysis of 40 placental samples representing different gestational stages showed (1) no significant differences in beclin-1, DRAM, and LC3B-II levels in placentas between early and mid-gestation, and late gestation with vaginal delivery; (2) placentas from late gestation with cesarean section had lower levels of LC3B-II compared to early and mid-gestation, and late gestation with vaginal delivery; levels of DRAM were also lower compared to placentas from early and mid-gestation; and (3) using explant cultures, villous tissues from early and late gestation had similar rates of autophagic flux under physiological oxygen concentrations. Knockdown of BECN1, DRAM, and LC3B had no effects on viability and invasion activity of JEG-3 cells. On the other hand, OGD caused a significant increase in the levels of LC3B-II in primary cytotrophoblasts, while re-supplementation of oxygen and glucose reduced these changes. Furthermore, there were differential changes in levels of beclin-1, DRAM, and LC3B-II in response to changes in oxygen and glucose levels. Our results indicate that autophagy is involved in development of the human placenta and that changes in oxygen and glucose levels participate in regulation of

  5. Influenza A virus infection of human Schwann cells in vitro.

    Science.gov (United States)

    Levine, Joshua; Buchman, Craig A; Fregien, Nevis

    2003-01-01

    Sudden sensorineural hearing loss, vestibular neuronitis, vocal fold paralysis and Bell's palsy have been associated with a viral etiology, due to the infection of nerve cells. The goal of this research was to ascertain whether Schwann cells can support infection with human influenza A virus and thereby represent a plausible alternative site for virus-host interaction. Viral infection of Schwann cells may lead to secretion of inflammatory mediators, leukocyte recruitment, demyelination and nerve damage. Cultured human Schwann cells were exposed to human influenza A virus. Infection was assayed at various times post-inoculation (0, 24, 48 and 72 h) using light microscopy, immunocytochemistry and influenza A virus-specific reverse transcriptase polymerase chain reaction (RT-PCR). A group of unexposed cells served as controls. Following exposure to the virus, vacuolization, cellular expansion and detachment from the dish were seen as early as 24 h post-inoculation. The exposed cells demonstrated positive immunocytochemical staining for influenza A virus antigen at 24, 48 and 72 h. Using RT-PCR, a sharp rise in influenza A virus-specific mRNA was detected. Human Schwann cells can be infected with human influenza A virus. Further studies will assess the inflammatory response in this model.

  6. NFKB1 regulates human NK cell maturation and effector functions.

    Science.gov (United States)

    Lougaris, Vassilios; Patrizi, Ornella; Baronio, Manuela; Tabellini, Giovanna; Tampella, Giacomo; Damiati, Eufemia; Frede, Natalie; van der Meer, Jos W M; Fliegauf, Manfred; Grimbacher, Bodo; Parolini, Silvia; Plebani, Alessandro

    2017-02-01

    NFKB1, a component of the canonical NF-κB pathway, was recently reported to be mutated in a limited number of CVID patients. CVID-associated mutations in NFKB2 (non-canonical pathway) have previously been shown to impair NK cell cytotoxic activity. Although a biological function of NFKB1 in non-human NK cells has been reported, the role of NFKB1 mutations for human NK cell biology and disease has not been investigated yet. We decided therefore to evaluate the role of monoallelic NFKB1 mutations in human NK cell maturation and functions. We show that NFKB1 mutated NK cells present impaired maturation, defective cytotoxicity and reduced IFN-γ production upon in vitro stimulation. Furthermore, human IL-2 activated NFKB1 mutated NK cells fail to up-regulate the expression of the activating marker NKp44 and show reduced proliferative capacity. These data suggest that NFKB1 plays an essential novel role for human NK cell maturation and effector functions. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. The safety of human pluripotent stem cells in clinical treatment.

    Science.gov (United States)

    Simonson, Oscar E; Domogatskaya, Anna; Volchkov, Pavel; Rodin, Sergey

    2015-01-01

    Human pluripotent stem cells (hPSCs) have practically unlimited proliferation potential and a capability to differentiate into any cell type in the human body. Since the first derivation in 1998, they have been an attractive source of cells for regenerative medicine. Numerous ethical, technological, and regulatory complications have been hampering hPSC use in clinical applications. Human embryonic stem cells (ESCs), parthenogenetic human ESCs, human nuclear transfer ESCs, and induced pluripotent stem cells are four types of hPSCs that are different in many clinically relevant features such as propensity to epigenetic abnormalities, generation methods, and ability for development of autologous cell lines. Propensity to genetic mutations and tumorigenicity are common features of all pluripotent cells that complicate hPSC-based therapies. Several recent advances in methods of derivation, culturing, and monitoring of hPSCs have addressed many ethical concerns and technological challenges in development of clinical-grade hPSC lines. Generation of banks of such lines may be useful to minimize immune rejection of hPSC-derived allografts. In this review, we discuss different sources of hPSCs available at the moment, various safety risks associated with them, and possible solutions for successful use of hPSCs in the clinic. We also discuss ongoing clinical trials of hPSC-based treatments.

  8. Alteration of Cell Cycle Mediated by Zinc in Human Bronchial ...

    Science.gov (United States)

    Zinc (Zn2+), a ubiquitous ambient air contaminant, presents an oxidant challenge to the human lung and is linked to adverse human health effects. To further elucidate the adaptive and apoptotic cellular responses of human airway cells to Zn2+, we performed pilot studies to examine cell cycle perturbation upon exposure using a normal human bronchial epithelial cell culture (BEAS-2B). BEAS-2B cells were treated with low (0, 1, 2 µM) and apoptotic (3 µM) doses of Zn2+ plus 1 µM pyrithione, a Zn2+-specific ionophore facilitating cellular uptake, for up to 24 h. Fixed cells were then stained with propidium iodine (PI) and cell cycle phase was determined by fluorescent image cytometry. Initial results report the percentage of cells in the S phase after 18 h exposure to 1, 2, and 3 µM Zn2+ were similar (8%, 7%, and 12%, respectively) compared with 7% in controls. Cells exposed to 3 µM Zn2+ increased cell populations in G2/M phase (76% versus 68% in controls). Interestingly, exposure to 1 µM Zn2+ resulted in decreased (59%) cells in G2/M. While preliminary, these pilot studies suggest Zn2+ alters cell cycle in BEAS-2B cells, particularly in the G2/M phase. The G2/M checkpoint maintains DNA integrity by enabling initiation of DNA repair or apoptosis. Our findings suggest that the adaptive and apoptotic responses to Zn2+ exposure may be mediated via perturbation of the cell cycle at the G2/M checkpoint. This work was a collaborative summer student project. The st

  9. Human tissue legislation in South Africa: Focus on stem cell ...

    African Journals Online (AJOL)

    2015-08-04

    Aug 4, 2015 ... under the Human Tissue Act 65 of 1983, which was repealed with the enactment of the final sections of the NHA in 2012.[5]. Chapter 8 of the NHA covers seven areas that are relevant to human tissues. These include: • Blood and blood products. • Assisted reproductive technology. • Cell-based therapy.

  10. Humanizing recombinant glycoproteins from Chinese hamster ovary cells

    DEFF Research Database (Denmark)

    Hansen, Anders Holmgaard; Amann, Thomas; Kol, Stefan

    hamster ovary (CHO) cells are making a very heterogeneous mixture of NGlycans. We speculate that the CHO pattern of N-Glycans would affect half-life and/or efficacy of the glycoprotein in the bloodstream making it unsuitable for human intravenous use, whereas our humanized version would be identical...

  11. DIVERSITY OF ARSENIC METABOLISM IN CULTURED HUMAN CANCER CELL LINES

    Science.gov (United States)

    Diversity of arsenic metabolism in cultured human cancer cell lines. Arsenic has been known to cause a variety of malignancies in human. Pentavalent As (As 5+) is reduced to trivalent As (As3+) which is further methylated by arsenic methyltransferase(s) to monomethylarson...

  12. Protein dynamics in individual human cells: experiment and theory.

    Directory of Open Access Journals (Sweden)

    Ariel Aharon Cohen

    Full Text Available A current challenge in biology is to understand the dynamics of protein circuits in living human cells. Can one define and test equations for the dynamics and variability of a protein over time? Here, we address this experimentally and theoretically, by means of accurate time-resolved measurements of endogenously tagged proteins in individual human cells. As a model system, we choose three stable proteins displaying cell-cycle-dependant dynamics. We find that protein accumulation with time per cell is quadratic for proteins with long mRNA life times and approximately linear for a protein with short mRNA lifetime. Both behaviors correspond to a classical model of transcription and translation. A stochastic model, in which genes slowly switch between ON and OFF states, captures measured cell-cell variability. The data suggests, in accordance with the model, that switching to the gene ON state is exponentially distributed and that the cell-cell distribution of protein levels can be approximated by a Gamma distribution throughout the cell cycle. These results suggest that relatively simple models may describe protein dynamics in individual human cells.

  13. Hexavalent Chromium Induces Chromosome Instability in Human Urothelial Cells

    Science.gov (United States)

    Wise, Sandra S.; Holmes, Amie L.; Liou, Louis; Adam, Rosalyn M.; Wise, John Pierce

    2016-01-01

    Numerous metals are well-known human bladder carcinogens. Despite the significant occupational and public health concern of metals and bladder cancer, the carcinogenic mechanisms remain largely unknown. Chromium, in particular, is a metal of concern as incidences of bladder cancer have been found elevated in chromate workers, and there is an increasing concern for patients with metal hip implants. However, the impact of Cr(VI) on bladder cells has not been studied. We compared chromate toxicity in two bladder cell lines; primary human urothelial cells and hTERT-immortalized human urothelial cells. Hexavalent chromium (Cr(VI)) induced a concentration- and time-dependent increase in chromosome damage in both cell lines, with the hTERT-immortalized cells exhibiting more chromosome damage than the primary cells. Chronic exposure to Cr(VI) also induced a concentration-dependent increase in aneuploid metaphases in both cell lines which was not observed after a 24 h exposure. Aneuploidy induction was higher in the hTERT-immortalized cells. When we correct for uptake, Cr(VI) induces a similar amount of chromosome damage and aneuploidy suggesting that the differences in Cr(VI) sensitivity between the two cells lines were due to differences in uptake. The increase in chromosome instability after chronic chromate treatment suggests this may be a mechanism for chromate-induced bladder cancer specifically and may be a mechanism for metal-induced bladder cancer in general. PMID:26908176

  14. A Novel Class of Human Cardiac Stem Cells.

    Science.gov (United States)

    Moccetti, Tiziano; Leri, Annarosa; Goichberg, Polina; Rota, Marcello; Anversa, Piero

    2015-01-01

    Following the recognition that hematopoietic stem cells improve the outcome of myocardial infarction in animal models, bone marrow mononuclear cells, CD34-positive cells, and mesenchymal stromal cells have been introduced clinically. The intracoronary or intramyocardial injection of these cell classes has been shown to be safe and to produce a modest but significant enhancement in systolic function. However, the identification of resident cardiac stem cells in the human heart (hCSCs) has created great expectation concerning the potential implementation of this category of autologous cells for the management of the human disease. Although phase 1 clinical trials have been conducted with encouraging results, the search for the most powerful hCSC for myocardial regeneration is in its infancy. This manuscript discusses the efforts performed in our laboratory to characterize the critical biological variables that define the growth reserve of hCSCs. Based on the theory of the immortal DNA template, we propose that stem cells retaining the old DNA represent 1 of the most powerful cells for myocardial regeneration. Similarly, the expression of insulin-like growth factor-1 receptors in hCSCs recognizes a cell phenotype with superior replicating reserve. However, the impressive recovery in ventricular hemodynamics and anatomy mediated by clonal hCSCs carrying the "mother" DNA underscores the clinical relevance of this hCSC class for the treatment of human heart failure.

  15. Hexavalent chromium induces chromosome instability in human urothelial cells.

    Science.gov (United States)

    Wise, Sandra S; Holmes, Amie L; Liou, Louis; Adam, Rosalyn M; Wise, John Pierce

    2016-04-01

    Numerous metals are well-known human bladder carcinogens. Despite the significant occupational and public health concern of metals and bladder cancer, the carcinogenic mechanisms remain largely unknown. Chromium, in particular, is a metal of concern as incidences of bladder cancer have been found elevated in chromate workers, and there is an increasing concern for patients with metal hip implants. However, the impact of hexavalent chromium (Cr(VI)) on bladder cells has not been studied. We compared chromate toxicity in two bladder cell lines; primary human urothelial cells and hTERT-immortalized human urothelial cells. Cr(VI) induced a concentration- and time-dependent increase in chromosome damage in both cell lines, with the hTERT-immortalized cells exhibiting more chromosome damage than the primary cells. Chronic exposure to Cr(VI) also induced a concentration-dependent increase in aneuploid metaphases in both cell lines which was not observed after a 24h exposure. Aneuploidy induction was higher in the hTERT-immortalized cells. When we correct for uptake, Cr(VI) induces a similar amount of chromosome damage and aneuploidy suggesting that the differences in Cr(VI) sensitivity between the two cells lines were due to differences in uptake. The increase in chromosome instability after chronic chromate treatment suggests this may be a mechanism for chromate-induced bladder cancer, specifically, and may be a mechanism for metal-induced bladder cancer, in general. Copyright © 2016. Published by Elsevier Inc.

  16. Comparison of the Gene Expression Profiles of Human Hematopoietic Stem Cells between Humans and a Humanized Xenograft Model.

    Science.gov (United States)

    Matsuzawa, Hideyuki; Matsushita, Hiromichi; Yahata, Takashi; Tanaka, Masayuki; Ando, Kiyoshi

    2017-04-20

    The aim of this study is to evaluate the feasibility of NOD/Shi-scid-IL2Rγnull(NOG) mice transplanted with human CD34+/CD38-/Lin-/low hematopoietic cells from cord blood (CB) as an experimental model of the gene expression in human hematopoiesis. We compared the gene expressions of human CD34+/CD38-/Lin-/low cells from human bone marrow (BM) and in xenograft models. The microarray data revealed that 25 KEGG pathways were extracted from the comparison of human CD34+/CD38-/Lin-/low HSCs between CB and BM, and that 17 of them--which were mostly related to cellular survival, RNA metabolism and lymphoid development--were shared with the xenograft model. When the probes that were commonly altered in CD34+/CD38-/Lin-/low cells from both human and xenograft BM were analyzed, most of them, including the genes related hypoxia, hematopoietic differentiation, epigenetic modification, translation initiation, and RNA degradation, were downregulated. These alterations of gene expression suggest a reduced differentiation capacity and likely include key alterations of gene expression for settlement of CB CD34+/CD38-/Lin-/low cells in BM. Our findings demonstrate that the xenograft model of human CB CD34+/CD38-/Lin-/low cells using NOG mice was useful, at least in part, for the evaluation of the gene expression profile of human hematopoietic stem cells.

  17. New Monoclonal Antibodies to Defined Cell Surface Proteins on Human Pluripotent Stem Cells.

    Science.gov (United States)

    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. © 2016 The Authors Stem Cells published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

  18. Endogenous collagen influences differentiation of human multipotent mesenchymal stromal cells

    NARCIS (Netherlands)

    Fernandes, H.; Mentink, A.; Bank, R.; Stoop, R.; Blitterswijk, C. van; Boer, J. de

    2010-01-01

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix

  19. Endogenous Collagen Influences Differentiation of Human Multipotent Mesenchymal Stromal Cells

    NARCIS (Netherlands)

    Fernandes, H.A.M.; Mentink-Leusink, Anouk; Bank, Ruud; Stoop, Reinout; van Blitterswijk, Clemens; de Boer, Jan

    2010-01-01

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix

  20. Endogenous Collagen Influences Differentiation of Human Multipotent Mesenchymal Stromal Cells

    NARCIS (Netherlands)

    Fernandes, Hugo; Mentink, Anouk; Bank, Ruud; Stoop, Reinout; van Blitterswijk, Clemens; de Boer, Jan

    Human multipotent mesenchymal stromal cells (hMSCs) are multipotent cells that, in the presence of appropriate stimuli, can differentiate into different lineages such as the osteogenic, chondrogenic, and adipogenic lineages. In the presence of ascorbic acid, MSCs secrete an extracellular matrix

  1. In vitro radiosensitivity of human leukemia cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Weichselbaum, R.R.; Greenberger, J.S.; Schmidt, A.; Karpas, A.; Moloney, W.C.; Little, J.B.

    1981-05-01

    The in vitro radiobiologic survival values (anti n, D/sub 0/) of four tumor lines derived from human hematopoietic tumors were studied. These cell lines were HL60 promyelocytic leukemia; K562 erythroleukemia; 45 acute lymphocytic leukemia; and 176 acute monomyelogenous leukemia. More cell lines must be examined before the exact relationship between in vitro radiosensitivity and clinical radiocurability is firmly established.

  2. Green tea polyphenol induces significant cell death in human lung ...

    African Journals Online (AJOL)

    Green tea polyphenol induces significant cell death in human lung cancer cells. Jie Huang, Fa-jiu Li, Shi Chen, Yi Shi, Xiao-jiang Wang, Chuan-hai Wang, Qing- ..... method for the determination of green and black tea polyphenols in biomatrices by high-performance liquid chromatography with coulometric array detection.

  3. Phosphorylation dynamics during early differentiation of human embryonic stem cells

    DEFF Research Database (Denmark)

    Van Hoof, Dennis; Muñoz, Javier; Braam, Stefan R

    2009-01-01

    Pluripotent stem cells self-renew indefinitely and possess characteristic protein-protein networks that remodel during differentiation. How this occurs is poorly understood. Using quantitative mass spectrometry, we analyzed the (phospho)proteome of human embryonic stem cells (hESCs) during...

