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Sample records for fetal human pancreatic

  1. Isolation, Culture, and Imaging of Human Fetal Pancreatic Cell Clusters

    Science.gov (United States)

    Lopez, Ana D.; Kayali, Ayse G.; Hayek, Alberto; King, Charles C.

    2014-01-01

    For almost 30 years, scientists have demonstrated that human fetal ICCs transplanted under the kidney capsule of nude mice matured into functioning endocrine cells, as evidenced by a significant increase in circulating human C-peptide following glucose stimulation1-9. However in vitro, genesis of insulin producing cells from human fetal ICCs is low10; results reminiscent of recent experiments performed with human embryonic stem cells (hESC), a renewable source of cells that hold great promise as a potential therapeutic treatment for type 1 diabetes. Like ICCs, transplantation of partially differentiated hESC generate glucose responsive, insulin producing cells, but in vitro genesis of insulin producing cells from hESC is much less robust11-17. A complete understanding of the factors that influence the growth and differentiation of endocrine precursor cells will likely require data generated from both ICCs and hESC. While a number of protocols exist to generate insulin producing cells from hESC in vitro11-22, far fewer exist for ICCs10,23,24. Part of that discrepancy likely comes from the difficulty of working with human fetal pancreas. Towards that end, we have continued to build upon existing methods to isolate fetal islets from human pancreases with gestational ages ranging from 12 to 23 weeks, grow the cells as a monolayer or in suspension, and image for cell proliferation, pancreatic markers and human hormones including glucagon and C-peptide. ICCs generated by the protocol described below result in C-peptide release after transplantation under the kidney capsule of nude mice that are similar to C-peptide levels obtained by transplantation of fresh tissue6. Although the examples presented here focus upon the pancreatic endoderm proliferation and β cell genesis, the protocol can be employed to study other aspects of pancreatic development, including exocrine, ductal, and other hormone producing cells. PMID:24895054

  2. Isolation, culture, and imaging of human fetal pancreatic cell clusters.

    Science.gov (United States)

    Lopez, Ana D; Kayali, Ayse G; Hayek, Alberto; King, Charles C

    2014-05-18

    For almost 30 years, scientists have demonstrated that human fetal ICCs transplanted under the kidney capsule of nude mice matured into functioning endocrine cells, as evidenced by a significant increase in circulating human C-peptide following glucose stimulation(1-9). However in vitro, genesis of insulin producing cells from human fetal ICCs is low(10); results reminiscent of recent experiments performed with human embryonic stem cells (hESC), a renewable source of cells that hold great promise as a potential therapeutic treatment for type 1 diabetes. Like ICCs, transplantation of partially differentiated hESC generate glucose responsive, insulin producing cells, but in vitro genesis of insulin producing cells from hESC is much less robust(11-17). A complete understanding of the factors that influence the growth and differentiation of endocrine precursor cells will likely require data generated from both ICCs and hESC. While a number of protocols exist to generate insulin producing cells from hESC in vitro(11-22), far fewer exist for ICCs(10,23,24). Part of that discrepancy likely comes from the difficulty of working with human fetal pancreas. Towards that end, we have continued to build upon existing methods to isolate fetal islets from human pancreases with gestational ages ranging from 12 to 23 weeks, grow the cells as a monolayer or in suspension, and image for cell proliferation, pancreatic markers and human hormones including glucagon and C-peptide. ICCs generated by the protocol described below result in C-peptide release after transplantation under the kidney capsule of nude mice that are similar to C-peptide levels obtained by transplantation of fresh tissue(6). Although the examples presented here focus upon the pancreatic endoderm proliferation and β cell genesis, the protocol can be employed to study other aspects of pancreatic development, including exocrine, ductal, and other hormone producing cells.

  3. In vitro cultivation of human fetal pancreatic ductal stem cells and their differentiation into insulin-producing cells

    Institute of Scientific and Technical Information of China (English)

    Zhong-Xiang Yao; Mao-Lin Qin; Jian-Jun Liu; Xing-Shu Chen; De-Shan Zhou

    2004-01-01

    AIM: To isolate, culture and identify the human fetal pancreatic ductal stem cells in vitro, and to observe the potency of these multipotential cells differentiation into insulin-producing cells.METHODS: The human fetal pancreas was digested by 1 g/L collagease type Ⅳ and then 2.5 g/L trypsin was used to isolate the pancreatic ducta stem cells, followed by culture in serum-free, glucose-free DMEM media with some additional chemical substrates in vitro (according to the different Stage). The cells were induced by glucose-free (control),5 mmol/L, 17.8 mmol/L and 25 mmol/L glucose, respectively.The cell types of differentiated cells were identified using immunocytochemical staining.RESULTS: The shape of human fetal pancreatic ductal stem cells culturedin vitro was firstly fusiform in the first 2 wk,and became monolayer and cobblestone pattern after another 3 to 4 wk. After induced and differentiated by the glucose of different concentrations for another 1 to 2 wk,the cells formed the pancreatic islet-like structures. The identification and potency of these cells were then identified by using the pancreatic ductal stem cell marker, cytokeratin-19 (CK-19), pancreatic β cell marker, insulin and pancreatic α cell marker, glucagons with immunocytochemical staining.At the end of the second week, 95.2% of the cells were positive for CK-19 immunoreactivity. Up to 22.7% of the cells induced by glucose were positive for insulin immunoreactivity, and less than 3.8% of the cells were positive for glucagon immunoreactivity in pancreatic isletlike structures. The positive ratio of immunoreactive staining was dependent on the concentration of glucose, and it was observed that the 17.8 mmol/L glucose stimulated effectively to produce insulin- and glucagons-producing cells.CONCLUSION: The human fetal pancreatic ductal stem cells are capable of proliferation in vitro. These cells have multidifferentiation potential and can be induced by glucose and differentiated into insulin

  4. PDZ-domain containing-2 (PDZD2) drives the maturity of human fetal pancreatic progenitor-derived islet-like cell clusters with functional responsiveness against membrane depolarization.

    Science.gov (United States)

    Leung, Kwan Keung; Suen, Po Man; Lau, Tse Kin; Ko, Wing Hung; Yao, Kwok Ming; Leung, Po Sing

    2009-09-01

    We recently reported the isolation and characterization of a population of pancreatic progenitor cells (PPCs) from early trimester human fetal pancreata. The PPCs, being the forerunners of adult pancreatic cell lineages, were amenable to growth and differentiation into insulin-secreting islet-like cell clusters (ICCs) upon stimulation by adequate morphogens. Of note, a novel morphogenic factor, PDZ-domain containing-2 (PDZD2) and its secreted form (sPDZD2) were ubiquitously expressed in the PPCs. Our goals for this study were to evaluate the potential role of sPDZD2 in stimulating PPC differentiation and to establish the optimal concentration for such stimulation. We found that 10(-9)M sPDZD2 promoted PPC differentiation, as evidenced by the upregulation of the pancreatic endocrine markers (PDX-1, NGN3, NEURO-D, ISL-1, NKX 2.2, NKX 6.1) and INSULIN mRNA. Inhibited endogenous production of sPDZD2 suppressed expression of these factors. Secreted PDZD2 treatment significantly elevated the C-peptide content of the ICCs and increased the basal rate of insulin secretion. However, they remained unresponsive to glucose stimulation, reflected by a minimal increase in GLUT-2 and GLUCOKINASE mRNA expression. Interestingly, sPDZD2 treatment induced increased expression of the L-type voltage-gated calcium channel (Ca(v)1.2) in the ICCs, triggering calcium ion influx under KCl stimulation and conferring an ability to secrete insulin in response to KCl. Pancreatic progenitor cells from 10- and 13-week fetal pancreata showed peak expression of endogenous sPDZD2, implying that sPDZD2 has a specific role in islet development during the first trimester. In conclusion, our data suggest that sPDZD2 promotes functional maturation of human fetal PPC-derived ICCs, thus enhancing its transplanting potentials.

  5. The SDF-1α/CXCR4 axis is required for proliferation and maturation of human fetal pancreatic endocrine progenitor cells.

    Directory of Open Access Journals (Sweden)

    Ayse G Kayali

    Full Text Available The chemokine receptor CXCR4 and ligand SDF-1α are expressed in fetal and adult mouse islets. Neutralization of CXCR4 has previously been shown to diminish ductal cell proliferation and increase apoptosis in the IFNγ transgenic mouse model in which the adult mouse pancreas displays islet regeneration. Here, we demonstrate that CXCR4 and SDF-1α are expressed in the human fetal pancreas and that during early gestation, CXCR4 colocalizes with neurogenin 3 (ngn3, a key transcription factor for endocrine specification in the pancreas. Treatment of islet like clusters (ICCs derived from human fetal pancreas with SDF-1α resulted in increased proliferation of epithelial cells in ICCs without a concomitant increase in total insulin expression. Exposure of ICCs in vitro to AMD3100, a pharmacological inhibitor of CXCR4, did not alter expression of endocrine hormones insulin and glucagon, or the pancreatic endocrine transcription factors PDX1, Nkx6.1, Ngn3 and PAX4. However, a strong inhibition of β cell genesis was observed when in vitro AMD3100 treatment of ICCs was followed by two weeks of in vivo treatment with AMD3100 after ICC transplantation into mice. Analysis of the grafts for human C-peptide found that inhibition of CXCR4 activity profoundly inhibits islet development. Subsequently, a model pancreatic epithelial cell system (CFPAC-1 was employed to study the signals that regulate proliferation and apoptosis by the SDF-1α/CXCR4 axis. From a selected panel of inhibitors tested, both the PI 3-kinase and MAPK pathways were identified as critical regulators of CFPAC-1 proliferation. SDF-1α stimulated Akt phosphorylation, but failed to increase phosphorylation of Erk above the high basal levels observed. Taken together, these results indicate that SDF-1α/CXCR4 axis plays a critical regulatory role in the genesis of human islets.

  6. Vitamin D and vitamin A receptor expression and the proliferative effects of ligand activation of these receptors on the development of pancreatic progenitor cells derived from human fetal pancreas.

    Science.gov (United States)

    Ng, Ka Yan; Ma, Man Ting; Leung, Kwan Keung; Leung, Po Sing

    2011-03-01

    The growth and development of pancreatic islet cells are regulated by various morphogens. Vitamin A modulates in vitro differentiation of islet cells and vitamin D affects beta-cell insulin secretion, while both vitamin ligands act through heterodimerization with the retinoid X receptor (RXR). However, their effects in modulating pancreatic development have not been determined. In this study, cultured human pancreatic progenitor cells (PPCs) isolated from human fetal pancreas were stimulated to differentiate into islet-like cell clusters (ICCs). RT-PCR, Western blotting and immunocytochemistry were used to examine the expression and localization of vitamin D receptor (VDR), retinoic acid receptor (RAR), and RXR in PPCs. The effects of added all-trans retinoic acid (atRA, a form of vitamin A), calcitriol (activated vitamin D) and of these ligands together on PPC cell viability, proliferation and apoptosis were assessed by MTT, BrdU and ELISA assays, respectively. Post-treatment neurogenin-3 (NGN3) expression, necessary for islet-cell lineage development, was examined by real-time RT-PCR. Results showed that RAR, RXR and VDR were expressed in PPCs. RAR and RXR were localized in nuclei, and the VDR in nuclei, cytoplasm and plasma membrane. atRA and calcitriol each increased PPC viability and proliferation; atRA additionally decreased PPC apoptosis. Co-addition of atRA and calcitriol had no additive effects on cell viability but did increase ngn3 responses. In conclusion, RAR, RXR and VDR are expressed in human fetal PPCs and PPC proliferation can be promoted by calcitriol, atRA or both together, data valuable for elucidating mechanisms underlying islet development and for developing clinical islet transplantation.

  7. Human fetal mesenchymal stem cells.

    Science.gov (United States)

    O'Donoghue, Keelin; Chan, Jerry

    2006-09-01

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

  8. Fetal endocannabinoids orchestrate the organization of pancreatic islet microarchitecture.

    Science.gov (United States)

    Malenczyk, Katarzyna; Keimpema, Erik; Piscitelli, Fabiana; Calvigioni, Daniela; Björklund, Peyman; Mackie, Kenneth; Di Marzo, Vincenzo; Hökfelt, Tomas G M; Dobrzyn, Agnieszka; Harkany, Tibor

    2015-11-10

    Endocannabinoids are implicated in the control of glucose utilization and energy homeostasis by orchestrating pancreatic hormone release. Moreover, in some cell niches, endocannabinoids regulate cell proliferation, fate determination, and migration. Nevertheless, endocannabinoid contributions to the development of the endocrine pancreas remain unknown. Here, we show that α cells produce the endocannabinoid 2-arachidonoylglycerol (2-AG) in mouse fetuses and human pancreatic islets, which primes the recruitment of β cells by CB1 cannabinoid receptor (CB1R) engagement. Using subtractive pharmacology, we extend these findings to anandamide, a promiscuous endocannabinoid/endovanilloid ligand, which impacts both the determination of islet size by cell proliferation and α/β cell sorting by differential activation of transient receptor potential cation channel subfamily V member 1 (TRPV1) and CB1Rs. Accordingly, genetic disruption of TRPV1 channels increases islet size whereas CB1R knockout augments cellular heterogeneity and favors insulin over glucagon release. Dietary enrichment in ω-3 fatty acids during pregnancy and lactation in mice, which permanently reduces endocannabinoid levels in the offspring, phenocopies CB1R(-/-) islet microstructure and improves coordinated hormone secretion. Overall, our data mechanistically link endocannabinoids to cell proliferation and sorting during pancreatic islet formation, as well as to life-long programming of hormonal determinants of glucose homeostasis.

  9. Differentiation of fetal pancreatic stem cells into neuron-like and islet-like cells in vitro

    Institute of Scientific and Technical Information of China (English)

    Xiufeng Hua; Yanwei Wang; Peiwen Lian; Shouxin Zhang; Jianyuan Li; Haiyan Wang; Shulin Chen; Wei Gao

    2012-01-01

    Pancreatic stem cells were isolated and cultured from aborted human fetal pancreases of gestational age 14-20 weeks.They were seeded at a density of 1 × 104 in serum-free media for differentiation into neuron-like cells, expressing β-tubulin III and glial fibrillary acidic protein.These neuron-like cells displayed a synapse-like morphology and appeared to form a neuronal network.Pancreatic stem cells were also seeded at a density of 1 × 105 for differentiation into islet-like cells, expressing insulin and glucagon, with an islet-like morphology.These cells had glucose-stimulated secretion of human insulin and C-peptide.Results suggest that pancreatic stem cells can be differentiated into neuron-like and islet-like cells.

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

    Science.gov (United States)

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

    2014-02-25

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

  11. Ex vivo culture of human fetal gonads

    DEFF Research Database (Denmark)

    Jørgensen, A; Nielsen, J.E.; Perlman, S

    2015-01-01

    STUDY QUESTION: What are the effects of experimentally manipulating meiosis signalling by addition of retinoic acid (RA) in cultured human fetal gonads? SUMMARY ANSWER: RA-treatment accelerated meiotic entry in cultured fetal ovary samples, while addition of RA resulted in a dysgenetic gonadal...... phenotype in fetal testis cultures. WHAT IS KNOWN ALREADY: One of the first manifestations of sex differentiation is the initiation of meiosis in fetal ovaries. In contrast, meiotic entry is actively prevented in the fetal testis at this developmental time-point. It has previously been shown that RA......-treatment mediates initiation of meiosis in human fetal ovary ex vivo. STUDY DESIGN, SIZE, DURATION: This was a controlled ex vivo study of human fetal gonads treated with RA in 'hanging-drop' tissue cultures. The applied experimental set-up preserves germ cell-somatic niche interactions and the investigated...

  12. Rapid Evolution from the First Episode of Acute Pancreatitis to Chronic Pancreatitis in Human Subjects

    OpenAIRE

    Elie Aoun; Adam Slivka; Papachristou, Dionysios J.; Whitcomb, David C.; Gleeson, Ferga C; Papachristou, Georgios I

    2007-01-01

    Context Growing evidence suggests that recurrent acute pancreatitis leads to chronic pancreatitis, but this sequence is seldom reported in human subjects. The sentinel acute pancreatitis event hypothesis suggests that an initial episode of acute pancreatitis is the first step in a complicated series of events ultimately leading to chronic pancreatitis. Objective To identify patients who evolved from recurrent acute pancreatitis to chronic pancreatitis. Setting The Severity of Acute Pancreatit...

  13. Generation of pancreatic islet cells from human embryonic stem cells

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Efficiently obtaining functional pancreatic islet cells derived from human embryonic stem(hES) cells not only provides great potential to solve the shortage of islets sources for type I diabetes cell therapy,but also benefits the study of the development of the human pancreas and diabetes pathology.In 2001,hES cells were reported to have the capacity to generate insulin-producing cells by spontaneous differentiation in vitro.Since then,many strategies(such as overexpression of key transcription factors,delivery of key proteins for pancreatic development,co-transplantation of differentiated hES cells along with fetal pancreas,stepwise differentiation by mimicking in vivo pancreatic development) have been employed in order to induce the differentiation of pancreatic islet cells from hES cells.Moreover,patient-specific induced pluripotent stem(iPS) cells can be generated by reprogramming somatic cells.iPS cells have characteristics similar to those of ES cells and offer a new cell source for type I diabetes cell therapy that reduces the risk of immunologic rejection.In this review,we summarize the recent progress made in the differentiation of hES and iPS cells into functional pancreatic islet cells and discuss the challenges for their future study.

  14. Chronically Increased Amino Acids Improve Insulin Secretion, Pancreatic Vascularity, and Islet Size in Growth-Restricted Fetal Sheep.

    Science.gov (United States)

    Brown, Laura D; Davis, Melissa; Wai, Sandra; Wesolowski, Stephanie R; Hay, William W; Limesand, Sean W; Rozance, Paul J

    2016-10-01

    Placental insufficiency is associated with reduced supply of amino acids to the fetus and leads to intrauterine growth restriction (IUGR). IUGR fetuses are characterized by lower glucose-stimulated insulin secretion, smaller pancreatic islets with less β-cells, and impaired pancreatic vascularity. To test whether supplemental amino acids infused into the IUGR fetus could improve these complications of IUGR we used acute (hours) and chronic (11 d) direct fetal amino acid infusions into a sheep model of placental insufficiency and IUGR near the end of gestation. IUGR fetuses had attenuated acute amino acid-stimulated insulin secretion compared with control fetuses. These results were confirmed in isolated IUGR pancreatic islets. After the chronic fetal amino acid infusion, fetal glucose-stimulated insulin secretion and islet size were restored to control values. These changes were associated with normalization of fetal pancreatic vascularity and higher fetal pancreatic vascular endothelial growth factor A protein concentrations. These results demonstrate that decreased fetal amino acid supply contributes to the pathogenesis of pancreatic islet defects in IUGR. Moreover, the results show that pancreatic islets in IUGR fetuses retain their ability to respond to increased amino acids near the end of gestation after chronic fetal growth restriction.

  15. Generation of pancreatic islet cells from human embryonic stem cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG DongHui; JIANG Wei; SHI Yan; DENG HongKui

    2009-01-01

    Efficiently obtaining functional pancreaUc islet cells derived from human embryonic stem (hES) cells not only provides great potential to solve the shortage of islets sources for type I diabetes cell therapy,but also benefits the study of the development of the human pancreas and diabetes pathology. In 2001,hES cells were reported to have the capacity to generate insulin-producing cells by spontaneous differentiation in vitro. Since then, many strategies (such as overexpression of key transcription factors,delivery of key proteins for pancreatic development, co-transplantation of differentiated hES cells along with fetal pancreas, stepwise differentiation by mimicking in vivo pancreatic development) have been employed in order to induce the differentiation of pancreatic islet cells from hES cells. Moreover, patient-specific induced pluripotent stem (iPS) cells can be generated by reprogramming somatic cells.iPS cells have characteristics similar to those of ES cells and offer a new cell source for type I diabetes cell therapy that reduces the risk of immunologic rejection. In this review, we summarize the recent progress made in the differentiation of hES and iPS cells into functional pancreatic islet cells and discuss the challenges for their future study.

  16. Fetal Exposure to Sertraline Hydrochloride Impairs Pancreatic β-Cell Development.

    Science.gov (United States)

    De Long, Nicole E; Gutgesell, Marie K; Petrik, James J; Holloway, Alison C

    2015-06-01

    Ten percent to 15% of women take selective serotonin reuptake inhibitor (SSRI) antidepressants during pregnancy. Offspring exposed to SSRIs are more likely to have low birth weight; this is associated with an increased risk of development of diabetes in adulthood in part due to altered pancreatic development. The effects of perinatal exposure to SSRIs on pancreatic development are unknown. Therefore, the objective of this study was to determine the effect of fetal exposure to sertraline hydrochloride on pregnancy outcomes and pancreatic development. Wistar rats were given vehicle (n = 5) or sertraline hydrochloride (10 mg/kg/d; n = 8) via daily subcutaneous injection from the confirmation of mating until parturition. Results from this animal model demonstrated that offspring born to sertraline-exposed dams have no changes in birth weight but had a reduction in pancreatic β-cell area. The altered pancreatic islet development was a result of altered gene expression regulating islet development and survival. Therefore, fetal exposure to sertraline reduces β-cell capacity at birth, raising concerns regarding the long-term metabolic sequelae of such exposures.

  17. Isolation, Culture and Induced Differentiation of Fetal Porcine Islet Derived Pancreatic Stem Cell

    Institute of Scientific and Technical Information of China (English)

    FENG Ruo-peng; ZHANG Hui-ru; WANG Yun; QIAO Hai; ZHAO Ting; SHEN Wen-zheng; DOU Zhong-ying

    2007-01-01

    To isolate and culture the porcine pancreatic stem cells and investigate their function, the fetal porcine pancreatic stem cells were isolated by the method of suspending plus adhering culture. The isolated cells were then identified by irnmunohistochemical staining, and their culture viability measured through the MTT method in vitro. This induced them to differentiate into endocrine cells and detect their function. The isolated IPSCS did not express nestin, but expressed CK-19, a marker of ductal epithelia cells and oc-actin, a smooth muscle marker, demonstrating the growth characteristics of ES-like cells, and strong proliferative ability, after 18 passages. They could excrete insulin, and showed ultrastructure changes after being induced. Porcine pancreatic stem cells can be isolated by this method, induced to form islet-like clusters, and can secret insulin.

  18. Comparative proteomic analysis of human pancreatic juice : Methodological study

    NARCIS (Netherlands)

    Zhou, Lu; Lu, ZhaoHui; Yang, AiMing; Deng, RuiXue; Mai, CanRong; Sang, XinTing; Faber, Klaas Nico; Lu, XingHua

    Pancreatic cancer is the most lethal of all the common malignancies. Markers for early detection of this disease are urgently needed. Here, we optimized and applied a proteome analysis of human pancreatic juice to identify biomarkers for pancreatic cancer. Pancreatic juice samples, devoid of blood

  19. Comparative proteomic analysis of human pancreatic juice : Methodological study

    NARCIS (Netherlands)

    Zhou, Lu; Lu, ZhaoHui; Yang, AiMing; Deng, RuiXue; Mai, CanRong; Sang, XinTing; Faber, Klaas Nico; Lu, XingHua

    2007-01-01

    Pancreatic cancer is the most lethal of all the common malignancies. Markers for early detection of this disease are urgently needed. Here, we optimized and applied a proteome analysis of human pancreatic juice to identify biomarkers for pancreatic cancer. Pancreatic juice samples, devoid of blood o

  20. Comparative proteomic analysis of human pancreatic juice : Methodological study

    NARCIS (Netherlands)

    Zhou, Lu; Lu, ZhaoHui; Yang, AiMing; Deng, RuiXue; Mai, CanRong; Sang, XinTing; Faber, Klaas Nico; Lu, XingHua

    2007-01-01

    Pancreatic cancer is the most lethal of all the common malignancies. Markers for early detection of this disease are urgently needed. Here, we optimized and applied a proteome analysis of human pancreatic juice to identify biomarkers for pancreatic cancer. Pancreatic juice samples, devoid of blood o

  1. Comparative proteomic analysis of human pancreatic juice: Methodological study

    NARCIS (Netherlands)

    Zhou, Lu; Lu, Z.H.; Yang, A.M.; Deng, R.X.; Mai, C.R.; Sang, X.T.; Faber, Klaas Nico; Lu, X.H.