  4. Wnt signaling inhibits osteogenic differentiation of human mesenchymal stem cells

    NARCIS (Netherlands)

    de Boer, Jan; Siddappa, R.; Gaspar, Claudia; van Apeldoorn, Aart A.; Fodde, Riccardo; van Blitterswijk, Clemens

    2004-01-01

    Human mesenchymal stem cells (hMSCs) from the bone marrow represent a potential source of pluripotent cells for autologous bone tissue engineering. We previously discovered that over activation of the Wnt signal transduction pathway by either lithium or Wnt3A stimulates hMSC proliferation while

  5. Phosphorylation dynamics during early differentiation of human embryonic stem cells

    NARCIS (Netherlands)

    van Hoof, D.; Munoz, J.; Braam, S.R.; Pinkse, M.W.H.; Linding, R.; Heck, A.J.R.; Mummery, C.L.; Krijgsveld, J.

    2009-01-01

    Pluripotent stem cells self-renew indefinitely and possess characteristic protein-protein networks that remodel during differentiation. How this occurs is poorly understood. Using quantitative mass spectrometry, we analyzed the (phospho)proteome of human embryonic stem cells (hESCs) during

  6. Improved genetic manipulation of human embryonic stem cells.

    NARCIS (Netherlands)

    Braam, S.R.; Denning, C.; van den Brink, S.; Kats, P.; Hochstenbach, R.; Passier, R.; Mummery, C.L.

    2008-01-01

    Low efficiency of transfection limits the ability to genetically manipulate human embryonic stem cells (hESCs), and differences in cell derivation and culture methods require optimization of transfection protocols. We transiently transferred multiple independent hESC lines with different growth

  7. Tangeretin sensitises human lung cancer cells to TRAIL- induced ...

    African Journals Online (AJOL)

    necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in human lung cancer cells. (H1299 and H1975). Methods: ... Western blotting was performed to assess the expression of death receptors, apoptosis pathway proteins, JNK and ERK1/2. ...... upregulation in hepatocellular carcinoma cells. World J.

  8. (Ezh2) Enhancement in Human Esophageal Cell Lines

    African Journals Online (AJOL)

    Purpose: To investigate the inhibitory effect of berberine treatment on enhancement of zeste of homolog 2 (Ezh2) expressions in KYSE450 human esophageal cancer cells. Methods: Transwell motility chambers were used to analyze cell migration and invasion. Bio-Rad protein assay was used for the determination of ...

  9. Human Pluripotent Stem Cells to Engineer Blood Vessels.

    Science.gov (United States)

    Chan, Xin Yi; Elliott, Morgan B; Macklin, Bria; Gerecht, Sharon

    2017-11-01

    Development of pluripotent stem cells (PSCs) is a remarkable scientific advancement that allows scientists to harness the power of regenerative medicine for potential treatment of disease using unaffected cells. PSCs provide a unique opportunity to study and combat cardiovascular diseases, which continue to claim the lives of thousands each day. Here, we discuss the differentiation of PSCs into vascular cells, investigation of the functional capabilities of the derived cells, and their utilization to engineer microvascular beds or vascular grafts for clinical application. Graphical Abstract Human iPSCs generated from patients are differentiated toward ECs and perivascular cells for use in disease modeling, microvascular bed development, or vascular graft fabrication.

  10. Renal cell apoptosis in human lupus nephritis: a histological study

    DEFF Research Database (Denmark)

    Faurschou, M; Penkowa, Milena; Andersen, C B

    2009-01-01

    Nuclear autoantigens from apoptotic cells are believed to drive the immunological response in systemic lupus erythematosus (SLE). Conflicting data exist as to the possible renal origin of apoptotic cells in SLE patients with nephritis. We assessed the level of renal cell apoptosis in kidney...... biopsies from 35 patients with lupus nephritis by means of terminal deoxynucleotidyl-transferase (TdT)-mediated deoxyuridine triphosphate (dUTP)-digoxigenin nick end labeling (TUNEL). Five samples of normal kidney tissue served as control specimens. We did not observe apoptotic glomerular cells in any...... cells constitute a quantitatively important source of auto-antibody-inducing nuclear auto-antigens in human lupus nephritis....

  11. Low dose perfluorooctanoate exposure promotes cell proliferation in a human non-tumor liver cell line

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Hongxia; Cui, Ruina [Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China); Guo, Xuejiang [State Key Laboratory of Reproductive Medicine, Nanjing Medical University, Nanjing 210029 (China); Hu, Jiayue [Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China); Dai, Jiayin, E-mail: daijy@ioz.ac.cn [Key Laboratory of Animal Ecology and Conservation Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101 (China)

    2016-08-05

    Highlights: • Differential expression of proteins induced by PFOA in HL-7702 was identified. • Most of the differentially expressed proteins are related to cell proliferation. • A low dose of PFOA stimulates HL-7702 cell proliferation. • A high dose of PFOA inhibits HL-7702 cell proliferation. - Abstract: Perfluorooctanoate (PFOA) is a well-known persistent organic pollutant widely found in the environment, wildlife and humans. Medical surveillance and experimental studies have investigated the potential effects of PFOA on human livers, but the hepatotoxicity of PFOA on humans and its underlying mechanism remain to be clarified. We exposed a human liver cell line (HL-7702) to 50 μM PFOA for 48 h and 96 h, and identified 111 significantly differentially expressed proteins by iTRAQ analysis. A total of 46 proteins were related to cell proliferation and apoptosis. Through further analysis of the cell cycle, apoptosis and their related proteins, we found that low doses of PFOA (50–100 μM) promoted cell proliferation and numbers by promoting cells from the G1 to S phases, whereas high doses of PFOA (200–400 μM) led to reduced HL-7702 cell numbers compared with that of the control mainly due to cell cycle arrest in the G0/G1 phase. To our knowledge, this is the first report on the promotion of cell cycle progression in human cells following PFOA exposure.

  12. Growth in Agarose of Human Cells Infected with Cytomegalovirus

    Science.gov (United States)

    Lang, David J.; Montagnier, Luc; Latarjet, Raymond

    1974-01-01

    After infection by human cytomegalovirus (CMV), human diploid fibroblasts could grow in agarose medium for several generations. Clones of infected cells grew for weeks, although in every case they ultimately underwent lysis owing to the cytopathic effect of the virus. Virus was inoculated at high dilution and after UV irradiation in an effort to derive cells infected with noninfectious defective particles still capable of inducing cell stimulation. Dilute or irradiated virus occasionally yielded large colonies of replicating cells, although permanent transformation was not observed. One clone derived from UV-CMV-infected cells was passaged four times before undergoing lysis. During these passages the cells exhibited alterations in morphology and orientation. Images PMID:4367907

  13. Long-term maintenance of human induced pluripotent stem cells by automated cell culture system.

    Science.gov (United States)

    Konagaya, Shuhei; Ando, Takeshi; Yamauchi, Toshiaki; Suemori, Hirofumi; Iwata, Hiroo

    2015-11-17

    Pluripotent stem cells, such as embryonic stem cells and induced pluripotent stem (iPS) cells, are regarded as new sources for cell replacement therapy. These cells can unlimitedly expand under undifferentiated conditions and be differentiated into multiple cell types. Automated culture systems enable the large-scale production of cells. In addition to reducing the time and effort of researchers, an automated culture system improves the reproducibility of cell cultures. In the present study, we newly designed a fully automated cell culture system for human iPS maintenance. Using an automated culture system, hiPS cells maintained their undifferentiated state for 60 days. Automatically prepared hiPS cells had a potency of differentiation into three germ layer cells including dopaminergic neurons and pancreatic cells.

  14. Identification and quantitation of morphological cell types in electrophoretically separated human embryonic kidney cell cultures

    Science.gov (United States)

    Williams, K. B.; Kunze, M. E.; Todd, P. W.

    1985-01-01

    Four major cell types were identified by phase microscopy in early passage human embryonic kidney cell cultures. They are small and large epithelioid, domed, and fenestrated cells. Fibroblasts are also present in some explants. The percent of each cell type changes with passage number as any given culture grows. As a general rule, the fraction of small epithelioid cells increases, while the fraction of fenestrated cells, always small, decreases further. When fibroblasts are present, they always increase in percentage of the total cell population. Electrophoretic separation of early passage cells showed that the domed cells have the highest electrophoretic mobility, fibroblasts have an intermediate high mobility, small epithelioid cells have a low mobility, broadly distributed, and fenestrated cells have the lowest mobility. All cell types were broadly distributed among electrophoretic subfractions, which were never pure but only enriched with respect to a given cell type.

  15. Generating a non-integrating human induced pluripotent stem cell bank from urine-derived cells.

    Directory of Open Access Journals (Sweden)

    Yanting Xue

    Full Text Available Induced pluripotent stem cell (iPS cell holds great potential for applications in regenerative medicine, drug discovery, and disease modeling. We describe here a practical method to generate human iPS cells from urine-derived cells (UCs under feeder-free, virus-free, serum-free condition and without oncogene c-MYC. We showed that this approach could be applied in a large population with different genetic backgrounds. UCs are easily accessible and exhibit high reprogramming efficiency, offering advantages over other cell types used for the purpose of iPS generation. Using the approach described in this study, we have generated 93 iPS cell lines from 20 donors with diverse genetic backgrounds. The non-viral iPS cell bank with these cell lines provides a valuable resource for iPS cells research, facilitating future applications of human iPS cells.

  16. Purified Human Dental Pulp Stem Cells Promote Osteogenic Regeneration.

    Science.gov (United States)

    Yasui, T; Mabuchi, Y; Toriumi, H; Ebine, T; Niibe, K; Houlihan, D D; Morikawa, S; Onizawa, K; Kawana, H; Akazawa, C; Suzuki, N; Nakagawa, T; Okano, H; Matsuzaki, Y

    2016-02-01

    Human dental pulp stem/progenitor cells (hDPSCs) are attractive candidates for regenerative therapy because they can be easily expanded to generate colony-forming unit-fibroblasts (CFU-Fs) on plastic and the large cell numbers required for transplantation. However, isolation based on adherence to plastic inevitably changes the surface marker expression and biological properties of the cells. Consequently, little is currently known about the original phenotypes of tissue precursor cells that give rise to plastic-adherent CFU-Fs. To better understand the in vivo functions and translational therapeutic potential of hDPSCs and other stem cells, selective cell markers must be identified in the progenitor cells. Here, we identified a dental pulp tissue-specific cell population based on the expression profiles of 2 cell-surface markers LNGFR (CD271) and THY-1 (CD90). Prospectively isolated, dental pulp-derived LNGFR(Low+)THY-1(High+) cells represent a highly enriched population of clonogenic cells--notably, the isolated cells exhibited long-term proliferation and multilineage differentiation potential in vitro. The cells also expressed known mesenchymal cell markers and promoted new bone formation to heal critical-size calvarial defects in vivo. These findings suggest that LNGFR(Low+)THY-1(High+) dental pulp-derived cells provide an excellent source of material for bone regenerative strategies. © International & American Associations for Dental Research 2015.

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

    Science.gov (United States)

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

    2018-01-30

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

  18. DNA Repair in Human Pluripotent Stem Cells Is Distinct from That in Non-Pluripotent Human Cells

    Science.gov (United States)

    Luo, Li Z.; Park, Sang-Won; Bates, Steven E.; Zeng, Xianmin; Iverson, Linda E.; O'Connor, Timothy R.

    2012-01-01

    The potential for human disease treatment using human pluripotent stem cells, including embryonic stem cells and induced pluripotent stem cells (iPSCs), also carries the risk of added genomic instability. Genomic instability is most often linked to DNA repair deficiencies, which indicates that screening/characterization of possible repair deficiencies in pluripotent human stem cells should be a necessary step prior to their clinical and research use. In this study, a comparison of DNA repair pathways in pluripotent cells, as compared to those in non-pluripotent cells, demonstrated that DNA repair capacities of pluripotent cell lines were more heterogeneous than those of differentiated lines examined and were generally greater. Although pluripotent cells had high DNA repair capacities for nucleotide excision repair, we show that ultraviolet radiation at low fluxes induced an apoptotic response in these cells, while differentiated cells lacked response to this stimulus, and note that pluripotent cells had a similar apoptotic response to alkylating agent damage. This sensitivity of pluripotent cells to damage is notable since viable pluripotent cells exhibit less ultraviolet light-induced DNA damage than do differentiated cells that receive the same flux. In addition, the importance of screening pluripotent cells for DNA repair defects was highlighted by an iPSC line that demonstrated a normal spectral karyotype, but showed both microsatellite instability and reduced DNA repair capacities in three out of four DNA repair pathways examined. Together, these results demonstrate a need to evaluate DNA repair capacities in pluripotent cell lines, in order to characterize their genomic stability, prior to their pre-clinical and clinical use. PMID:22412831

  19. Human Blood Cell Sensing with Platinum Black Electroplated Impedance Sensor

    OpenAIRE

    Zheng, Siyang; Nandra, Mandheerej S.; Tai, Yu-Chong

    2007-01-01

    AC impedance sensing is an important method for biological cell analysis in flow cytometry. For micro impedance cell sensors, downsizing electrodes increases the double layer impedance of the metal-electrolyte interface, thus leaves no sensing zone in frequency domain and reduces the sensitivity significantly. We proposed using platinum black electroplated electrodes to solve the problem. In this paper, using this technique we demonstrated human blood cell sensing with high signal to noise ra...

  20. Property Of Human Dental Pulp Stem Cells And Peripheral Blood Hematopoietic Stem Cells That Differentiated Both Group To Cardiac Cells

    OpenAIRE

    Jabari F; Mohammadnejad J; Yavari K

    2013-01-01

    Dental pulp is the soft live tissue inside a tooth. Dental pulp contains stem cells, known as Dental Pulp Stem Cells. The finest Dental Pulp Stem Cells are found in a baby teeth or milk teeth. The stem cells from the milk teeth are ‘mesenchymal’ type of cells. cells that have the ability to generate a wide variety of cell types like chondrocytes, osteoblasts and adipocytes. To isolate high-quality human dental pulp stem cells from accessible resources is an important goal for stem-cell resear...

  1. Gut microbiota modulate T cell trafficking into human colorectal cancer.

    Science.gov (United States)

    Cremonesi, Eleonora; Governa, Valeria; Garzon, Jesus Francisco Glaus; Mele, Valentina; Amicarella, Francesca; Muraro, Manuele Giuseppe; Trella, Emanuele; Galati-Fournier, Virginie; Oertli, Daniel; Däster, Silvio Raffael; Droeser, Raoul A; Weixler, Benjamin; Bolli, Martin; Rosso, Raffaele; Nitsche, Ulrich; Khanna, Nina; Egli, Adrian; Keck, Simone; Slotta-Huspenina, Julia; Terracciano, Luigi M; Zajac, Paul; Spagnoli, Giulio Cesare; Eppenberger-Castori, Serenella; Janssen, Klaus-Peter; Borsig, Lubor; Iezzi, Giandomenica

    2018-02-06

    Tumour-infiltrating lymphocytes (TILs) favour survival in human colorectal cancer (CRC). Chemotactic factors underlying their recruitment remain undefined. We investigated chemokines attracting T cells into human CRCs, their cellular sources and microenvironmental triggers. Expression of genes encoding immune cell markers, chemokines and bacterial 16S ribosomal RNA (16SrRNA) was assessed by quantitative reverse transcription-PCR in fresh CRC samples and corresponding tumour-free tissues. Chemokine receptor expression on TILs was evaluated by flow cytometry on cell suspensions from digested tissues. Chemokine production by CRC cells was evaluated in vitro and in vivo, on generation of intraperitoneal or intracecal tumour xenografts in immune-deficient mice. T cell trafficking was assessed on adoptive transfer of human TILs into tumour-bearing mice. Gut flora composition was analysed by 16SrRNA sequencing. CRC infiltration by distinct T cell subsets was associated with defined chemokine gene signatures, including CCL5, CXCL9 and CXCL10 for cytotoxic T lymphocytes and T-helper (Th)1 cells; CCL17, CCL22 and CXCL12 for Th1 and regulatory T cells; CXCL13 for follicular Th cells; and CCL20 and CCL17 for interleukin (IL)-17-producing Th cells. These chemokines were expressed by tumour cells on exposure to gut bacteria in vitro and in vivo. Their expression was significantly higher in intracecal than in intraperitoneal xenografts and was dramatically reduced by antibiotic treatment of tumour-bearing mice. In clinical samples, abundance of defined bacteria correlated with high chemokine expression, enhanced T cell infiltration and improved survival. Gut microbiota stimulate chemokine production by CRC cells, thus favouring recruitment of beneficial T cells into tumour tissues. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  2. CD34-positive interstitial cells of the human detrusor

    DEFF Research Database (Denmark)

    Rasmussen, Helle; Hansen, Alastair; Smedts, Frank

    2007-01-01

    Interstitial cells of Cajal (ICC) are well described in the bowel wall. They are c-kit positive and play a role as pacemaker cells. Similar c-kit-positive cells have recently been described in the human bladder. The aim of this study was to characterize interstitial cells of the bladder detrusor...... using a panel of antibodies directed against CD117/c-kit, CD34, CD31, S100, tryptase, neurofilament, NSE, Factor-VIII and GFAP. A striking finding was an interstitial type of cell which is CD34 immunoreactive (CD34-ir) but CD117/c-kit negative. The cells have a tentacular morphology, enveloping...... and intermingling with individual muscle fasicles. Morphologically and immunohistochemically, they show no neurogenic, endothelial or mast cell differentiation. Transmission electron microscopy (TEM) showed the presence of interstitial cells with a round-to-oval nucleus, sparse perinuclear cytoplasm and long...

  3. Vulnerability of Normal Human Mammary Epithelial Cells to Oncogenic Transformation

    Science.gov (United States)

    2012-04-01

    escape senescence and acquire genomic changes. Nature 2001;409:633–7. 10. Olsen CL, Gardie B, Yaswen P, Stampfer MR. Raf-1- induced growth arrest in...p16INK4a. Cell 88:593–602. 10. Olsen CL, Gardie B, Yaswen P, Stampfer MR (2002) Raf-1-induced growth arrest in human mammary epithelial cells is p16...Cycle 3, 244–246. Olsen CL, Gardie B, Yaswen P, Stampfer MR (2002) Raf-1-induced growth arrest in human mammary epithelial cells is p16-independent and

  4. Human thymic epithelial cells express functional HLA-DP molecules

    DEFF Research Database (Denmark)

    Jørgensen, A; Röpke, C; Nielsen, M

    1996-01-01

    T lymphocytes, we examined whether human thymic epithelial cells (TEC) expressed HLA-DP molecules. We present evidence that TEC obtained from short time culture express low but significant levels of HLA-DP molecules. The expression of HLA-DP molecules was comparable to or higher than the expression...... of HLA-DP allospecific primed lymphocyte typing (PLT) CD4 T cell lines. IFN-gamma treatment strongly upregulated the HLA-DP allospecific PLT responses whereas other PLT responses remained largely unchanged. In conclusion, these data indicate that human thymus epithelial cells express significant levels...