    2007-01-01

    Pancreatic cancer is the most lethal of all the common malignancies. Markers for early detection of this disease are urgently needed. Here, we optimized and applied a proteome analysis of human pancreatic juice to identify biomarkers for pancreatic cancer. Pancreatic juice samples, devoid of blood o

  2. Establishing a human pancreatic stem cell line and transplanting induced pancreatic islets to reverse experimental diabetes in rats

    Institute of Scientific and Technical Information of China (English)

    XIAO Mei; DOU ZhongYing; AN LiLong; YANG XueYi; GE Xin; QIAO Hai; ZHAO Ting; MA XiaoFei; FAN JingZhua; ZHU MengYang

    2008-01-01

    The major obstacle in using pancreatic islet transplantation to cure type Ⅰ and some type Ⅱ diabetes is the shortage of the donors. One of ways to overcome such obstacle is to isolate and clone pancreatic stem cells as "seed cells" and induce their differentiation into functional islets as an abundant trans-plantation source. In this study, a monoclonal human pancreatic stem cell (mhPSC) line was obtained from abortive fetal pancreatic tissues. Pancreatic tissues were taken from abortive fetus by sterile procedures, and digested into single cells and cell clusters with 0.1% type Ⅳ collagenase. Cultured in modified glucose-low DMEM with 10% fetal bovine serum (FBS), these single cells and cell clusters adhered to culture dishes, and then primary epidermal-like pancreatic stem ceils started to clone. After digesting with 0.25% trypsin and 0.04% EDTA, fibroblasts and other cells were gradually eliminated and epithelioid pancreatic stem cells were gradually purified during generations. Using clone-ring selection, the mhPSCs were obtained. After addition of 10 ng/mL epidermal growth factor (EGF) in cell culture medium, the mhPSCs quickly grew and formed a gravelstone-like monolayer. Continuously proliferated, a mhPSC line, which was derived from a male abortive fetus of 4 months old, has been passed through 50 generations. More than 1×109 mhPSCs were cryo-preserved in liquid nitrogen. Karyotype analysis showed that the chromosome set of the mhPSC line was normal diploid. Immunocytochemistry results demonstrated that the mhPSC line was positive for the pdxl, glucagon, nestin and CK19, and negative for the insulin, CD34, CD44 and CD45 protein expression. RT-PCR revealed further that the mhPSCs expressed transcription factors of the pdx1, glucagon, nestin and CK19. Also, in vitro induced with β-mercaptoethanol, the mhPSCs differentiated into nerve cells that expressed the NF protein. Induced with nicotinamide, the mhPSCs differentiated into functional islet

  3. Establishing a human pancreatic stem cell line and transplanting induced pancreatic islets to reverse experimental diabetes in rats

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    The major obstacle in using pancreatic islet transplantation to cure type I and some type II diabetes is the shortage of the donors. One of ways to overcome such obstacle is to isolate and clone pancreatic stem cells as "seed cells" and induce their differentiation into functional islets as an abundant trans-plantation source. In this study, a monoclonal human pancreatic stem cell (mhPSC) line was obtained from abortive fetal pancreatic tissues. Pancreatic tissues were taken from abortive fetus by sterile procedures, and digested into single cells and cell clusters with 0.1% type IV collagenase. Cultured in modified glucose-low DMEM with 10% fetal bovine serum (FBS), these single cells and cell clusters adhered to culture dishes, and then primary epidermal-like pancreatic stem cells started to clone. After digesting with 0.25% trypsin and 0.04% EDTA, fibroblasts and other cells were gradually eliminated and epithelioid pancreatic stem cells were gradually purified during generations. Using clone-ring selection, the mhPSCs were obtained. After addition of 10 ng/mL epidermal growth factor (EGF) in cell culture medium, the mhPSCs quickly grew and formed a gravelstone-like monolayer. Continuously proliferated, a mhPSC line, which was derived from a male abortive fetus of 4 months old, has been passed through 50 generations. More than 1×109 mhPSCs were cryo-preserved in liquid nitrogen. Karyotype analysis showed that the chromosome set of the mhPSC line was normal diploid. Immunocytochemistry results demonstrated that the mhPSC line was positive for the pdx1, glucagon, nestin and CK19, and negative for the insulin, CD34, CD44 and CD45 protein expression. RT-PCR revealed further that the mhPSCs expressed transcription factors of the pdx1, glucagon, nestin and CK19. Also, in vitro induced with β-mercaptoethanol, the mhPSCs differentiated into nerve cells that expressed the NF protein. Induced with nicotinamide, the mhPSCs differentiated into functional islet

  4. Cyclooxygenase-2 Expression in Hamster and Human Pancreatic Neoplasia

    Directory of Open Access Journals (Sweden)

    Pamela L. Crowell

    2006-06-01

    Full Text Available Cyclooxygenase-2 (COX-2 has been implicated in the development of gastrointestinal malignancies. The aim of the present study was to determine COX-2 expression/activity throughout stages of experimental and human pancreatic neoplasia. COX-2 immunohistochemistry was performed in pancreata of hamsters subjected to the carcinogen N-nitrosobis-(2-oxopropylamine (BOP and in human pancreatic tumors. COX-2 activity was determined by prostaglandin E2 assay in tumor versus matched normal pancreatic tissues. The activity of the COX inhibitor sulindac was tested in the PC-1 hamster pancreatic cancer model. COX-2 expression was elevated in all pancreatic intraepithelial neoplasias (PanINs and adenocarcinomas. In BOP-treated hamsters, there were significant progressive elevations in COX-2 expression throughout pancreatic tumorigenesis. In human samples, peak COX-2 expression occurred in PanIN2 lesions and remained moderately elevated in PanIN3 and adenocarcinoma tissues. COX-2 activity was significantly elevated in hamster and human pancreatic cancers compared to pair-matched normal pancreas. Furthermore, hamster pancreatic tumor engraftment/formation in the PC-1 hamster pancreatic cancer model was reduced 4.9-fold by oral administration of sulindac. Increased COX-2 expression is an early event in pancreatic carcinogeneses. The BOP-induced hamster carcinogenesis model is a representative model used to study the role of COX-2 in well-differentiated pancreatic tumorigenesis. COX inhibitors may have a role in preventing tumor engraftment/formation.

  5. Distribution of melatonin receptor in human fetal brain

    Institute of Scientific and Technical Information of China (English)

    WANG Guo-quan; SHAO Fu-yuan; ZHAO Ying; LIU Zhi-min

    2001-01-01

    Objective: To study the distribution of 2 kinds of melatonin receptor subtypes (mtl and MT2) in human fetal brain. Methods: The fetal brain tissues were sliced and the distribution ofmelatonin receptors in human fetal brain were detected using immunohistochemistry and in situ hybridization. Results: Melatonin receptor mtl existed in the cerebellun and hypothalamus, melatonin receptor MT2 exists in hypothalamus, occipital and medulla. Conclusion: Two kinds of melatonin receptors, mtl and MT2 exist in the membrane and cytosol of brain cells, indicating that human fetal brain is a target organ of melatonin.

  6. Human pancreatic cell autotransplantation following total pancreatectomy.

    Science.gov (United States)

    Traverso, L W; Abou-Zamzam, A M; Longmire, W P

    1981-01-01

    During total pancreaticoduodenectomy for chronic pancreatitis, four patients received an intraportal pancreatic mixed-cell autograft prepared by collagenase digestion. The technique of this autotransplantation procedure was successfully developed using a normal canine pancreas, but has proved difficult to apply in the human chronic pancreatitis model. Our four patients became insulin-dependent, with proof of intrahepatic insulin production in only one patient. Three factors have contributed to the lack of graft success: 1) the preoperative endocrine status, 2) systemic hypotension and portal hypertension secondary to graft infusion, and 3) difficulty applying the successful technique in a normal dog pancreas to an extensively scarred human pancreas. The preoperative insulin response during a glucose tolerance test was blunted or delayed in the three patients tested. An immediate decrease in blood pressure and rise in portal pressure occurred in every patient and prevented infusion of the entire graft (30-50%) in three patients. Unfortunately, the patient with the most compromised insulin status was the only patient able to receive the entire graft. Our experience would indicate that further refinements in technique are necessary to prevent the vascular reaction and allow infusion of the entire graft. Furthermore, normal islet cell function is necessary before a successful graft can be expected. PMID:6781424

  7. Fetal magnetic resonance imaging and human genetics

    Energy Technology Data Exchange (ETDEWEB)

    Hengstschlaeger, Markus [Medical Genetics, Obstetrics and Gynecology, Medical University of Vienna, Waehringer Guertel 18-20, 1090 Vienna (Austria)]. E-mail: markus.hengstschlaeger@meduniwien.ac.at

    2006-02-15

    The use of fetal magnetic resonance imaging (MRI), in addition to prenatal genetic testing and sonography, has the potential to improve prenatal diagnosis of genetic disorders. MRI plays an important role in the evaluation of fetal abnormalities and malformations. Fetal MRI often enables a differential diagnosis, a determination of the extent of the disorder, the prognosis, and an improvement in therapeutic management. For counseling of parents, as well as to basically understand how genetic aberrations affect fetal development, it is of great importance to correlate different genotypes with fetal MRI data.

  8. Fetal Growth and Timing of Parturition in Humans

    Science.gov (United States)

    Sundaram, Rajeshwari; Sun, Wenyu; Troendle, James

    2008-01-01

    Animal studies indicate that either the fetus or the intrauterine environment, both of which set the pattern for fetal growth, may affect the timing of parturition. The authors examined the association between fetal growth and timing of spontaneous onset of labor in humans among low-risk white US women with singleton pregnancies (1987–1991). They restricted the data to pregnancies which had a reliable date of the last menstrual period, normal fetal growth in the first half of pregnancy, and no history of or current pregnancy complications that might have impaired fetal growth (n = 3,360). Subjects received ultrasound examinations at 15–22 and 31–35 weeks’ gestation. Fetal growth was adjusted for parity, fetal sex, and maternal prepregnancy weight and height. Results showed that slower or faster fetal growth in the second half of pregnancy resulted in substantially lower or higher birth weight, respectively. However, fetal growth in the second half of pregnancy, even at extremes (2 standard deviations below or above the mean), did not have a meaningful impact on the timing of parturition; neither did fetal growth acceleration or deceleration in late pregnancy. Thus, in low-risk pregnancies where fetal growth is normal in early gestation, fetal growth in the second half of pregnancy does not affect the timing of normal parturition. PMID:18775925

  9. Fetal growth and timing of parturition in humans.

    Science.gov (United States)

    Zhang, Jun; Sundaram, Rajeshwari; Sun, Wenyu; Troendle, James

    2008-10-15

    Animal studies indicate that either the fetus or the intrauterine environment, both of which set the pattern for fetal growth, may affect the timing of parturition. The authors examined the association between fetal growth and timing of spontaneous onset of labor in humans among low-risk white US women with singleton pregnancies (1987-1991). They restricted the data to pregnancies which had a reliable date of the last menstrual period, normal fetal growth in the first half of pregnancy, and no history of or current pregnancy complications that might have impaired fetal growth (n = 3,360). Subjects received ultrasound examinations at 15-22 and 31-35 weeks' gestation. Fetal growth was adjusted for parity, fetal sex, and maternal prepregnancy weight and height. Results showed that slower or faster fetal growth in the second half of pregnancy resulted in substantially lower or higher birth weight, respectively. However, fetal growth in the second half of pregnancy, even at extremes (2 standard deviations below or above the mean), did not have a meaningful impact on the timing of parturition; neither did fetal growth acceleration or deceleration in late pregnancy. Thus, in low-risk pregnancies where fetal growth is normal in early gestation, fetal growth in the second half of pregnancy does not affect the timing of normal parturition.

  10. Characterization of insulin-like growth factor I produced by fetal rat pancreatic islets

    Energy Technology Data Exchange (ETDEWEB)

    Scharfmann, R.; Corvol, M.; Czernichow, P. (Institut National de la Sante et de la Recherche Medicale (Unit 30), Paris (France))

    1989-06-01

    Pancreatic islets were prepared from 22-day-old rat fetuses. After 5 days of culture in dishes allowing cell attachment, neoformed islets were kept free floating in RPMI-1640 medium (16.5 mM glucose, 1% fetal calf serum). The islets were then pulsed with ({sup 3}H)leucine and ({sup 35}S)methionine for 24 h. The conditioned medium was acidified with acetic acid (final pH 2.7), desalted, concentrated, and gel filtered on Bio-Gel P100 in acid conditions. The radioactive material that comigrated with immunoreactive insulinlike growth factor I (IGF-I) produced by the islets was pooled, concentrated, and further characterized by reverse-phase high-performance liquid chromatography on a C18 Bondapak column with a linear gradient of acetonitrile (20-80%). The radioactive material that eluted as pure IGF-I (40% acetonitrile) was further studied by chromatofocusing on a Pharmacia PBE 94 column. A sharp radioactive peak containing ({sup 3}H)leucine and ({sup 35}S)methionine was eluted at pH 8.55. This material was immunoprecipitated with an antiserum to IGF-I. This study demonstrated that fetal islet cells synthesize molecules that are, by several criteria, equivalent to native IGF-I.

  11. Mouse Model of Human Hereditary Pancreatitis

    Science.gov (United States)

    2016-09-01

    models that recapitulate the human disease . Therefore, we introduced mutations in the endogenous mouse T7 cationic trypsinogen gene and obtained several...ACCOMPLISHMENTS: What were the major goals of the project? Our original proposal had three specific aims. Aim 1. Identify and biochemically characterize...pancreatitis in mutant mice which do not develop spontaneous disease (strains T7-D23del-Cre, T7-D23del-Neo, T7-K24R-Cre and T7- K24R-Neo), will be

  12. Apoptosis of human pancreatic cancer cells induced by Triptolide

    Institute of Scientific and Technical Information of China (English)

    Guo-Xiong Zhou; Xiao-Ling Ding; Jie-Fei Huang; Hong Zhang; Sheng-Bao Wu; Jian-Ping Cheng; Qun Wei

    2008-01-01

    AIM:To investigate apoptosis in human pancreatic cancer ceils induced by Triptolide (TL),and the relationship between this apoptosis and expression of caspase-3' bcl-2 and bax.METHODS:Human pancreatic cancer cell line SW1990 was cultured in DIEM media for this study.MTT assay was used to determine the cell growth inhibitory rate in vitro.Flow cytometry and TUNEL assay were used to detect the apoptosis of human pancreatic cancer cells before and after TL treatment.RT-PCR was used to detect the expression of apoptosis-associated gene caspase-3' bcl-2 and bax.RESULTS:TL inhibited the growth of human pancreatic cancer cells in a dose-and time-dependent manner.TL induced human pancreatic cancer cells to undergo apoptosis with typically apoptotic characteristics.TUNEL assay showed that after the treatment of human pancreatic cancer cells with 40 ng/mL TL for 12 h and 24 h,the apoptotic rates of human pancreatic cancer cells increased significantly.RT-PCR demonstrated that caspase-3 and bax were significantly up-regulated in SW1990 cells treated with TL while bcl-2 mRNA was not.CONCLUSION:TL is able to induce the apoptosis in human pancreatic cancer cells.This apoptosis may be mediated by up-regulating the expression of apoptosisassociated caspase-3 and bax gene.

  13. [Biomechanical characteristics of human fetal membranes. Preterm fetal membranes are stronger than term fetal membranes].

    Science.gov (United States)

    Rangaswamy, N; Abdelrahim, A; Moore, R M; Uyen, L; Mercer, B M; Mansour, J M; Kumar, D; Sawady, J; Moore, J J

    2011-06-01

    The purpose of this study was to determine the biomechanical characteristics of human fetal membranes (FM) throughout gestation. Biomechanical properties were determined for 115 FM of 23-41 weeks gestation using our previously described methodology. The areas of membrane immediately adjacent to the strongest and weakest tested spots were sampled for histomorphometric analysis. Clinical data on the patients whose FM were examined were also collected. FM less than 28 weeks gestation were associated with higher incidence of abruption and chorioamnionitis. Topographically FM at all gestations had heterogeneous biomechanical characteristics over their surfaces with distinct weak areas. The most premature membranes were the strongest. FM strength represented by rupture force and work to rupture decreased with increasing gestation in both weak and strong regions of FM. This decrease in FM strength was most dramatic at more than 38 weeks gestation. The FM component amnion-chorion sublayers were thinner in the weak areas compared to strong areas. Compared to term FM, preterm FM are stronger but have similar heterogeneous weak and strong areas. Following a gradual increase in FM weakness with increasing gestation, there is a major drop-off at term 38 weeks gestation. The FM weak areas are thinner than the stronger areas. Whether the difference in thickness is enough to account for the strength differences is unknown.

  14. Blood pressure estimation in the human fetal descending aorta.

    NARCIS (Netherlands)

    Struijk, P.C.; Mathews, V.J.; Loupas, T.; Stewart, P.A.; Clark, E.B.; Steegers, E.A.P.; Wladimiroff, J.W.

    2008-01-01

    OBJECTIVES: The objectives of this study were to estimate fetal blood pressure non-invasively from two-dimensional color Doppler-derived aortic blood flow and diameter waveforms, and to compare the results with invasively derived human fetal blood pressures available from the literature. METHODS:

  15. Blood pressure estimation in the human fetal descending aorta

    NARCIS (Netherlands)

    P.C. Struijk (Pieter); V.J. Mathews; T. Loupas; P.A. Stewart (Patricia); E.B. Clark; R.P.M. Steegers-Theunissen (Régine); J.W. Wladimiroff (Juriy)

    2008-01-01

    textabstractObjectives: The objectives of this study were to estimate fetal blood pressure non-invasively from two-dimensional color Doppler-derived aortic blood flow and diameter waveforms, and to compare the results with invasively derived human fetal blood pressures available from the literature.

  16. Differentiation and functional regulation of human fetal NK cells.

    Science.gov (United States)

    Ivarsson, Martin A; Loh, Liyen; Marquardt, Nicole; Kekäläinen, Eliisa; Berglin, Lena; Björkström, Niklas K; Westgren, Magnus; Nixon, Douglas F; Michaëlsson, Jakob

    2013-09-01

    The human fetal immune system is naturally exposed to maternal allogeneic cells, maternal antibodies, and pathogens. As such, it is faced with a considerable challenge with respect to the balance between immune reactivity and tolerance. Here, we show that fetal natural killer (NK) cells differentiate early in utero and are highly responsive to cytokines and antibody-mediated stimulation but respond poorly to HLA class I-negative target cells. Strikingly, expression of killer-cell immunoglobulin-like receptors (KIRs) did not educate fetal NK cells but rendered them hyporesponsive to target cells lacking HLA class I. In addition, fetal NK cells were highly susceptible to TGF-β-mediated suppression, and blocking of TGF-β signaling enhanced fetal NK cell responses to target cells. Our data demonstrate that KIR-mediated hyporesponsiveness and TGF-β-mediated suppression are major factors determining human fetal NK cell hyporesponsiveness to HLA class I-negative target cells and provide a potential mechanism for fetal-maternal tolerance in utero. Finally, our results provide a basis for understanding the role of fetal NK cells in pregnancy complications in which NK cells could be involved, for example, during in utero infections and anti-RhD-induced fetal anemia.

  17. Effect of phorbol and glucose on insulin secretion from the human fetal pancreas.

    Science.gov (United States)

    Tuch, B E; Williams, P F; Handelsman, D; Dunlop, M; Grigoriou, S; Turtle, J R

    1987-04-01

    It has been reported previously that 12-0-tetradecanoylphorbol-13-acetate is capable of stimulating the release of insulin from adult and neonatal pancreatic tissue. The data from this study show that this agent at a concentration of 1.3 uM, in the presence of 2.8 mM glucose, was unable to cause significant secretion of insulin from cultured human fetal pancreatic explants. By contrast 20 mM glucose was able to cause a small but significant immediate increase in secretion of insulin, but was unable to maintain this response beyond ten minutes. When the two agents were combined, a synergistic effect was seen throughout the entire 50 minute period of stimulation. The reason for this synergism is unclear since, whilst both secretagogues were able to cause a rise in the levels of diacylglycerol, together no extra effect was observed.

  18. Experimental Models in Syrian Golden Hamster Replicate Human Acute Pancreatitis.

    Science.gov (United States)

    Wang, Yunan; Kayoumu, Abudurexiti; Lu, Guotao; Xu, Pengfei; Qiu, Xu; Chen, Liye; Qi, Rong; Huang, Shouxiong; Li, Weiqin; Wang, Yuhui; Liu, George

    2016-06-15

    The hamster has been shown to share a variety of metabolic similarities with humans. To replicate human acute pancreatitis with hamsters, we comparatively studied the efficacy of common methods, such as the peritoneal injections of caerulein, L-arginine, the retrograde infusion of sodium taurocholate, and another novel model with concomitant administration of ethanol and fatty acid. The severity of pancreatitis was evaluated by serum amylase activity, pathological scores, myeloperoxidase activity, and the expression of inflammation factors in pancreas. The results support that the severity of pathological injury is consistent with the pancreatitis induced in mice and rat using the same methods. Specifically, caerulein induced mild edematous pancreatitis accompanied by minimal lung injury, while L-arginine induced extremely severe pancreatic injury including necrosis and neutrophil infiltration. Infusion of Na-taurocholate into the pancreatic duct induced necrotizing pancreatitis in the head of pancreas and lighter inflammation in the distal region. The severity of acute pancreatitis induced by combination of ethanol and fatty acids was between the extent of caerulein and L-arginine induction, with obvious inflammatory cells infiltration. In view of the advantages in lipid metabolism features, hamster models are ideally suited for the studies of pancreatitis associated with altered metabolism in humans.

  19. Translating discovery in zebrafish pancreatic development to human pancreatic cancer: biomarkers, targets, pathogenesis, and therapeutics.

    Science.gov (United States)

    Yee, Nelson S; Kazi, Abid A; Yee, Rosemary K

    2013-06-01

    Abstract Experimental studies in the zebrafish have greatly facilitated understanding of genetic regulation of the early developmental events in the pancreas. Various approaches using forward and reverse genetics, chemical genetics, and transgenesis in zebrafish have demonstrated generally conserved regulatory roles of mammalian genes and discovered novel genetic pathways in exocrine pancreatic development. Accumulating evidence has supported the use of zebrafish as a model of human malignant diseases, including pancreatic cancer. Studies have shown that the genetic regulators of exocrine pancreatic development in zebrafish can be translated into potential clinical biomarkers and therapeutic targets in human pancreatic adenocarcinoma. Transgenic zebrafish expressing oncogenic K-ras and zebrafish tumor xenograft model have emerged as valuable tools for dissecting the pathogenetic mechanisms of pancreatic cancer and for drug discovery and toxicology. Future analysis of the pancreas in zebrafish will continue to advance understanding of the genetic regulation and biological mechanisms during organogenesis. Results of those studies are expected to provide new insights into how aberrant developmental pathways contribute to formation and growth of pancreatic neoplasia, and hopefully generate valid biomarkers and targets as well as effective and safe therapeutics in pancreatic cancer.

  20. Cerebral Organoids Recapitulate Epigenomic Signatures of the Human Fetal Brain

    Directory of Open Access Journals (Sweden)

    Chongyuan Luo

    2016-12-01

    Full Text Available Organoids derived from human pluripotent stem cells recapitulate the early three-dimensional organization of the human brain, but whether they establish the epigenomic and transcriptional programs essential for brain development is unknown. We compared epigenomic and regulatory features in cerebral organoids and human fetal brain, using genome-wide, base resolution DNA methylome and transcriptome sequencing. Transcriptomic dynamics in organoids faithfully modeled gene expression trajectories in early-to-mid human fetal brains. We found that early non-CG methylation accumulation at super-enhancers in both fetal brain and organoids marks forthcoming transcriptional repression in the fully developed brain. Demethylated regions (74% of 35,627 identified during organoid differentiation overlapped with fetal brain regulatory elements. Interestingly, pericentromeric repeats showed widespread demethylation in multiple types of in vitro human neural differentiation models but not in fetal brain. Our study reveals that organoids recapitulate many epigenomic features of mid-fetal human brain and also identified novel non-CG methylation signatures of brain development.

  1. Gene profile identifies zinc transporters differentially expressed in normal human organs and human pancreatic cancer.

    Science.gov (United States)

    Yang, J; Zhang, Y; Cui, X; Yao, W; Yu, X; Cen, P; Hodges, S E; Fisher, W E; Brunicardi, F C; Chen, C; Yao, Q; Li, M

    2013-03-01

    Deregulated expression of zinc transporters was linked to several cancers. However, the detailed expression profile of all human zinc transporters in normal human organs and in human cancer, especially in pancreatic cancer is not available. The objectives of this study are to investigate the complete expression patterns of 14 ZIP and 10 ZnT transporters in a large number of normal human organs and in human pancreatic cancer tissues and cell lines. We examined the expression patterns of ZIP and ZnT transporters in 22 different human organs and tissues, 11 pairs of clinical human pancreatic cancer specimens and surrounding normal/benign tissues, as well as 10 established human pancreatic cancer cell lines plus normal human pancreatic ductal epithelium (HPDE) cells, using real time RT-PCR and immunohistochemistry. The results indicate that human zinc transporters have tissue specific expression patterns, and may play different roles in different organs or tissues. Almost all the ZIPs except for ZIP4, and most ZnTs were down-regulated in human pancreatic cancer tissues compared to the surrounding benign tissues. The expression patterns of individual ZIPs and ZnTs are similar among different pancreatic cancer lines. Those results and our previous studies suggest that ZIP4 is the only zinc transporter that is significantly up-regulated in human pancreatic cancer and might be the major zinc transporter that plays an important role in pancreatic cancer growth. ZIP4 might serve as a novel molecular target for pancreatic cancer diagnosis and therapy.

  2. Distinct functional programming of human fetal and adult monocytes.

    Science.gov (United States)

    Krow-Lucal, Elisabeth R; Kim, Charles C; Burt, Trevor D; McCune, Joseph M

    2014-03-20

    Preterm birth affects 1 out of 9 infants in the United States and is the leading cause of long-term neurologic handicap and infant mortality, accounting for 35% of all infant deaths in 2008. Although cytokines including interferon-γ (IFN-γ), interleukin-10 (IL-10), IL-6, and IL-1 are produced in response to in utero infection and are strongly associated with preterm labor, little is known about how human fetal immune cells respond to these cytokines. We demonstrate that fetal and adult CD14(+)CD16(-) classical monocytes are distinct in terms of basal transcriptional profiles and in phosphorylation of signal transducers and activators of transcription (STATs) in response to cytokines. Fetal monocytes phosphorylate canonical and noncanonical STATs and respond more strongly to IFN-γ, IL-6, and IL-4 than adult monocytes. We demonstrate a higher ratio of SOCS3 to IL-6 receptor in adult monocytes than in fetal monocytes, potentially explaining differences in STAT phosphorylation. Additionally, IFN-γ signaling results in upregulation of antigen presentation and costimulatory machinery in adult, but not fetal, monocytes. These findings represent the first evidence that primary human fetal and adult monocytes are functionally distinct, potentially explaining how these cells respond differentially to cytokines implicated in development, in utero infections, and the pathogenesis of preterm labor.

  3. Comprehensive proteomic analysis of human pancreatic juice

    DEFF Research Database (Denmark)

    Grønborg, Mads; Bunkenborg, Jakob; Kristiansen, Troels Zakarias

    2004-01-01

    ). In addition, we identified a number of proteins that have not been previously described in pancreatic juice (e.g., tumor rejection antigen (pg96) and azurocidin). Interestingly, a novel protein that is 85% identical to HIP/PAP was identified, which we have designated as PAP-2. The proteins identified...... in this study could be directly assessed for their potential as biomarkers for pancreatic cancer by quantitative proteomics methods or immunoassays....

  4. Human pancreatic islet progenitor cells demonstrate phenotypic plasticity in vitro

    Indian Academy of Sciences (India)

    Maithili P Dalvi; Malati R Umrani; Mugdha V Joglekar; Anandwardhan A Hardikar

    2009-10-01

    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 diabetes. The phenotypic plasticity exhibited by pancreatic progenitors during reversible epithelial-to-mesenchymal transition (EMT) and possible role of microRNAs in regulation of this process is also presented herein.

  5. Human fetal bone cells in delivery systems for bone engineering.

    Science.gov (United States)

    Tenorio, Diene M H; Scaletta, Corinne; Jaccoud, Sandra; Hirt-Burri, Nathalie; Pioletti, Dominique P; Jaques, Bertrand; Applegate, Lee Ann

    2011-11-01

    The aim of this study was to culture human fetal bone cells (dedicated cell banks of fetal bone derived from 14 week gestation femurs) within both hyaluronic acid gel and collagen foam, to compare the biocompatibility of both matrices as potential delivery systems for bone engineering and particularly for oral application. Fetal bone cell banks were prepared from one organ donation and cells were cultured for up to 4 weeks within hyaluronic acid (Mesolis®) and collagen foams (TissueFleece®). Cell survival and differentiation were assessed by cell proliferation assays and histology of frozen sections stained with Giemsa, von Kossa and ALP at 1, 2 and 4 weeks of culture. Within both materials, fetal bone cells could proliferate in three-dimensional structure at ∼70% capacity compared to monolayer culture. In addition, these cells were positive for ALP and von Kossa staining, indicating cellular differentiation and matrix production. Collagen foam provides a better structure for fetal bone cell delivery if cavity filling is necessary and hydrogels would permit an injectable technique for difficult to treat areas. In all, there was high biocompatibility, cellular differentiation and matrix deposition seen in both matrices by fetal bone cells, allowing for easy cell delivery for bone stimulation in vivo. Copyright © 2011 John Wiley & Sons, Ltd.

  6. Materno-Fetal Transmission of Human Immune Deficiency Virus

    Directory of Open Access Journals (Sweden)

    Axel Schäfer

    1997-01-01

    Full Text Available Mother-to-child transmission of human immune deficiency virus (HIV is a multifactorial event highly associated with advanced maternal HIV disease and obstetric incidents taking place during parturition. Thus, various approaches to prevention may be beneficial. Although the time and the route of materno-fetal HIV transmission are still not sufficiently clear, much speaks in favor of a late HIV transmission, most probably taking place during parturition or the phase before the delivery. The fetus is remarkably protected by the placenta and the intact fetal membranes against many viral infections during gestation. These conditions change at parturition and the chance for a transition of HIV-infected carrier cells or virus into the fetal compartment increases. Proinflammatory cytokines secreted at the materno-fetal interface accumulate in amniotic fluid and may chemoattract and stimulate potentially HIV-infected immunocytes. After rupture of membranes, maternal cells of the decidua are directly exposed to the amniotic fluid. Aside from the contamination of the fetal skin at vaginal delivery as a debatable route of infection, blood-to-blood contacts and the fetal swallowing of contaminated amniotic fluid may be the major path of fetal HIV infection. For the fetal prophylaxis of an intrauterine infection, the application of zidovudine is recommended. However, cesarian section before the onset of labor leads also to a diminution of the transmission rate. As the transmission seems to have both systemic and local causes, it makes sense to combine different intervention strategies. Whether a combination of zidovudine and elective cesarean section can lower the transmission risk further has to be evaluated.

  7. Activation of Raf-1 in human pancreatic adenocarcinoma.

    Science.gov (United States)

    Berger, D H; Jardines, L A; Chang, H; Ruggeri, B

    1997-04-01

    Point mutations in the Ras oncogene cause Ras to remain in its active GTP-bound state sending signals downstream continuously. Since 75 to 90% of all human pancreatic ductal adenocarcinomas harbor activating mutations at codon 12 of the K-ras oncogene it was our belief that Raf-1-MEK-MAPK will be activated in the majority of human pancreatic cancers. The aim of this study was to confirm activation of Raf-1 in K-ras mutant human pancreatic cancer. Additionally, we sought to determine if Raf-1 activation differed in K-ras mutant and nonmutant pancreatic cancer. Furthermore, we were interested in determining if Raf-1 activation in pancreatic cancer led to subsequent activation of downstream effectors such as MAP kinase. The presence of mutations in codon 12 of the K-ras oncogene in 14 human pancreatic adenocarcinoma cell lines was determined by use of mutant allele-specific PCR restriction fragment length polymorphism analysis. Raf-1 expression of quiescent cells was determined by immunoblotting using a rabbit anti-human polyclonal antibody and enhanced chemiluminescence. MAP kinase activity was determined by measuring the incorporation of phosphate into Myelin Basic Protein. Seven cell lines were noted to have mutations in codon 12 of K-ras while seven cell lines did not. There was no difference in expression of the 74 kDa-activated form of Raf-1 in K-ras mutant vs K-ras nonmutant cell lines. However, there was a significant increase in MAP kinase activity in the nonmutant cell lines compared to the cell lines with Ras mutations (P = 0.026). We conclude that Raf-1 is expressed in its active form in human pancreatic cancer regardless of K-ras status. However, signalling downstream of Raf-1 differs in cell lines with K-ras mutations compared to those cell lines without K-ras mutations.

  8. Fetal and neonatal nicotine exposure in Wistar rats causes progressive pancreatic mitochondrial damage and beta cell dysfunction.

    Directory of Open Access Journals (Sweden)

    Jennifer E Bruin

    Full Text Available Nicotine replacement therapy (NRT is currently recommended as a safe smoking cessation aid for pregnant women. However, fetal and neonatal nicotine exposure in rats causes mitochondrial-mediated beta cell apoptosis at weaning, and adult-onset dysglycemia, which we hypothesize is related to progressive mitochondrial dysfunction in the pancreas. Therefore in this study we examined the effect of fetal and neonatal exposure to nicotine on pancreatic mitochondrial structure and function during postnatal development. Female Wistar rats were given saline (vehicle control or nicotine bitartrate (1 mg/kg/d via subcutaneous injection for 2 weeks prior to mating until weaning. At 3-4, 15 and 26 weeks of age, oral glucose tolerance tests were performed, and pancreas tissue was collected for electron microscopy, enzyme activity assays and islet isolation. Following nicotine exposure mitochondrial structural abnormalities were observed beginning at 3 weeks and worsened with advancing age. Importantly the appearance of these structural defects in nicotine-exposed animals preceded the onset of glucose intolerance. Nicotine exposure also resulted in significantly reduced pancreatic respiratory chain enzyme activity, degranulation of beta cells, elevated islet oxidative stress and impaired glucose-stimulated insulin secretion compared to saline controls at 26 weeks of age. Taken together, these data suggest that maternal nicotine use during pregnancy results in postnatal mitochondrial dysfunction that may explain, in part, the dysglycemia observed in the offspring from this animal model. These results clearly indicate that further investigation into the safety of NRT use during pregnancy is warranted.

  9. Pancreatitis

    Science.gov (United States)

    ... the hormones insulin and glucagon into the bloodstream. Pancreatitis is inflammation of the pancreas. It happens when digestive enzymes start digesting the pancreas itself. Pancreatitis can be acute or chronic. Either form is ...

  10. Inhibitory effect of calcitonin on pure human pancreatic secretion.

    Directory of Open Access Journals (Sweden)

    Tanaka,Juntaro

    1989-06-01

    Full Text Available The inhibitory effect of calcitonin on human pancreatic secretion was evaluated to examine whether the different results reported earlier between humans, cats and dogs can be ascribed to the different sensitivity of these species to calcitonin, as suggested by some investigators. Pancreatic juice was obtained by endoscopic cannulation of the pancreatic duct from 11 patients with relapsing pancreatitis during intravenous infusion of secretin (1 U/kg/h plus caerulein (0.04 microgram/kg/h. After steady secretion was attained 20 min after the beginning of collection, five 2-min fractions were obtained before, and ten 2-min fractions were obtained after intravenous infusion of calcitonin (1 IU/kg/h. The pre- and post-calcitonin fractions from each patient were compared by Student's t-test. Calcitonin inhibited the secretory volume (26.8 to 65.6% and bicarbonate secretion (21.4 to 62.0% in 8 patients, and amylase (48.4 to 89.5% and lipase secretion (47.4 to 90.5% in all patients. The present studies reconfirmed that prominent inhibition of enzyme secretion occurs in humans. A new finding was that significant inhibition of the secretory volume and bicarbonate secretion occurs in humans. The inhibitory effects of calcitonin in humans did not appear to differ from those in cats and dogs, when evaluated similarly with the use of pure pancreatic juice.

  11. Pancreatitis

    Institute of Scientific and Technical Information of China (English)

    1997-01-01

    970359 CT diagnosis of pancreatic carcinoma andchronic pancreatitis. LUAN Baoqing(栾宝庆), et al,Dept Radiol, Beijing Friendship Hosp, Capital Med U-niv, Beijing, 100050. Chin J Radiol 1997; 31(2): 114-118. Objective: To improve the diagnostic accuracy ofpancreatic carcinoma and chronic pancreatitis. Materi-

  12. Generation of glucose-responsive functional islets with a three-dimensional structure from mouse fetal pancreatic cells and iPS cells in vitro.

    Directory of Open Access Journals (Sweden)

    Hiroki Saito

    Full Text Available Islets of Langerhans are a pancreatic endocrine compartment consisting of insulin-producing β cells together with several other hormone-producing cells. While some insulin-producing cells or immature pancreatic cells have been generated in vitro from ES and iPS cells, islets with proper functions and a three-dimensional (3D structure have never been successfully produced. To test whether islets can be formed in vitro, we first examined the potential of mouse fetal pancreatic cells. We found that E16.5 pancreatic cells, just before forming islets, were able to develop cell aggregates consisting of β cells surrounded by glucagon-producing α cells, a structure similar to murine adult islets. Moreover, the transplantation of these cells improved blood glucose levels in hyperglycemic mice. These results indicate that functional islets are formed in vitro from fetal pancreatic cells at a specific developmental stage. By adopting these culture conditions to the differentiation of mouse iPS cells, we developed a two-step system to generate islets, i.e. immature pancreatic cells were first produced from iPS cells, and then transferred to culture conditions that allowed the formation of islets from fetal pancreatic cells. The islets exhibited distinct 3D structural features similar to adult pancreatic islets and secreted insulin in response to glucose concentrations. Transplantation of the islets improved blood glucose levels in hyperglycemic mice. In conclusion, the two-step culture system allows the generation of functional islets with a 3D structure from iPS cells.

  13. Pancreatitis

    Institute of Scientific and Technical Information of China (English)

    1995-01-01

    950347 Pancreatic endorcine response to parenteralnutrition in experimental acute pancreatitis.SUN Xi-aoguang(孙晓光),et al.Dept Nucl Med,ZhongshanHosp,Shanghai Med Univ,Shanghai.Shanghai Med J1995;18(2),74-70.In order to study the pancreatic endocrine responseto parenteral nutrition (PN) in acute pancreatitis,thedisease was induced in dogs by injecting 4% tauro-cholate sodium 0.5ml/kg plus trypsin 0.5mg/kg into the pancreatic duct.Intravenous infusion of PN wasinitiated one hour after the establishment of the dis-

  14. Nestin-positive progenitor cells isolated from human fetal pancreas have phenotypic markers identical to mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    Ling Zhang; Tian-Pei Hong; Jiang Hu; Yi-Nan Liu; Yong-Hua Wu; Ling-Song Li

    2005-01-01

    AIM: To isolate nestin-positive progenitor cells from human fetal pancreas and to detect their surface markers and their capability of proliferation and differentiation into pancreatic islet endocrine cells in vitro.METHODS: Islet-like cell clusters (ICCs) were isolated from human fetal pancreas by using collagenase digestion. The free-floating ICCs were handpicked and cultured in a new dish. After the ICCs developed into monolayer epithelium-like cells, they were passaged and induced for differentiation. Reverse transcription polymerase chain reaction (RT-PCR), immunofluorescence stain, fluorescenceactivated cell sorting (FACS) and radioimmunoassay (RIA)were used to detect the expression of cell markers. RESULTS: (1) The monolayer epithelium-like cells had highly proliferative potential and could be passaged more than 16 timesin vitro; (2) RT-PCR analysis and immunofluorescence stain showed that these cells expressed both nestin and ABCG2, two of stem cellmarkers; (3) FACS analysis revealed that CD44, CD90and CD147 were positive, whereas CD34, CD38, CD45, CD71, CD117, CD133 and HLA-DR were negative on the nestin-positive cells; (4) RT-PCR analysis showed that the mRNA expression of insulin, glucagon and pancreaticduodenal homeobox gene-1 was detected, whereas the expression of nestin and neurogenin 3 disappeared in these cells treated with serum-free media supplemented with the cocktail of growth factors. Furthermore, the intracellular insulin content was detected by RIA after the induction culture.CONCLUSION: Nestin-positive cells isolated from human fetal pancreas possess the characteristics of pancreatic progenitor cells since they have highly proliferative potential and the capability of differentiation into insulinproducing cells in vitro. Interestingly, the nestin-positive pancreatic progenitor cells share many phenotypic markers with mesenchymal stem cells derived from bone marrow.

  15. Enabling research with human embryonic and fetal tissue resources

    Science.gov (United States)

    Gerrelli, Dianne; Lisgo, Steven; Copp, Andrew J.; Lindsay, Susan

    2015-01-01

    Summary Congenital anomalies are a significant burden on human health. Understanding the developmental origins of such anomalies is key to developing potential therapies. The Human Developmental Biology Resource (HDBR), based in London and Newcastle UK, was established to provide embryonic and fetal material for a variety of human studies ranging from single gene expression analysis to large scale genomic/transcriptomic studies. Increasingly HDBR material is enabling the derivation of stem cell lines and contributing towards developments in tissue engineering. Use of the HDBR and other fetal tissue resources discussed here will contribute to the long term aims of understanding the causation and pathogenesis of congenital anomalies, and developing new methods for their treatment and prevention. PMID:26395135

  16. Human embryonic stem cell-derived pancreatic endoderm alleviates diabetic pathology and improves reproductive outcome in C57BL/KsJ-Lep(db/+) gestational diabetes mellitus mice.

    Science.gov (United States)

    Xing, Baoheng; Wang, Lili; Li, Qin; Cao, Yalei; Dong, Xiujuan; Liang, Jun; Wu, Xiaohua

    2015-07-01

    Gestational diabetes mellitus is a condition commonly encountered during mid to late pregnancy with pathologic manifestations including hyperglycemia, hyperinsulinemia, insulin resistance, and fetal maldevelopment. The cause of gestational diabetes mellitus can be attributed to both genetic and environmental factors, hence complicating its diagnosis and treatment. Pancreatic progenitors derived from human embryonic stem cells were shown to be able to effectively treat diabetes in mice. In this study, we have developed a system of treating diabetes using human embryonic stem cell-derived pancreatic endoderm in a mouse model of gestational diabetes mellitus. Human embryonic stem cells were differentiated in vitro into pancreatic endoderm, which were then transplanted into db/+ mice suffering from gestational diabetes mellitus. The transplant greatly improved glucose metabolism and reproductive outcome of the females compared with the control groups. Our findings support the feasibility of using differentiated human embryonic stem cells for treating gestational diabetes mellitus patients.

  17. Prognostic value of metastin expression in human pancreatic cancer

    OpenAIRE

    Kawaguchi Yoshiya; Masui Toshihiko; Koizumi Masayuki; Kida Atsushi; Ito Tatsuo; Katagiri Fumihiko; Doi Ryuichiro; Nagai Kazuyuki; Tomita Kenji; Oishi Shinya; Fujii Nobutaka; Uemoto Shinji

    2009-01-01

    Abstract Background KiSS-1 was identified as a metastasis-suppressing gene in melanoma cells. The KiSS-1 gene product (metastin) was isolated from human placenta as the ligand of GPR54, a G-protein-coupled receptor. The role of metastin and GPR54 in tumor progression is not fully understood. Methods We investigated the clinical significance of metastin and GPR54 expression in pancreatic cancer. We evaluated immunohistochemical expression of metastin and GPR54 in pancreatic ductal adenocarcino...

  18. Prognostic value of metastin expression in human pancreatic cancer

    OpenAIRE

    Nagai, Kazuyuki; Doi, Ryuichiro; Katagiri, Fumihiko; Ito, Tatsuo; Kida, Atsushi; Koizumi, Masayuki; Masui, Toshihiko; Kawaguchi, Yoshiya; Tomita, Kenji; Oishi, Shinya; Fujii, Nobutaka; Uemoto, Shinji

    2009-01-01

    Background KiSS-1 was identified as a metastasis-suppressing gene in melanoma cells. The KiSS-1 gene product (metastin) was isolated from human placenta as the ligand of GPR54, a G-protein-coupled receptor. The role of metastin and GPR54 in tumor progression is not fully understood. Methods We investigated the clinical significance of metastin and GPR54 expression in pancreatic cancer. We evaluated immunohistochemical expression of metastin and GPR54 in pancreatic ductal adenocarcinoma tissue...

  19. Prognostic value of metastin expression in human pancreatic cancer.

    OpenAIRE

    Nagai, Kazuyuki; Doi, Ryuichiro; Katagiri, Fumihiko; Ito, Tatsuo; Kida, Atsushi; Koizumi, Masayuki; Masui, Toshihiko; Kawaguchi, Yoshiya; Tomita, Kenji; Oishi, Shinya; Fujii, Nobutaka; Uemoto, Shinji

    2009-01-01

    [Background]KiSS-1 was identified as a metastasis-suppressing gene in melanoma cells. The KiSS-1 gene product (metastin) was isolated from human placenta as the ligand of GPR54, a G-protein-coupled receptor. The role of metastin and GPR54 in tumor progression is not fully understood. [Methods]We investigated the clinical significance of metastin and GPR54 expression in pancreatic cancer. We evaluated immunohistochemical expression of metastin and GPR54 in pancreatic ductal adenocarcinoma tiss...

  20. Perivascular mesenchymal progenitors in human fetal and adult liver.

    Science.gov (United States)

    Gerlach, Jörg C; Over, Patrick; Turner, Morris E; Thompson, Robert L; Foka, Hubert G; Chen, William C W; Péault, Bruno; Gridelli, Bruno; Schmelzer, Eva

    2012-12-10

    The presence of mesenchymal stem cells (MSCs) has been described in various organs. Pericytes possess a multilineage differentiation potential and have been suggested to be one of the developmental sources for MSCs. In human liver, pericytes have not been defined. Here, we describe the identification, purification, and characterization of pericytes in human adult and fetal liver. Flow cytometry sorting revealed that human adult and fetal liver contains 0.56%±0.81% and 0.45%±0.39% of CD146(+)CD45(-)CD56(-)CD34(-) pericytes, respectively. Of these, 41% (adult) and 30% (fetal) were alkaline phosphatase-positive (ALP(+)). In situ, pericytes were localized around periportal blood vessels and were positive for NG2 and vimentin. Purified pericytes could be cultured extensively and had low population doubling times. Immunofluorescence of cultures demonstrated that cells were positive for pericyte and mesenchymal cell markers CD146, NG2, CD90, CD140b, and vimentin, and negative for endothelial, hematopoietic, stellate, muscle, or liver epithelial cell markers von Willebrand factor, CD31, CD34, CD45, CD144, CD326, CK19, albumin, α-fetoprotein, CYP3A7, glial fibrillary acid protein, MYF5, and Pax7 by gene expression; myogenin and alpha-smooth muscle actin expression were variable. Fluorescence-activated cell sorting analysis of cultures confirmed surface expression of CD146, CD73, CD90, CD10, CD13, CD44, CD105, and ALP and absence of human leukocyte antigen-DR. In vitro differentiation assays demonstrated that cells possessed robust osteogenic and myogenic, but low adipogenic and low chondrogenic differentiation potentials. In functional in vitro assays, cells had typical mesenchymal strong migratory and invasive activity. In conclusion, human adult and fetal livers harbor pericytes that are similar to those found in other organs and are distinct from hepatic stellate cells.

  1. Identification of insulin in the human pancreatic juice.

    Science.gov (United States)

    Ishii, H; Sato, K; Murozono, S

    1986-12-01

    In this study, we purified insulin-like substance (ILS) in the human pancreatic juice by the combined use of affinity chromatography and radioimmunoassay (RIA). The amino acid sequence of ILS in the N-terminal region is the same as that of human insulin. The influence of the enzymes present in the pancreatic juice on the RIA procedure, was examined. Trypsin, chymotrypsin and amylase showed steep influences on radioactivity. The addition of enzyme inhibitors could not reduce pseudo-activity, but the elimination of enzymes in the pancreatic juice by ultrafiltration with the Mole-Cut (Millipore, Japan) resulted in a lowering of the pseudo-insulin activity. Affinity chromatography on Sepharose 4B coupled with anti-porcine insulin was used to capture ILS. ILS was eluted by 1 M acetic acid from the affinity column monitoring pH and the insulin activity by RIA. The amino acid sequences of two components of ILS in amino terminal region were Phe-Val and Gly-Ile-Val. This indicates that ILS obtained from human pancreatic juice was the insulin derived from endocrine secretion of pancreas.

  2. Perinatal Oxidative Stress May Affect Fetal Ghrelin Levels in Humans

    OpenAIRE

    Zhong-Cheng Luo; Jean-François Bilodeau; Anne Monique Nuyt; Fraser, William D; Pierre Julien; Francois Audibert; Lin Xiao; Carole Garofalo; Emile Levy

    2015-01-01

    In vitro cell model studies have shown that oxidative stress may affect beta-cell function. It is unknown whether oxidative stress may affect metabolic health in human fetuses/newborns. In a singleton pregnancy cohort (n = 248), we studied maternal (24–28 weeks gestation) and cord plasma biomarkers of oxidative stress [malondialdehyde (MDA), F2-isoprostanes] in relation to fetal metabolic health biomarkers including cord plasma glucose-to-insulin ratio (an indicator of insulin sensitivity), p...

  3. Schwann cell cultures from human fetal dorsal root ganglia

    Institute of Scientific and Technical Information of China (English)

    Yaping Feng; Hui Zhu; Jiang Hao; Xinmin Wang; Shengping Wu; Li Bai; Xiangming Li; Yun Zha

    2009-01-01

    BACKGROUND:Previous studies have used many methods for in vitro Schwann cells (SCs) cul-tures and purification,such as single cell suspension and cytosine arabinoside.However,it has been difficult to obtain sufficient cellular density,and the procedures have been quite tedious.OBJECTIVE:To investigate the feasibility of culturing high-density SCs using fetal human dorsal root ganglion tissue explants.DESIGN,TIME AND SETTING:Cell culture and immunohistochemistry were performed at the Cen-tral Laboratory of Kunming General Hospital of Chinese PLA between March 2001 and October 2008.MATERIALS:Culture media containing 10% fetal bovine serum,as well as 0.2% collagenase and 0.25% trypsin were purchased from Gibco,USA;mouse anti-human S-100 monoclonal antibody and goat anti-mouse IgG labeled with horseradish peroxidase were provided by Beijing Institute of Bi-ological Products,China.METHODS:Primarily cultured SCs were dissociated from dorsal root ganglia of human aborted fe-tuses at 4-6 months pregnancy.Following removal of the dorsal root ganglion perineurium,the gan-glia were dissected into tiny pieces and digested with 0.2% collagenase and 0.25% trypsin (volume ratio 1:1),then explanted and cultured.SC purification was performed with 5 mL 10% fetal bovine serum added to the culture media,followed by differential adhesion.MAIN OUTCOME MEASURES:SCs morphology was observed under inverted phase contrast light microscopy.SC purity was evaluated according to percentage of S-100 immunostained cells.RESULTS:SCs were primarily cultured for 5-6 days and then subcultured for 4-5 passages.The highly enriched SC population reached > 95% purity and presented with normal morphology.CONCLUSION:A high purity of SCs was obtained with culture methods using human fetal dorsal root ganglion tissue explants.