  5. [Development of human embryonic stem cell platforms for human health-safety evaluation].

    Science.gov (United States)

    Yu, Guang-yan; Cao, Tong; Zou, Xiao-hui; Zhang, Xue-hui; Fu, Xin; Peng, Shuang-qing; Deng, Xu-liang; Li, Sheng-lin; Liu, He; Xiao, Ran; Ouyang, Hong-wei; Peng, Hui; Chen, Xiao; Zhao, Zeng-ming; Wang, Xiao-ying; Fang, Hai-qin; Lu, Lu; Ren, Yu-lan; Xu, Ming-ming

    2016-02-18

    The human embryonic stem cells (hESCs) serve as a self-renewable, genetically-healthy, pluripotent and single source of all body cells, tissues and organs. Therefore, it is considered as the good standard for all human stem cells by US, Europe and international authorities. In this study, the standard and healthy human mesenchymal progenitors, ligament tissues, cardiomyocytes, keratinocytes, primary neurons, fibroblasts, and salivary serous cells were differentiated from hESCs. The human cellular health-safety of NaF, retinoic acid, 5-fluorouracil, dexamethasone, penicillin G, adriamycin, lead acetate PbAc, bisphenol A-biglycidyl methacrylate (Bis-GMA) were evaluated selectively on the standardized platforms of hESCs, hESCs-derived cardiomyocytes, keratinocytes, primary neurons, and fibroblasts. The evaluations were compared with those on the currently most adopted cellular platforms. Particularly, the sensitivity difference of PM2.5 toxicity on standardized and healthy hESCs derived fibroblasts, currently adopted immortalized human bronchial epithelial cells Beas-2B and human umbilical vein endothelial cells (HUVECs) were evaluated. The RESULTS showed that the standardized hESCs cellular platforms provided more sensitivity and accuracy for human cellular health-safety evaluation.

  6. Yeast cell-based analysis of human lactate dehydrogenase isoforms.

    Science.gov (United States)

    Mohamed, Lulu Ahmed; Tachikawa, Hiroyuki; Gao, Xiao-Dong; Nakanishi, Hideki

    2015-12-01

    Human lactate dehydrogenase (LDH) has attracted attention as a potential target for cancer therapy and contraception. In this study, we reconstituted human lactic acid fermentation in Saccharomyces cerevisiae, with the goal of constructing a yeast cell-based LDH assay system. pdc null mutant yeast (mutated in the endogenous pyruvate decarboxylase genes) are unable to perform alcoholic fermentation; when grown in the presence of an electron transport chain inhibitor, pdc null strains exhibit a growth defect. We found that introduction of the human gene encoding LDHA complemented the pdc growth defect; this complementation depended on LDHA catalytic activity. Similarly, introduction of the human LDHC complemented the pdc growth defect, even though LDHC did not generate lactate at the levels seen with LDHA. In contrast, the human LDHB did not complement the yeast pdc null mutant, although LDHB did generate lactate in yeast cells. Expression of LDHB as a red fluorescent protein (RFP) fusion yielded blebs in yeast, whereas LDHA-RFP and LDHC-RFP fusion proteins exhibited cytosolic distribution. Thus, LDHB exhibits several unique features when expressed in yeast cells. Because yeast cells are amenable to genetic analysis and cell-based high-throughput screening, our pdc/LDH strains are expected to be of use for versatile analyses of human LDH. © The Authors 2015. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

  7. Cytokine activation induces human memory-like NK cells.

    Science.gov (United States)

    Romee, Rizwan; Schneider, Stephanie E; Leong, Jeffrey W; Chase, Julie M; Keppel, Catherine R; Sullivan, Ryan P; Cooper, Megan A; Fehniger, Todd A

    2012-12-06

    Natural killer (NK) cells are lymphocytes that play an important role in the immune response to infection and malignancy. Recent studies in mice have shown that stimulation of NK cells with cytokines or in the context of a viral infection results in memory-like properties. We hypothesized that human NK cells exhibit such memory-like properties with an enhanced recall response after cytokine preactivation. In the present study, we show that human NK cells preactivated briefly with cytokine combinations including IL-12, IL-15, and IL-18 followed by a 7- to 21-day rest have enhanced IFN-γ production after restimulation with IL-12 + IL-15, IL-12 + IL-18, or K562 leukemia cells. This memory-like phenotype was retained in proliferating NK cells. In CD56(dim) NK cells, the memory-like IFN-γ response was correlated with the expression of CD94, NKG2A, NKG2C, and CD69 and a lack of CD57 and KIR. Therefore, human NK cells have functional memory-like properties after cytokine activation, which provides a novel rationale for integrating preactivation with combinations of IL-12, IL-15, and IL-18 into NK cell immunotherapy strategies.

  8. Subpopulations of stem-like cells in side population cells from the human bladder transitional cell cancer cell line T24.

    Science.gov (United States)

    Ning, Z-F; Huang, Y-J; Lin, T-X; Zhou, Y-X; Jiang, C; Xu, K-W; Huang, H; Yin, X-B; Huang, J

    2009-01-01

    Cancer stem cells can be isolated from human tumours using specific cell surface markers. Bladder cancer cells, however, lack specific cell surface markers, making this approach impracticable. In this study an alternative method was used, involving isolation of side population cells to explore the stem cell characteristics of bladder cancer. Side population cells were isolated from the bladder transitional cell cancer cell line T24 and examined for potential stem cell characteristics related to proliferation, cell cycle distribution, self-renewal and differentiation. It was observed that T24 side population cells have stronger proliferative and colony formation abilities than non-side population cells. Side population cells were also more resistant to chemotherapy and radiotherapy, which may be due to expression of the ATP-binding cassette half-transporter, sub-family G, member 2 protein. Overall, the results suggest that side population cells from the human bladder transitional cell cancer cell line T24 harbour stem-like cells.

  9. Probing Human NK Cell Biology Using Human Immune System (HIS) Mice.

    Science.gov (United States)

    Li, Yan; Di Santo, James P

    2016-01-01

    Our incomplete understanding of the mechanisms that orchestrate human lymphocyte differentiation and condition human immune responses is in part due to the limited access to normal human tissue samples that can inform on these complex processes. In addition, in vitro culture conditions fail to recapitulate the three-dimensional microenvironments that influence cell-cell interactions and impact on immune outcomes. Small animals provide a preclinical model to dissect and probe immunity and over the past decades, development of immunodeficient hosts that can be engrafted with human hematopoietic precursors and mature cells have led to the development of new in vivo models to study human lymphocyte development and function. Natural killer (NK) cells are implicated in the recognition and elimination of pathogen-infected and transformed cells and belong to a family of diverse innate lymphoid cells (ILCs) that provide early immune defense against disease. Here, we summarize the use of humanized mouse models for the study of NK cell and group 1 ILCs and their respective roles in immunity and tissue homeostasis.

  10. Fraction against Human Cancer Cell Lines

    African Journals Online (AJOL)

    Purpose: To investigate the anti-proliferative and apoptotic activity of crude and dichloromethane fraction of A. sieberi against seven cancer cell lines (Colo20, HCT116, DLD, MCF7, Jurkat, HepG2 and. L929). Methods: A. sieberi was extracted with methanol and further purification was carried out using liquid-.

  11. Genetic Expeditions with Haploid Human Cells

    NARCIS (Netherlands)

    Jae, L.T.

    2015-01-01

    Random mutagenesis followed by phenotypic selection (forward genetics) is among the most powerful tools to elucidate the molecular basis of intricate biological processes and has been used in a suite of model organisms throughout the last century. However, its application to cultured mammalian cells

  12. Epigenetic Classification of Human Mesenchymal Stromal Cells

    Directory of Open Access Journals (Sweden)

    Danilo Candido de Almeida

    2016-02-01

    Full Text Available Standardization of mesenchymal stromal cells (MSCs is hampered by the lack of a precise definition for these cell preparations; for example, there are no molecular markers to discern MSCs and fibroblasts. In this study, we followed the hypothesis that specific DNA methylation (DNAm patterns can assist classification of MSCs. We utilized 190 DNAm profiles to address the impact of tissue of origin, donor age, replicative senescence, and serum supplements on the epigenetic makeup. Based on this, we elaborated a simple epigenetic signature based on two CpG sites to classify MSCs and fibroblasts, referred to as the Epi-MSC-Score. Another two-CpG signature can distinguish between MSCs from bone marrow and adipose tissue, referred to as the Epi-Tissue-Score. These assays were validated by site-specific pyrosequencing analysis in 34 primary cell preparations. Furthermore, even individual subclones of MSCs were correctly classified by our epigenetic signatures. In summary, we propose an alternative concept to use DNAm patterns for molecular definition of cell preparations, and our epigenetic scores facilitate robust and cost-effective quality control of MSC cultures.

  13. Vascular potential of human pluripotent stem cells

    Science.gov (United States)

    Cardiovascular disease is the number one cause of death and disability in the US. Understanding the biological activity of stem and progenitor cells, and their ability to contribute to the repair, regeneration and remodeling of the heart and blood vessels affected by pathological processes is an ess...

  14. Epitope tagging of endogenous genes in diverse human cell lines.

    Science.gov (United States)

    Kim, Jung-Sik; Bonifant, Challice; Bunz, Fred; Lane, William S; Waldman, Todd

    2008-11-01

    Epitope tagging is a powerful and commonly used approach for studying the physical properties of proteins and their functions and localization in eukaryotic cells. In the case of Saccharomyces cerevisiae, it has been possible to exploit the high efficiency of homologous recombination to tag proteins by modifying their endogenous genes, making it possible to tag virtually every endogenous gene and perform genome-wide proteomics experiments. However, due to the relative inefficiency of homologous recombination in cultured human cells, epitope-tagging approaches have been limited to ectopically expressed transgenes, with the attendant limitations of their nonphysiological transcriptional regulation and levels of expression. To overcome this limitation, a modification and extension of adeno-associated virus-mediated human somatic cell gene targeting technology is described that makes it possible to simply and easily create an endogenous epitope tag in the same way that it is possible to knock out a gene. Using this approach, we have created and validated human cell lines with epitope-tagged alleles of two cancer-related genes in a variety of untransformed and transformed human cell lines. This straightforward approach makes it possible to study the physical and biological properties of endogenous proteins in human cells without the need for specialized antibodies for individual proteins of interest.

  15. French legal framework relating to human tissues and cells.

    Science.gov (United States)

    Pascal, P; Damour, O; Colpart, J J; Braye, F

    2000-03-01

    Thousands of patients receive human tissue grafts every year. Developments in cell and tissue engineering have also increased considerably the number of available products of human origin. France has very strict regulations, in part stimulated by problems of public health and ethics that have emerged in recent years and also in part as a result of a report by the 'Inspection Générale des Affaires Sociales' on the removal and grafting of human tissues in May 1993. These have resulted in two laws on bio-ethics being passed, in July 1994, that are the basis of current legislation and represent the first steps in differentiating between organs and tissues or cells. Henceforth, the French legal framework covering tissues and cells of human origin has been increased to include a large number of legislative texts and regulations. The fundamental ethical principles that are consent, free donation, anonymity, no publicity and respect for public health have become a major ethical imperative that applies to all products originating from the human body including tissue and cells. In addition, specific provisions have been made covering: removal (conditions for removal and system for authorization); conservation, transformation, distribution, packaging, import and export of tissues and cells; and tissue and cell grafts. Finally, penal and administrative sanctions have been foreseen where there is non-compliance with these regulations.

  16. Transcriptome coexpression map of human embryonic stem cells

    Directory of Open Access Journals (Sweden)

    Mattson Mark P

    2006-05-01

    Full Text Available Abstract Background Human embryonic stem (ES cells hold great promise for medicine and science. The transcriptome of human ES cells has been studied in detail in recent years. However, no systematic analysis has yet addressed whether gene expression in human ES cells may be regulated in chromosomal domains, and no chromosomal domains of coexpression have been identified. Results We report the first transcriptome coexpression map of the human ES cell and the earliest stage of ES differentiation, the embryoid body (EB, for the analysis of how transcriptional regulation interacts with genomic structure during ES self-renewal and differentiation. We determined the gene expression profiles from multiple ES and EB samples and identified chromosomal domains showing coexpression of adjacent genes on the genome. The coexpression domains were not random, with significant enrichment in chromosomes 8, 11, 16, 17, 19, and Y in the ES state, and 6, 11, 17, 19 and 20 in the EB state. The domains were significantly associated with Giemsa-negative bands in EB, yet showed little correlation with known cytogenetic structures in ES cells. Different patterns of coexpression were revealed by comparative transcriptome mapping between ES and EB. Conclusion The findings and methods reported in this investigation advance our understanding of how genome organization affects gene expression in human ES cells and help to identify new mechanisms and pathways controlling ES self-renewal or differentiation.

  17. Human platelet lysate: Replacing fetal bovine serum as a gold standard for human cell propagation?

    Science.gov (United States)

    Burnouf, Thierry; Strunk, Dirk; Koh, Mickey B C; Schallmoser, Katharina

    2016-01-01

    The essential physiological role of platelets in wound healing and tissue repair builds the rationale for the use of human platelet derivatives in regenerative medicine. Abundant growth factors and cytokines stored in platelet granules can be naturally released by thrombin activation and clotting or artificially by freeze/thaw-mediated platelet lysis, sonication or chemical treatment. Human platelet lysate prepared by the various release strategies has been established as a suitable alternative to fetal bovine serum as culture medium supplement, enabling efficient propagation of human cells under animal serum-free conditions for a multiplicity of applications in advanced somatic cell therapy and tissue engineering. The rapidly increasing number of studies using platelet derived products for inducing human cell proliferation and differentiation has also uncovered a considerable variability of human platelet lysate preparations which limits comparability of results. The main variations discussed herein encompass aspects of donor selection, preparation of the starting material, the possibility for pooling in plasma or additive solution, the implementation of pathogen inactivation and consideration of ABO blood groups, all of which can influence applicability. This review outlines the current knowledge about human platelet lysate as a powerful additive for human cell propagation and highlights its role as a prevailing supplement for human cell culture capable to replace animal serum in a growing spectrum of applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

  18. The ethics of patenting human embryonic stem cells.

    Science.gov (United States)

    Chapman, Audrey R

    2009-09-01

    Just as human embryonic stem cell research has generated controversy about the uses of human embryos for research and therapeutic applications, human embryonic stem cell patents raise fundamental ethical issues. The United States Patent and Trademark Office has granted foundational patents, including a composition of matter (or product) patent to the Wisconsin Alumni Research Foundation (WARF), the University of Wisconsin-Madison's intellectual property office. In contrast, the European Patent Office rejected the same WARF patent application for ethical reasons. This article assesses the appropriateness of these patents placing the discussion in the context of the deontological and consequentialist ethical issues related to human embryonic stem cell patenting. It advocates for a patent system that explicitly takes ethical factors into account and explores options for new types of intellectual property arrangements consistent with ethical concerns.

  19. Human pluripotent stem cells: an emerging model in developmental biology.

    Science.gov (United States)

    Zhu, Zengrong; Huangfu, Danwei

    2013-02-01

    Developmental biology has long benefited from studies of classic model organisms. Recently, human pluripotent stem cells (hPSCs), including human embryonic stem cells and human induced pluripotent stem cells, have emerged as a new model system that offers unique advantages for developmental studies. Here, we discuss how studies of hPSCs can complement classic approaches using model organisms, and how hPSCs can be used to recapitulate aspects of human embryonic development 'in a dish'. We also summarize some of the recently developed genetic tools that greatly facilitate the interrogation of gene function during hPSC differentiation. With the development of high-throughput screening technologies, hPSCs have the potential to revolutionize gene discovery in mammalian development.

  20. Generation of neuropeptidergic hypothalamic neurons from human pluripotent stem cells

    Science.gov (United States)

    Merkle, Florian T.; Maroof, Asif; Wataya, Takafumi; Sasai, Yoshiki; Studer, Lorenz; Eggan, Kevin; Schier, Alexander F.

    2015-01-01

    Hypothalamic neurons orchestrate many essential physiological and behavioral processes via secreted neuropeptides, and are relevant to human diseases such as obesity, narcolepsy and infertility. We report the differentiation of human pluripotent stem cells into many of the major types of neuropeptidergic hypothalamic neurons, including those producing pro-opiolemelanocortin, agouti-related peptide, hypocretin/orexin, melanin-concentrating hormone, oxytocin, arginine vasopressin, corticotropin-releasing hormone (CRH) or thyrotropin-releasing hormone. Hypothalamic neurons can be generated using a ‘self-patterning’ strategy that yields a broad array of cell types, or via a more reproducible directed differentiation approach. Stem cell-derived human hypothalamic neurons share characteristic morphological properties and gene expression patterns with their counterparts in vivo, and are able to integrate into the mouse brain. These neurons could form the basis of cellular models, chemical screens or cellular therapies to study and treat common human diseases. PMID:25670790

  1. Exclusive destruction of mitotic spindles in human cancer cells.

    Science.gov (United States)

    Visochek, Leonid; Castiel, Asher; Mittelman, Leonid; Elkin, Michael; Atias, Dikla; Golan, Talia; Izraeli, Shai; Peretz, Tamar; Cohen-Armon, Malka

    2017-03-28

    We identified target proteins modified by phenanthrenes that cause exclusive eradication of human cancer cells. The cytotoxic activity of the phenanthrenes in a variety of human cancer cells is attributed by these findings to post translational modifications of NuMA and kinesins HSET/kifC1 and kif18A. Their activity prevented the binding of NuMA to α-tubulin and kinesins in human cancer cells, and caused aberrant spindles. The most efficient cytotoxic activity of the phenanthridine PJ34, caused significantly smaller aberrant spindles with disrupted spindle poles and scattered extra-centrosomes and chromosomes. Concomitantly, PJ34 induced tumor growth arrest of human malignant tumors developed in athymic nude mice, indicating the relevance of its activity for cancer therapy.