  4. Changes of multiple biotransformation phase Ⅰ and phase Ⅱ enzyme activities in human fetal adrenals during fetal development

    Institute of Scientific and Technical Information of China (English)

    Hui WANG; Jie PING; Ren-xiu PENG; Jiang YUE; Xue-yan XIA; Qi-xiong LI; Rui KONG; Jun-yan HONG

    2008-01-01

    Aim: Fetal adrenal, which synthesizes steroid hormones, is critical to fetal growth and development. Our recent research showed that some xenobiotics could inter-fere with steroidogenesis and induce intrauterine growth retardation in rats. The study on the characteristics of biotransformation enzymes in fetal adrenals still seems to be important with respect to possible significance in xenobiotic-induced fetal development toxicity. In this study, the activities of several important xenobiotic-related phase Ⅰ and phase Ⅱ enzymes in human fetal adrenals were examined and compared with those in fetal livers. Methods: The activity and mRNA expression were determined by enzymatic analysis and RT-PCR. Results: The levels of cytochrome (CYP)2A6, CYP2E1, and CYP3A7 isozymes in fetal adrenals were 82%, 92%, and 33% of those in fetal livers, respectively. There was a good positive correlation between adrenal CYP2A6 activity and gestational time. The values of α glutathione S-transferase (GST), πGST, and μGST in adrenals were 0.5, 4.4, and 8.3-fold of those in the livers, respectively, and the activity of adrenal πGST was negatively correlated with gestational time. The uridine diphosphoglucuronyl transferase activities, which were measured using p-hydroxy-biphenyl and 7-hydroxy-4-methylcoumarin as substrates, were 9% and 3%, respectively, of those in the fetal livers. Conclusion: Our investiga-tion suggested that adrenal could be an important xenobiotic-metabolizing or-gan in fetal development and may play a potential role in xenobiotic-induced fetal development toxicity.

  5. Ibuprofen results in alterations of human fetal testis development

    Science.gov (United States)

    Ben Maamar, Millissia; Lesné, Laurianne; Hennig, Kristin; Desdoits-Lethimonier, Christèle; Kilcoyne, Karen R.; Coiffec, Isabelle; Rolland, Antoine D.; Chevrier, Cécile; Kristensen, David M.; Lavoué, Vincent; Antignac, Jean-Philippe; Le Bizec, Bruno; Dejucq-Rainsford, Nathalie; Mitchell, Rod T.; Mazaud-Guittot, Séverine; Jégou, Bernard

    2017-01-01

    Among pregnant women ibuprofen is one of the most frequently used pharmaceutical compounds with up to 28% reporting use. Regardless of this, it remains unknown whether ibuprofen could act as an endocrine disruptor as reported for fellow analgesics paracetamol and aspirin. To investigate this, we exposed human fetal testes (7–17 gestational weeks (GW)) to ibuprofen using ex vivo culture and xenograft systems. Ibuprofen suppressed testosterone and Leydig cell hormone INSL3 during culture of 8–9 GW fetal testes with concomitant reduction in expression of the steroidogenic enzymes CYP11A1, CYP17A1 and HSD17B3, and of INSL3. Testosterone was not suppressed in testes from fetuses younger than 8 GW, older than 10–12 GW, or in second trimester xenografted testes (14–17 GW). Ex vivo, ibuprofen also affected Sertoli cell by suppressing AMH production and mRNA expression of AMH, SOX9, DHH, and COL2A1. While PGE2 production was suppressed by ibuprofen, PGD2 production was not. Germ cell transcripts POU5F1, TFAP2C, LIN28A, ALPP and KIT were also reduced by ibuprofen. We conclude that, at concentrations relevant to human exposure and within a particular narrow ‘early window’ of sensitivity within first trimester, ibuprofen causes direct endocrine disturbances in the human fetal testis and alteration of the germ cell biology. PMID:28281692

  6. DNA Methylation Landscapes of Human Fetal Development

    NARCIS (Netherlands)

    Slieker, Roderick C.; Roost, Matthias S.; van Iperen, Liesbeth; Suchiman, H. Eka D; Tobi, Elmar W.; Carlotti, Françoise; de Koning, Eelco J P; Slagboom, P. Eline; Heijmans, Bastiaan T.; Chuva de Sousa Lopes, Susana M.

    2015-01-01

    Remodelling the methylome is a hallmark of mammalian development and cell differentiation. However, current knowledge of DNA methylation dynamics in human tissue specification and organ development largely stems from the extrapolation of studies in vitro and animal models. Here, we report on the DNA

  7. Impact of global Fxr deficiency on experimental acute pancreatitis and genetic variation in the FXR locus in human acute pancreatitis.

    Directory of Open Access Journals (Sweden)

    Rian M Nijmeijer

    Full Text Available Infectious complications often occur in acute pancreatitis, related to impaired intestinal barrier function, with prolonged disease course and even mortality as a result. The bile salt nuclear receptor farnesoid X receptor (FXR, which is expressed in the ileum, liver and other organs including the pancreas, exhibits anti-inflammatory effects by inhibiting NF-κB activation and is implicated in maintaining intestinal barrier integrity and preventing bacterial overgrowth and translocation. Here we explore, with the aid of complementary animal and human experiments, the potential role of FXR in acute pancreatitis.Experimental acute pancreatitis was induced using the CCK-analogue cerulein in wild-type and Fxr-/- mice. Severity of acute pancreatitis was assessed using histology and a semi-quantitative scoring system. Ileal permeability was analyzed in vitro by Ussing chambers and an in vivo permeability assay. Gene expression of Fxr and Fxr target genes was studied by quantitative RT-PCR. Serum FGF19 levels were determined by ELISA in acute pancreatitis patients and healthy volunteers. A genetic association study in 387 acute pancreatitis patients and 853 controls was performed using 9 tagging single nucleotide polymorphisms (SNPs covering the complete FXR gene and two additional functional SNPs.In wild-type mice with acute pancreatitis, ileal transepithelial resistance was reduced and ileal mRNA expression of Fxr target genes Fgf15, SHP, and IBABP was decreased. Nevertheless, Fxr-/- mice did not exhibit a more severe acute pancreatitis than wild-type mice. In patients with acute pancreatitis, FGF19 levels were lower than in controls. However, there were no associations of FXR SNPs or haplotypes with susceptibility to acute pancreatitis, or its course, outcome or etiology.We found no evidence for a major role of FXR in acute human or murine pancreatitis. The observed altered Fxr activity during the course of disease may be a secondary phenomenon.

  8. Efficient Generation of Glucose-Responsive Beta Cells from Isolated GP2+ Human Pancreatic Progenitors

    DEFF Research Database (Denmark)

    Ameri, Jacqueline; Borup, Rehannah; Prawiro, Christy

    2017-01-01

    cell manufacturing. Comparative gene expression analysis revealed glycoprotein 2 (GP2) as a specific cell surface marker for isolating pancreatic endoderm cells (PECs) from differentiated hESCs and human fetal pancreas. Isolated GP2+ PECs efficiently differentiated into glucose responsive insulin......-producing cells in vitro. We found that in vitro PEC proliferation declines due to enhanced expression of the cyclin-dependent kinase (CDK) inhibitors CDKN1A and CDKN2A. However, we identified a time window when reducing CDKN1A or CDKN2A expression increased proliferation and yield of GP2+ PECs. Altogether, our...... results contribute tools and concepts toward the isolation and use of PECs as a source for the safe production of hPSC-derived β cells....

  9. Prognostic value of metastin expression in human pancreatic cancer

    Directory of Open Access Journals (Sweden)

    Kawaguchi Yoshiya

    2009-01-01

    Full Text Available Abstract Background KiSS-1 was identified as a metastasis-suppressing gene in melanoma cells. The KiSS-1 gene product (metastin was isolated from human placenta as the ligand of GPR54, a G-protein-coupled receptor. The role of metastin and GPR54 in tumor progression is not fully understood. Methods We investigated the clinical significance of metastin and GPR54 expression in pancreatic cancer. We evaluated immunohistochemical expression of metastin and GPR54 in pancreatic ductal adenocarcinoma tissues obtained from 53 consecutive patients who underwent resection between July 2003 and May 2007 at Kyoto University Hospital. In 23 consecutive patients, the plasma metastin level was measured before surgery by enzyme immunoassay. Results Strong immunohistochemical expression of metastin was detected in 13 tumors (24.5%, while strong expression of GPR54 was detected in 30 tumors (56.6%. Tumors that were negative for both metastin and GPR54 expression were significantly larger than tumors that were positive for either metastin or GPR54 (p = 0.047. Recurrence was less frequent in patients who had metastin-positive tumors compared with those who had metastin-negative tumors (38.5% versus 70.0%, p = 0.04. Strong expression of metastin and GPR54 was significantly correlated with longer survival (p = 0.02. Metastin expression by pancreatic cancer was an independent prognostic factor for longer survival (hazard ratio, 2.1; 95% confidence interval, 1.1–4.7; p = 0.03, and the patients with a high plasma metastin level (n = 6 did not die after surgical resection. Conclusion Strong expression of metastin and GPR54 by pancreatic cancer is associated with longer survival. Metastin expression is an independent prognostic factor for the survival of pancreatic cancer patients. The plasma metastin level could become a noninvasive prognostic factor for the assessment of pancreatic cancer.

  10. Organization of human hypothalamus in fetal development.

    Science.gov (United States)

    Koutcherov, Yuri; Mai, Jürgen K; Ashwell, Ken W S; Paxinos, George

    2002-05-13

    The organization of the human hypothalamus was studied in 33 brains aged from 9 weeks of gestation (w.g.) to newborn, using immunohistochemistry for parvalbumin, calbindin, calretinin, neuropeptide Y, neurophysin, growth-associated protein (GAP)-43, synaptophysin, and the glycoconjugate 3-fucosyl- N-acetyl-lactosamine. Developmental stages are described in relation to obstetric trimesters. The first trimester (morphogenetic periods 9-10 w.g. and 11-14 w.g.) is characterized by differentiating structures of the lateral hypothalamic zone, which give rise to the lateral hypothalamus (LH) and posterior hypothalamus. The PeF differentiates at 18 w.g. from LH neurons, which remain anchored in the perifornical position, whereas most of the LH cells are displaced laterally. A transient supramamillary nucleus was apparent at 14 w.g. but not after 16 w.g. As the ventromedial nucleus differentiated at 13-16 w.g., three principal parts, the ventrolateral part, the dorsomedial part, and the shell, were revealed by distribution of calbindin, calretinin, and GAP43 immunoreactivity. The second trimester (morphogenetic periods 15-17 w.g., 18-23 w.g., and 24-33 w.g.) is characterized by differentiation of the hypothalamic core, in which calbindin- positive neurons revealed the medial preoptic nucleus at 16 w.g. abutted laterally by the intermediate nucleus. The dorsomedial nucleus was clearly defined at 10 w.g. and consisted of compact and diffuse parts, an organization that was lost after 15 w.g. Differentiation of the medial mamillary body into lateral and medial was seen at 13-16 w.g. Late second trimester was marked by differentiation of periventricular zone structures, including suprachiasmatic, arcuate, and paraventricular nuclei. The subnuclear differentiation of these nuclei extends into the third trimester. The use of chemoarchitecture in the human fetus permitted the identification of interspecies nuclei homologies, which otherwise remain concealed in the cytoarchitecture.

  11. Efficient Generation of Glucose-Responsive Beta Cells from Isolated GP2+ Human Pancreatic Progenitors

    Directory of Open Access Journals (Sweden)

    Jacqueline Ameri

    2017-04-01

    Full Text Available Stem cell-based therapy for type 1 diabetes would benefit from implementation of a cell purification step at the pancreatic endoderm stage. This would increase the safety of the final cell product, allow the establishment of an intermediate-stage stem cell bank, and provide a means for upscaling β cell manufacturing. Comparative gene expression analysis revealed glycoprotein 2 (GP2 as a specific cell surface marker for isolating pancreatic endoderm cells (PECs from differentiated hESCs and human fetal pancreas. Isolated GP2+ PECs efficiently differentiated into glucose responsive insulin-producing cells in vitro. We found that in vitro PEC proliferation declines due to enhanced expression of the cyclin-dependent kinase (CDK inhibitors CDKN1A and CDKN2A. However, we identified a time window when reducing CDKN1A or CDKN2A expression increased proliferation and yield of GP2+ PECs. Altogether, our results contribute tools and concepts toward the isolation and use of PECs as a source for the safe production of hPSC-derived β cells.

  12. Cytokeratin expression in human fetal tongue and buccal mucosa

    Indian Academy of Sciences (India)

    M M Vaidya; Sharda S Sawant; Anita M Borges; N K Naresh; Manda C Purandare; A N Bhisey

    2000-09-01

    Expression of cytokeratins (CK), a subset of intermediate filament (IF) proteins in epithelia, is developmentally regulated. CK expression may also change after malignant transformation. Our earlier studies on CK expression in human oral tumours and pre-cancerous lesions have shown specific changes in CK expression. We analysed CK expression in human tongue and buccal mucosa (BM) in fetuses in the embryonic age group of 16 to 27 weeks using biochemical and immunohistochemical techniques to find out whether there is any similarity in CK expression in human oral squamous cell carcinomas (SCC) and fetal oral tissues. CK 1, 8 and 18 were detected in a majority of samples using both techniques. Our earlier studies had shown aberrant expression of CK 1 and 18 in many of the oral SCC and leukoplakias. Studies by immunohistochemistry showed that these different CK antigens were expressed in different cell layers. CK 1(2) were present in the stratified epithelial layers whereas CK 8 and 18 were restricted to glandular epithelium. Till 27 weeks of gestation, both tongue and BM expressed CK 1, 8 and 18 along with CK 6 and 16. Thus, fetal tissues showed some similarities in CK pattern with their respective SCC.

  13. Pancreatitis

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    2008318 Proteomics of hyperlipidemia-associated pancreatitis using differential gel electrophoresis and tandem mass spectrometry: experiment with rats. ZHANG Wei(张伟), et al. Dept Gastroenterol, Shanghai 1st Hosp, Shanghai Jiaotong Univ, Shanghai 200080. Natl Med J China 2008;88(16):1132-1131.Objective To analyze the injury mechanismof hyperlipidemia-associated acute pancreatitis utilizing pro-teomics.Methods Ten SD rats were fed with high fat feed to establish hyperlipidemic models,and 10 SD rats were fed with normal feed to be used as control group.

  14. Induction of pancreatic duct cells of neonatal rats into insulin-producing cells with fetal bovine serum: A natural protocol and its use for patch clamp experiments

    Institute of Scientific and Technical Information of China (English)

    San-Hua Leng; Fu-Er Lu

    2005-01-01

    AIM: To induce the pancreatic duct cells into endocrine cells with a new natural protocol for electrophysiological study.METHODS: The pancreatic duct cells of neonatal rats were isolated, cultured and induced into endocrine oells with 15% fetal bovine serum for a period of 20 d. During this period, insulin secretion, MTT value, and morphological change of neonatal and adult pancreatic islet cells were comparatively investigated. Pancreatic β-cells were identified by morphological and electrophysiological characteristics, while ATP sensitive potassium channels(KATP), voltage-dependent potassium channels (KV), and voltage-dependent calcium channels (KCA) in β-cells were identified by patch clamp technique.RESULTS: After incubation with fetal bovine serum, the neonatal duct cells budded out, changed from duct-like cells into islet clusters. In the first 4 d, MTT value and insulin secretion increased slowly (MTT value from 0.024±0.003 to0.028±0.003, insulin secretion from 2.6±0.6to 3.1±0.8 mIU/L). Then MTT value and insulin secretion increased quickly from d 5 to d 10 (MTT value from 0.028±0.003 to 0.052±0.008, insulin secretion from 3.1±0.8to 18.3±2.6 mIU/L), then reached high plateau (MTT value >0.052±0.008, insulin secretion >18.3±2.6 mIU/L).In contrast, for the isolated adult pancreatic islet cells,both insulin release and MTT value were stable in the first 4 d (MTT value from 0.029±0.01 to 0.031±0.011,insulin secretion from 13.9±3.1 to 14.3±3.3 mIU/L), but afterwards they reduced gradually (MTT value <0.031±0.011, insulin secretion <8.2±1.5 mIU/L), and the pancrearic islet cells became dispersed, broken or atrophied correspondingly. The differentiated neonatal cells were identified as pancreatic islet cells by dithizone staining method, and pancreatic β-cells were further identified by both morphological features and electrophysiological characteristics, i.e. the existence of recording currents from KATP KV, and KCA.CONCLUSION: Islet

  15. Crystalline structures in human pancreatic beta cell adenoma.

    Science.gov (United States)

    Mori, H; Kawai, T; Tanaka, T; Fujii, M; Takahashi, M; Miyashita, T

    1978-05-01

    An electron microscopic observation on a pancreatic tumor removed from a 34-year-old woman revealed the fine structural morphology of a functional beta cell adenoma. Characteristic PAS positive crystalline structures were frequently observed in the cytoplasm of the tumor cells. They were not bounded by a membrane and had a rectangular or irregular hexagonal shape. Highly regular patterns were seen as such as lattice or honeycomb and parallel ripple structures. They are similar to the Reinke's crystal or crystalline structures reported in human hepatocytes suffering from several different diseases and considered as a protein-carbohydrate complex. Occasionally, small paracrystalline structures appeared to indicate an immature type of these structures in the opaque fine fibrillar mass. Crystalline or paracrystalline structures were not detected in the normal pancreatic tissue removed with the tumor from the patient.

  16. Pancreatitis

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    2009216 Relation of inositol 1,4,5-trisphosphate with calcium metabolism in rats with severe acute pancreatitis.SHI Chengxian(石承先),et al.Dept Live Bili Pancre Surg,Guizhou Prov Hosp,Guiyang 550002.World Chin J Digestol,2009;17(6):598-601.

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

    Science.gov (United States)

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

    2003-06-01

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

  18. ULTRASTRUCTURAL STUDY OF THE HEMOPOIETIC MICROENVIRONMENT IN HUMAN FETAL SPLEEN

    Institute of Scientific and Technical Information of China (English)

    刘爱莲; 朱萍; 李学均; 张丽; 迟凤琴; 吴振铎; 宋雁南; 张亚坤

    1994-01-01

    Reciprocal interations between hemopoietic stromal cells and immature hemopoietic cells in human spleens obtained from 20 fetuses of 10-28 weeks gestation were observed by transmission electron microscopy and scanning electron microscopy.The close association of stromal cells with immature hemopoietic cells was confirmed under the electron microscope and a preasumptive HIM(Hemopoietic inductive mocroenvironment) was visualized.In regions of immature hemopoietice cell-reticular cell,endothelial cell,macrophage and interdigitating cell contact,some communicating structures were found between the plasma membranes of adjacent cells.moreovcer,the cytoplasm of these four stromal cells were full of various kinds of organelles.These results suggest that reticular cells,endothelial cells,macrophages and interdigitating cells are component parts of the HIM of human fetal spleen and that these cells have a nurturing function in relation to hemopoietic cells.

  19. An Insight into the Role of Human Pancreatic Lithostathine

    Directory of Open Access Journals (Sweden)

    Patard L

    2003-03-01

    Full Text Available Human lithostathine was initially isolated from pancreatic stones in patients with alcoholic calcifying chronic pancreatitis. It is secreted into the pancreatic juice where it was believed to inhibit stone formation. The N-terminal undecapeptide was assumed to play an important role in the mechanism, by adsorption to the crystal surface. Later, the role of lithostathine in calcite formation and growth was questioned, together with the associated mechanism of action. In particular, although lithostathine adsorbs on calcite crystal, this property does not now seem to be specific. Moreover, the N-terminal undecapeptide is not likely to have, by itself, the function of the entire protein. The different aspects of this controversy are reviewed and discussed, particularly in the light of recent structural biology. Comparative biological data now available allow us to draw an interesting parallel between lithostathine and other related proteins. Finally, lithostathine might affect stone formation and may also have another function which could be investigated in the other proteins belonging to the same structural family.

  20. Extracellular calcium sensing receptor in human pancreatic cells

    Science.gov (United States)

    Rácz, G Z; Kittel, Á; Riccardi, D; Case, R M; Elliott, A C; Varga, G

    2002-01-01

    Background and aims: The extracellular calcium sensing receptor (CaR) plays a key role in the calcium homeostatic system and is therefore widely expressed in tissues involved in calcium metabolism. However, the CaR has also been identified in other tissues where its role is less clear. We have investigated the presence of the CaR in the human pancreas. Methods: Messenger RNA for the CaR was detected by reverse transcription-polymerase chain reaction and the protein was localised by immunostaining. CaR function was assayed in Capan-1 cells by measuring intracellular calcium and [3H] thymidine incorporation. Results: The receptor was highly expressed in human pancreatic ducts. It was also expressed in exocrine acinar cells, in islets of Langerhans, and in intrapancreatic nerves and blood vessels. The CaR was expressed in both normal and neoplastic human tissue samples but was detected in only one of five ductal adenocarcinoma cells lines examined. Experiments on the CaR expressing adenocarcinoma cell line Capan-1 showed that the CaR was functional and was linked to mobilisation of intracellular calcium. Stimulation of the CaR reduced Capan-1 cell proliferation. Conclusions: We propose that the CaR may play multiple functional roles in the human pancreas. In particular, the CaR on the duct luminal membrane may monitor and regulate the Ca2+ concentration in pancreatic juice by triggering ductal electrolyte and fluid secretion. This could help to prevent precipitation of calcium salts in the duct lumen. The CaR may also help to regulate the proliferation of pancreatic ductal cells. PMID:12377811

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

    DEFF Research Database (Denmark)

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

    2008-01-01

    Hedgehog (Hh) signaling controls pancreatic development and homeostasis; aberrant Hh signaling is associated with several pancreatic diseases. Here we investigated the link between Hh signaling and primary cilia in the human developing pancreatic ducts and in cultures of human pancreatic duct...

  2. Human embryonic growth trajectories and associations with fetal growth and birthweight

    NARCIS (Netherlands)

    van Uitert, Evelyne M.; Exalto, Niek; Burton, Graham J.; Willemsen, Sten P.; Koning, Anton H. J.; Eilers, Paul H. C.; Laven, Joop S. E.; Steegers, Eric A. P.; Steegers-Theunissen, Regine P. M.

    2013-01-01

    How do human embryonic growth trajectories evolve in the first trimester, and is first-trimester embryonic growth associated with fetal growth and birthweight (BW)? Human embryonic growth rates increase between 9 and 10 weeks of gestation and are associated with mid-pregnancy fetal growth and BW. Fe

  3. Embryologic and Fetal Development of the Human Eyelid

    Science.gov (United States)

    Abdulhafez, Mohamed H.; Fouad, Yousef A.; Dutton, Jonathan J.

    2016-01-01

    Purpose: To review the recent data about eyelid morphogenesis, and outline a timeline for eyelid development from the very early stages during embryonic life till final maturation of the eyelid late in fetal life. Methods: The authors extensively review major studies detailing human embryologic and fetal eyelid morphogenesis. These studies span almost a century and include some more recent cadaver studies. Numerous studies in the murine model have helped to better understand the molecular signals that govern eyelid embryogenesis. The authors summarize the current findings in molecular biology, and highlight the most significant studies in mice regarding the multiple and interacting signaling pathways involved in regulating normal eyelid morphogenesis. Results: Eyelid morphogenesis involves a succession of subtle yet strictly regulated morphogenetic episodes of tissue folding, proliferation, contraction, and even migration, which may occur simultaneously or in succession. Conclusions: Understanding the extraordinary process of building eyelid tissue in embryonic life, and deciphering its underlying signaling machinery has far reaching clinical implications beyond understanding the developmental abnormalities involving the eyelids, and may pave the way for achieving scar-reducing therapies in adult mammalian wounds, or control the spread of malignancies. PMID:27124372

  4. TEAD and YAP regulate the enhancer network of human embryonic pancreatic progenitors.

    Science.gov (United States)

    Cebola, Inês; Rodríguez-Seguí, Santiago A; Cho, Candy H-H; Bessa, José; Rovira, Meritxell; Luengo, Mario; Chhatriwala, Mariya; Berry, Andrew; Ponsa-Cobas, Joan; Maestro, Miguel Angel; Jennings, Rachel E; Pasquali, Lorenzo; Morán, Ignasi; Castro, Natalia; Hanley, Neil A; Gomez-Skarmeta, Jose Luis; Vallier, Ludovic; Ferrer, Jorge

    2015-05-01

    The genomic regulatory programmes that underlie human organogenesis are poorly understood. Pancreas development, in particular, has pivotal implications for pancreatic regeneration, cancer and diabetes. We have now characterized the regulatory landscape of embryonic multipotent progenitor cells that give rise to all pancreatic epithelial lineages. Using human embryonic pancreas and embryonic-stem-cell-derived progenitors we identify stage-specific transcripts and associated enhancers, many of which are co-occupied by transcription factors that are essential for pancreas development. We further show that TEAD1, a Hippo signalling effector, is an integral component of the transcription factor combinatorial code of pancreatic progenitor enhancers. TEAD and its coactivator YAP activate key pancreatic signalling mediators and transcription factors, and regulate the expansion of pancreatic progenitors. This work therefore uncovers a central role for TEAD and YAP as signal-responsive regulators of multipotent pancreatic progenitors, and provides a resource for the study of embryonic development of the human pancreas.

  5. Clonal preservation of human pancreatic cell line derived from primary pancreatic adenocarcinoma.

    Science.gov (United States)

    Mohammad, R M; Li, Y; Mohamed, A N; Pettit, G R; Adsay, V; Vaitkevicius, V K; Al-Katib, A M; Sarkar, F H

    1999-11-01

    Adenocarcinoma of the pancreas generally remains an incurable disease by available treatment modalities, demanding the development of a suitable cell-culture/animal model and the discovery and evaluation of novel therapeutic agents. We report the clonal preservation of a human pancreatic cell line (KCI-MOH1) established from a 74-year-old African-American man diagnosed with pancreatic cancer. Initially the human primary tumor was grown as a xenograft in SCID mice and, subsequently, a cell line was established from tumors grown as a xenograft as reported in our earlier publication. The molecular characterization of the primary tumor, the tumors grown as xenograft, and the cell line all revealed similar genotypic properties. By using an automated DNA sequencer, a K-ras mutation (codon 12, GGT to CGT, Gly to Arg) was detected in the pancreatic tumor tissue taken from the patient, whereas no p53 mutation was detected. The same K-ras mutation and unaltered p53 was also found in the xenograft tumor and in the KCI-MOH1 cell line. Chromosome analysis of the cultured cells revealed: 42,XY,add(3)(p11.2),der(7)t(7;12) (p22;q12),-10,-12,add (14)(p11),-18,add (20)(q13),-22/84, idemx2, which is the same chromosome complement found in xenograft tumors. The KCI-MOH1 cell line grows well in tissue culture and forms tumors in the SCID mice when implanted subcutaneously, as well as in orthotopic sites. The KCI-MOH1 cell line-derived SCID mouse xenograft model was used for efficacy evaluation of bryostatin 1, auristatin-PE, spongistatin 1, and gemcitabine alone and in combination. Tumor growth inhibition (T/C expressed as percentage), tumor growth delay (T - C), and log 10 kill for these agents were 38%, 22 days, and 0.53; 15%, 30 days, and 0.80; 24%, 25 days, and 0.66; and 10%, 33 days, and 0.90, respectively. When given in combination, two of seven gemcitabine + auristatin-PE-treated animals were free of tumors for 150 days and were considered cured. Animals treated with a

  6. Arterial flow regulator enables transplantation and growth of human fetal kidneys in rats.

    Science.gov (United States)

    Chang, N K; Gu, J; Gu, S; Osorio, R W; Concepcion, W; Gu, E

    2015-06-01

    Here we introduce a novel method of transplanting human fetal kidneys into adult rats. To overcome the technical challenges of fetal-to-adult organ transplantation, we devised an arterial flow regulator (AFR), consisting of a volume adjustable saline-filled cuff, which enables low-pressure human fetal kidneys to be transplanted into high-pressure adult rat hosts. By incrementally withdrawing saline from the AFR over time, blood flow entering the human fetal kidney was gradually increased until full blood flow was restored 30 days after transplantation. Human fetal kidneys were shown to dramatically increase in size and function. Moreover, rats which had all native renal mass removed 30 days after successful transplantation of the human fetal kidney were shown to have a mean survival time of 122 days compared to 3 days for control rats that underwent bilateral nephrectomy without a prior human fetal kidney transplant. These in vivo human fetal kidney models may serve as powerful platforms for drug testing and discovery.

  7. Ex-vivo evaluation of gene therapy vectors in human pancreatic (cancer) tissue slices

    NARCIS (Netherlands)

    van Geer, M.A.; Kuhlmann, K.F.D.; Bakker, C.T.; ten Kate, F.J.W.; Oude Elferink, R.P.J.; Bosma, P.J.

    2009-01-01

    AIM: To culture human pancreatic tissue obtained from small resection specimens as a pre-clinical model for examining virus-host interactions. METHODS: Human pancreatic tissue samples (malignant and normal) were obtained from surgical specimens and processed immediately to tissue slices. Tissue slic

  8. Ex-vivo evaluation of gene therapy vectors in human pancreatic (cancer) tissue slices

    NARCIS (Netherlands)

    van Geer, M.A.; Kuhlmann, K.F.D.; Bakker, C.T.; ten Kate, F.J.W.; Oude Elferink, R.P.J.; Bosma, P.J.

    2009-01-01

    AIM: To culture human pancreatic tissue obtained from small resection specimens as a pre-clinical model for examining virus-host interactions. METHODS: Human pancreatic tissue samples (malignant and normal) were obtained from surgical specimens and processed immediately to tissue slices. Tissue slic

  9. Fetal developmental programing: insights from human studies and experimental models.

    Science.gov (United States)

    Lopes, Gisele Aparecida Dionísio; Ribeiro, Vinícius Luís Bertotti; Barbisan, Luís Fernando; Marchesan Rodrigues, Maria Aparecida

    2017-03-01

    Environmental factors, particularly nutrition during pregnancy and early life can influence the risk of chronic diseases in later life. The underlying mechanism, termed "programing", postulates that an environmental stimulus during a critical window of time, early in life, has a permanent effect on subsequent structure and function of the organism. In this study we review the concept of fetal programing on chronic diseases and the proposed hypotheses for the association between early development and later disease, including epigenetic variation. We concentrate on specific aspects of maternal nutrition, particularly under-nutrition and over-nutrition, in humans and animal models. An adequate maternal nutrition during pregnancy is crucial for the health outcome of the offspring at adulthood.

  10. Trisomy 21 enhances human fetal erythro-megakaryocytic development

    Science.gov (United States)

    Chou, Stella T.; Opalinska, Joanna B.; Yao, Yu; Fernandes, Myriam A.; Kalota, Anna; Brooks, John S. J.; Choi, John K.; Gewirtz, Alan M.; Danet-Desnoyers, Gwenn-ael; Nemiroff, Richard L.

    2008-01-01

    Children with Down syndrome exhibit 2 related hematopoietic diseases: transient myeloproliferative disorder (TMD) and acute megakaryoblastic leukemia (AMKL). Both exhibit clonal expansion of blasts with biphenotypic erythroid and megakaryocytic features and contain somatic GATA1 mutations. While altered GATA1 inhibits erythro-megakaryocytic development, less is known about how trisomy 21 impacts blood formation, particularly in the human fetus where TMD and AMKL originate. We used in vitro and mouse transplantation assays to study hematopoiesis in trisomy 21 fetal livers with normal GATA1 alleles. Remarkably, trisomy 21 progenitors exhibited enhanced production of erythroid and megakaryocytic cells that proliferated excessively. Our findings indicate that trisomy 21 itself is associated with cell-autonomous expansion of erythro-megakaryocytic progenitors. This may predispose to TMD and AMKL by increasing the pool of cells susceptible to malignant transformation through acquired mutations in GATA1 and other cooperating genes. PMID:18812473

  11. Heme oxygenase-1 accelerates tumor angiogenesis of human pancreatic cancer.

    Science.gov (United States)

    Sunamura, Makoto; Duda, Dan G; Ghattas, Maivel H; Lozonschi, Lucian; Motoi, Fuyuhiko; Yamauchi, Jun-Ichiro; Matsuno, Seiki; Shibahara, Shigeki; Abraham, Nader G

    2003-01-01

    Angiogenesis is necessary for the continued growth of solid tumors, invasion and metastasis. Several studies clearly showed that heme oxygenase-1 (HO-1) plays an important role in angiogenesis. In this study, we used the vital microscope system, transparent skinfold model, lung colonization model and transduced pancreatic cancer cell line (Panc-1)/human heme oxygenase-1 (hHO-1) cells, to precisely analyze, for the first time, the effect of hHO-1 gene on tumor growth, angiogenesis and metastasis. Our results revealed that HO-1 stimulates angiogenesis of pancreatic carcinoma in severe combined immune deficient mice. Overexpression of human hHO-1 after its retroviral transfer into Panc-1 cells did not interfere with tumor growth in vitro. While in vivo the development of tumors was accelerated upon transfection with hHO-1. On the other hand, inhibition of heme oxygenase (HO) activity by stannous mesoporphyrin was able transiently to delay tumor growth in a dose dependent manner. Tumor angiogenesis was markedly increased in Panc-1/hHO-1 compared to mock transfected and wild type. Lectin staining and Ki-67 proliferation index confirmed these results. In addition hHO-1 stimulated in vitro tumor angiogenesis and increased endothelial cell survival. In a lung colonization model, overexpression of hHO-1 increased the occurrence of metastasis, while inhibition of HO activity by stannous mesoporphyrin completely inhibited the occurrence of metastasis. In conclusion, overexpression of HO-1 genes potentiates pancreatic cancer aggressiveness, by increasing tumor growth, angiogenesis and metastasis and that the inhibition of the HO system may be of useful benefit for the future treatment of the disease.

  12. Influence of nutrient restriction and melatonin supplementation of pregnant ewes on maternal and fetal pancreatic digestive enzymes and insulin-containing clusters.

    Science.gov (United States)

    Keomanivong, F E; Lemley, C O; Camacho, L E; Yunusova, R; Borowicz, P P; Caton, J S; Meyer, A M; Vonnahme, K A; Swanson, K C

    2016-03-01

    Primiparous ewes (n=32) were assigned to dietary treatments in a 2×2 factorial arrangement to determine effects of nutrient restriction and melatonin supplementation on maternal and fetal pancreatic weight, digestive enzyme activity, concentration of insulin-containing clusters and plasma insulin concentrations. Treatments consisted of nutrient intake with 60% (RES) or 100% (ADQ) of requirements and melatonin supplementation at 0 (CON) or 5 mg/day (MEL). Treatments began on day 50 of gestation and continued until day 130. On day 130, blood was collected under general anesthesia from the uterine artery, uterine vein, umbilical artery and umbilical vein for plasma insulin analysis. Ewes were then euthanized and the pancreas removed from the ewe and fetus, trimmed of mesentery and fat, weighed and snap-frozen until enzyme analysis. In addition, samples of pancreatic tissue were fixed in 10% formalin solution for histological examination including quantitative characterization of size and distribution of insulin-containing cell clusters. Nutrient restriction decreased (P⩽0.001) maternal pancreatic mass (g) and α-amylase activity (U/g, kU/pancreas, U/kg BW). Ewes supplemented with melatonin had increased pancreatic mass (P=0.03) and α-amylase content (kU/pancreas and U/kg BW). Melatonin supplementation decreased (P=0.002) maternal pancreatic insulin-positive tissue area (relative to section of tissue), and size of the largest insulin-containing cell cluster (P=0.04). Nutrient restriction decreased pancreatic insulin-positive tissue area (P=0.03) and percent of large (32 001 to 512 000 µm2) and giant (⩾512 001 µm2) insulin-containing cell clusters (P=0.04) in the fetus. Insulin concentrations in plasma from the uterine vein, umbilical artery and umbilical vein were greater (P⩽0.01) in animals receiving 100% requirements. When comparing ewes to fetuses, ewes had a greater percentage of medium insulin-containing cell clusters (2001 to 32 000 µm2) while fetuses

  13. Human pancreatic triglyceride lipase expressed in yeast cells: purification and characterization.

    Science.gov (United States)

    Yang, Y; Lowe, M E

    1998-06-01

    A cDNA clone encoding human pancreatic triglyceride lipase was cloned into a yeast expression vector so that the yeast PHO1 signal peptide replaced the native signal peptide. Pichia pastoris cells were transfected with the vector, and clones expressing human pancreatic triglyceride lipase were isolated. Recombinant human pancreatic lipase was expressed in broth cultures and was purified from the medium by DEAE blue Sepharose and hydroxyapatite chromatography. The highly purified lipase had specific activities for various triglyceride substrates identical to those of tissue-purified human pancreatic triglyceride lipase; it was inhibited by bile salts, required colipase for activity, and demonstrated interfacial activation. This expression system is suitable for the rapid, efficient production of human pancreatic triglyceride lipase in amounts adequate for biophysical studies.

  14. Pancreatic duct replication is increased with obesity and type 2 diabetes in humans.

    Science.gov (United States)

    Butler, A E; Galasso, R; Matveyenko, A; Rizza, R A; Dry, S; Butler, P C

    2010-01-01

    In a high-fat-fed rat model of type 2 diabetes we noted increased exocrine duct replication. This is a predisposing factor for pancreatitis and pancreatic cancer, both of which are more common in type 2 diabetes. The aim of the study reported here was to establish if obesity and/or type 2 diabetes are associated with increased pancreatic ductal replication in humans. We obtained pancreas at autopsy from 45 humans, divided into four groups: lean (BMI obese (BMI >27 kg/m(2)); non-diabetic; and with type 2 diabetes. Pancreases were evaluated after immunostaining for the duct cell marker cytokeratin and Ki67 for replication. We show for the first time that both obesity and type 2 diabetes in humans are associated with increased pancreatic ductal replication. Specifically, we report that (1) replication of pancreatic duct cells is increased tenfold by obesity, and (2) lean subjects with type 2 diabetes demonstrate a fourfold increase in replication of pancreatic duct cells compared with their lean non-diabetic controls. Pancreatic duct cell replication is increased in humans in response to both obesity and type 2 diabetes, potentially providing a mechanism for the increased risk of pancreatitis and pancreatic cancer in those with obesity and/or type 2 diabetes.

  15. Retinoic Acid signalling and the control of meiotic entry in the human fetal gonad.

    Directory of Open Access Journals (Sweden)

    Andrew J Childs

    Full Text Available The development of mammalian fetal germ cells along oogenic or spermatogenic fate trajectories is dictated by signals from the surrounding gonadal environment. Germ cells in the fetal testis enter mitotic arrest, whilst those in the fetal ovary undergo sex-specific entry into meiosis, the initiation of which is thought to be mediated by selective exposure of fetal ovarian germ cells to mesonephros-derived retinoic acid (RA. Aspects of this model are hard to reconcile with the spatiotemporal pattern of germ cell differentiation in the human fetal ovary, however. We have therefore examined the expression of components of the RA synthesis, metabolism and signalling pathways, and their downstream effectors and inhibitors in germ cells around the time of the initiation of meiosis in the human fetal gonad. Expression of the three RA-synthesising enzymes, ALDH1A1, 2 and 3 in the fetal ovary and testis was equal to or greater than that in the mesonephros at 8-9 weeks gestation, indicating an intrinsic capacity within the gonad to synthesise RA. Using immunohistochemistry to detect RA receptors RARα, β and RXRα, we find germ cells to be the predominant target of RA signalling in the fetal human ovary, but also reveal widespread receptor nuclear localization indicative of signalling in the testis, suggesting that human fetal testicular germ cells are not efficiently shielded from RA by the action of the RA-metabolising enzyme CYP26B1. Consistent with this, expression of CYP26B1 was greater in the human fetal ovary than testis, although the sexually-dimorphic expression patterns of the germ cell-intrinsic regulators of meiotic initiation, STRA8 and NANOS2, appear conserved. Finally, we demonstrate that RA induces a two-fold increase in STRA8 expression in cultures of human fetal testis, but is not sufficient to cause widespread meiosis-associated gene expression. Together, these data indicate that while local production of RA within the fetal ovary may

  16. ABSORPTION-SPECTRA OF HUMAN FETAL AND ADULT OXYHEMOGLOBIN, DE-OXYHEMOGLOBIN, CARBOXYHEMOGLOBIN, AND METHEMOGLOBIN

    NARCIS (Netherlands)

    ZIJLSTRA, WG; MEEUWSENVANDERROEST, WP

    1991-01-01

    We determined the millimolar absorptivities of the four clinically relevant derivatives of fetal and adult human hemoglobin in the visible and near-infrared spectral range (450-1000 nm). As expected, spectral absorption curves of similar shape were found, but the small differences between fetal and

  17. Differential role of Hedgehog signaling in human pancreatic (patho-) physiology: An up to date review.

    Science.gov (United States)

    Klieser, Eckhard; Swierczynski, Stefan; Mayr, Christian; Jäger, Tarkan; Schmidt, Johanna; Neureiter, Daniel; Kiesslich, Tobias; Illig, Romana

    2016-05-15

    Since the discovery of the Hedgehog (Hh) pathway in drosophila melanogaster, our knowledge of the role of Hh in embryonic development, inflammation, and cancerogenesis in humans has dramatically increased over the last decades. This is the case especially concerning the pancreas, however, real therapeutic breakthroughs are missing until now. In general, Hh signaling is essential for pancreatic organogenesis, development, and tissue maturation. In the case of acute pancreatitis, Hh has a protective role, whereas in chronic pancreatitis, Hh interacts with pancreatic stellate cells, leading to destructive parenchym fibrosis and atrophy, as well as to irregular tissue remodeling with potency of initiating cancerogenesis. In vitro and in situ analysis of Hh in pancreatic cancer revealed that the Hh pathway participates in the development of pancreatic precursor lesions and ductal adenocarcinoma including critical interactions with the tumor microenvironment. The application of specific inhibitors of components of the Hh pathway is currently subject of ongoing clinical trials (phases 1 and 2). Furthermore, a combination of Hh pathway inhibitors and established chemotherapeutic drugs could also represent a promising therapeutic approach. In this review, we give a structured survey of the role of the Hh pathway in pancreatic development, pancreatitis, pancreatic carcinogenesis and pancreatic cancer as well as an overview of current clinical trials concerning Hh pathway inhibitors and pancreas cancer.

  18. Single-Cell Sequencing of Human Pancreatic Islets-New Kids on the Block.

    Science.gov (United States)

    Prasad, Rashmi B; Groop, Leif

    2016-10-11

    RNA sequencing of human pancreatic islets has provided important insights into the islet transcriptome but little information on the specific cells. In this issue, Segerstolpe et al. (2016) and Xin et al. (2016b) report on the transcriptome of single pancreatic cells from non-diabetic and type 2 diabetic donors. Copyright © 2016. Published by Elsevier Inc.

  19. Microarray-Based Differential Expression Monitoring of 79 Novel Genes in Human Fetal Tissues

    Institute of Scientific and Technical Information of China (English)

    Ma; Shu-hua; Wang; Dun-cheng; 等

    2003-01-01

    79 ESTs fragments with represents corresponding novel genes were obtained by sequencing and bioinformatics analysis of human fetal kidney cDNA library. Microarray was prepared by using these novel EST fragments by automatic spotting. Expression patters of 79 ESTs of novel genes from human fetal kidney were analyzed in fetal brain and fetal heart tissues of 20-week-and 26-week-age fetus by performing of cDNA chip hybridization. This provides clues for studying exact functions of the novel genes. 8 genes were obtained which were expressed differentially in the fetal brain and heart of 20-week-and 26-week-age respectively. Then differentially expressed genes were identified by Northern analysis. The more exact function of the novel genes is under study.

  20. Prolonged exposure to acetaminophen reduces testosterone production by the human fetal testis in a xenograft model

    DEFF Research Database (Denmark)

    van den Driesche, Sander; Macdonald, Joni; Anderson, Richard A

    2015-01-01

    Most common male reproductive disorders are linked to lower testosterone exposure in fetal life, although the factors responsible for suppressing fetal testosterone remain largely unknown. Protracted use of acetaminophen during pregnancy is associated with increased risk of cryptorchidism in sons......, but effects on fetal testosterone production have not been demonstrated. We used a validated xenograft model to expose human fetal testes to clinically relevant doses and regimens of acetaminophen. Exposure to a therapeutic dose of acetaminophen for 7 days significantly reduced plasma testosterone (45......% reduction; P = 0.025) and seminal vesicle weight (a biomarker of androgen exposure; 18% reduction; P = 0.005) in castrate host mice bearing human fetal testis xenografts, whereas acetaminophen exposure for just 1 day did not alter either parameter. Plasma acetaminophen concentrations (at 1 hour after...

  1. Myocardial bridges of the coronary arteries in the human fetal heart.

    Science.gov (United States)

    Cakmak, Yusuf Ozgür; Cavdar, Safiye; Yalin, Aymelek; Yener, Nuran; Ozdogmus, Omer

    2010-09-01

    During the last century, many investigators reported on myocardial bridges in the adult human heart. In the present study, 39 human fetal hearts (the mean gestastional age was 30 weeks) were studied for myocardial bridging, and the results were correlated with adult data. Among the 39 (27 male and 12 female) fetal hearts studied, 26 bridges were observed on 18 fetal hearts (46.2%). Ten of the bridges had one myocardial bridge, whereas double myocardial bridges were observed in eight fetal hearts. The most frequent myocardial bridges were observed on the left anterior descending artery (LAD), which had 13 bridges (50%). Eight (30.7%) myocardial bridges were on the diagonal artery, and on the posterior descending artery there were five (19.3%). Myocardial bridges were not observed on the circumflex artery. The data presented in this study may provide potentially useful information for the preoperative evaluation of the newborn and may have a clinical implication for sudden fetal death.

  2. Effects of insulin on human pancreatic cancer progression modeled in vitro.

    Science.gov (United States)

    Chan, Michelle T; Lim, Gareth E; Skovsø, Søs; Yang, Yu Hsuan Carol; Albrecht, Tobias; Alejandro, Emilyn U; Hoesli, Corinne A; Piret, James M; Warnock, Garth L; Johnson, James D

    2014-11-06

    Pancreatic adenocarcinoma is one of the most lethal cancers, yet it remains understudied and poorly understood. Hyperinsulinemia has been reported to be a risk factor of pancreatic cancer, and the rapid rise of hyperinsulinemia associated with obesity and type 2 diabetes foreshadows a rise in cancer incidence. However, the actions of insulin at the various stages of pancreatic cancer progression remain poorly defined. Here, we examined the effects of a range of insulin doses on signalling, proliferation and survival in three human cell models meant to represent three stages in pancreatic cancer progression: primary pancreatic duct cells, the HPDE immortalized pancreatic ductal cell line, and the PANC1 metastatic pancreatic cancer cell line. Cells were treated with a range of insulin doses, and their proliferation/viability were tracked via live cell imaging and XTT assays. Signal transduction was assessed through the AKT and ERK signalling pathways via immunoblotting. Inhibitors of AKT and ERK signalling were used to determine the relative contribution of these pathways to the survival of each cell model. While all three cell types responded to insulin, as indicated by phosphorylation of AKT and ERK, we found that there were stark differences in insulin-dependent proliferation, cell viability and cell survival among the cell types. High concentrations of insulin increased PANC1 and HPDE cell number, but did not alter primary duct cell proliferation in vitro. Cell survival was enhanced by insulin in both primary duct cells and HPDE cells. Moreover, we found that primary cells were more dependent on AKT signalling, while HPDE cells and PANC1 cells were more dependent on RAF/ERK signalling. Our data suggest that excessive insulin signalling may contribute to proliferation and survival in human immortalized pancreatic ductal cells and metastatic pancreatic cancer cells, but not in normal adult human pancreatic ductal cells. These data suggest that signalling pathways

  3. Culture and purification of human fetal olfactory bulb ensheathing cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective: To obtain high purity of human fetal olfactory bulb ensheathing cells (OB-hOECs) in vitro and to develop a simple and effective method for primary culture of OB-hOECs. Methods: OB-hOECs were cultured based on the differential rates of attachment of the various harvested cell types. Then the method was combined with arabinoside cytosine (Ara-C)inhibition, serum-free starvation or intermittent neurotrophin 3 (NT3) nutrition method to observe cell states in different cultural environments. The purity of OB-hOECs was assessed with immunocytochemical analysis. Results: OB-hOECs appeared bipolar and tripolar shape, with slender processes forming network. The purity of OECs reached 88% with the selective attachment method on day 6, and then fibroblast proliferated quickly and reduced the purity. When combined with the starvation method, the purity of OECs was 91% on day 6 and 86% on day 9, however, OECs were in a poor state. While combined with the NT3 method, the purity reached 95% on day 9 and 83% on day 12, respectively. The cells still remained in a good state. Conclusion: A combination of selective attachment and intermittent NT3 nutrition is an effective method to obtain OECs with higher purity and quality.

  4. Human Fetal Liver: An In Vitro Model of Erythropoiesis

    Directory of Open Access Journals (Sweden)

    Guillaume Pourcher

    2011-01-01

    Full Text Available We previously described the large-scale production of RBCs from hematopoietic stem cells (HSCs of diverse sources. Our present efforts are focused to produce RBCs thanks to an unlimited source of stem cells. Human embryonic stem (ES cells or induced pluripotent stem cell (iPS are the natural candidates. Even if the proof of RBCs production from these sources has been done, their amplification ability is to date not sufficient for a transfusion application. In this work, our protocol of RBC production was applied to HSC isolated from fetal liver (FL as an intermediate source between embryonic and adult stem cells. We studied the erythroid potential of FL-derived CD34+ cells. In this in vitro model, maturation that is enucleation reaches a lower level compared to adult sources as observed for embryonic or iP, but, interestingly, they (i displayed a dramatic in vitro expansion (100-fold more when compared to CB CD34+ and (ii 100% cloning efficiency in hematopoietic progenitor assays after 3 days of erythroid induction, as compared to 10–15% cloning efficiency for adult CD34+ cells. This work supports the idea that FL remains a model of study and is not a candidate for ex vivo RBCS production for blood transfusion as a direct source of stem cells but could be helpful to understand and enhance proliferation abilities for primitive cells such as ES cells or iPS.