  2. Gene expression signatures of human cell and tissue longevity.

    Science.gov (United States)

    Seim, Inge; Ma, Siming; Gladyshev, Vadim N

    2016-01-01

    Different cell types within the body exhibit substantial variation in the average time they live, ranging from days to the lifetime of the organism. The underlying mechanisms governing the diverse lifespan of different cell types are not well understood. To examine gene expression strategies that support the lifespan of different cell types within the human body, we obtained publicly available RNA-seq data sets and interrogated transcriptomes of 21 somatic cell types and tissues with reported cellular turnover, a bona fide estimate of lifespan, ranging from 2 days (monocytes) to a lifetime (neurons). Exceptionally long-lived neurons presented a gene expression profile of reduced protein metabolism, consistent with neuronal survival and similar to expression patterns induced by longevity interventions such as dietary restriction. Across different cell lineages, we identified a gene expression signature of human cell and tissue turnover. In particular, turnover showed a negative correlation with the energetically costly cell cycle and factors supporting genome stability, concomitant risk factors for aging-associated pathologies. In addition, the expression of p53 was negatively correlated with cellular turnover, suggesting that low p53 activity supports the longevity of post-mitotic cells with inherently low risk of developing cancer. Our results demonstrate the utility of comparative approaches in unveiling gene expression differences among cell lineages with diverse cell turnover within the same organism, providing insights into mechanisms that could regulate cell longevity.

  3. Telmisartan inhibits human urological cancer cell growth through early apoptosis

    Science.gov (United States)

    MATSUYAMA, MASAHIDE; FUNAO, KIYOAKI; KURATSUKURI, KATSUYUKI; TANAKA, TOMOAKI; KAWAHITO, YUTAKA; SANO, HAJIME; CHARGUI, JAMEL; TOURAINE, JEAN-LOUIS; YOSHIMURA, NORIO; YOSHIMURA, RIKIO

    2010-01-01

    Angiotensin II receptor blockers (ARBs) are widely used as hypertensive therapeutic agents. In addition, studies have provided evidence that ARBs have the potential to inhibit the growth of several types of cancer cells. It was reported that telmisartan (a type of ARB) has peroxisome proliferator-activated receptor (PPAR)-γ activation activity. We previously reported that the PPAR-γ ligand induces growth arrest in human urological cancer cells through apoptosis. In this study, we evaluated the effects of telmisartan and other ARBs on cell proliferation in renal cell carcinoma (RCC), bladder cancer (BC), prostate cancer (PC) and testicular cancer (TC) cell lines. The inhibitory effects of telmisartan and other ARBs (candesartan, valsartan, irbesartan and losartan) on the growth of the RCC, BC, PC and TC cell lines was investigated using an MTT assay. Flow cytometry and Hoechst staining were used to determine whether the ARBs induced apoptosis. Telmisartan caused marked growth inhibition in the urological cancer cells in a dose- and time-dependent manner. Urological cancer cells treated with 100 μM telmisartan underwent early apoptosis and DNA fragmentation. However, the other ARBs had no effect on cell proliferation in any of the urological cancer cell lines. Telmisartan may mediate potent anti-proliferative effects in urological cancer cells through PPAR-γ. Thus, telmisartan is a potent target for the prevention and treatment of human urological cancer. PMID:22993542

  4. Comparison of Human T Cell Repertoire Generated in Xenogeneic Porcine and Human Thymus Grafts1

    Science.gov (United States)

    Shimizu, Ichiro; Fudaba, Yasuhiro; Shimizu, Akira; Yang, Yong-Guang; Sykes, Megan

    2009-01-01

    Background Xenogeneic thymus transplantation is an effective approach to achieving T cell tolerance across highly disparate xenogeneic species barriers. We have previously demonstrated that phenotypically normal, specifically tolerant human T cells are generated in porcine thymic grafts. In this study, we assessed the diversity of the human T cell repertoire generated in porcine thymic xenografts. We also examined the ability of porcine thymus grafts to coexist with human thymus grafts. Methods Fetal swine (SW) or human (HU) thymus with human fetal liver (FL) fragments were transplanted under the kidney capsule of 3Gy irradiated NOD/SCID mouse recipients. Thymus tissue was harvested approximately 16 weeks post-transplant for analysis of mixed lymphocyte reactions and spectratyping of human CD4 and CD8 single positive (SP) thymocytes. Results TCR β genes of human CD4 and CD8 SP cells developing in HU and SW thymus grafts showed similar, normal CDR3 length distributions. Human T cells developing in SW thymus grafts showed specific unresponsiveness to the MHC of the donor swine, in MLR assays. In 2 of 3 animals receiving SW and HU thymus grafts under opposite kidney capsules, both grafts functioned. In animals with surviving SW grafts, thymocytes from the SW but not the HU grafts showed specific unresponsiveness to the SW donor. Conclusion SW thymus grafts support generation of human T cells with a diverse TCR repertoire. Human thymocytes in human thymus grafts are not tolerized by the presence of an additional porcine thymus, but tolerance might be achieved by post-thymic encounter with porcine antigens. PMID:18724231

  5. Alteration of Cell Cycle Mediated by Zinc in Human Bronchial Epithelial Cells In Vitro

    Science.gov (United States)

    Zinc (Zn2+), a ubiquitous ambient air contaminant, presents an oxidant challenge to the human lung and is linked to adverse human health effects. To further elucidate the adaptive and apoptotic cellular responses of human airway cells to Zn2+, we performed pilot studies to examin...

  6. Mercury induces inflammatory mediator release from human mast cells

    Directory of Open Access Journals (Sweden)

    Peterson Erika

    2010-03-01

    Full Text Available Abstract Background Mercury is known to be neurotoxic, but its effects on the immune system are less well known. Mast cells are involved in allergic reactions, but also in innate and acquired immunity, as well as in inflammation. Many patients with Autism Spectrum Disorders (ASD have "allergic" symptoms; moreover, the prevalence of ASD in patients with mastocytosis, characterized by numerous hyperactive mast cells in most tissues, is 10-fold higher than the general population suggesting mast cell involvement. We, therefore, investigated the effect of mercuric chloride (HgCl2 on human mast cell activation. Methods Human leukemic cultured LAD2 mast cells and normal human umbilical cord blood-derived cultured mast cells (hCBMCs were stimulated by HgCl2 (0.1-10 μM for either 10 min for beta-hexosaminidase release or 24 hr for measuring vascular endothelial growth factor (VEGF and IL-6 release by ELISA. Results HgCl2 induced a 2-fold increase in β-hexosaminidase release, and also significant VEGF release at 0.1 and 1 μM (311 ± 32 pg/106 cells and 443 ± 143 pg/106 cells, respectively from LAD2 mast cells compared to control cells (227 ± 17 pg/106 cells, n = 5, p 2 (0.1 μM to the proinflammatory neuropeptide substance P (SP, 0.1 μM had synergestic action in inducing VEGF from LAD2 mast cells. HgCl2 also stimulated significant VEGF release (360 ± 100 pg/106 cells at 1 μM, n = 5, p 6 cells, and IL-6 release (466 ± 57 pg/106 cells at 0.1 μM compared to untreated cells (13 ± 25 pg/106 cells, n = 5, p 2 (0.1 μM to SP (5 μM further increased IL-6 release. Conclusions HgCl2 stimulates VEGF and IL-6 release from human mast cells. This phenomenon could disrupt the blood-brain-barrier and permit brain inflammation. As a result, the findings of the present study provide a biological mechanism for how low levels of mercury may contribute to ASD pathogenesis.

  7. Human natural killer cell development in secondary lymphoid tissues

    Science.gov (United States)

    Freud, Aharon G.; Yu, Jianhua; Caligiuri, Michael A.

    2014-01-01

    For nearly a decade it has been appreciated that critical steps in human natural killer (NK) cell development likely occur outside of the bone marrow and potentially necessitate distinct microenvironments within extramedullary tissues. The latter include the liver and gravid uterus as well as secondary lymphoid tissues such as tonsils and lymph nodes. For as yet unknown reasons these tissues are naturally enriched with NK cell developmental intermediates (NKDI) that span a maturation continuum starting from an oligopotent CD34+CD45RA+ hematopoietic precursor cell to a cytolytic mature NK cell. Indeed despite the detection of NKDI within the aforementioned tissues, relatively little is known about how, why, and when these tissues may be most suited to support NK cell maturation and how this process fits in with other components of the human immune system. With the discovery of other innate lymphoid subsets whose immunophenotypes overlap with those of NKDI, there is also need to revisit and potentially re-characterize the basic immunophenotypes of the stages of the human NK cell developmental pathway in vivo. In this review, we provide an overview of human NK cell development in secondary lymphoid tissues and discuss the many questions that remain to be answered in this exciting field. PMID:24661538

  8. Mutation of human cells by kerosene soot

    Energy Technology Data Exchange (ETDEWEB)

    Skopek, T.R. (Massachusetts Inst. of Tech., Cambridge, MA); Liber, H.L.; Kaden, D.A.; Hites, R.A.; Thilly, W.G.

    1979-08-01

    The polycyclic aromatic hydrocarbon fraction of a kerosene soot induced forward mutation in human diploid lymphoblasts when coincubated with coincubated with Sprague-Dawley rat liver postmitochondrial supematant. Two components of the kerosene soot extract, benzo(a)pyrene (BP) and cyclopenta(cd)pyrene (CP), were also tested. BP was not mutagenic at the concentration found in the soot extract, although it was active at higher concentrations. The amount of CP present could account for approximately 8% of the total mutation observed with the soot. The results were compared to data obtained previously in a similar mutation assay in Salmonella typhimurium. the protocol described permits the facile assay of mutation at the hgprt locus in human lymphoblasts; such mutation is induced by compounds or complex mixtures requiring mixed-function oxygenase activity for metabolism to genetically active derivatives.

  9. Towards expansion of human hair follicle stem cells in vitro.

    Science.gov (United States)

    Oh, J H; Mohebi, P; Farkas, D L; Tajbakhsh, J

    2011-06-01

    Multipotential human hair follicle stem cells can differentiate into various cell lineages and thus are investigated here as potential autologous sources for regenerative medicine. Towards this end, we have attempted to expand these cells, directly isolated from minimal amounts of hair follicle explants, to numbers more suitable for stem-cell therapy. Two types of human follicle stem cells, commercially available and directly isolated, were cultured using an in-house developed medium. The latter was obtained from bulge areas of hair follicles by mechanical and enzymatic dissociation, and was magnetically enriched for its CD200(+) fraction. Isolated cells were cultured for up to 4 weeks, on different supports: blank polystyrene, laminin- and Matrigel(TM) -coated surfaces. Two-fold expansion was found, highlighting the slow-cycling nature of these cells. Flow cytometry characterization revealed: magnetic enrichment increased the proportion of CD200(+) cells from initially 43.3% (CD200+, CD34: 25.8%; CD200+, CD34+: 17.5%) to 78.2% (CD200+, CD34: 41.5%; CD200+, CD34+: 36.7%). Enriched cells seemed to have retained and passed on their morphological and molecular phenotypes to their progeny, as isolated CD200(+) presenting cells expanded in our medium to a population with 80% of cells being CD200(+): 51.5% (CD200(+), CD34(-)) and 29.6% (CD200(+), CD34(+)). This study demonstrates the possibility of culturing human hair follicle stem cells without causing any significant changes to phenotypes of the cells. © 2011 Blackwell Publishing Ltd.

  10. Delocalized Claudin-1 promotes metastasis of human osteosarcoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Jian, Yuekui; Chen, Changqiong; Li, Bo; Tian, Xiaobin, E-mail: drtxb_guiyang@sina.com

    2015-10-23

    Tight junction proteins (TJPs) including Claudins, Occludin and tight junction associated protein Zonula occludens-1 (ZO-1), are the most apical component of junctional complex that mediates cell–cell adhesion in epithelial and endothelial cells. In human malignancies, TJPs are often deregulated and affect cellular behaviors of tumor cells. In this study, we investigated alternations of TJPs and related biological characteristics in human osteosarcoma (OS). Claudin1 was increased in the metastatic OS cells (KRIB and KHOS) compared with the normal osteoblast cells (hFOB1.19) or primary tumor cells (HOS and U2OS), whereas no significant difference was found in Occludin and ZO-1. Immunohistochemistry, immunofluorescence and Western blotting revealed that Claudin1 was initially localized at cell junctions of normal osteoblasts, but substantially delocalized to the nucleus of metastatic OS cells. Phenotypically, inhibition of the nucleus Claudin1 expression compromised the metastatic potential of KRIB and KHOS cells. Moreover, we found that protein kinase C (PKC) but not PKA phosphorylation influenced Claudin1 expression and cellular functions, as PKC inhibitor (Go 6983 and Staurosporine) or genetic silencing of PKC reduced Claudin1 expression and decreased the motility of KRIB and KHOS cells. Taken together, our study implied that delocalization of claudin-1 induced by PKC phosphorylation contributes to metastatic capacity of OS cells. - Highlights: • Claudin1 is increased during the malignant transformation of human OS. • Delocalization of Claudin1 in metastatic OS cells. • Silencing nuclear Claudin1 expression inhibits cell invasion of OS. • Deregulated Claudin1 is regulated by PKC.

  11. Inflammatory Alteration of Human T Cells Exposed Continuously to Asbestos.

    Science.gov (United States)

    Kumagai-Takei, Naoko; Yamamoto, Shoko; Lee, Suni; Maeda, Megumi; Masuzzaki, Hidenori; Sada, Nagisa; Yu, Min; Yoshitome, Kei; Nishimura, Yasumitsu; Otsuki, Takemi

    2018-02-08

    Asbestos is a known carcinogen and exposure can lead to lung cancer and malignant mesothelioma. To examine the effects of asbestos fibers on human immune cells, the human T cell leukemia/lymphoma virus (HTLV)-1 immortalized human T cell line MT-2 was employed. Following continuous exposure to asbestos fibers for more than eight months, MT-2 sublines showed acquisition of resistance to asbestos-induced apoptosis with decreased death signals and increased surviving signals. These sublines showed various characteristics that suggested a reduction in anti-tumor immunity. On the other hand, inflammatory changes such as expression of MMP7, CXCR5, CXCL13 and CD44 was found to be markedly higher in sublines continuously exposed to asbestos compared with original MT-2 cells. All of these molecules contribute to lung inflammation, T and B cell interactions and connections between mesothelial cells and T cells. Thus, further investigation focusing on these molecules may shed light on the role of chronic inflammation caused by asbestos exposure and the occurrence of malignant mesothelioma. Finally, regarding peripheral T cells from healthy donors (HD) and asbestos-exposed patients with pleural plaque (PP) or malignant pleural mesothelioma (MPM), following stimulation of CD4+ T cells, T cells from MPM patients showed reduced potential of interferon (IFN)-γ expression. Moreover, levels of interleukin (IL)-6, one of the most important cytokines in chronic inflammation, in cultured supernatants were higher in PP and MPM patients compared with HD. Overall, asbestos-induced chronic inflammation in the lung as well as the pleural cavity may facilitate the onset of asbestos-induced cancers due to alterations in the interactions among fibers, immune cells such as T and B cells and macrophages, and mesothelial and lung epithelial cells. Further investigations regarding chronic inflammation caused by asbestos fibers may assist in identifying molecular targets for preventive and

  12. Tim-3 expression defines regulatory T cells in human tumors.

    Directory of Open Access Journals (Sweden)

    Jing Yan

    Full Text Available Tim-3, a member of the novel Tim (T cell immunoglobulin and mucin domain family, has been reported to negatively regulate the immune responses against viral infection and had implications for autoimmune disease. However, the nature and role of Tim-3(+ CD4 T cells in human tumors remain largely unknown. In the present study, we characterized Tim-3(+ CD4 T cells in 100 specimens from human hepatocellular, cervical, colorectal and ovarian carcinoma patients. Compared with peripheral blood and nontumor-infiltrating lymphocytes, the lymphocytes isolated from the corresponding tumor tissues of hepatocellular, cervical, colorectal and ovarian carcinoma patients contained significantly greater proportion of Tim-3(+ CD4 T cells. The majority of tumor-derived Tim-3(+ CD4 T cells exhibited an impaired capacity to produce IFN-γ and IL-2, but expressed higher levels of CD25, Foxp3, CTLA-4 and GITR than their Tim-3(- CD4 T cell counterparts. In contrast, most Tim-3(+ CD4 T cells isolated from the paired nontumor tissues and peripheral blood did not express these molecules. Moreover, tumor-derived Tim-3(+ CD4 T cells, but not tumor-derived Tim-3(- CD4 T cells, significantly suppressed the proliferation of autologous CD8(+ T cells in vitro. Notably, multi-color immunofluorescence and confocal microscopy demonstrated that Tim-3(+Foxp3(+CD4(+ cells were preferentially distributed in the tumor nest rather than the peritumoral stroma of hepatocellular carcinoma. Together, our data indicate that Tim-3-expressing CD4 T cells in human tumors could represent the functional regulatory T cells which contribute to the formation of the immune-suppressive tumor micromilieu.

  13. MUC1 in human milk blocks transmission of human immunodeficiency virus from dendritic cells to T cells

    NARCIS (Netherlands)

    Saeland, Eirikur; de Jong, Marein A. W. P.; Nabatov, Alexey A.; Kalay, Hakan; Geijtenbeek, Teunis B. H.; van Kooyk, Yvette

    2009-01-01

    Mother-to-child transmission of human immunodeficiency virus-1 (HIV-1) occurs frequently via breast-feeding. HIV-1 targets DC-SIGN+ dendritic cells (DCs) in mucosal areas that allow efficient transmission of the virus to T cells. Here, we demonstrate that the epithelial mucin MUC1, abundant in milk,

  14. Differentiation of reprogrammed human adipose mesenchymal stem cells toward neural cells with defined transcription factors.