  5. The human protooncogene product p33pim is expressed during fetal hematopoiesis and in diverse leukemias.

    OpenAIRE

    Amson, R; Sigaux, F; Przedborski, S; Flandrin, G; Givol, D; Telerman, A

    1989-01-01

    We measured the human pim-1 protooncogene (PIM) expression during fetal development and in hematopoietic malignancies. Our data indicate that during human fetal hematopoiesis the 33-kDa pim product, p33pim, is highly expressed in the liver and spleen. In contrast, at the adult stage it is only slightly expressed in circulating granulocytes. Out of 70 hematopoietic malignancies analyzed, 51 patients and 19 cell lines, p33pim was overexpressed in approximately 30% of the samples, particularly i...

  6. Blood flow in the human fetal descending aorta : a pulsed Doppler study

    OpenAIRE

    Pijpers, Leendert

    1985-01-01

    textabstractIn 1628 William Harvey introduced his concept ofthe human circulation. Although a lot of studies concerning the fetal circulation were done before it was not until the 1930s that Barcroft (1934, 1939) and associates performed radiograpbic studies on the feta! goal and lamb to establish the feta! circulation. Later in 1964 Lind, Stern and Wegelius used cine-angiographic studies to describe the human fetal circulation. Volume flow measurements were already carried out in 1884 by Coh...

  7. Human fetal cardiac progenitors: The role of stem cells and progenitors in the fetal and adult heart.

    Science.gov (United States)

    Bulatovic, Ivana; Månsson-Broberg, Agneta; Sylvén, Christer; Grinnemo, Karl-Henrik

    2016-02-01

    The human fetal heart is formed early during embryogenesis as a result of cell migrations, differentiation, and formative blood flow. It begins to beat around gestation day 22. Progenitor cells are derived from mesoderm (endocardium and myocardium), proepicardium (epicardium and coronary vessels), and neural crest (heart valves, outflow tract septation, and parasympathetic innervation). A variety of molecular disturbances in the factors regulating the specification and differentiation of these cells can cause congenital heart disease. This review explores the contribution of different cardiac progenitors to the embryonic heart development; the pathways and transcription factors guiding their expansion, migration, and functional differentiation; and the endogenous regenerative capacity of the adult heart including the plasticity of cardiomyocytes. Unfolding these mechanisms will become the basis for understanding the dynamics of specific congenital heart disease as well as a means to develop therapy for fetal as well as postnatal cardiac defects and heart failure.

  8. GLI3 Links Environmental Arsenic Exposure and Human Fetal Growth

    Directory of Open Access Journals (Sweden)

    Emily F. Winterbottom

    2015-06-01

    Full Text Available Although considerable evidence suggests that in utero arsenic exposure affects children's health, these data are mainly from areas of the world where groundwater arsenic levels far exceed the World Health Organization limit of 10 μg/L. We, and others, have found that more common levels of in utero arsenic exposure may also impact children's health. However, the underlying molecular mechanisms are poorly understood. To address this issue, we analyzed the expression of key developmental genes in fetal placenta in a birth cohort of women using unregulated water supplies in a US region with elevated groundwater arsenic. We identified several genes whose expression associated with maternal arsenic exposure in a fetal sex-specific manner. In particular, expression of the HEDGEHOG pathway component, GLI3, in female placentae was both negatively associated with arsenic exposure and positively associated with infant birth weight. This suggests that modulation of GLI3 in the fetal placenta, and perhaps in other fetal tissues, contributes to arsenic's detrimental effects on fetal growth. We showed previously that arsenic-exposed NIH3T3 cells have reduced GLI3 repressor protein. Together, these studies identify GLI3 as a key signaling node that is affected by arsenic, mediating a subset of its effects on developmental signaling and fetal health.

  9. GLI3 Links Environmental Arsenic Exposure and Human Fetal Growth.

    Science.gov (United States)

    Winterbottom, Emily F; Fei, Dennis L; Koestler, Devin C; Giambelli, Camilla; Wika, Eric; Capobianco, Anthony J; Lee, Ethan; Marsit, Carmen J; Karagas, Margaret R; Robbins, David J

    2015-06-01

    Although considerable evidence suggests that in utero arsenic exposure affects children's health, these data are mainly from areas of the world where groundwater arsenic levels far exceed the World Health Organization limit of 10 μg/L. We, and others, have found that more common levels of in utero arsenic exposure may also impact children's health. However, the underlying molecular mechanisms are poorly understood. To address this issue, we analyzed the expression of key developmental genes in fetal placenta in a birth cohort of women using unregulated water supplies in a US region with elevated groundwater arsenic. We identified several genes whose expression associated with maternal arsenic exposure in a fetal sex-specific manner. In particular, expression of the HEDGEHOG pathway component, GLI3, in female placentae was both negatively associated with arsenic exposure and positively associated with infant birth weight. This suggests that modulation of GLI3 in the fetal placenta, and perhaps in other fetal tissues, contributes to arsenic's detrimental effects on fetal growth. We showed previously that arsenic-exposed NIH3T3 cells have reduced GLI3 repressor protein. Together, these studies identify GLI3 as a key signaling node that is affected by arsenic, mediating a subset of its effects on developmental signaling and fetal health.

  10. Comparison of Oct4, Sox2 and Nanog Expression in Pancreatic Cancer Cell Lines and Human Pancreatic Tumor

    Directory of Open Access Journals (Sweden)

    Vahideh Assadollahi

    2015-12-01

    Full Text Available Background: Genes are involved in the control of stem cell self-renewal as a new class of molecular markers of cancer. Objectives: In this study, the expression of Oct4, Nanog and Sox2 in cell lines MIA Paca-2, PA-TU-8902 and AsPC-1 and pancreatic cancer tissue were examined. Materials and Methods: In this experimental study, cell lines, MIA Paca-2, PA-TU-8902 and AsPC-1, were cultured in DMEM (Dulbecco’s Modified Eagles Medium and RPMI-1640 (Roswell Park Memorial Institute containing FBS 10% (fetal bovine serum in a 37°C incubator containing Co2 5% and humidity 90%. Samples of tumor and non-cancer pancreatic tumor were purchased Iran tumor bank. Extraction of RNA and synthesis of cDNA was performed. Expression levels of Oct4, Nanog and Sox2 were determined using Real-time PCR. The protein expression levels of target genes in the cell lines were studied by flow cytometry and immunocytochemistry. Results: The expression rate of Oct4, Nanog and Sox2 is more in the cancer cell lines than those in the control (normal tissue samples. The protein expression levels of target genes in the cell lines were confirmed by flow cytometry and immunocytochemistry. Conclusions: The genes are involved in stem cell self-renewal as a new class of molecular markers of cancer that detected in the pancreatic cell lines. Maybe, these genes play important role in the uncontrolled proliferation of cancer cells.

  11. ROLE OF PANCREATIC STELLATE CELLS AND GALECTIN-3 ON PROLIFERATION AND INFILTRATION OF HUMAN PANCREATIC CANCER CELL LINE SW1990

    Institute of Scientific and Technical Information of China (English)

    JIANG Hai-biao; XU Ming; WANG Xing-peng

    2008-01-01

    Objective To investigate the role of pancreatic stellate cells (PSCs) and galectin-3 (GAL-3)on the proliferation and infiltration of pancreatic cancer cell line SW1990. Methods Human pancreatic cancercell line SW1990 and PSCs were cultured in vitro. Supernatant of cultured PSCs and SW1990 cells was collected.Expressions of GAL-3 in SW1990 cells and PSCs were detected by ELISA, RT-PCR and Western blot. Theproliferation of those cultured PSCs and SW1990 cells were measured by MTT assay and flowcytometry. Infiltrationof SW1990 cells was detected by cell infiltration kit. Results SW1990 cells expressed GAL-3 and the expressionwas up-regulated by the supernatant fluid of cultured PSCs. PSCs did not express GAL-3. SW1990 cells couldstimulate the proliferation of PSCs via GAL-3. GAL-3 antibody could inhibit SW1990 cells proliferation andinfiltration, which indicated that supernatant of PSCs might stimulate the proliferation of SW1990 cells through theinteraction with GAL-3 protein. The supernatant fluid of PSCs could enhance the invasiveness of SW1990 cellsthrough the interaction with GAL-3. Conclusion GAL-3 and PSCs was involved in the proliferation andinfiltration process of pancreatic cancer.

  12. Mitochondria-targeted antioxidant enzyme activity regulates radioresistance in human pancreatic cancer cells

    OpenAIRE

    Fisher, Carolyn J.; Goswami, Prabhat C.

    2008-01-01

    In recent years, cellular redox environment gained significant attention as a critical regulator of cellular responses to oxidative stress. Cellular redox environment is a balance between production of reactive oxygen species and their removal by antioxidant enzymes. We investigated the hypothesis that mitochondrial antioxidant enzyme activity regulates radioresistance in human pancreatic cancer cells. Vector-control and manganese superoxide dismutase (MnSOD) overexpressing human pancreatic c...

  13. Inhibition of human pancreatic and biliary output but not intestinal motility by physiological intraileal lipid loads

    DEFF Research Database (Denmark)

    Keller, Jutta; Holst, Jens Juul; Layer, Peter

    2005-01-01

    . Physiological postprandial ileal lipid concentrations dose dependently inhibited human digestive pancreatic protease and bile acid output, but not intestinal motor activity. Thus physiological postprandial ileal nutrient exposure may be of importance for the termination of digestive secretory responses......Lipid perfusion into the distal ileal lumen at supraphysiological loads inhibits pancreatic exocrine secretion and gastrointestinal motility in humans. In the present study, we sought to determine the effects of physiological postprandial intraileal lipid concentrations on endogenously stimulated...

  14. Pancreatic Steatosis and Its Relationship to β-Cell Dysfunction in Humans

    Science.gov (United States)

    Szczepaniak, Lidia S.; Victor, Ronald G.; Mathur, Ruchi; Nelson, Michael D.; Szczepaniak, Edward W.; Tyer, Nicole; Chen, Ida; Unger, Roger H.; Bergman, Richard N.; Lingvay, Ildiko

    2012-01-01

    OBJECTIVE To evaluate racial/ethnic differences in pancreatic triglyceride (TG) levels and their relationship to β-cell dysfunction in humans. RESEARCH DESIGN AND METHODS We studied black, Hispanic, and white adults who completed three research visits: screening and an oral glucose tolerance test; frequently sampled intravenous glucose tolerance tests for evaluation of β-cell function and insulin resistance; and proton magnetic resonance spectroscopy for evaluation of pancreatic and hepatic TG levels. RESULTS Pancreatic TG levels were higher in Hispanics and whites than in blacks (P = 0.006). Hepatic TG levels were highest in Hispanics (P = 0.004). Compensatory insulin secretion and disposition index were higher in blacks (P = 0.003 and P = 0.024, respectively). Insulin sensitivity was comparable between Hispanics and blacks and was lower than in whites (P = 0.005). In blacks, compensatory insulin secretion increased steeply with small increments in pancreatic TG levels (R2 = 0.45, slope = 247). In whites, the range of pancreatic TG levels was higher, and the slope was less steep than in blacks (R2 = 0.27, slope = 27). In Hispanics, pancreatic TG levels were similar to those of whites, but compensatory insulin secretion was described by a combination of pancreatic and hepatic TG levels and visceral fat mass ( R2 = 0.32). CONCLUSIONS In a multiethnic sample of adults with mild obesity and without diabetes, we found striking ethnic differences in the levels of pancreatic TGs and in the relationship between pancreatic TGs and β-cell dysfunction. Our data implicate pancreatic TG content measured by proton magnetic resonance spectroscopy as a noninvasive novel biomarker for pancreatic β-cell dysfunction, especially in the Hispanic population. PMID:22968187

  15. MORPHOLOGICAL FEATURES AND MORPHOMETRIC PARAMETERS OF HUMAN FETAL THYMUS GLANDS

    Directory of Open Access Journals (Sweden)

    S. Havila Hasini

    2014-03-01

    Full Text Available Introduction: Thymus is one of the central lymphoid organs. It plays an important role in the differentiation, selection and maturation of T-lymphocytes. In the recent years morphology and morphometry of the thymus gland in the newborn is gaining significance as it demonstrates great variability between individual infants and in the same infant at different times. Materials and methods: In the present study 45 thymus specimens from aborted human fetuses of 16 to 40 weeks gestational age and both sexes were studied by autopsy for morphological and morphometric features. The morphometric parameters were measured using pachymeter. Results: The thymus gland was located in the superior mediastinum. 60% (27/45 specimens showed cervical extensions. Brachiocephalic vein anterior to thymus was observed in 3 cases which is an important anomaly to be observed in thymectomy procedure. Thymuses were greyish pink to greyish brown in colour. Variations were also observed in the number of lobes of glands in which one is single lobed, most of the glands are bilobed and few are trilobed. There is progressive increase in all morphometric dimensions of the thymus in relation to gestational age. Most of the specimens were less than 4cm in length. Half of the specimens were below 2cm in width and other half were 2.0 to 5.0 cm in width. For 90% of the specimens thickness of the organ was less than 0.5cm. The thymus gland was 0.2% of fetal body weight. Conclusion: The morphological observations of thymus gland shows great variations which has to be considered in thymectomy. In addition to anthropometric parameters of fetus, morphometric parameters of thymus glands present significant relation to the gestational age of fetuses. It is possible to determine the thymic morphometric parameters in relation to gestational age.

  16. Organoid Models of Human and Mouse Ductal Pancreatic Cancer

    NARCIS (Netherlands)

    Boj, Sylvia F.; Hwang, Chang-Il; Baker, Lindsey A.; Chio, Iok In Christine; Engle, Dannielle D.; Corbo, Vincenzo; Jager, Myrthe; Ponz-Sarvise, Mariano; Tiriac, Herve; Spector, Mona S.; Gracanin, Ana; Oni, Tobiloba; Yu, Kenneth H.; van Boxtel, Ruben; Huch, Meritxell; Rivera, Keith D.; Wilson, John P.; Feigin, Michael E.; Oehlund, Daniel; Handly-Santana, Abram; Ardito-Abraham, Christine M.; Ludwig, Michael; Elyada, Ela; Alagesan, Brinda; Biffi, Giulia; Yordanov, Georgi N.; Delcuze, Bethany; Creighton, Brianna; Wright, Kevin; Park, Youngkyu; Morsink, Folkert H. M.; Molenaar, IQ; Borel Rinkes, Inne H.; Cuppen, Edwin; Hao, Yuan; Jin, Ying; Nijman, Isaac J.; Iacobuzio-Donahue, Christine; Leach, Steven D.; Pappin, Darryl J.; Hammell, Molly; Klimstra, David S.; Basturk, Olca; Hruban, Ralph H.; Offerhaus, George Johan; Vries, Robert G. J.; Clevers, Hans; Tuveson, David A.

    2015-01-01

    Pancreatic cancer is one of the most lethal malignancies due to its late diagnosis and limited response to treatment. Tractable methods to identify and interrogate pathways involved in pancreatic tumorigenesis are urgently needed. We established organoid models from normal and neoplastic murine and

  17. Organoid models of human and mouse ductal pancreatic cancer

    NARCIS (Netherlands)

    Boj, Sylvia F; Hwang, Chang-Il; Baker, Lindsey A; Chio, Iok In Christine; Engle, Dannielle D; Corbo, Vincenzo; Jager, Myrthe; Ponz-Sarvise, Mariano; Tiriac, Hervé; Spector, Mona S; Gracanin, Ana; Oni, Tobiloba; Yu, Kenneth H; van Boxtel, Ruben; Huch, Meritxell; Rivera, Keith D; Wilson, John P; Feigin, Michael E; Öhlund, Daniel; Handly-Santana, Abram; Ardito-Abraham, Christine M; Ludwig, Michael; Elyada, Ela; Alagesan, Brinda; Biffi, Giulia; Yordanov, Georgi N; Delcuze, Bethany; Creighton, Brianna; Wright, Kevin; Park, Youngkyu; Morsink, Folkert H M; Molenaar, I Quintus; Borel Rinkes, Inne H; Cuppen, Edwin; Hao, Yuan; Jin, Ying; Nijman, Isaac J; Iacobuzio-Donahue, Christine; Leach, Steven D; Pappin, Darryl J; Hammell, Molly; Klimstra, David S; Basturk, Olca; Hruban, Ralph H; Offerhaus, George Johan; Vries, Robert G J; Clevers, Hans; Tuveson, David A

    2015-01-01

    Pancreatic cancer is one of the most lethal malignancies due to its late diagnosis and limited response to treatment. Tractable methods to identify and interrogate pathways involved in pancreatic tumorigenesis are urgently needed. We established organoid models from normal and neoplastic murine and

  18. Organoid Models of Human and Mouse Ductal Pancreatic Cancer

    NARCIS (Netherlands)

    Boj, Sylvia F.; Hwang, Chang-Il; Baker, Lindsey A.; Chio, Iok In Christine; Engle, Dannielle D.; Corbo, Vincenzo; Jager, Myrthe; Ponz-Sarvise, Mariano; Tiriac, Herve; Spector, Mona S.; Gracanin, Ana; Oni, Tobiloba; Yu, Kenneth H.; van Boxtel, Ruben; Huch, Meritxell; Rivera, Keith D.; Wilson, John P.; Feigin, Michael E.; Oehlund, Daniel; Handly-Santana, Abram; Ardito-Abraham, Christine M.; Ludwig, Michael; Elyada, Ela; Alagesan, Brinda; Biffi, Giulia; Yordanov, Georgi N.; Delcuze, Bethany; Creighton, Brianna; Wright, Kevin; Park, Youngkyu; Morsink, Folkert H. M.; Molenaar, IQ; Borel Rinkes, Inne H.; Cuppen, Edwin; Hao, Yuan; Jin, Ying; Nijman, Isaac J.; Iacobuzio-Donahue, Christine; Leach, Steven D.; Pappin, Darryl J.; Hammell, Molly; Klimstra, David S.; Basturk, Olca; Hruban, Ralph H.; Offerhaus, George Johan; Vries, Robert G. J.; Clevers, Hans; Tuveson, David A.

    2015-01-01

    Pancreatic cancer is one of the most lethal malignancies due to its late diagnosis and limited response to treatment. Tractable methods to identify and interrogate pathways involved in pancreatic tumorigenesis are urgently needed. We established organoid models from normal and neoplastic murine and

  19. Organoid models of human and mouse ductal pancreatic cancer

    NARCIS (Netherlands)

    Boj, Sylvia F; Hwang, Chang-Il; Baker, Lindsey A; Chio, Iok In Christine; Engle, Dannielle D; Corbo, Vincenzo; Jager, Myrthe; Ponz-Sarvise, Mariano; Tiriac, Hervé; Spector, Mona S; Gracanin, Ana; Oni, Tobiloba; Yu, Kenneth H; van Boxtel, Ruben; Huch, Meritxell; Rivera, Keith D; Wilson, John P; Feigin, Michael E; Öhlund, Daniel; Handly-Santana, Abram; Ardito-Abraham, Christine M; Ludwig, Michael; Elyada, Ela; Alagesan, Brinda; Biffi, Giulia; Yordanov, Georgi N; Delcuze, Bethany; Creighton, Brianna; Wright, Kevin; Park, Youngkyu; Morsink, Folkert H M; Molenaar, I Quintus; Borel Rinkes, Inne H; Cuppen, Edwin; Hao, Yuan; Jin, Ying; Nijman, Isaac J; Iacobuzio-Donahue, Christine; Leach, Steven D; Pappin, Darryl J; Hammell, Molly; Klimstra, David S; Basturk, Olca; Hruban, Ralph H; Offerhaus, George Johan; Vries, Robert G J; Clevers, Hans; Tuveson, David A

    2015-01-01

    Pancreatic cancer is one of the most lethal malignancies due to its late diagnosis and limited response to treatment. Tractable methods to identify and interrogate pathways involved in pancreatic tumorigenesis are urgently needed. We established organoid models from normal and neoplastic murine and

  20. Pancreatitis

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    2008459 Screening of proteins binding to hepatitis B core antigen from human pancreas cDNA library by yeast two-hybrid system. Zhang Weiyan(张维燕), et al.Instit Infect Dis, Beijiug Ditan Hosp, Beijing 100011. World Chin J Digestol 2008;16(16):1746-1750. Objective To screen proteins in human pancreas cDNA library interacting with hepatitis B core antigen (HbcAg) by yeast two-hybrid system.

  1. Effect of Supracervical Apposition and Spontaneous Labour on Apoptosis and Matrix Metalloproteinases in Human Fetal Membranes

    Directory of Open Access Journals (Sweden)

    Mahalia Chai

    2013-01-01

    Full Text Available Background. Apoptosis and matrix metalloproteinase (MMP-9 are capable of hydrolysing components of the extracellular matrix and weakening the fetal membranes which leads to eventual rupture, a key process of human parturition. The aim of this study was to determine the effect of supracervical apposition and spontaneous labour on apoptosis and MMP-9 in human fetal membranes at term. Methods. Fetal membranes were obtained from term non-labouring supracervical site (SCS and compared to (i a paired distal site (DS or (ii site of rupture (SOR after spontaneous labour onset. Results. The expression of the proapoptotic markers Bax, Smac, Fas, FasL, caspase-3, and PARP, was significantly higher in the non-labouring SCS chorion compared to paired DS. Bax, Smac, FasL, caspase-3, and PARP staining was higher in the non-labouring SCS fetal membranes than that in the post-labour SOR. MMP-9 expression and activity were higher in the post-labour SOR fetal membranes compared to non-labouring SCS fetal membranes. Conclusion. Components of the apoptotic signalling pathways and MMP-9 may play a role in rupture and labour. Non-labouring SCS fetal membranes display altered morphology and altered apoptotic biochemical characteristics in preparation for labour, while the laboured SOR displays unique MMP characteristics.

  2. CCDC88A, a prognostic factor for human pancreatic cancers, promotes the motility and invasiveness of pancreatic cancer cells

    OpenAIRE

    Tanouchi, Aki; Taniuchi, Keisuke; Furihata, Mutsuo; NAGANUMA, Seiji; Dabanaka, Ken; Kimura, Masashi; Watanabe, Ryohei; Kohsaki, Takuhiro; Shimizu, Takahiro; Saito, Motoaki; Hanazaki, Kazuhiro; Saibara, Toshiji

    2016-01-01

    Background Coiled-Coil Domain Containing 88A (CCDC88A) was identified as a substrate of the serine/threonine kinase Akt that is capable of binding to the actin cytoskeleton. The aim of this study was to investigate the potential role of CCDC88A in the migration and invasiveness of pancreatic ductal adenocarcinoma (PDAC) cells. Methods Immunohistochemistry was performed to determine whether high CCDC88A expression in human PDAC tissues is correlated with poor prognosis. Immunoprecipitation, im...

  3. Identification of CYP3A7 for Glyburide Metabolism in Human Fetal Livers

    Science.gov (United States)

    Shuster, Diana L.; Risler, Linda J.; Prasad, Bhagwat; Calamia, Justina C.; Voellinger, Jenna L.; Kelly, Edward J.; Unadkat, Jashvant D.; Hebert, Mary F.; Shen, Danny D.; Thummel, Kenneth E.; Mao, Qingcheng

    2014-01-01

    Glyburide is commonly prescribed for the treatment of gestational diabetes mellitus; however, fetal exposure to glyburide is not well understood and may have short- and long-term consequences for the health of the child. Glyburide can cross the placenta; fetal concentrations at term are nearly comparable to maternal levels. Whether or not glyburide is metabolized in the fetus and by what mechanisms has yet to be determined. In this study, we determined the kinetic parameters for glyburide depletion by CYP3A isoenzymes; characterized glyburide metabolism by human fetal liver tissues collected during the first or early second trimester of pregnancy; and identified the major enzyme responsible for glyburide metabolism in human fetal livers. CYP3A4 had the highest metabolic capacity towards glyburide, followed by CYP3A7 and CYP3A5 (Clint,u = 37.1, 13.0, and 8.7 ml/min/nmol P450, respectively). M5 was the predominant metabolite generated by CYP3A7 and human fetal liver microsomes (HFLMs) with approximately 96% relative abundance. M5 was also the dominant metabolite generated by CYP3A4, CYP3A5, and adult liver microsomes; however, M1-M4 were also present, with up to 15% relative abundance. CYP3A7 protein levels in HFLMs were highly correlated with glyburide Clint, 16α-OH DHEA formation, and 4′-OH midazolam formation. Likewise, glyburide Clint was highly correlated with 16α-OH DHEA formation. Fetal demographics as well as CYP3A5 and CYP3A7 genotype did not alter CYP3A7 protein levels or glyburide Clint. These results indicate that human fetal livers metabolize glyburide predominantly to M5 and that CYP3A7 is the major enzyme responsible for glyburide metabolism in human fetal livers. PMID:25450675

  4. Diabetes mellitus is associated with an increased expression of resistin in human pancreatic islet cells.

    Science.gov (United States)

    Al-Salam, Suhail; Rashed, Hameed; Adeghate, Ernest

    2011-01-01

    The pattern of distribution of resistin in the pancreas of diabetic patients was investigated to determine whether diabetes mellitus influences the expression of resistin. Pancreatic tissue samples retrieved, during pancreatectomy for pancreatic cancer, from cancer patients with and without type 2 diabetes were processed for immunohistochemistry. The pancreatic tissue samples were retrieved from non-cancerous and clear margins. An immunofluorescence technique was used to examine the expression of resistin and its co-localization with insulin and glucagon in pancreatic islet cells. Resistin was observed in many cells located in the central region of pancreatic islet. The expression of resistin increased significantly (p diabetic patients compared to control. Resistin co-localized with insulin but not glucagon in pancreatic islet cells of both normal and diabetic patients. However, the degree of co-localization was higher in pancreata of diabetic patients compared to normal. The number of human pancreatic islet cells expressing resistin increased significantly after the onset of type 2 diabetes. In conclusion, resistin may play a role in the regulation of pancreatic β-cell function.