    Science.gov (United States)

    Qu, Xinjian; Liu, Tianqing; Song, Kedong; Li, Xiangqin; Ge, Dan

    2013-10-04

    Somatic cell reprogramming may become a powerful approach to generate specific human cell types for cell-fate determination studies and potential transplantation therapies of neurological diseases. Here we report a reprogramming methodology with which human adipose stem cells (hADSCs) can be differentiated into neural cells. After being reprogrammed with polycistronic plasmid carrying defined factor OCT3/4, SOX2, KLF4 and c-MYC, and further treated with neural induce medium, the hADSCs switched to differentiate toward neural cell lineages. The generated cells had normal karyotypes and exogenous vector sequences were not inserted in the genomes. Therefore, this cell lineage conversion methodology bypasses the risk of mutation and gene instability, and provides a novel strategy to obtain patient-specific neural cells for basic research and therapeutic application. Copyright © 2013 Elsevier Inc. All rights reserved.

  15. Lipopolysaccharide induces IFN-γ production in human NK cells

    Directory of Open Access Journals (Sweden)

    Leonid M Kanevskiy

    2013-01-01

    Full Text Available NK cells have been shown to play a regulatory role in sepsis. According to the current view, NK cells become activated via macrophages or dendritic cells primed by lipopolysaccharide (LPS. Recently TLR4 gene expression was detected in human NK cells suggesting the possibility of a direct action of LPS on NK cells. In this study, effects of LPS on NK cell cytokine production and cytotoxicity were studied using highly purified human NK cells. LPS induced IFN-γ production in the presence of IL-2 in cell populations containing >98% CD56+ cells. Surprisingly, in the same experiments LPS decreased NK cell degranulation. No significant expression of markers related to blood dendritic cells, monocytes or T or B lymphocytes in the NK cell preparations was observed; the portions of HLA-DRbright, CD14+, CD3+ and CD20+ cells amounted to less than 0.1% within the cell populations. No more than 0.2% of NK cells were shown to be slightly positive for surface TLR4 in our experimental system, although intracellular staining revealed moderate amounts of TLR4 inside the NK cell population. These cells were negative for surface CD14, the receptor participating in LPS recognition by TLR4. Incubation of NK cells with IL-2 or/and LPS did not lead to an increase in TLR4 surface expression. TLR4–CD56+ NK cells isolated by cell sorting secreted IFN-γ in response to LPS. Antibody to TLR4 did not block the LPS-induced increase in IFN-γ production. We have also shown that Re-form of LPS lacking outer core oligosaccharide and O-antigen induces less cytokine production in NK cells than full length LPS. We speculate that the polysaccharide fragments of LPS molecule may take part in LPS-induced IFN-γ production by NK cells. Collectively our data suggest the existence of a mechanism of LPS direct action on NK cells distinct from established TLR4-mediated signaling.

  16. Secretory products of breast cancer cells specifically affect human osteoblastic cells: partial characterization of active factors.

    Science.gov (United States)

    Siwek, B; Lacroix, M; De Pollak, C; Marie, P; Body, J J

    1997-04-01

    The pathogenesis of tumor-induced osteolysis (TIO) following breast cancer metastases in bone remains unclear. We postulated that osteoblasts could be target cells for the secretory products of breast cancer cells. We previously showed that serum-free conditioned medium (CM) of the breast cancer cell line MCF-7 inhibits DNA synthesis by 75% of control values in osteoblast-like cells SaOS-2 and that this effect is only in a minor part due to transforming growth factor beta secretion. To establish the specificity of our observations and to look for other biologically active factors, we have tested the effects of medium conditioned by several cancer and noncancer cell lines (breast, colon, placenta, or fibrosarcoma) on the proliferation of osteoblast-like cells (SaOS-2, MG-63), normal human osteoblasts, human fibrosarcoma cells, and normal human fibroblasts. Culture medium (1:2) of the breast cancer cell lines MCF-7, T-47D, MDA-MB-231, and SK-BR-3 inhibited by 25-50% the proliferation of osteoblast-like cells SaOS-2, MG-63, and normal osteoblasts as evaluated by the MTT survival test or [3H]thymidine incorporation. MCF-7 cells completely inhibited the proliferation of normal human osteoblasts in coculture. This inhibitory effect was reversible and not due to cytotoxicity. Moreover, the cyclic adenosine monophosphate (cAMP) response to parathyroid hormone (PTH) of osteoblast-like cells SaOS-2 was also increased by 100-240% by the same CM. Such activities were, however, not detected in medium from the breast noncancer cell line HBL-100 or in the medium conditioned by non-breast cancer cell lines (COLO 320DM, HT-29, JAR, or HT-1080). Medium from the breast cancer cells had no effect on normal human fibroblasts or fibrosarcoma cells (HT-1080), suggesting the specificity of their action on human osteoblasts. After partial purification by ultrafiltration and size-exclusion chromatography, we found that medium of T-47D cells contained at least three nonprostanoid factors of

  17. Generation of mature T cells from human hematopoietic stem and progenitor cells in artificial thymic organoids.

    Science.gov (United States)

    Seet, Christopher S; He, Chongbin; Bethune, Michael T; Li, Suwen; Chick, Brent; Gschweng, Eric H; Zhu, Yuhua; Kim, Kenneth; Kohn, Donald B; Baltimore, David; Crooks, Gay M; Montel-Hagen, Amélie

    2017-05-01

    Studies of human T cell development require robust model systems that recapitulate the full span of thymopoiesis, from hematopoietic stem and progenitor cells (HSPCs) through to mature T cells. Existing in vitro models induce T cell commitment from human HSPCs; however, differentiation into mature CD3(+)TCR-αβ(+) single-positive CD8(+) or CD4(+) cells is limited. We describe here a serum-free, artificial thymic organoid (ATO) system that supports efficient and reproducible in vitro differentiation and positive selection of conventional human T cells from all sources of HSPCs. ATO-derived T cells exhibited mature naive phenotypes, a diverse T cell receptor (TCR) repertoire and TCR-dependent function. ATOs initiated with TCR-engineered HSPCs produced T cells with antigen-specific cytotoxicity and near-complete lack of endogenous TCR Vβ expression, consistent with allelic exclusion of Vβ-encoding loci. ATOs provide a robust tool for studying human T cell differentiation and for the future development of stem-cell-based engineered T cell therapies.

  18. Meta-analysis reveals conserved cell cycle transcriptional network across multiple human cell types.

    Science.gov (United States)

    Giotti, Bruno; Joshi, Anagha; Freeman, Tom C

    2017-01-05

    Cell division is central to the physiology and pathology of all eukaryotic organisms. The molecular machinery underpinning the cell cycle has been studied extensively in a number of species and core aspects of it have been found to be highly conserved. Similarly, the transcriptional changes associated with this pathway have been studied in different organisms and different cell types. In each case hundreds of genes have been reported to be regulated, however there seems to be little consensus in the genes identified across different studies. In a recent comparison of transcriptomic studies of the cell cycle in different human cell types, only 96 cell cycle genes were reported to be the same across all studies examined. Here we perform a systematic re-examination of published human cell cycle expression data by using a network-based approach to identify groups of genes with a similar expression profile and therefore function. Two clusters in particular, containing 298 transcripts, showed patterns of expression consistent with cell cycle occurrence across the four human cell types assessed. Our analysis shows that there is a far greater conservation of cell cycle-associated gene expression across human cell types than reported previously, which can be separated into two distinct transcriptional networks associated with the G 1 /S-S and G 2 -M phases of the cell cycle. This work also highlights the benefits of performing a re-analysis on combined datasets.

  19. EXPLANTATION OF MESANGIAL CELL HILLOCKS - A METHOD FOR OBTAINING HUMAN MESANGIAL CELLS IN CULTURE

    NARCIS (Netherlands)

    MULLER, EW; KIM, Y; MICHAEL, AF; VERNIER, RL; VANDERHEM, GK; VANDERWOUDE, FJ

    A simple method is presented for selective cell culture of human mesangial cells using explanatation of mesangial cell hillocks. Glomeruli which had been incubated with collagenase were explanted on plastic tissue culture flasks. Three to 6 weeks after explantation, a rapidly growing multilayer of

  20. Differential expression of cell-cycle regulators in human beta-cells derived from insulinoma tissue.

    Science.gov (United States)

    Ueberberg, Sandra; Tannapfel, Andrea; Schenker, Peter; Viebahn, Richard; Uhl, Waldemar; Schneider, Stephan; Meier, Juris J

    2016-05-01

    The low frequency of beta-cell replication in the adult human pancreas limits beta-cell regeneration. A better understanding of the regulation of human beta-cell proliferation is crucial to develop therapeutic strategies aiming to enhance beta-cell mass. To identify factors that control beta-cell proliferation, cell-cycle regulation was examined in human insulinomas as a model of increased beta-cell proliferation (n=11) and healthy pancreatic tissue from patients with benign pancreatic tumors (n=9). Tissue sections were co-stained for insulin and cell-cycle proteins. Transcript levels of selected cell-cycle factors in beta-cells were determined by qRT-PCR after performing laser-capture microdissection. The frequency of beta-cell replication was 3.74±0.92% in the insulinomas and 0.11±0.04% in controls (p=0.0016). p21 expression was higher in insulinomas (p=0.0058), and Rb expression was higher by trend (p=0.085), whereas p16 (pcell-cycle factors in beta-cells derived from insulinomas and healthy adults differs markedly. Targeting such differentially regulated cell-cycle proteins may evolve as a future strategy to enhance beta-cell regeneration. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. Reprogramming Malignant Cancer Cells toward a Benign Phenotype following Exposure to Human Embryonic Stem Cell Microenvironment

    Science.gov (United States)

    Arena, Vincenzo; Arena, Manuel; Arena, Goffredo Orazio

    2017-01-01

    The embryonic microenvironment is well known to be non-permissive for tumor development because early developmental signals naturally suppress the expression of proto-oncogenes. In an analogous manner, mimicking an early embryonic environment during embryonic stem cell culture has been shown to suppress oncogenic phenotypes of cancer cells. Exosomes derived from human embryonic stem cells harbor substances that mirror the content of the cells of origin and have been reported to reprogram hematopoietic stem/progenitor cells via horizontal transfer of mRNA and proteins. However, the possibility that these embryonic stem cells-derived exosomes might be the main effectors of the anti-tumor effect mediated by the embryonic stem cells has not been explored yet. The present study aims to investigate whether exosomes derived from human embryonic stem cells can reprogram malignant cancer cells to a benign stage and reduce their tumorigenicity. We show that the embryonic stem cell-conditioned medium contains factors that inhibit cancer cell growth and tumorigenicity in vitro and in vivo. Moreover, we demonstrate that exosomes derived from human embryonic stem cells display anti-proliferation and pro-apoptotic effects, and decrease tumor size in a xenograft model. These exosomes are also able to transfer their cargo into target cancer cells, inducing a dose-dependent increase in SOX2, OCT4 and Nanog proteins, leading to a dose-dependent decrease of cancer cell growth and tumorigenicity. This study shows for the first time that human embryonic stem cell-derived exosomes play an important role in the tumor suppressive activity displayed by human embryonic stem cells. PMID:28068409

  2. Genome editing of human pluripotent stem cells to generate human cellular disease models

    Directory of Open Access Journals (Sweden)

    Kiran Musunuru

    2013-07-01

    Full Text Available Disease modeling with human pluripotent stem cells has come into the public spotlight with the awarding of the Nobel Prize in Physiology or Medicine for 2012 to Drs John Gurdon and Shinya Yamanaka for the discovery that mature cells can be reprogrammed to become pluripotent. This discovery has opened the door for the generation of pluripotent stem cells from individuals with disease and the differentiation of these cells into somatic cell types for the study of disease pathophysiology. The emergence of genome-editing technology over the past few years has made it feasible to generate and investigate human cellular disease models with even greater speed and efficiency. Here, recent technological advances in genome editing, and its utility in human biology and disease studies, are reviewed.

  3. Dissection of a stem cell hierarchy in the human breast

    DEFF Research Database (Denmark)

    Rubner Fridriksdottir, Agla Jael

    where senescence is bypassed. These findings identify an adult human breast ductal stem cell activity and the earliest descendants thereof. Further characterization of the ductal stem cell niche, especially in terms of possible interaction with surrounding stromal cells (Sigurdsson and Fridriksdottir et......The human breast is a unique organ in that it undergoes most of its development after birth under the control of systemic hormones. Throughout the reproductive life of women, the breast gland undergoes changes in morphogenesis as evidenced by continuous cell proliferation, differentiation...... and apoptosis during each menstrual cycle. These changes are most prominent during pregnancy, lactation and involution after breast feeding. These highly dynamic changes are thought to rely on the presence of a breast epithelial stem cell population (reviewed in (Fridriksdottir et al. 2005)). Nevertheless...

  4. Biology and relevance of human acute myeloid leukemia stem cells.

    Science.gov (United States)

    Thomas, Daniel; Majeti, Ravindra

    2017-03-23

    Evidence of human acute myeloid leukemia stem cells (AML LSCs) was first reported nearly 2 decades ago through the identification of rare subpopulations of engrafting cells in xenotransplantation assays. These AML LSCs were shown to reside at the apex of a cellular hierarchy that initiates and maintains the disease, exhibiting properties of self-renewal, cell cycle quiescence, and chemoresistance. This cancer stem cell model offers an explanation for chemotherapy resistance and disease relapse and implies that approaches to treatment must eradicate LSCs for cure. More recently, a number of studies have both refined and expanded our understanding of LSCs and intrapatient heterogeneity in AML using improved xenotransplant models, genome-scale analyses, and experimental manipulation of primary patient cells. Here, we review these studies with a focus on the immunophenotype, biological properties, epigenetics, genetics, and clinical associations of human AML LSCs and discuss critical questions that need to be addressed in future research. © 2017 by The American Society of Hematology.

  5. Dipeptidyl peptidase IV in two human glioma cell lines

    Directory of Open Access Journals (Sweden)

    A Sedo

    2009-12-01

    Full Text Available There is growing evidence that dipeptidyl peptidase IV [DPP-IV, EC 3.4.14.5] takes part in the metabolism of biologically active peptides participating in the regulation of growth and transformation of glial cells. However, the knowledge on the DPP-IV expression in human glial and glioma cells is still very limited. In this study, using histochemical and biochemical techniques, the DPP-IV activity was demonstrated in two commercially available human glioma cell lines of different transformation degree, as represented by U373 astrocytoma (Grade III and U87 glioblastoma multiforme (Grade IV lines. Higher total activity of the enzyme, as well as its preferential localisation in the plasma membrane, was observed in U87 cells. Compared to U373 population, U87 cells were morphologically more pleiomorphic, they were cycling at lower rate and expressing less Glial Fibrillary Acidic Protein. The data revealed positive correlation between the degree of transformation of cells and activity of DPP-IV. Great difference in expression of this enzyme, together with the phenotypic differences of cells, makes these lines a suitable standard model for further 57 studies of function of this enzyme in human glioma cells.

  6. Immunohistochemistry of Programmed Cell Death in Archival Human Pathology Specimens

    Directory of Open Access Journals (Sweden)

    Takami Matsuyama

    2012-05-01

    Full Text Available Immunohistochemistry (IHC for detecting key signal molecules involved in programmed cell death (PCD in archival human pathology specimens is fairly well established. Detection of cleaved caspase-3 in lymphocytes in rheumatoid arthritis (RA and gastric surface foveolar glandular epithelia but not in synoviocytes in RA, gastric fundic glandular epithelia, or nasal NK/T-cell lymphoma (NKTCL cells suggests anti-apoptotic mechanisms in cell differentiation and in oncogenesis such as the induction of survivin. Enzymatically pretreated and ultra-super sensitive detection of beclin-1 in synoviocytes in RA and gastric fundic glandular epithelia suggests enhanced autophagy. The deposition of beclin-1 in fibrinoid necrosis in RA and expression of beclin-1 in detached gastric fundic glandular cells suggest that enhanced autophagy undergoes autophagic cell death (ACD. NKTCL exhibited enhanced autophagy through LC3 labeling and showed densely LC3 labeled cell-debris in regions of peculiar necrosis without deposition of beclin-1, indicating massive ACD in NKTCL and the alternative pathway enhancing autophagy following autophagic vesicle nucleation. Autophagy progression was monitored by labeling aggregated mitochondria and cathepsin D. The cell-debris in massive ACD in NKTCL were positive for 8-hydroxydeoxyguanosine, suggesting DNA oxidation occurred in ACD. Immunohistochemical autophagy and PCD analysis in archival human pathology specimens may offer new insights into autophagy in humans.

  7. Human T Cell Development, Localization, and Function throughout Life.

    Science.gov (United States)

    Kumar, Brahma V; Connors, Thomas J; Farber, Donna L

    2018-02-20

    Throughout life, T cells coordinate multiple aspects of adaptive immunity, including responses to pathogens, allergens, and tumors. In mouse models, the role of T cells is studied in the context of a specific type of pathogen, antigen, or disease condition over a limited time frame, whereas in humans, T cells control multiple insults simultaneously throughout the body and maintain immune homeostasis over decades. In this review, we discuss how humancells develop and provide essential immune protection at different life stages and highlight tissue localization and subset delineation as key determinants of the T cell functional role in immune responses. We also discuss how anatomic compartments undergo distinct age-associated changes in T cell subset composition and function over a lifetime. It is important to consider age and tissue influences on humancells when developing targeted strategies to modulate T cell-mediated immunity in vaccines and immunotherapies. Copyright © 2018 Elsevier Inc. All rights reserved.