  5. SIRT1 promotes the proliferation and metastasis of human pancreatic cancer cells.

    Science.gov (United States)

    Jin, Jianguang; Chu, Zhijie; Ma, Pengfei; Meng, Yuanpu; Yang, Yanhui

    2017-03-01

    SIRT1 plays an important role in human malignant progression, inducing cancer cell proliferation and metastasis by regulating downstream gene expressions. However, little is known about the underlying mechanisms in which SIRT1 promotes pancreatic cancer tumorigenesis. The aim of this study is to investigate the SIRT1 expression levels and biological functions in promoting pancreatic cancer progression. We first investigated the expression of SIRT1 in a series of pancreatic cancer tissues as well as in a panel of pancreatic cancer cell lines. The effect of SIRT1 on cell activity was explored by knockdown experiments. Cell growth was measured using the MTT assay and colony-formation assay. Migration and invasion were tested using transwell assay. Our results showed that the expression of SIRT1 was significantly up-regulated both in pancreatic cancer tissues and cell lines. Knockdown of SIRT1 suppressed cell proliferation and migration of pancreatic cancer cells. This is the first report to disclose the role of SIRT1 in regulation of pancreatic cancer cell proliferation and migration, which may provide a potential therapeutic target for pancreatic cancer patients.

  6. Expression of stem cell markers in the human fetal kidney.

    Directory of Open Access Journals (Sweden)

    Sally Metsuyanim

    Full Text Available In the human fetal kidney (HFK self-renewing stem cells residing in the metanephric mesenchyme (MM/blastema are induced to form all cell types of the nephron till 34(th week of gestation. Definition of useful markers is crucial for the identification of HFK stem cells. Because wilms' tumor, a pediatric renal cancer, initiates from retention of renal stem cells, we hypothesized that surface antigens previously up-regulated in microarrays of both HFK and blastema-enriched stem-like wilms' tumor xenografts (NCAM, ACVRIIB, DLK1/PREF, GPR39, FZD7, FZD2, NTRK2 are likely to be relevant markers. Comprehensive profiling of these putative and of additional stem cell markers (CD34, CD133, c-Kit, CD90, CD105, CD24 in mid-gestation HFK was performed using immunostaining and FACS in conjunction with EpCAM, an epithelial surface marker that is absent from the MM and increases along nephron differentiation and hence can be separated into negative, dim or bright fractions. No marker was specifically localized to the MM. Nevertheless, FZD7 and NTRK2 were preferentially localized to the MM and emerging tubules (50% of HFK cells and predominantly co-express EpCAM(bright, indicating they are mostly markers of differentiation. Furthermore, localization of NCAM exclusively in the MM and in its nephron progenitor derivatives but also in stroma and the expression pattern of significantly elevated renal stem/progenitor genes Six2, Wt1, Cited1, and Sall1 in NCAM(+EpCAM(- and to a lesser extent in NCAM(+EpCAM(+ fractions confirmed regional identity of cells and assisted us in pinpointing the presence of subpopulations that are putative MM-derived progenitor cells (NCAM(+EpCAM(+FZD7(+, MM stem cells (NCAM(+EpCAM(-FZD7(+ or both (NCAM(+FZD7(+. These results and concepts provide a framework for developing cell selection strategies for human renal cell-based therapies.

  7. Maturation of the human fetal startle response: evidence for sex-specific maturation of the human fetus.

    Science.gov (United States)

    Buss, Claudia; Davis, Elysia Poggi; Class, Quetzal A; Gierczak, Matt; Pattillo, Carol; Glynn, Laura M; Sandman, Curt A

    2009-10-01

    Despite the evidence for early fetal experience exerting programming influences on later neurological development and health risk, very few prospective studies of human fetal behavior have been reported. In a prospective longitudinal study, fetal nervous system maturation was serially assessed by monitoring fetal heart rate (FHR) responses to vibroacoustic stimulation (VAS) in 191 maternal/fetal dyads. Responses were not detected at 26 weeks gestational age (GA). Sex-specific, age-characteristic changes in the FHR response to VAS were observed by 31 weeks' GA. Males showed larger responses and continued to exhibit maturational changes until 37 weeks' GA, females however, presented with a mature FHR startle response by 31 weeks' GA. The results indicate that there are different rates of maturation in the male and female fetuses that may have implications for sex-specific programming influences.

  8. Maturation of the human fetal startle response: Evidence for sex-specific maturation of the human fetus1

    Science.gov (United States)

    Buss, Claudia; Davis, Elysia Poggi; Class, Quetzal A.; Gierczak, Matt; Pattillo, Carol; Glynn, Laura M.; Sandman, Curt A.

    2009-01-01

    Despite the evidence for early fetal experience exerting programming influences on later neurological development and health risk, very few prospective studies of human fetal behavior have been reported. In a prospective longitudinal study, fetal nervous system maturation was serially assessed by monitoring fetal heart rate (FHR) responses to vibroacoustic stimulation (VAS) in 191 maternal/fetal dyads. Responses were not detected at 26 weeks gestational age (GA). Sex-specific, age-characteristic changes in the FHR response to VAS were observed by 31 weeks’ GA. Males showed larger responses and continued to exhibit maturational changes until 37 weeks’ GA, females however, presented with a mature FHR startle response by 31 weeks’ GA. The results indicate that there are different rates of maturation in the male and female fetus that may have implications for sex-specific programming influences. PMID:19726143

  9. Human chorionic gonadotropin (hCG) concentrations during the late first trimester are associated with fetal growth in a fetal sex-specific manner.

    Science.gov (United States)

    Barjaktarovic, Mirjana; Korevaar, Tim I M; Jaddoe, Vincent W V; de Rijke, Yolanda B; Visser, Theo J; Peeters, Robin P; Steegers, Eric A P

    2017-02-01

    Human chorionic gonadotropin (hCG) is a pregnancy-specific hormone that regulates placental development. hCG concentrations vary widely throughout gestation and differ based on fetal sex. Abnormal hCG concentrations are associated with adverse pregnancy outcomes including fetal growth restriction. We studied the association of hCG concentrations with fetal growth and birth weight. In addition, we investigated effect modification by gestational age of hCG measurement and fetal sex. Total serum hCG (median 14.4 weeks, 95 % range 10.1-26.2), estimated fetal weight (measured by ultrasound during 18-25th weeks and >25th weeks) and birth weight were measured in 7987 mother-child pairs from the Generation R cohort and used to establish fetal growth. Small for gestational age (SGA) was defined as a standardized birth weight lower than the 10th percentile of the study population. There was a non-linear association of hCG with birth weight (P = 0.009). However, only low hCG concentrations measured during the late first trimester (11th and 12th week) were associated with birth weight and SGA. Low hCG concentrations measured in the late first trimester were also associated with decreased fetal growth (P = 0.0002). This was the case for both male and female fetuses. In contrast, high hCG concentrations during the late first trimester were associated with increased fetal growth amongst female, but not male fetuses. Low hCG in the late first trimester is associated with lower birth weight due to a decrease in fetal growth. Fetal sex differences exist in the association of hCG concentrations with fetal growth.

  10. Influence of maternal folate status on human fetal growth parameters

    NARCIS (Netherlands)

    van Uitert, Evelyne M.; Steegers-Theunissen, Regine P. M.

    2013-01-01

    Worldwide periconceptional folic acid supplement use is recommended to prevent neural tube defects. This also stimulated research on maternal folate status in association with fetal growth, an important predictor of perinatal and future development and health. We provide an overview of literature on

  11. Adenosine receptors in rat and human pancreatic ducts stimulate chloride transport

    DEFF Research Database (Denmark)

    Novak, Ivana; Hede, Susanne; Hansen, Mette

    2007-01-01

    these could be involved in secretory processes, which involve cystic fibrosis transmembrane regulator (CFTR) Cl(-) channels or Ca(2+)-activated Cl(-) channels and [Formula: see text] transporters. Reverse transcriptase polymerase chain reaction analysis on rat pancreatic ducts and human duct cell......, plasma membrane of many PANC-1 cells, but only a few CFPAC-1 cells. Taken together, our data indicate that A(2A) receptors open Cl(-) channels in pancreatic ducts cells with functional CFTR. We propose that adenosine can stimulate pancreatic secretion and, thereby, is an active player in the acini...

  12. Adenoviral transfer of human interleukin-10 gene in lethal pancreatitis

    Institute of Scientific and Technical Information of China (English)

    Zi-Qian Chen; Yao-Qing Tang; Yi Zhang; Zhi-Hong Jiang; En-Qiang Mao; Wei-Guo Zou; Ruo-Qing Lei; Tian-Quan Han; Sheng-Dao Zhang

    2004-01-01

    AIM: To evaluate the therapeutic effect of adenoviral-vectordelivered human interleukin-10 (hIL-10) gene on severe acute pancreatitis (SAP) rats.METHODS: Healthy Sprague-Dawley (SD) rats were intraperitoneally injected with adenoviral IL-10 gene (AdvhIL-10), empty vector (Adv0) or PBS solution. Blood,liver, pancreas and lung were harvested on the second day to examine hIL-10 level by ELISA and serum amylase by enzymatic assay. A SAP model was induced by retrograde injection of sodium taurocholate through pancreatic duct.SAP rats were then administered with AdvhIL-10, Adv0 and PBS solution by a single intraperitoneal injection 20 min after SAP induction. In addition to serum amylase assay,levels of hIL-10 and tumor necrosis factor-α (TNF-α) were detected by RT-PCR, ELISA and histological study. The mortality rate was studied and analyzed by Kaplan-Meier and log rank analysis.RESULTS: The levels of hIL-10 in the pancreas, liver and lung of healthy rats increased significantly after AdvhIL-10injection (1.42 ng/g in liver, 0.91 ng/g in pancreas); while there was no significant change of hIL-10 in the other two control groups. The concentration of hIL-10 was increased significantly in the SAP rats after AdvhIL-10 injection (1.68 ng/g in liver, 1.12 ng/g in pancreas) compared to the other two SAP groups with blank vector or PBS treatment (P<0.05). The serum amylase levels remained normal in the AdvhIL-10 transfected healthy rats. However,the serum amylase level was significantly elevated in the other two control SAP rats. In contrast, serum amylase was down-regulated in the AdvhIL-10 treated SAP groups.The TNF-α expression in the AdvhIL-10 treated SAP rats was significantly lower compared to the other two control SAP groups. The pathohistological changes in the AdvhIL-10 treated group were better than those in the other two control groups. Furthermore, the mortality of the AdvhIL-10 treated group was significantly reduced compared to the other two control groups (P

  13. Differential role of Hedgehog signaling in human pancreatic (patho-) physiology: An up to date review

    OpenAIRE

    Klieser, Eckhard; SWIERCZYNSKI, STEFAN; Mayr, Christian; Jäger, Tarkan; Schmidt, Johanna; Neureiter, Daniel; KIESSLICH, TOBIAS; Illig, Romana

    2016-01-01

    Since the discovery of the Hedgehog (Hh) pathway in drosophila melanogaster, our knowledge of the role of Hh in embryonic development, inflammation, and cancerogenesis in humans has dramatically increased over the last decades. This is the case especially concerning the pancreas, however, real therapeutic breakthroughs are missing until now. In general, Hh signaling is essential for pancreatic organogenesis, development, and tissue maturation. In the case of acute pancreatitis, Hh has a prote...

  14. Pancreatic Steatosis and Its Relationship to β-Cell Dysfunction in Humans

    OpenAIRE

    Szczepaniak, Lidia S.; Victor, Ronald G.; Mathur, Ruchi; Nelson, Michael D; Edward W Szczepaniak; Tyer, Nicole; Chen, Ida; Unger, Roger H.; Bergman, Richard N.; Lingvay, Ildiko

    2012-01-01

    OBJECTIVE To evaluate racial/ethnic differences in pancreatic triglyceride (TG) levels and their relationship to β-cell dysfunction in humans. RESEARCH DESIGN AND METHODS We studied black, Hispanic, and white adults who completed three research visits: screening and an oral glucose tolerance test; frequently sampled intravenous glucose tolerance tests for evaluation of β-cell function and insulin resistance; and proton magnetic resonance spectroscopy for evaluation of pancreatic and hepatic T...

  15. The expression pattern of two novel cytokines (IL-24 and IL-29) in human fetal membranes.

    Science.gov (United States)

    Nace, Judith; Fortunato, Stephen J; Maul, Holger; Menon, Ramkumar

    2010-11-01

    interleukin (IL)-24 and -29 are novel cytokines, produced by immune cells in response to microbial antigens. The functions of these cytokines in the reproductive system are unknown. We examined the expression pattern of IL-24 and IL-29 in human fetal membranes from preterm and term births and in in vitro in response to microbial antigens. fetal membranes collected from cesarean sections at term (normal, not in labor) were placed in culture for 48 h. These membranes were then stimulated with bacterial lipopolysaccharide (LPS) or viral antigen poly-inosinic and cytidylic acid (polyIC) for an additional 24 h. Amniotic fluids (AF) and fetal membranes were also collected from preterm and term deliveries. IL-24 and IL-29 expressions were studied by RT-PCR. ELISA documented culture media and AF cytokine concentrations. IL-24 and IL-29 expressions were seen in cultured fetal membranes regardless of stimulation. Expressions were also found in preterm and term labor membranes, but not in non-labor tissues at term. IL-24 concentrations were higher after LPS stimulation whereas IL-29 concentrations were higher after polyIC-stimulation. AF analysis did not detect either of the cytokines either preterm or term. this is the first study to report IL-24 and IL-29 expressions in human fetal membranes. Higher concentrations of these cytokines in response to distinct infectious stimuli suggest different pathways for fetal immune response during infection.

  16. Ethical issues surrounding the transplantation of human fetal tissues.

    Science.gov (United States)

    Hurd, R E

    1992-12-01

    Organ transplants have been one of the greatest advances in medicine. However, organs from living relatives or cadavers are in short supply, and many people die awaiting a donor organ. Increasing the donor pool by using organs from aborted fetuses has been proposed to increase the supply. In addition, there are benefits of using fetal tissue including its particular usefulness in children, the fact that it is not readily rejected, and its potential for growth. Guidelines for fetal research were issued in 1975, but a research moratorium was imposed in 1988 to allow study of ethical and legal issues. While the federal government delays in lifting the ban, several states have written laws governing experimentation with fetuses. Ethical arguments against using fetal tissue for organ transplant include a concern that this would create a branch of biomedicine which depends on the continuation of induced abortions. This could lead to neglect of research for other therapies. The timing and type of abortion should continue to benefit the mother, rather than the organ recipient. Ethicists debate whether or not use of aborted tissue implies complicity in the abortion process beyond that which exists for all members of a society which permits abortion. They also wonder whether knowing that some good could come of an abortion would influence a woman's decision to have one. Proposals to keep the use of fetal tissue ethical include banning the commercial use of sale of tissues, forbidding designation of the tissue recipient (to prevent harvesting fetal tissue for a relative), separating abortion counseling and management from harvesting of the tissue, and obtaining informed consent (perhaps from a proxy surrogate rather than from the mother) for the use of fetal tissue. When the medical and ethical communities have reached some consensus on these issues, crafted safeguards, and precluded conflicts of interest, then restrictions on government funding should be lifted. Whereas it

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

    Science.gov (United States)

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

    2012-02-01

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

  18. KeyGenes, a Tool to Probe Tissue Differentiation Using a Human Fetal Transcriptional Atlas

    NARCIS (Netherlands)

    Roost, M.S.; Iperen, L. van; Ariyurek, Y.; Buermans, H.P.; Arindrarto, W.; Devalla, H.D.; Passier, R.; Mummery, C.L.; Carlotti, F.; Koning, E.J. de; Zwet, E.W. van; Goeman, J.J.; Chuva de Sousa Lopes, S.M.

    2015-01-01

    Differentiated derivatives of human pluripotent stem cells in culture are generally phenotypically immature compared to their adult counterparts. Their identity is often difficult to determine with certainty because little is known about their human fetal equivalents in vivo. Cellular identity and s

  19. KeyGenes, a Tool to Probe Tissue Differentiation Using a Human Fetal Transcriptional Atlas

    NARCIS (Netherlands)

    Roost, Matthias S; van Iperen, Liesbeth; Ariyurek, Yavuz; Buermans, Henk P; Arindrarto, Wibowo; Devalla, Harsha D; Passier, Robert; Mummery, Christine L; Carlotti, Françoise; de Koning, Eelco J P; van Zwet, Erik W; Goeman, Jelle J; Chuva de Sousa Lopes, Susana M

    2015-01-01

    Differentiated derivatives of human pluripotent stem cells in culture are generally phenotypically immature compared to their adult counterparts. Their identity is often difficult to determine with certainty because little is known about their human fetal equivalents in vivo. Cellular identity and

  20. SIRT1 Disruption in Human Fetal Hepatocytes Leads to Increased Accumulation of Glucose and Lipids.

    Directory of Open Access Journals (Sweden)

    Takamasa Tobita

    Full Text Available There are unprecedented epidemics of obesity, such as type II diabetes and non-alcoholic fatty liver diseases (NAFLD in developed countries. A concerning percentage of American children are being affected by obesity and NAFLD. Studies have suggested that the maternal environment in utero might play a role in the development of these diseases later in life. In this study, we documented that inhibiting SIRT1 signaling in human fetal hepatocytes rapidly led to an increase in intracellular glucose and lipids levels. More importantly, both de novo lipogenesis and gluconeogenesis related genes were upregulated upon SIRT1 inhibition. The AKT/FOXO1 pathway, a major negative regulator of gluconeogenesis, was decreased in the human fetal hepatocytes inhibited for SIRT1, consistent with the higher level of gluconeogenesis. These results indicate that SIRT1 is an important regulator of lipid and carbohydrate metabolisms within human fetal hepatocytes, acting as an adaptive transcriptional response to environmental changes.

  1. Clinicopathological significance of p53 and mdm2 protein expression in human pancreatic cancer

    Institute of Scientific and Technical Information of China (English)

    Ming Dong; Gang Ma; Wei Tu; Ke-Jian Guo; Yu-Lin Tian; Yu-Ting Dong

    2005-01-01

    AIM: To study the clinicopathological significance of p53 and mdm2 protein expression in human pancreatic cancer. METHODS: To investigate the expression of p53 and mdm2 in pancreatic cancer by immunohistochemistry, and the relationships between the p53 and mdm2 protein expression and clinicopathological parameters in pancreatic cancer.RESULTS: The positive expression of p53 protein was found in 40 of 59 patients (67.8%) and that of mdm2 protein in 17 of 59 patients (28.8%). No obvious relationships were found between p53 as well as mdm2 expression and sex, tumor site, TNM staging and histological differentiation. p53 expression was increased in patients younger than 65 years old, while mdm2 had no relationship with age. The survival time of the patients with the positive expression of p53 and mdm2 proteins was obviously shorter than the other groups. CONCLUSION: Both p53 and mdm2 presented relatively high expression in human pancreatic cancer. The overexpression of p53 and mdm2 might reflect the malignant proliferation of pancreatic cancer and their co-expression might be helpful to evaluate the prognosis of the patients with pancreatic cancer.

  2. Ex-vivo evaluation of gene therapy vectors in human pancreatic (cancer) tissue slices

    Institute of Scientific and Technical Information of China (English)

    Michael A van Geer; Koert FD Kuhlmann; Conny T Bakker; Fibo JW ten Kate; Ronald PJ Oude Elferink; Piter J Bosma

    2009-01-01

    AIM: To culture human pancreatic tissue obtained from small resection specimens as a pre-clinical model for examining virus-host interactions.METHODS: Human pancreatic tissue samples (malignant and normal) were obtained from surgical specimens and processed immediately to tissue slices.Tissue slices were cultured ex vivo for 1-6 d in an incubator using 95% O2. Slices were subsequently analyzed for viability and morphology. In addition the slices were incubated with different viral vectors expressing the repor ter genes GFP or DsRed.Expression of these reporter genes was measured at 72 h after infection.RESULTS: With the Krumdieck tissue slicer, uniform slices could be generated from pancreatic tissue but only upon embedding the tissue in 3% low melting agarose. Immunohistological examination showed the presence of all pancreatic cell types. Pancreatic normal and cancer tissue slices could be cultured for up to 6 d, while retaining viability and a moderate to good morphology. Reporter gene expression indicated that the slices could be infected and transduced efficiently by adenoviral vectors and by adeno associated viral vectors, whereas transduction with lentiviral vectors was limited. For the adenoviral vector, the transduction seemed limited to the peripheral layers of the explants.CONCLUSION: The presented sys tem al lows reproducible processing of minimal amounts of pancreatic tissue into slices uniform in size, suitable for pre-clinical evaluation of gene therapy vectors.

  3. Expression, biosynthesis and release of preadipocyte factor-1/ delta-like protein/fetal antigen-1 in pancreatic -cells

    DEFF Research Database (Denmark)

    Friedrichsen, B N; Carlsson, C; Møldrup, A

    2003-01-01

    Preadipocyte factor-1 (Pref-1)/delta-like protein/fetal antigen-1 (FA1) is a member of the epidermal growth factor-like family. It is widely expressed in embryonic tissues, whereas in adults it is confined to the adrenal gland, the anterior pituitary, the endocrine pancreas, the testis and the ov......Preadipocyte factor-1 (Pref-1)/delta-like protein/fetal antigen-1 (FA1) is a member of the epidermal growth factor-like family. It is widely expressed in embryonic tissues, whereas in adults it is confined to the adrenal gland, the anterior pituitary, the endocrine pancreas, the testis...

  4. Binding of furosemide to albumin isolated from human fetal and adult serum.

    Science.gov (United States)

    Viani, A; Cappiello, M; Silvestri, D; Pacifici, G M

    1991-01-01

    Albumin was isolated from pooled fetal serum from 58 placentas obtained at normal delivery at term and from pooled adult plasma from 8 individuals. Albumin isolation was carried out by means of PEG precipitation followed by ion-exchange chromatography on DEAE-Sephadex A 50 and then on SP-Sephadex C 50. The electrophoresis on SDS-polyacrylamide gels showed only one spot that comigrated with commercial human albumin. Binding to albumin was measured by equilibrium dialysis of an aliquot of albumin solution (0.7 ml) against the same volume of 0.13 M sodium orthophosphate buffer (pH 7.4). At a total concentration of 2 micrograms/ml (therapeutic range), the unbound fraction of furosemide was 2.71% (fetal albumin) and 2.51% (adult albumin). Two classes of binding sites for furosemide were observed in fetal and adult albumin. The number of binding sites (moles of furosemide per mole of albumin) was 1.22 (fetal albumin) and 1.58 (adult albumin) for the high-affinity site and 2.97 (fetal albumin) and 3.25 (adult albumin) for the low-affinity site. The association constants (M-1) were 3.1 X 10(4) (fetal albumin) and 2.6 X 10(4) (adult albumin) for the high-affinity set of sites and 0.83 X 10(4) (fetal albumin) and 1.0 X 10(4) (adult albumin) low-affinity site. The displacement of furosemide from albumin was studied with therapeutic concentrations of several drugs. Valproic acid, salicylic acid, azapropazone and tolbutamide had the highest displacing effects which were significantly higher with fetal than with adult albumin.

  5. A targeting ligand enhances infectivity and cytotoxicity of an oncolytic adenovirus in human pancreatic cancer tissues.

    Science.gov (United States)

    Yamamoto, Yuki; Hiraoka, Nobuyoshi; Goto, Naoko; Rin, Yosei; Miura, Kazuki; Narumi, Kenta; Uchida, Hiroaki; Tagawa, Masatoshi; Aoki, Kazunori

    2014-10-28

    The addition of a targeting strategy is necessary to enhance oncolysis and secure safety of a conditionally replicative adenovirus (CRAd). We have constructed an adenovirus library displaying random peptides on the fiber, and have successfully identified a pancreatic cancer-targeting ligand (SYENFSA). Here, the usefulness of cancer-targeted CRAd for pancreatic cancer was examined as a preclinical study. First, we constructed a survivin promoter-regulated CRAd expressing enhanced green fluorescent protein gene (EGFP), which displayed the identified targeting ligand (AdSur-SYE). The AdSur-SYE resulted in higher gene transduction efficiency and oncolytic potency than the untargeted CRAd (AdSur) in several pancreatic cancer cell lines. An intratumoral injection of AdSur-SYE significantly suppressed the growth of subcutaneous tumors, in which AdSur-SYE effectively proliferated and spread. An ectopic infection in adjacent tissues and organs of intratumorally injected AdSur-SYE was decreased compared with AdSur. Then, to examine whether the targeting ligand actually enhanced the infectivity of CRAd in human pancreatic cancer tissues, tumor cells prepared from surgical specimens were infected with viruses. The AdSur-SYE increased gene transduction efficiency 6.4-fold higher than did AdSur in single cells derived from human pancreatic cancer, whereas the infectivity of both vectors was almost the same in the pancreas and other cancers. Immunostaining showed that most EGFP(+) cells were cytokeratin-positive in the sliced tissues, indicating that pancreatic cancer cells but not stromal cells were injected with AdSur-SYE. AdSur-SYE resulted in a stronger oncolysis in the primary pancreatic cancer cells co-cultured with mouse embryonic fibroblasts than AdSur did. CRAd in combination with a tumor-targeting ligand is promising as a next-generation of oncolytic virotherapy for pancreatic cancer.