  8. Concise review: programming human pluripotent stem cells into blood.

    Science.gov (United States)

    Easterbrook, Jennifer; Fidanza, Antonella; Forrester, Lesley M

    2016-06-01

    Blood disorders are treated with cell therapies including haematopoietic stem cell (HSC) transplantation as well as platelet and red blood cell transfusions. However the source of cells is entirely dependent on donors, procedures are susceptible to transfusion-transmitted infections and serious complications can arise in recipients due to immunological incompatibility. These problems could be alleviated if it was possible to produce haematopoietic cells in vitro from an autologous and renewable cell source. The production of haematopoietic cells in the laboratory from human induced pluripotent stem cells (iPSCs) may provide a route to realize this goal but it has proven challenging to generate long-term reconstituting HSCs. To date, the optimization of differentiation protocols has mostly relied on the manipulation of extrinsic signals to mimic the in vivo environment. We review studies that have taken an alternative approach to modulate intrinsic signals by enforced expression of transcription factors. Single and combinations of multiple transcription factors have been used in a variety of contexts to enhance the production of haematopoietic cells from human pluripotent stem cells. This programming approach, together with the recent advances in the production and use of synthetic transcription factors, holds great promise for the production of fully functional HSCs in the future. © 2016 The Authors. British Journal of Haematology published by John Wiley & Sons Ltd.

  9. Characterization of cancer stem-like cells in the side population cells of human gastric cancer cell line MKN-45.

    Science.gov (United States)

    Zhang, Hai-hong; Cai, Ai-zhen; Wei, Xue-ming; Ding, Li; Li, Feng-zhi; Zheng, Ai-ming; Dai, Da-jiang; Huang, Rong-rong; Cao, Hou-jun; Zhou, Hai-yang; Wang, Jian-mei; Wang, Xue-jing; Shi, Wei; Zhu, Heng; Yuan, Xiao-ying; Chen, Lin

    2013-03-01

    Side population (SP) cells may play a crucial role in tumorigenesis and the recurrence of cancer. Many kinds of cell lines and tissues have demonstrated the presence of SP cells, including several gastric cancer cell lines. This study is aimed to identify the cancer stem-like cells in the SP of gastric cancer cell line MKN-45. We used fluorescence activated cell sorting (FACS) to sort SP cells in the human gastric carcinoma cell line MKN-45 (cells labeled with Hoechst 33342) and then characterized the cancer stem-like properties of SP cells. This study found that the SP cells had higher clone formation efficiency than major population (MP) cells. Five stemness-related gene expression profiles, including OCT-4, SOX-2, NANOG, CD44, and adenosine triphosphate (ATP)-binding cassette transporters gene ABCG2, were tested in SP and MP cells using quantitative real-time reverse transcription polymerase chain reaction (RT-PCR). Western blot was used to show the difference of protein expression between SP and MP cells. Both results show that there was significantly higher protein expression in SP cells than in MP cells. When inoculated into non-obese diabetic/severe combined immunodeficiency (NOD/SCID) mice, SP cells show higher tumorigenesis tendency than MP cells. These results indicate that SP cells possess cancer stem cell properties and prove that SP cells from MKN-45 are gastric cancer stem-like cells.

  10. Generation of human female reproductive tract epithelium from human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Louie Ye

    Full Text Available BACKGROUND: Recent studies have identified stem/progenitor cells in human and mouse uterine epithelium, which are postulated to be responsible for tissue regeneration and proliferative disorders of human endometrium. These progenitor cells are thought to be derived from Müllerian duct (MD, the primordial female reproductive tract (FRT. METHODOLOGY/PRINCIPAL FINDINGS: We have developed a model of human reproductive tract development in which inductive neonatal mouse uterine mesenchyme (nMUM is recombined with green fluorescent protein (GFP-tagged human embryonic stem cells (hESCs; GFP-hESC (ENVY. We demonstrate for the first time that hESCs can be differentiated into cells with a human FRT epithelial cell phenotype. hESC derived FRT epithelial cells emerged from cultures containing MIXL1(+ mesendodermal precursors, paralleling events occurring during normal organogenesis. Following transplantation, nMUM treated embryoid bodies (EBs generated epithelial structures with a typical MD phenotype that expressed the MD markers PAX2, HOXA10. Functionally, the hESCs derived FRT epithelium responded to exogenous estrogen by proliferating and secreting uterine-specific glycodelin A (GdA. CONCLUSIONS/SIGNIFICANCE: These data show nMUM can induce differentiation of hESC to form the FRT epithelium. This may provide a model to study early developmental events of the human FRT.

  11. [Stimulation of cholinogenesis in the human fetal nerve cells culture].

    Science.gov (United States)

    Tsymbaliuk, V I; Vasyl'ieva, I H; Oleksenko, N P; Chopyk, N H; Tsiubko, O I; Halanta, O S

    2013-01-01

    The aim of the research was to establish cultured population of nerve cells reached by cholinergic neurons and their determinative precursors. The most effective combination of neuroinductors which stimulated cholinergic cells differentiation from the nerve stem cells was retinoic acid and acetylcholine. During the period of culturing the amount of ChAT+ cells reliably increased from 5.3 +/- 2.9% to 21.1 +/- 6.2%. At the same time in the control samples their concentration was 9.1 +/- 4.8% of total cell count. Enrichment of cell population by cholinergic neurons and their determinative precursors correlated with increasing of AChE-activity level. So, addition of retinoic acid and acetylcholine stimulate both neurogenesis and cholinogenesis in the culture of human fetal nerve cells.

  12. Electrical Guidance of Human Stem Cells in the Rat Brain

    Directory of Open Access Journals (Sweden)

    Jun-Feng Feng

    2017-07-01

    Full Text Available Limited migration of neural stem cells in adult brain is a roadblock for the use of stem cell therapies to treat brain diseases and injuries. Here, we report a strategy that mobilizes and guides migration of stem cells in the brain in vivo. We developed a safe stimulation paradigm to deliver directional currents in the brain. Tracking cells expressing GFP demonstrated electrical mobilization and guidance of migration of human neural stem cells, even against co-existing intrinsic cues in the rostral migration stream. Transplanted cells were observed at 3 weeks and 4 months after stimulation in areas guided by the stimulation currents, and with indications of differentiation. Electrical stimulation thus may provide a potential approach to facilitate brain stem cell therapies.

  13. Genetic engineering of human embryonic stem cells with lentiviral vectors.

    Science.gov (United States)

    Xiong, Chen; Tang, Dong-Qi; Xie, Chang-Qing; Zhang, Li; Xu, Ke-Feng; Thompson, Winston E; Chou, Wayne; Gibbons, Gary H; Chang, Lung-Ji; Yang, Li-Jun; Chen, Yuqing E

    2005-08-01

    Human embryonic stem (hES) cells present a valuable source of cells with a vast therapeutic potential. However, the low efficiency of directed differentiation of hES cells remains a major obstacle in their uses for regenerative medicine. While differentiation may be controlled by the genetic manipulation, effective and efficient gene transfer into hES cells has been an elusive goal. Here, we show stable and efficient genetic manipulations of hES cells using lentiviral vectors. This method resulted in the establishment of stable gene expression without loss of pluripotency in hES cells. In addition, lentiviral vectors were effective in conveying the expression of an U6 promoter-driven small interfering RNA (siRNA), which was effective in silencing its specific target. Taken together, our results suggest that lentiviral gene delivery holds great promise for hES cell research and application.

  14. [Increase in cell metabolism in normal, diploid human glial cells in stationary cell cultures induced by meclofenoxate].

    Science.gov (United States)

    Ludwig-Festl, M; Gräter, B; Bayreuther, K

    1983-01-01

    Quantitative biochemical studies were undertaken in order to examine the influence of the accumulation of lipofuscin in secondary lysosomes on cell metabolic activities of normal diploid human glia cells in a stationary cell culture system. Glia cells accumulate lipofuscin as a function of the duration of the stationary cultivation in vitro. The accumulation of lipofuscin can be decreased by the long-term treatment with the pharmacon meclofenoxate (centrophenoxine, Helfergin). Concomitant with the reduction of the accumulated lipofuscin, meclofenoxate-treated glia cells show enhanced rates of RNA synthesis, protein synthesis and glucose uptake. Most likely, in meclofenoxate-treated normal diploid human glia cells in vitro, the utilisation of glucose is shifted from glycolysis to the pentose phosphate pathway. The data suggest that the meclofenoxate-induced reduction of lipofuscin accumulation has a positive effect on cell metabolic functions and causes a delay of the cellular aging of the human glia cells in vitro.

  15. Expression of the melanoma cell adhesion molecule in human mesenchymal stromal cells regulates proliferation, differentiation, and maintenance of hematopoietic stem and progenitor cells.

    Science.gov (United States)

    Stopp, Sabine; Bornhäuser, Martin; Ugarte, Fernando; Wobus, Manja; Kuhn, Matthias; Brenner, Sebastian; Thieme, Sebastian

    2013-04-01

    The melanoma cell adhesion molecule defines mesenchymal stromal cells in the human bone marrow that regenerate bone and establish a hematopoietic microenvironment in vivo. The role of the melanoma cell adhesion molecule in primary human mesenchymal stromal cells and the maintenance of hematopoietic stem and progenitor cells during ex vivo culture has not yet been demonstrated. We applied RNA interference or ectopic overexpression of the melanoma cell adhesion molecule in human mesenchymal stromal cells to evaluate the effect of the melanoma cell adhesion molecule on their proliferation and differentiation as well as its influence on co-cultivated hematopoietic stem and progenitor cells. Knockdown and overexpression of the melanoma cell adhesion molecule affected several characteristics of human mesenchymal stromal cells related to osteogenic differentiation, proliferation, and migration. Furthermore, knockdown of the melanoma cell adhesion molecule in human mesenchymal stromal cells stimulated the proliferation of hematopoietic stem and progenitor cells, and strongly reduced the formation of long-term culture-initiating cells. In contrast, melanoma cell adhesion molecule-overexpressing human mesenchymal stromal cells provided a supportive microenvironment for hematopoietic stem and progenitor cells. Expression of the melanoma cell adhesion molecule increased the adhesion of hematopoietic stem and progenitor cells to human mesenchymal stromal cells and their migration beneath the monolayer of human mesenchymal stromal cells. Our results demonstrate that the expression of the melanoma cell adhesion molecule in human mesenchymal stromal cells determines their fate and regulates the maintenance of hematopoietic stem and progenitor cells through direct cell-cell contact.

  16. Progress towards human primordial germ cell specification in vitro.

    Science.gov (United States)

    Canovas, S; Campos, R; Aguilar, E; Cibelli, J B

    2017-01-01

    Primordial germ cells (PGCs) have long been considered the link between one generation and the next. PGC specification begins in the early embryo as a result of a highly orchestrated combination of transcriptional and epigenetic mechanisms. Understanding the molecular events that lead to proper PGC development will facilitate the development of new treatments for human infertility as well as species conservation. This article describes the latest, most relevant findings about the mechanisms of PGC formation, emphasizing human PGC. It also discusses our own laboratory's progress in using transdifferentiation protocols to derive human PGCs (hPGCs). Our preliminary results arose from our pursuit of a sequential hPGC induction strategy that starts with the repression of lineage-specific factors in the somatic cell, followed by the reactivation of germ cell-related genes using specific master regulators, which can indeed reactivate germ cell-specific genes in somatic cells. While it is still premature to assume that fully functional human gametes can be obtained in a dish, our results, together with those recently published by others, provide strong evidence that generating their precursors, PGCs, is within reach. © The Author 2016. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  17. De Novo Generated Human Red Blood Cells in Humanized Mice Support Plasmodium falciparum Infection.

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    Anburaj Amaladoss

    Full Text Available Immunodeficient mouse-human chimeras provide a powerful approach to study host specific pathogens like Plasmodium (P. falciparum that causes human malaria. Existing mouse models of P. falciparum infection require repeated injections of human red blood cells (RBCs. In addition, clodronate lipsomes and anti-neutrophil antibodies are injected to suppress the clearance of human RBCs by the residual immune system of the immunodeficient mice. Engraftment of NOD-scid Il2rg-/- mice with human hematopoietic stem cells leads to reconstitution of human immune cells. Although human B cell reconstitution is robust and T cell reconstitution is reasonable in the recipient mice, human RBC reconstitution is generally poor or undetectable. The poor reconstitution is mainly the result of a deficiency of appropriate human cytokines that are necessary for the development and maintenance of these cell lineages. Delivery of plasmid DNA encoding human erythropoietin and interleukin-3 into humanized mice by hydrodynamic tail-vein injection resulted in significantly enhanced reconstitution of erythrocytes. With this improved humanized mouse, here we show that P. falciparum infects de novo generated human RBCs, develops into schizonts and causes successive reinvasion. We also show that different parasite strains exhibit variation in their ability to infect these humanized mice. Parasites could be detected by nested PCR in the blood samples of humanized mice infected with P. falciparum K1 and HB3 strains for 3 cycles, whereas in other strains such as 3D7, DD2, 7G8, FCR3 and W2mef parasites could only be detected for 1 cycle. In vivo adaptation of K1 strain further improves the infection efficiency and parasites can be detected by microscopy for 3 cycles. The parasitemia ranges between 0.13 and 0.25% at the first cycle of infection, falls between 0.08 and 0.15% at the second cycle, and drops to barely detectable levels at the third cycle of infection. Compared to existing

  18. Derivation and characterization of hepatic progenitor cells from human embryonic stem cells.

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    Dongxin Zhao

    Full Text Available The derivation of hepatic progenitor cells from human embryonic stem (hES cells is of value both in the study of early human liver organogenesis and in the creation of an unlimited source of donor cells for hepatocyte transplantation therapy. Here, we report for the first time the generation of hepatic progenitor cells derived from hES cells. Hepatic endoderm cells were generated by activating FGF and BMP pathways and were then purified by fluorescence activated cell sorting using a newly identified surface marker, N-cadherin. After co-culture with STO feeder cells, these purified hepatic endoderm cells yielded hepatic progenitor colonies, which possessed the proliferation potential to be cultured for an extended period of more than 100 days. With extensive expansion, they co-expressed the hepatic marker AFP and the biliary lineage marker KRT7 and maintained bipotential differentiation capacity. They were able to differentiate into hepatocyte-like cells, which expressed ALB and AAT, and into cholangiocyte-like cells, which formed duct-like cyst structures, expressed KRT19 and KRT7, and acquired epithelial polarity. In conclusion, this is the first report of the generation of proliferative and bipotential hepatic progenitor cells from hES cells. These hES cell-derived hepatic progenitor cells could be effectively used as an in vitro model for studying the mechanisms of hepatic stem/progenitor cell origin, self-renewal and differentiation.

  19. Hedgehog signaling regulates telomerase reverse transcriptase in human cancer cells.

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    Tapati Mazumdar

    Full Text Available The Hedgehog (HH signaling pathway is critical for normal embryonic development, tissue patterning and cell differentiation. Aberrant HH signaling is involved in multiple human cancers. HH signaling involves a multi-protein cascade activating the GLI proteins that transcriptionally regulate HH target genes. We have previously reported that HH signaling is essential for human colon cancer cell survival and inhibition of this signal induces DNA damage and extensive cell death. Here we report that the HH/GLI axis regulates human telomerase reverse transcriptase (hTERT, which determines the replication potential of cancer cells. Suppression of GLI1/GLI2 functions by a C-terminus truncated GLI3 repressor mutant (GLI3R, or by GANT61, a pharmacological inhibitor of GLI1/GLI2, reduced hTERT protein expression in human colon cancer, prostate cancer and Glioblastoma multiforme (GBM cell lines. Expression of an N-terminus deleted constitutively active mutant of GLI2 (GLI2ΔN increased hTERT mRNA and protein expression and hTERT promoter driven luciferase activity in human colon cancer cells while GANT61 inhibited hTERT mRNA expression and hTERT promoter driven luciferase activity. Chromatin immunoprecipitation with GLI1 or GLI2 antibodies precipitated fragments of the hTERT promoter in human colon cancer cells, which was reduced upon exposure to GANT61. In contrast, expression of GLI1 or GLI2ΔN in non-malignant 293T cells failed to alter the levels of hTERT mRNA and protein, or hTERT promoter driven luciferase activity. Further, expression of GLI2ΔN increased the telomerase enzyme activity, which was reduced by GANT61 administration in human colon cancer, prostate cancer, and GBM cells. These results identify hTERT as a direct target of the HH signaling pathway, and reveal a previously unknown role of the HH/GLI axis in regulating the replication potential of cancer cells. These findings are of significance in understanding the important regulatory

  20. Cryopreservation of Human Pluripotent Stem Cells in Defined Medium

    Science.gov (United States)

    Liu, Weiwei; Chen, Guokai

    2014-01-01

    This protocol describes a cryopreservation procedure using an enzyme-free dissociation method to harvest cells and preserve cells in albumin-free chemically defined E8 medium for human pluripotent stem cells (hPSCs). The dissociation by EDTA/PBS produces small cell aggregates that allow high survival efficiency in passaging and cryopreservation. The preservation in E8 medium eliminates serum or other animal products, and is suitable for the increasing demand for high quality hPSCs in translational research. In combination with the special feature of EDTA/PBS dissociation, this protocol allows efficient cryopreservation in more time-saving manner. PMID:25366897

  1. Human Urine as a Noninvasive Source of Kidney Cells

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    Fanny Oliveira Arcolino

    2015-01-01

    Full Text Available Urine represents an unlimited source of patient-specific kidney cells that can be harvested noninvasively. Urine derived podocytes and proximal tubule cells have been used to study disease mechanisms and to screen for novel drug therapies in a variety of human kidney disorders. The urinary kidney stem/progenitor cells and extracellular vesicles, instead, might be promising for therapeutic treatments of kidney injury. The greatest advantages of urine as a source of viable cells are the easy collection and less complicated ethical issues. However, extensive characterization and in vivo studies still have to be performed before the clinical use of urine-derived kidney progenitors.

  2. Features of Human CD3+CD20+ T Cells.

    Science.gov (United States)

    Schuh, Elisabeth; Berer, Kerstin; Mulazzani, Matthias; Feil, Katharina; Meinl, Ingrid; Lahm, Harald; Krane, Markus; Lange, Rüdiger; Pfannes, Kristina; Subklewe, Marion; Gürkov, Robert; Bradl, Monika; Hohlfeld, Reinhard; Kümpfel, Tania; Meinl, Edgar; Krumbholz, Markus

    2016-08-15

    Monoclonal Abs against CD20 reduce the number of relapses in multiple sclerosis (MS); commonly this effect is solely attributed to depletion of B cells. Recently, however, a subset of CD3(+)CD20(+) T cells has been described that is also targeted by the anti-CD20 mAb rituximab. Because the existence of cells coexpressing CD3 and CD20 is controversial and features of this subpopulation are poorly understood, we studied this issue in detail. In this study, we confirm that 3-5% of circulating human T cells display CD20 on their surface and transcribe both CD3 and CD20. We report that these CD3(+)CD20(+) T cells pervade thymus, bone marrow, and secondary lymphatic organs. They are found in the cerebrospinal fluid even in the absence of inflammation; in the cerebrospinal fluid of MS patients they occur at a frequency similar to B cells. Phenotypically, these T cells are enriched in CD8(+) and CD45RO(+) memory cells and in CCR7(-) cells. Functionally, they show a higher frequency of IL-4-, IL-17-, IFN-γ-, and TNF-α-producing cells compared with T cells lacking CD20. CD20-expressing T cells respond variably to immunomodulatory treatments given to MS patients: they are reduced by fingolimod, alemtuzumab, and dimethyl fumarate, whereas natalizumab disproportionally increases them in the blood. After depletion by rituximab, they show earlier and higher repopulation than CD20(+) B cells. Taken together, human CD3(+)CD20(+) T cells pervade lymphatic organs and the cerebrospinal fluid, have a strong ability to produce different cytokines, and respond to MS disease modifying drugs. Copyright © 2016 by The American Association of Immunologists, Inc.

  3. INTERNALIZATION OF ANTIMICROBIAL PEPTIDE ACIPENSIN 1 INTO HUMAN TUMOR CELLS

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    E. S. Umnyakova

    2016-01-01

    Full Text Available Search for new compounds providing delivery of drugs into infected or neoplastic cells, is an important direction of biomedical research. Cell-penetrating peptides are among those compounds, due to their ability to translocate through membranes of eukaryotic cells, serving as potential carriers of various therapeutic agents to the target cells. The aim of present work was to investigate the ability of acipensin 1, an antimicrobial peptide of innate immune system, for in vitro penetration into human tumor cells. Acipensin 1 is a cationic peptide that we have previously isolated from leukocytes of the Russian sturgeon, Acipenser gueldenstaedtii. Capability of acipensin 1 to enter the human erytroleukemia K-562 cells has been investigated for the first time. A biotechnological procedure for producing a recombinant acipensin 1 peptide has been developed. The obtained peptide was conjugated with a fluorescent probe BODIPY FL. By means of confocal microscopy, we have shown that the tagged acipensin 1 rapidly enters into K-562 cells and can be detected in the intracellular space within 5 min after its addition to the cell culture. Using flow cytometry technique, penetration kinetics of the labeled peptide into K-562 cells (at nontoxic micromolar concentrations has been studied. We have observed a rapid internalization of the peptide to the target cells, thus confirming the results of microscopic analysis, i.e, the labeled acipensin was detectable in K-562 cells as soon as wihin 2-3 seconds after its addition to the incubation medium. The maximum of fluorescence was reached within a period of approx. 45 seconds, with further “plateau” at the terms of >100 seconds following cell stimulation with the test compound. These data support the concept, that the antimicrobial peptides of innate immunity system possess the features of cell-penetrating peptides, and allow us to consider the studied sturgeon peptide a promising template for development of new

  4. Triazole fungicide tebuconazole disrupts human placental trophoblast cell functions.

    Science.gov (United States)

    Zhou, Jinghua; Zhang, Jianyun; Li, Feixue; Liu, Jing

    2016-05-05

    Triazole fungicides are one of the top ten classes of current-use pesticides. Although exposure to triazole fungicides is associated with reproductive toxicity in mammals, limited information is available regarding the effects of triazole fungicides on human placental trophoblast function. Tebuconazole (TEB) is a common triazole fungicide that has been extensively used for fungi control. In this work, we showed that TEB could reduce cell viability, disturb normal cell cycle distribution and induce apoptosis of human placental trophoblast cell line HTR-8/SVneo (HTR-8). Bcl-2 protein expression decreased and the level of Bax protein increased after TEB treatment in HTR-8 cells. The results demonstrated that this fungicide induced apoptosis of trophoblast cells via mitochondrial pathway. Importantly, we found that the invasive and migratory capacities of HTR-8 cells decreased significantly after TEB administration. TEB altered the expression of key regulatory genes involved in the modulation of trophoblast functions. Taken together, TEB suppressed human trophoblast invasion and migration through affecting the expression of protease, hormones, angiogenic factors, growth factors and cytokines. As the invasive and migratory abilities of trophoblast are essential for successful placentation and fetus development, our findings suggest a potential risk of triazole fungicides to human pregnancy. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. Modelling IRF8 Deficient Human Hematopoiesis and Dendritic Cell Development with Engineered iPS Cells.

    Science.gov (United States)

    Sontag, Stephanie; Förster, Malrun; Qin, Jie; Wanek, Paul; Mitzka, Saskia; Schüler, Herdit M; Koschmieder, Steffen; Rose-John, Stefan; Seré, Kristin; Zenke, Martin

    2017-04-01

    Human induced pluripotent stem (iPS) cells can differentiate into cells of all three germ layers, including hematopoietic stem cells and their progeny. Interferon regulatory factor 8 (IRF8) is a transcription factor, which acts in hematopoiesis as lineage determining factor for myeloid cells, including dendritic cells (DC). Autosomal recessive or dominant IRF8 mutations occurring in patients cause severe monocytic and DC immunodeficiency. To study IRF8 in human hematopoiesis we generated human IRF8-/- iPS cells and IRF8-/- embryonic stem (ES) cells using RNA guided CRISPR/Cas9n genome editing. Upon induction of hematopoietic differentiation, we demonstrate that IRF8 is dispensable for iPS cell and ES cell differentiation into hemogenic endothelium and for endothelial-to-hematopoietic transition, and thus development of hematopoietic progenitors. We differentiated iPS cell and ES cell derived progenitors into CD141+ cross-presenting cDC1 and CD1c+ classical cDC2 and CD303+ plasmacytoid DC (pDC). We found that IRF8 deficiency compromised cDC1 and pDC development, while cDC2 development was largely unaffected. Additionally, in an unrestricted differentiation regimen, IRF8-/- iPS cells and ES cells exhibited a clear bias toward granulocytes at the expense of monocytes. IRF8-/- DC showed reduced MHC class II expression and were impaired in cytokine responses, migration, and antigen presentation. Taken together, we engineered a human IRF8 knockout model that allows studying molecular mechanisms of human immunodeficiencies in vitro, including the pathophysiology of IRF8 deficient DC. Stem Cells 2017;35:898-908. © 2017 The Authors Stem Cells published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

  6. Protective Effects of Memantine on Hydroquinone-Treated Human Retinal Pigment Epithelium Cells and Human Retinal Müller Cells.

    Science.gov (United States)

    Moustafa, Mohamed Tarek; Ramirez, Claudio; Schneider, Kevin; Atilano, Shari R; Limb, Gloria Astrid; Kuppermann, Baruch D; Kenney, Maria Cristina

    2017-10-01

    Memantine (MEM) acts on the glutamatergic system by blocking N-methyl-d-aspartate (NMDA) glutamate receptors. The role that MEM plays in protecting retinal cells is unknown. Hydroquinone (HQ) is one of the cytotoxic components in cigarette smoke. In the present study, we tested whether pretreatment with MEM could protect against the cytotoxic effects of HQ on human retinal pigment epithelium cells (ARPE-19) and human retinal Müller cells (MIO-M1) in vitro. Cells were plated, pretreated for 6 h with 30 μM of MEM, and then exposed for 24 h to 200, 100, 50, and 25 μM of HQ while MEM was still present. Cell viability (CV), reactive oxygen species (ROS), mitochondrial membrane potential (ΔΨm), and lactate dehydrogenase (LDH) release assays were performed. HQ-treated cells showed a dose-dependent decrease in CV and ΔΨm, but an increase in ROS production and LDH levels in both cell lines. MEM pretreatment reversed the CV in 50, 100, and 200 μM doses in ARPE-19 cells and at all HQ concentrations in MIO-M1 cells compared to HQ-treated cultures. ROS production was reversed in all HQ concentrations in both cell lines. ΔΨm was significantly increased after MEM pretreatment only in 50 μM HQ concentration for both cell lines. LDH levels were decreased at 50 and 25 μM HQ in ARPE-19 and MIO-M1 cells, respectively. HQ-induced toxicity is concentration dependent in ARPE-19 and MIO-M1 cultures. MEM exerts protective effects against HQ-induced toxicity on human retinal pigment epithelial and Müller cells in vitro.

  7. Human skeletal muscle-derived stem cells retain stem cell properties after expansion in myosphere culture

    Energy Technology Data Exchange (ETDEWEB)

    Wei, Yan [Department of Otolaryngology, Head and Neck Surgery Charite-Universitaetsmedizin Berlin, Berlin (Germany); Department of Otolaryngology, Head and Neck Surgery, The Third Affiliated Hospital of Sun Yat-sen University, Guang Zhou (China); Li, Yuan [Department of Otolaryngology, Head and Neck Surgery, The Third Affiliated Hospital of Sun Yat-sen University, Guang Zhou (China); Chen, Chao; Stoelzel, Katharina [Department of Otolaryngology, Head and Neck Surgery Charite-Universitaetsmedizin Berlin, Berlin (Germany); Kaufmann, Andreas M. [Clinic for Gynecology CCM/CBF, Charite-Universitaetsmedizin Berlin, Berlin (Germany); Albers, Andreas E., E-mail: andreas.albers@charite.de [Department of Otolaryngology, Head and Neck Surgery Charite-Universitaetsmedizin Berlin, Berlin (Germany)

    2011-04-15

    Human skeletal muscle contains an accessible adult stem-cell compartment in which differentiated myofibers are maintained and replaced by a self-renewing stem cell pool. Previously, studies using mouse models have established a critical role for resident stem cells in skeletal muscle, but little is known about this paradigm in human muscle. Here, we report the reproducible isolation of a population of cells from human skeletal muscle that is able to proliferate for extended periods of time as floating clusters of rounded cells, termed 'myospheres' or myosphere-derived progenitor cells (MDPCs). The phenotypic characteristics and functional properties of these cells were determined using reverse transcription-polymerase chain reaction (RT-PCR), flow cytometry and immunocytochemistry. Our results showed that these cells are clonogenic, express skeletal progenitor cell markers Pax7, ALDH1, Myod, and Desmin and the stem cell markers Nanog, Sox2, and Oct3/4 significantly elevated over controls. They could be maintained proliferatively active in vitro for more than 20 weeks and passaged at least 18 times, despite an average donor-age of 63 years. Individual clones (4.2%) derived from single cells were successfully expanded showing clonogenic potential and sustained proliferation of a subpopulation in the myospheres. Myosphere-derived cells were capable of spontaneous differentiation into myotubes in differentiation media and into other mesodermal cell lineages in induction media. We demonstrate here that direct culture and expansion of stem cells from human skeletal muscle is straightforward and reproducible with the appropriate technique. These cells may provide a viable resource of adult stem cells for future therapies of disease affecting skeletal muscle or mesenchymal lineage derived cell types.

  8. Generation of mature T cells from human hematopoietic stem/progenitor cells in artificial thymic organoids

    Science.gov (United States)

    Seet, Christopher S.; He, Chongbin; Bethune, Michael T.; Li, Suwen; Chick, Brent; Gschweng, Eric H.; Zhu, Yuhua; Kim, Kenneth; Kohn, Donald B.; Baltimore, David; Crooks, Gay M.; Montel-Hagen, Amélie

    2017-01-01

    Studies of human T cell development require robust model systems that recapitulate the full span of thymopoiesis, from hematopoietic stem and progenitor cells (HSPCs) through to mature T cells. Existing in vitro models induce T cell commitment from human HSPCs; however, differentiation into mature CD3+TCRab+ single positive (SP) CD8+ or CD4+ cells is limited. We describe here a serum-free, artificial thymic organoid (ATO) system that supports highly efficient and reproducible in vitro differentiation and positive selection of conventional human T cells from all sources of HSPCs. ATO-derived T cells exhibited mature naïve phenotypes, a diverse TCR repertoire, and TCR-dependent function. ATOs initiated with TCR-engineered HSPCs produced T cells with antigen specific cytotoxicity and near complete lack of endogenous TCR Vβ expression, consistent with allelic exclusion of Vβ loci. ATOs provide a robust tool for studying human T cell development and stem cell based approaches to engineered T cell therapies. PMID:28369043

  9. Overexpressed ubiquitin ligase Cullin7 in breast cancer promotes cell proliferation and invasion via down-regulating p53

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Hongsheng [Department of Histology and Embryology, Guangdong Medical College, Dongguan 523808, Guangdong (China); Wu, Fenping [The 7th People’s Hospital of Chengdu, Chengdu 610041, Sichuan (China); Wang, Yan [The Second School of Clinical Medicine, Guangdong Medical College, Dongguan 523808, Guangdong (China); Yan, Chong [School of Pharmacy, Guangdong Medical College, Dongguan 523808, Guangdong (China); Su, Wenmei, E-mail: wenmeisutg@126.com [Oncology of Affiliated Hospital Guangdong Medical College, Zhanjiang 524000, Guangdong (China)

    2014-08-08

    Highlights: • Cullin7 is overexpressed in human breast cancer samples. • Cullin7 stimulated proliferation and invasion of breast cancer cells. • Inhibition of p53 contributes to Cullin7-induced proliferation and invasion. - Abstract: Ubiquitin ligase Cullin7 has been identified as an oncogene in some malignant diseases such as choriocarcinoma and neuroblastoma. However, the role of Cullin7 in breast cancer carcinogenesis remains unclear. In this study, we compared Cullin7 protein levels in breast cancer tissues with normal breast tissues and identified significantly higher expression of Cullin7 protein in breast cancer specimens. By overexpressing Cullin7 in breast cancer cells HCC1937, we found that Cullin7 could promote cell growth and invasion in vitro. In contrast, the cell growth and invasion was inhibited by silencing Cullin7 in breast cancer cell BT474. Moreover, we demonstrated that Cullin7 promoted breast cancer cell proliferation and invasion via down-regulating p53 expression. Thus, our study provided evidence that Cullin7 functions as a novel oncogene in breast cancer and may be a potential therapeutic target for breast cancer management.

  10. Immortalization of human alveolar epithelial cells to investigate nanoparticle uptake.

    Science.gov (United States)

    Kemp, Sarah J; Thorley, Andrew J; Gorelik, Julia; Seckl, Michael J; O'Hare, Michael J; Arcaro, Alexandre; Korchev, Yuri; Goldstraw, Peter; Tetley, Teresa D

    2008-11-01

    Primary human alveolar type 2 (AT2) cells were immortalized by transduction with the catalytic subunit of telomerase and simian virus 40 large-tumor antigen. Characterization by immunochemical and morphologic methods demonstrated an AT1-like cell phenotype. Unlike primary AT2 cells, immortalized cells no longer expressed alkaline phosphatase, pro-surfactant protein C, and thyroid transcription factor-1, but expressed increased caveolin-1 and receptor for advanced glycation end products (RAGE). Live cell imaging using scanning ion conductance microscopy showed that the cuboidal primary AT2 cells were approximately 15 microm and enriched with surface microvilli, while the immortal AT1 cells were attenuated more than 40 microm, resembling these cells in situ. Transmission electron microscopy highlighted the attenuated morphology and showed endosomal vesicles in some immortal AT1 cells (but not primary AT2 cells) as found in situ. Particulate air pollution exacerbates cardiopulmonary disease. Interaction of ultrafine, nano-sized particles with the alveolar epithelium and/or translocation into the cardiovasculature may be a contributory factor. We hypothesized differential uptake of nanoparticles by AT1 and AT2 cells, depending on particle size and surface charge. Uptake of 50-nm and 1-microm fluorescent latex particles was investigated using confocal microscopy and scanning surface confocal microscopy of live cells. Fewer than 10% of primary AT2 cells internalized particles. In contrast, 75% immortal AT1 cells internalized negatively charged particles, while less than 55% of these cells internalized positively charged particles; charge, rather than size, mattered. The process was rapid: one-third of the total cell-associated negatively charged 50-nm particle fluorescence measured at 24 hours was internalized during the first hour. AT1 cells could be important in translocation of particles from the lung into the circulation.

  11. Application of human amniotic mesenchymal cells as an allogeneic transplantation cell source in bone regenerative therapy

    Energy Technology Data Exchange (ETDEWEB)

    Tsuno, Hiroaki [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Department of Oral and Maxillofacial Surgery, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Yoshida, Toshiko [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Nogami, Makiko [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Department of Orthopedic Surgery, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Koike, Chika; Okabe, Motonori [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Noto, Zenko [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Department of Oral and Maxillofacial Surgery, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Arai, Naoya; Noguchi, Makoto [Department of Oral and Maxillofacial Surgery, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan); Nikaido, Toshio, E-mail: tnikaido@med.u-toyama.ac.jp [Department of Regenerative Medicine, Graduate School of Medicine and Pharmaceutical Sciences for Research, University of Toyama, 2630 Sugitani Toyama, Toyama 930-0194 (Japan)

    2012-12-01

    Autogenous mesenchymal stem cells (MSCs) have therapeutic applications in bone regenerative therapy due to their pluripotency. However, the ability of MSCs to proliferate and differentiate varies between donors. Furthermore, alternative sources of MSCs are required for patients with contraindications to autogenous cell therapy. The aim of this study was to evaluate the potential of mesenchymal cells from the human amniotic membrane (HAM) as a source of cells for allogeneic transplantation in bone regenerative therapy. Cells that retained a proliferative capacity of more than 50 population doubling level were distinguished from other HAM cells as HAM{alpha} cells and induced to osteogenic status-their in vivo osteogenesis was subsequently investigated in rats. It was found that HAM{alpha} cells were spindle shaped and were positive for MSC markers and negative for hematopoietic stem cell markers. Alkaline phosphatase activity and calcium deposition increased with osteogenic status of HAM{alpha} cells. The expression of osteocalcin mRNA was increased in HAM{alpha} cells cultured on calcium phosphate scaffolds. Moreover, xenografted HAM{alpha} cells remained viable and produced extracellular matrix for several weeks. Thus, this study suggests that human amniotic mesenchymal cells possess osteogenic differentiation potential and could be applied to allogeneic transplantation in bone regenerative therapy. - Highlights: Black-Right-Pointing-Pointer Human amniotic mesenchymal cells include cells (HAM{alpha} cells) that have the properties of MSCs. Black-Right-Pointing-Pointer HAM{alpha} cells have excellent osteogenic differentiation potential. Black-Right-Pointing-Pointer Osteogenic differentiation ability of HAM{alpha} was amplified by calcium phosphate scaffolds. Black-Right-Pointing-Pointer HAM{alpha} cells can be applicable to allogeneic cell transplantation in bone regenerative therapy.

  12. Spermidine promotes human hair growth and is a novel modulator of human epithelial stem cell functions.

    Directory of Open Access Journals (Sweden)

    Yuval Ramot

    Full Text Available BACKGROUND: Rapidly regenerating tissues need sufficient polyamine synthesis. Since the hair follicle (HF is a highly proliferative mini-organ, polyamines may also be important for normal hair growth. However, the role of polyamines in human HF biology and their effect on HF epithelial stem cells in situ remains largely unknown. METHODS AND FINDINGS: We have studied the effects of the prototypic polyamine, spermidine (0.1-1 µM, on human scalp HFs and human HF epithelial stem cells in serum-free organ culture. Under these conditions, spermidine promoted hair shaft elongation and prolonged hair growth (anagen. Spermidine also upregulated expression of the epithelial stem cell-associated keratins K15 and K19, and dose-dependently modulated K15 promoter activity in situ and the colony forming efficiency, proliferation and K15 expression of isolated human K15-GFP+ cells in vitro. Inhibiting the rate-limiting enzyme of polyamine synthesis, ornithine decarboyxlase (ODC, downregulated intrafollicular K15 expression. In primary human epidermal keratinocytes, spermidine slightly promoted entry into the S/G2-M phases of the cell cycle. By microarray analysis of human HF mRNA extracts, spermidine upregulated several key target genes implicated e.g. in the control of cell adherence and migration (POP3, or endoplasmic reticulum and mitochondrial functions (SYVN1, NACA and SLC25A3. Excess spermidine may restrict further intrafollicular polyamine synthesis by inhibiting ODC gene and protein expression in the HF's companion layer in situ. CONCLUSIONS: These physiologically and clinically relevant data provide the first direct evidence that spermidine is a potent stimulator of human hair growth and a previously unknown modulator of human epithelial stem cell biology.

  13. Human immunodeficiencies related to APC/T cell interaction

    Directory of Open Access Journals (Sweden)

    Marinos eKallikourdis

    2015-08-01

    Full Text Available The primary event for initiating adaptive immune responses is the encounter between T lymphocytes and antigen presenting cells (APC in the T cell area of secondary lymphoid organs and the formation of highly organized inter-cellular junctions referred to as the immune synapses. In vivo live-cell imaging of APC-T cell interactions combined to functional studies unveiled that T cell fate is dictated, in large part, by the stability of the initial contact. Immune cell interaction is equally important during delivery of T cell help to B cells and for the killing of target cells by cytotoxic T cells and NK cells. The critical role of contact dynamics and synapse stability on the immune response is well illustrated by human immune deficiencies in which disease pathogenesis is linked to altered adhesion or defective cross-talk between the synaptic partners. Here we will discuss in details the mechanisms of defective APC-T cell communications in Wiskott-Aldrich syndrome (WAS and in warts, hypogammaglobulinemia, infections, myelokathexis syndrome (WHIM. In addition, we will summarize the evidences pointing to a compromised conjugate formation in WIP deficiency, DOCK8 deficiency and X-linked lymphoproliferative syndrome.

  14. Mast cells in human and experimental cardiometabolic diseases.

    Science.gov (United States)

    Shi, Guo-Ping; Bot, Ilze; Kovanen, Petri T

    2015-11-01

    Mast cells, like many other types of inflammatory cell, perform pleiotropic roles in cardiometabolic diseases such as atherosclerosis, abdominal aortic aneurysms, obesity, and diabetes mellitus, as well as complications associated with these diseases. Low numbers of mast cells are present in the heart, aorta, and adipose tissue of healthy humans, but patients with cardiometabolic diseases and animals with experimentally-induced cardiometabolic pathologies have high numbers of mast cells with increased activity in the affected tissues. Mediators released by the activated mast cells, such as chemokines, cytokines, growth factors, heparin, histamine, and proteases, not only function as biomarkers of cardiometabolic diseases, but might also directly contribute to the pathogenesis of such diseases. Mast-cell mediators impede the functions of vascular cells, the integrity of the extracellular matrix, and the activity of other inflammatory cells, thereby contributing to the pathobiology of the conditions at multiple levels. In mouse models, mast-cell activation aggravates the progression of various cardiometabolic pathologies, whereas a genetic deficiency or pharmacological stabilization of mast cells, or depletion or inhibition of specific mast-cell mediators, tends to delay the progression of such conditions. Pharmacological inhibition of mast-cell activation or their targeted effector functions offers potential novel therapeutic strategies for patients with cardiometabolic disorders.

  15. Phosphonium Salt Displays Cytotoxic Effects Against Human Cancer Cell Lines.

    Science.gov (United States)

    Dhanya, Dhanyalayam; Palma, Giuseppe; Cappello, AnnaRita; Mariconda, Annaluisa; Sinicropi, Maria Stefania; Giordano, Francesca; Vecchio, Vitale Del; Ramunno, Anna; Arra, Claudio; Longo, Pasquale; Saturnino, Carmela

    2017-07-19

    Aims/ Objective: Phosphonium salts are compounds whose structural characteristics enable them to cross the plasma and mitochondrial membrane with ease. Cancer cells have higher plasma membrane potentials than normal cells, phosphonium salts selectively accumulate in the mitochondria of neoplastic cells and inhibit mitochondrial function. In the presente work, we investigate the cytotoxic activity of lipophilic phosphonium salt (11-methoxy11-oxo-undecyl) triphenylphosphonium bromide (MUTP) as well as of two new phosphine oxide salts, 3,3'-(methylphosphoryl) dibenzenaminium chloride (SBAMPO) and 3,3' (phenylphosphoryl) dibenzenaminium chloride (SBAPPO) on the proliferation of breast cancer cell line (MCF-7) and human uterin cervix adenocarcinoma cells (HeLa). We show that only MUTP exhibits antiproliferative effects on both cell lines, without affecting normal breast epithelial cell proliferation. More specifically, we demonstrate that MUTP treatment of breast cancer cells is associated with impaired cell-cycle progression and metabolically induces mitochondrial damage and triggers apoptotic cell death in MCF-7 and HeLa cells. Taken together, these findings suggest that MUTP may be capable of selectively targeting neoplastic cell growth and therefore has potential applications as anticancer agent. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  16. [Effects of emdogain on human periodontal ligament cells in vitro].

    Science.gov (United States)

    Zhang, Feng-qiu; Meng, Huan-xin; Han, Jie; Liu, Kai-ning

    2012-02-18

    To investigate the effects of emdogain, enamel matrix derivative (EMD), on the proliferation, cell cycle, mineralization and ultrastructure of human periodontal ligament (PDL) cells in vitro. The influence of cell growth on PDL cells was evaluated by Cell Counting Kit-8 (CCK-8) in the presence and absence of emdogain, after 1, 3, and 5 d of culture. DNA synthesis and ultrastructure of PDL cells were observed by flow cytometry(FCM) and transmission electron microscopy (TEM) in the presence and absence of emdogain after 3 d of culture. The increasing of osteogenic capacity was verified by the expression changes of osteogenic differentiation markers of bone sialoprotein (BSP) and osteopontin (OPN) in emdogain-treated PDL cells by immunohistochemicl staining. Incubation of PDL cells with emdogain after 3 d significantly stimulated cell growth and DNA synthesis. Emdogain enhanced the osteogenic potential of PDL cells by high expression of osteogenic differentiation markers of BSP and OPN. The data indicate that Emdogain enhances cell proliferation and promotes differentiation of PDL cells, which contributes to periodontal tissue regeneration .

  17. DNA damage responses in human induced pluripotent stem cells and embryonic stem cells.

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    Olga Momcilovic

    2010-10-01

    Full Text Available Induced pluripotent stem (iPS cells have the capability to undergo self-renewal and differentiation into all somatic cell types. Since they can be produced through somatic cell reprogramming, which uses a defined set of transcription factors, iPS cells represent important sources of patient-specific cells for clinical applications. However, before these cells can be used in therapeutic designs, it is essential to understand their genetic stability.Here, we describe DNA damage responses in human iPS cells. We observe hypersensitivity to DNA damaging agents resulting in rapid induction of apoptosis after γ-irradiation. Expression of pluripotency factors does not appear to be diminished after irradiation in iPS cells. Following irradiation, iPS cells activate checkpoint signaling, evidenced by phosphorylation of ATM, NBS1, CHEK2, and TP53, localization of ATM to the double strand breaks (DSB, and localization of TP53 to the nucleus of NANOG-positive cells. We demonstrate that iPS cells temporary arrest cell cycle progression in the G(2 phase of the cell cycle, displaying a lack of the G(1/S cell cycle arrest similar to human embryonic stem (ES cells. Furthermore, both cell types remove DSB within six hours of γ-irradiation, form RAD51 foci and exhibit sister chromatid exchanges suggesting homologous recombination repair. Finally, we report elevated expression of genes involved in DNA damage signaling, checkpoint function, and repair of various types of DNA lesions in ES and iPS cells relative to their differentiated counterparts.High degrees of similarity in DNA damage responses between ES and iPS cells were found. Even though reprogramming did not alter checkpoint signaling following DNA damage, dramatic changes in cell cycle structure, including a high percentage of cells in the S phase, increased radiosensitivity and loss of DNA damage-induced G(1/S cell cycle arrest, were observed in stem cells generated by induced pluripotency.

  18. Telomere Transcripts Target Telomerase in Human Cancer Cells

    Directory of Open Access Journals (Sweden)

    Theresa Kreilmeier

    2016-08-01

    Full Text Available Long non-coding transcripts from telomeres, called telomeric repeat-containing RNA (TERRA, were identified as blocking telomerase activity (TA, a telomere maintenance mechanism (TMM, in tumors. We expressed recombinant TERRA transcripts in tumor cell lines with TA and with alternative lengthening of telomeres (ALT to study effects on TMM and cell growth. Adeno- and lentivirus constructs (AV and LV were established for transient and stable expression of approximately 130 units of telomere hexanucleotide repeats under control of cytomegalovirus (CMV and human RNase P RNA H1 (hH1 promoters with and without polyadenylation, respectively. Six human tumor cell lines either using telomerase or ALT were infected and analyzed for TA levels. Pre-infection cells using telomerase had 1%–3% of the TERRA expression levels of ALT cells. AV and LV expression of recombinant TERRA in telomerase positive cells showed a 1.3–2.6 fold increase in TERRA levels, and a decrease in TA of 25%–58%. Dominant-negative or small hairpin RNA (shRNA viral expression against human telomerase reverse transcriptase (hTERT results in senescence, not induced by TERRA expression. Population doubling time, cell viability and TL (telomere length were not impacted by ectopic TERRA expression. Clonal growth was reduced by TERRA expression in TA but not ALT cell lines. ALT cells were not affected by treatments applied. Established cell models and tools may be used to better understand the role of TERRA in the cell, especially for targeting telomerase.

  19. Integrating human stem cell expansion and neuronal differentiation in bioreactors

    Science.gov (United States)

    Serra, Margarida; Brito, Catarina; Costa, Eunice M; Sousa, Marcos FQ; Alves, Paula M

    2009-01-01

    Background Human stem cells are cellular resources with outstanding potential for cell therapy. However, for the fulfillment of this application, major challenges remain to be met. Of paramount importance is the development of robust systems for in vitro stem cell expansion and differentiation. In this work, we successfully developed an efficient scalable bioprocess for the fast production of human neurons. Results The expansion of undifferentiated human embryonal carcinoma stem cells (NTera2/cl.D1 cell line) as 3D-aggregates was firstly optimized in spinner vessel. The media exchange operation mode with an inoculum concentration of 4 × 105 cell/mL was the most efficient strategy tested, with a 4.6-fold increase in cell concentration achieved in 5 days. These results were validated in a bioreactor where similar profile and metabolic performance were obtained. Furthermore, characterization of the expanded population by immunofluorescence microscopy and flow cytometry showed that NT2 cells maintained their stem cell characteristics along the bioreactor culture time. Finally, the neuronal differentiation step was integrated in the bioreactor process, by addition of retinoic acid when cells were in the middle of the exponential phase. Neurosphere composition was monitored and neuronal differentiation efficiency evaluated along the culture time. The results show that, for bioreactor cultures, we were able to increase significantly the neuronal differentiation efficiency by 10-fold while reducing drastically, by 30%, the time required for the differentiation process. Conclusion The culture systems developed herein are robust and represent one-step-forward towards the development of integrated bioprocesses, bridging stem cell expansion and differentiation in fully controlled bioreactors. PMID:19772662

  20. Increasing cell culture population doublings for long-term growth of finite life span human cell cultures

    Energy Technology Data Exchange (ETDEWEB)

    Stampfer, Martha R.; Garbe, James C.

    2016-06-28

    Cell culture media formulations for culturing human epithelial cells are herein described. Also described are methods of increasing population doublings in a cell culture of finite life span human epithelial cells and prolonging the life span of human cell cultures. Using the cell culture media disclosed alone and in combination with addition to the cell culture of a compound associated with anti-stress activity achieves extended growth of pre-stasis cells and increased population doublings and life span in human epithelial cell cultures.

  1. Increasing cell culture population doublings for long-term growth of finite life span human cell cultures

    Science.gov (United States)

    Stampfer, Martha R; Garbe, James C

    2015-02-24

    Cell culture media formulations for culturing human epithelial cells are herein described. Also described are methods of increasing population doublings in a cell culture of finite life span human epithelial cells and prolonging the life span of human cell cultures. Using the cell culture media disclosed alone and in combination with addition to the cell culture of a compound associated with anti-stress activity achieves extended growth of pre-stasis cells and increased population doublings and life span in human epithelial cell cultures.

  2. Side population cells isolated from KATO III human gastric cancer cell line have cancer stem cell-like characteristics.

    Science.gov (United States)

    She, Jun-Jun; Zhang, Peng-Ge; Wang, Xuan; Che, Xiang-Ming; Wang, Zi-Ming

    2012-09-07

    To investigate whether the side population (SP) cells possess cancer stem cell-like characteristics in vitro and the role of SP cells in tumorigenic process in gastric cancer. We analyzed the presence of SP cells in different human gastric carcinoma cell lines, and then isolated and identified the SP cells from the KATO III human gastric cancer cell line by flow cytometry. The clonogenic ability and self-renewal were evaluated by clone and sphere formation assays. The related genes were determined by reverse transcription polymerase chain reaction. To compare tumorigenic ability, SP and non-side population (NSP) cells from the KATO III human gastric cancer cell line were subcutaneously injected into nude mice. SP cells from the total population accounted for 0.57% in KATO III, 1.04% in Hs-746T, and 0.02% in AGS (CRL-1739). SP cells could grow clonally and have self-renewal capability in conditioned media. The expression of ABCG2, MDRI, Bmi-1 and Oct-4 was different between SP and NSP cells. However, there was no apparent difference between SP and NSP cells when they were injected into nude mice. SP cells have some cancer stem cell-like characteristics in vitro and can be used for studying the tumorigenic process in gastric cancer.

  3. The pancreatic beta cells in human type 2 diabetes.

    Science.gov (United States)

    Marchetti, Piero; Bugliani, Marco; Boggi, Ugo; Masini, Matilde; Marselli, Lorella

    2012-01-01

    Bell-cell (beta-cell) impairment is central to the development and progression of human diabetes, as a result of the combined effects of genetic and acquired factors. Reduced islet number and/or reduced beta cells amount in the pancreas of individuals with Type 2 diabetes have been consistently reported. This is mainly due to increased beta cell death, not adequately compensated for by regeneration. In addition, several quantitative and/or qualitative defects of insulin secretion have been observed in Type 2 diabetes, both in vivo and ex vivo with isolated islets. All this is associated with modifications of islet cell gene and protein expression. With the identification of several susceptible Type 2 diabetes loci, the role of genotype in affecting beta-cell function and survival has been addressed in a few studies and the relationships between genotype and beta-cell phenotype investigated. Among acquired factors, the importance of metabolic insults (in particular glucotoxicity and lipotoxicity) in the natural history of beta-cell damage has been widely underlined. Continuous improvements in our knowledge of the beta cells in human Type 2 diabetes will lead to more targeted and effective strategies for the prevention and treatment of the disease.

  4. Characterization of pancreatic ductal cells in human islet preparations.

    Science.gov (United States)

    Ichii, Hirohito; Miki, Atsushi; Yamamoto, Toshiyuki; Molano, Ruth D; Barker, Scott; Mita, Atsuyoshi; Rodriguez-Diaz, Rayner; Klein, Dagmar; Pastori, Ricardo; Alejandro, Rodolfo; Inverardi, Luca; Pileggi, Antonello; Ricordi, Camillo

    2008-11-01

    Substantial amounts of nonendocrine cells are implanted as part of human islet grafts, and a possible influence of nonendocrine cells on clinical islet transplantation outcome has been postulated. There are currently no product release criteria specific for nonendocrine cells due to lack of available methods. The aims of this study were to develop a method for the evaluation of pancreatic ductal cells (PDCs) for clinical islet transplantation and to characterize them regarding phenotype, viability, and function. We assessed 161 human islet preparations using laser scanning cytometry (LSC/iCys) for phenotypic analysis of nonendocrine cells and flow cytometry (FACS) for PDC viability. PDC and beta-cells obtained from different density fractions during the islet cell purification were compared in terms of viability. Furthermore, we examined PDC ability to produce proinf