  6. Maternal endotoxemia, fetal anomalies, and central nervous system damage: a rat model of a human problem.

    Science.gov (United States)

    Ornoy, A; Altshuler, G

    1976-01-15

    Endotoxemia is a common consequence of the gram-negative urinary tract infections that complicate human pregnancies. Only rarely, however, have the effects of maternal endotoxemia been evaluated by animal experiments or by human investigations. Data of the Collaborative Perinatal Study suggest an association between maternal endotoxemia and fetal central nervous system damage. For these reasons we performed controlled studies of the fetal effects of treatment of pregnant rats, at appropriate gestational ages, with E. coli endotoxin. We found a maximum 7 per cent incidence of fetal anomalies in the treated animals but no anomalies in controls. Placental light microscopy examinations indicated the mechanism to include Shwartzman-lixemia produces periventricular leukomalacia. We obtained an incidence of neuronal necrosis in treated fetuses that was 10 times greater than in control fetuses. It is therefore of importance that additional studies of the pathologic effects of endotoxin be performed.

  7. Complex epithelial remodeling underlie the fusion event in early fetal development of the human penile urethra.

    Science.gov (United States)

    Shen, Joel; Overland, Maya; Sinclair, Adriane; Cao, Mei; Yue, Xuan; Cunha, Gerald; Baskin, Laurence

    We recently described a two-step process of urethral plate canalization and urethral fold fusion to form the human penile urethra. Canalization ("opening zipper") opens the solid urethral plate into a groove, and fusion ("closing zipper") closes the urethral groove to form the penile urethra. We hypothesize that failure of canalization and/or fusion during human urethral formation can lead to hypospadias. Herein, we use scanning electron microscopy (SEM) and analysis of transverse serial sections to better characterize development of the human fetal penile urethra as contrasted to the development of the human fetal clitoris. Eighteen 7-13 week human fetal external genitalia specimens were analyzed by SEM, and fifteen additional human fetal specimens were sectioned for histologic analysis. SEM images demonstrate canalization of the urethral/vestibular plate in the developing male and female external genitalia, respectively, followed by proximal to distal fusion of the urethral folds in males only. The fusion process during penile development occurs sequentially in multiple layers and through the interlacing of epidermal "cords". Complex epithelial organization is also noted at the site of active canalization. The demarcation between the epidermis of the shaft and the glans becomes distinct during development, and the epithelial tag at the distal tip of the penile and clitoral glans regresses as development progresses. In summary, SEM analysis of human fetal specimens supports the two-zipper hypothesis of formation of the penile urethra. The opening zipper progresses from proximal to distal along the shaft of the penis and clitoris into the glans in identical fashion in both sexes. The closing zipper mechanism is active only in males and is not a single process but rather a series of layered fusion events, uniquely different from the simple fusion of two epithelial surfaces as occurs in formation of the palate and neural tube. Copyright © 2016 International Society

  8. Complex epithelial remodeling underlie the fusion event in early fetal development of the human penile urethra

    Science.gov (United States)

    Sinclair, Adriane; Cao, Mei; Yue, Xuan; Cunha, Gerald; Baskin, Laurence

    2016-01-01

    We recently described a two-step process of urethral plate canalization and urethral fold fusion to form the human penile urethra. Canalization (“opening zipper”) opens the solid urethral plate into a groove, and fusion (“closing zipper”) closes the urethral groove to form the penile urethra. We hypothesize that failure of canalization and/or fusion during human urethral formation can lead to hypospadias. Herein, we use scanning electron microscopy (SEM) and analysis of transverse serial sections to better characterize development of the human fetal penile urethra as contrasted to the development of the human fetal clitoris. Eighteen 7-13 week human fetal external genitalia specimens were analyzed by SEM, and fifteen additional human fetal specimens were sectioned for histologic analysis. SEM images demonstrate canalization of the urethral/vestibular plate in the developing male and female external genitalia, respectively, followed by proximal to distal fusion of the urethral folds in males only. The fusion process during penile development occurs sequentially in multiple layers and through the interlacing of epidermal “cords”. Complex epithelial organization is also noted at the site of active canalization. The demarcation between the epidermis of the shaft and the glans becomes distinct during development, and the epithelial tag at the distal tip of the penile and clitoral glans regresses as development progresses. In summary, SEM analysis of human fetal specimens supports the two-zipper hypothesis of formation of the penile urethra. The opening zipper progresses from proximal to distal along the shaft of the penis and clitoris into the glans in identical fashion in both sexes. The closing zipper mechanism is active only in males and is not a single process but rather a series of layered fusion events, uniquely different from the simple fusion of two epithelial surfaces as occurs in formation of the palate and neural tube. PMID:27397682

  9. FA1 immunoreactivity in endocrine tumours and during development of the human fetal pancreas; negative correlation with glucagon expression

    DEFF Research Database (Denmark)

    Tornehave, D; Jensen, Charlotte Harken; Teisner, B;

    1996-01-01

    proteins delta and notch and to the murine preadipocyte differentiation factor Pref-1. These proteins participate in determining cell fate choices during differentiation. We now report that FA1 immunoreactivity is present in a number of neuroectodermally derived tumours as well as in pancreatic endocrine...... tumours. A negative correlation between FA1 and glucagon immunoreactants in these tumours prompted a reexamination of FA1 immunoreactants during fetal pancreatic development. At the earliest stages of development, FA1 was expressed by most of the non-endocrine parenchymal cells and, with ensuing...... development, gradually disappeared from these cells and became restricted to insulin-producing beta cells. Throughout development FA1 was not detected in endocrine glucagon, somatostatin or pancreatic polypeptide cells. Moreover, developing insulin cells that coexpressed glucagon were negative for FA1. Thus...

  10. Demonstration of pepsinogen C in human pancreatic islets

    DEFF Research Database (Denmark)

    Poulsen, Steen Seier

    1987-01-01

    Pancreatic tissue from 16 post mortem kidney donors have been examined for the content of pepsinogens. A zymogen with electrophoretic mobility, isoelectric point and molecular weight equal to that of pepsinogen C of gastric origin was found in all specimens. A comparison between pepsinogen C extr...

  11. A Novel Ras Inhibitor (MDC-1016 Reduces Human Pancreatic Tumor Growth in Mice

    Directory of Open Access Journals (Sweden)

    Gerardo G Mackenzie

    2013-10-01

    Full Text Available Pancreatic cancer has one of the poorest prognoses among all cancers partly because of its persistent resistance to chemotherapy. The currently limited treatment options for pancreatic cancer underscore the need for more efficient agents. Because activating Kras mutations initiate and maintain pancreatic cancer, inhibition of this pathway should have a major therapeutic impact. We synthesized phospho-farnesylthiosalicylic acid (PFTS; MDC-1016 and evaluated its efficacy, safety, and metabolism in preclinical models of pancreatic cancer. PFTS inhibited the growth of human pancreatic cancer cells in culture in a concentration- and time-dependent manner. In an MIA PaCa-2 xenograft mouse model, PFTS at a dose of 50 and 100 mg/kg significantly reduced tumor growth by 62% and 65% (P < .05 vs vehicle control. Furthermore, PFTS prevented pancreatitis-accelerated acinar-to-ductal metaplasia in mice with activated Kras. PFTS appeared to be safe, with the animals showing no signs of toxicity during treatment. Following oral administration, PFTS was rapidly absorbed, metabolized to FTS and FTS glucuronide, and distributed through the blood to body organs. Mechanistically, PFTS inhibited Ras-GTP, the active form of Ras, both in vitro and in vivo, leading to the inhibition of downstream effector pathways c-RAF/mitogen-activated protein-extracellular signal-regulated kinase (ERK kinase (MEK/ERK1/2 kinase and phosphatidylinositol 3-kinase/AKT. In addition, PFTS proved to be a strong combination partner with phospho-valproic acid, a novel signal transducer and activator of transcription 3 (STAT3 inhibitor, displaying synergy in the inhibition of pancreatic cancer growth. In conclusion, PFTS, a direct Ras inhibitor, is an efficacious agent for the treatment of pancreatic cancer in preclinical models, deserving further evaluation.

  12. Experimental Animal Models of Pancreatic Carcinogenesis for Prevention Studies and Their Relevance to Human Disease

    Directory of Open Access Journals (Sweden)

    Hitoshi Nakagama

    2011-02-01

    Full Text Available Pancreatic cancer is difficult to cure, so its prevention is very important. For this purpose, animal model studies are necessary to develop effective methods. Injection of N-nitrosobis(2-oxopropylamine (BOP into Syrian golden hamsters is known to induce pancreatic ductal adenocarcinomas, the histology of which is similar to human tumors. Moreover, K-ras activation by point mutations and p16 inactivation by aberrant methylation of 5’ CpG islands or by homozygous deletions have been frequently observed in common in both the hamster and humans. Thus, this chemical carcinogenesis model has an advantage of histopathological and genetic similarity to human pancreatic cancer, and it is useful to study promotive and suppressive factors. Syrian golden hamsters are in a hyperlipidemic state even under normal dietary conditions, and a ligand of peroxizome proliferator-activated receptor gamma was found to improve the hyperlipidemia and suppress pancreatic carcinogenesis. Chronic inflammation is a known important risk factor, and selective inhibitors of inducible nitric oxide synthase and cyclooxygenase-2 also have protective effects against pancreatic cancer development. Anti-inflammatory and anti-hyperlipidemic agents can thus be considered candidate chemopreventive agents deserving more attention.

  13. Experimental Animal Models of Pancreatic Carcinogenesis for Prevention Studies and Their Relevance to Human Disease

    Energy Technology Data Exchange (ETDEWEB)

    Takahashi, Mami, E-mail: mtakahas@ncc.go.jp; Hori, Mika; Mutoh, Michihiro [Division of Cancer Development System, Carcinogenesis Research Group, National Cancer Center Research Institute, 1-1, Tsukiji 5-chome, Chuo-ku, Tokyo 104-0045 (Japan); Wakabayashi, Keiji [Graduate School of Nutritional and Environmental Sciences, University of Shizuoka, Yada 52-1, Suruga-ku, Shizuoka 422-8526 (Japan); Nakagama, Hitoshi [Division of Cancer Development System, Carcinogenesis Research Group, National Cancer Center Research Institute, 1-1, Tsukiji 5-chome, Chuo-ku, Tokyo 104-0045 (Japan)

    2011-02-09

    Pancreatic cancer is difficult to cure, so its prevention is very important. For this purpose, animal model studies are necessary to develop effective methods. Injection of N-nitrosobis(2-oxopropyl)amine (BOP) into Syrian golden hamsters is known to induce pancreatic ductal adenocarcinomas, the histology of which is similar to human tumors. Moreover, K-ras activation by point mutations and p16 inactivation by aberrant methylation of 5′ CpG islands or by homozygous deletions have been frequently observed in common in both the hamster and humans. Thus, this chemical carcinogenesis model has an advantage of histopathological and genetic similarity to human pancreatic cancer, and it is useful to study promotive and suppressive factors. Syrian golden hamsters are in a hyperlipidemic state even under normal dietary conditions, and a ligand of peroxizome proliferator-activated receptor gamma was found to improve the hyperlipidemia and suppress pancreatic carcinogenesis. Chronic inflammation is a known important risk factor, and selective inhibitors of inducible nitric oxide synthase and cyclooxygenase-2 also have protective effects against pancreatic cancer development. Anti-inflammatory and anti-hyperlipidemic agents can thus be considered candidate chemopreventive agents deserving more attention.

  14. Herpes simplex virus thymidine kinase and ganciclovir suicide gene therapy for human pancreatic cancer

    Institute of Scientific and Technical Information of China (English)

    Jing Wang; Xiao-Xuan Lu; Dao-Zhen Chen; Shu-Feng Li; Li-Shan Zhang

    2004-01-01

    AIM: To investigate the in vitro effects of suicide gene therepy system of herpes simplex virus thymidine kinase gene (HSV-TK) in combination with the treatment of nucleotide analog-ganciclovir (GCV) on human pancreatic cancer, and to provide a novel clinical therapeutic method for human pancreatic cancer.METHODS: We used a replication defective recombinant retrovirus vector GINaTK (bearing HSV-TK gene) to make packaging cell PA317 produce progeny virions. We then transferred the HSV-TK gene to target cells SW1990 using these progeny virions, and treated these gene-modified tumor cells with GCV to study the sensitivity of the cells to GCV and their bystander effects by routine MTT-method.RESULTS: Packaging cell PA317/TK was successfully constructed, and we acquired SW1990/TK through virus progeny infection. These gene-modified pancreatic cancer cells were sensitive to the treatment of GCV compared with unmodified tumor cells (t=4.15, n=10, P<0.0025). We also observed a remarkable bystander effect by mixing two kinds of cells at different ratio.CONCLUSION: Our data demonstrate that HSV-TK/GCV suicide gene therapy system is effective for treating experimental human pancreatic cancer, which is largely resistant to the common therapies, so the suicide gene therapy system may be a potential treatment approach for pancreatic cancer.

  15. p-[{sup 123}I]iodo-l-phenylalanine for detection of pancreatic cancer: basic investigations of the uptake characteristics in primary human pancreatic tumour cells and evaluation in in vivo models of human pancreatic adenocarcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Samnick, Samuel; Hellwig, Dirk; Kirsch, Carl-Martin [Department of Nuclear Medicine, Saarland University Medical Center, 66421, Homburg/Saar (Germany); Romeike, Bernd F.M.; Feiden, Wolfgang [Department of Neuropathology, Saarland University Medical Center, Homburg/Saar (Germany); Kubuschok, Boris [Department of Internal Medicine I, Saarland University Medical Center, Homburg/Saar (Germany); Amon, Michaela; Menger, Michael D. [Department of Clinical Experimental Surgery, Saarland University Medical Center, Homburg/Saar (Germany)

    2004-04-01

    Pancreatic cancer is associated with the worst 5-year survival rate of any human cancer. This high mortality is due, in part, to difficulties in establishing early and accurate diagnosis. Because most tumours share the ability to accumulate amino acids more effectively than normal tissues and any other pathology, assessment of amino acid transport in tumour cells using radiolabelled amino acids has become one of the most promising tools for tumour imaging. This study investigated the potential of p-[{sup 123}I]iodo-l-phenylalanine (IPA) for detection of pancreatic cancer by single-photon emission tomography. IPA affinity for pancreatic tumour was investigated in human pancreatic adenocarcinoma PaCa44 and PanC1 cells, followed by analysis of the underlying mechanisms of tracer accumulation in neoplastic cells. Thereafter, IPA was evaluated for targeting of pancreatic tumours using SCID mice engrafted with primary human pancreatic adenocarcinoma cells, as well as in acute inflammation models in immunocompetent mice and rats. IPA accumulated intensively in human pancreatic tumour cells. Radioactivity accumulation in tumour cells following a 30-min incubation at 37 C/pH 7.4 varied from 41% to 58% of the total loaded activity per 10{sup 6} cells. The cellular uptake was temperature and pH dependent and predominantly mediated by specific carriers for neutral amino acids, namely the sodium-independent and l-leucine-preferring (L-system) transporter and the alanine-, serine- and cysteine-preferring (ASC-system) transporter. Protein incorporation was less than 8%. Biodistribution studies showed rapid localization of the tracer to tumours, reaching 10%{+-}2.5% to 15%{+-}3% of the injected dose per gram (I.D./g) in heterotopic tumours compared with 17%{+-}3.5% to 22%{+-}4.3% I.D./g in the orthotopic tumours, at 60 and 240 min post injection of IPA, respectively. In contrast, IPA uptake in the gastrointestinal tract and areas of inflammation remained moderate and decreased

  16. Diagnosis of the human fetal age based on the development of the normal kidney

    Directory of Open Access Journals (Sweden)

    Lizardo-Daudt Helena Maria

    2002-01-01

    Full Text Available Background and aims: The diagnosis of human fetal age is usually estimated based on the measurement of crown-rump length or crown-heel length and the weight of the fetus. However, this estimate is not totally accurate and sometimes is necessary to combine other data to determine the fetal age. An analysis of the normal embryological development of the kidney may assist in this determination. The histology of this process, although well described, lacks photographic documentation. We intend to fill this gap by providing histologists and pathologists, especially inexperienced ones, with information about the staging of the renal development through microphotography. The objective of the present study was to achieve greater accuracy for the diagnosis of human fetal age through the proposed classification and the photographic documentation presented. Material and methods: Normal embryological development of the human kidney was studied by light microscopy. The fetal period from 6 to 40 weeks of gestation was observed according the stage of maturity of glomeruli and tubules; localization of glomeruli, occurrence of nephrogenic tissue and cortico-medullary differentiation. At least 5 different exams were observed from each week of development. Two hundred four exams were analyzed in the whole study. The histological characteristics were quantified and the process was documented by microphotography. Results and final considerations: The fetal development of the kidney was divided into 8 stages, which was documented through microphotography. Nephron structural formation occurred until the 34th week of prenatal development. From the 35th week on, tubules and glomeruli continued to mature without the formation of new nephrons. The proposed classification intends to improve the accuracy of the fetal age diagnosis.

  17. Blood flow in the human fetal descending aorta : a pulsed Doppler study

    NARCIS (Netherlands)

    L. Pijpers (Leendert)

    1985-01-01

    textabstractIn 1628 William Harvey introduced his concept ofthe human circulation. Although a lot of studies concerning the fetal circulation were done before it was not until the 1930s that Barcroft (1934, 1939) and associates performed radiograpbic studies on the feta! goal and lamb to establish t

  18. Towards a New Study on Associative Learning in Human Fetuses: Fetal Associative Learning in Primates

    Science.gov (United States)

    Kawai, Nobuyuki

    2010-01-01

    Research has revealed that fetuses can learn from events in their environment. The most convincing evidence for fetal learning is habituation to vibroacoustic stimulation (VAS) in human fetuses and classical conditioning in rat fetuses. However, these two research areas have been independent of each other. There have been few attempts at classical…

  19. Reduced cell number in the neocortical part of the human fetal brain in Down syndrome

    DEFF Research Database (Denmark)

    Larsen, K.B.; Laursen, H.; Graem, N.

    2008-01-01

    Mental retardation is seen in all individuals with Down syndrome (DS) and different brain abnormalities are reported. The aim of this study was to investigate if mental retardation at least in part is a result of a lower cell number in the neocortical part of the human fetal forebrain. We therefore...

  20. Dissecting human cerebral organoids and fetal neocortex using single-cell RNAseq

    Science.gov (United States)

    Treutlein, Barbara

    Cerebral organoids - three-dimensional cultures of human cerebral tissue derived from pluripotent stem cells - have emerged as models of human cortical development. However, the extent to which in vitro organoid systems recapitulate neural progenitor cell proliferation and neuronal differentiation programs observed in vivo remains unclear. Here we use single-cell RNA sequencing (scRNA-seq) to dissect and compare cell composition and progenitor-to-neuron lineage relationships in human cerebral organoids and fetal neocortex. Covariation network analysis using the fetal neocortex data reveals known and novel interactions among genes central to neural progenitor proliferation and neuronal differentiation. In the organoid, we detect diverse progenitors and differentiated cell types of neuronal and mesenchymal lineages, and identify cells that derived from regions resembling the fetal neocortex. We find that these organoid cortical cells use gene expression programs remarkably similar to those of the fetal tissue in order to organize into cerebral cortex-like regions. Our comparison of in vivo and in vitro cortical single cell transcriptomes illuminates the genetic features underlying human cortical development that can be studied in organoid cultures.

  1. Platelet-rich plasma can replace fetal bovine serum in human meniscus cell cultures

    NARCIS (Netherlands)

    Gonzales, V.K.; Mulder, E.L.W. de; Boer, T. den; Hannink, G.; Tienen, T.G. van; Heerde, W.L. van; Buma, P.

    2013-01-01

    Concerns over fetal bovine serum (FBS) limit the clinical application of cultured tissue-engineered constructs. Therefore, we investigated if platelet-rich plasma (PRP) can fully replace FBS for meniscus tissue engineering purposes. Human PRP and platelet-poor plasma (PPP) were isolated from three h

  2. Resveratrol inhibits steroidogenesis in human fetal adrenocortical cells at the end of first trimester

    DEFF Research Database (Denmark)

    Savchuk, Iuliia; Morvan, Marie-Line; Søeborg, Tue

    2017-01-01

    steroidogenesis at gestational weeks (GW) 9-12. METHODS AND RESULTS: Adrenals from aborted fetuses (GW10-12) were used to prepare primary cultures of human fetal adrenocortical cells (HFAC). HFAC were treated in the presence or absence of ACTH (10 ng/ml) with or without resveratrol (10 μM) for 24 hours...

  3. An attempt to eliminate fibroblast-like cells from primary cultures of fetal human livers.

    Directory of Open Access Journals (Sweden)

    Tokiwa,Takayoshi

    1986-04-01

    Full Text Available The elimination of fibroblast-like cells from primary cultures of fetal human livers was studied. A fibroblast-like cell line (HuF, which was obtained by subculturing fetal human liver cells 4 or more times, was briefly treated with hydrocortisone (HC or putrescine (PUT. The growth of HuF cells was inhibited by HC at a concentration of 10(-2 M and by PUT at a concentration higher than 10(-3 M. Long-term treatment of HuF cells with 10(-3 M HC inhibited the growth of the cells. Primary cultures of fetal human livers were made in medium containing HC or PUT, and morphological and functional examinations were made. The cultures were predominantly composed of epithelial-like cells, with few fibroblast-like cells, when the HC concentration was 10(-5M to 10(-3 M. A high amount of albumin was secreted at these concentrations of HC. On the other hand, at 10(-3 M PUT, many epithelial-like cells were seen, but albumin was undetectable. The present results indicate that albumin-producing epithelial-like cells can be selectively maintained in medium containing HC, in primary cultures of fetal human livers.

  4. Towards a New Study on Associative Learning in Human Fetuses: Fetal Associative Learning in Primates

    Science.gov (United States)

    Kawai, Nobuyuki

    2010-01-01

    Research has revealed that fetuses can learn from events in their environment. The most convincing evidence for fetal learning is habituation to vibroacoustic stimulation (VAS) in human fetuses and classical conditioning in rat fetuses. However, these two research areas have been independent of each other. There have been few attempts at classical…

  5. Human cerebral organoids recapitulate gene expression programs of fetal neocortex development.

    Science.gov (United States)

    Camp, J Gray; Badsha, Farhath; Florio, Marta; Kanton, Sabina; Gerber, Tobias; Wilsch-Bräuninger, Michaela; Lewitus, Eric; Sykes, Alex; Hevers, Wulf; Lancaster, Madeline; Knoblich, Juergen A; Lachmann, Robert; Pääbo, Svante; Huttner, Wieland B; Treutlein, Barbara

    2015-12-22

    Cerebral organoids-3D cultures of human cerebral tissue derived from pluripotent stem cells-have emerged as models of human cortical development. However, the extent to which in vitro organoid systems recapitulate neural progenitor cell proliferation and neuronal differentiation programs observed in vivo remains unclear. Here we use single-cell RNA sequencing (scRNA-seq) to dissect and compare cell composition and progenitor-to-neuron lineage relationships in human cerebral organoids and fetal neocortex. Covariation network analysis using the fetal neocortex data reveals known and previously unidentified interactions among genes central to neural progenitor proliferation and neuronal differentiation. In the organoid, we detect diverse progenitors and differentiated cell types of neuronal and mesenchymal lineages and identify cells that derived from regions resembling the fetal neocortex. We find that these organoid cortical cells use gene expression programs remarkably similar to those of the fetal tissue to organize into cerebral cortex-like regions. Our comparison of in vivo and in vitro cortical single-cell transcriptomes illuminates the genetic features underlying human cortical development that can be studied in organoid cultures.

  6. Human fetal testis Leydig cell disruption by exposure to the pesticide dieldrin at low concentrations.

    Science.gov (United States)

    Fowler, Paul A; Abramovich, David R; Haites, Neva E; Cash, Phillip; Groome, Nigel P; Al-Qahtani, Ahmed; Murray, Tessa J; Lea, Richard G

    2007-11-01

    Declining human reproductive health over the last 60 years has been proposed to be due to effects of environmental chemicals, especially endocrine disrupting compounds, on fetal development. We investigated whether a model pesticide, dieldrin, at concentrations within both maternal circulation and environmental ranges (1 pmol/l = 0.0004 p.p.b. = 380.9 pg/l), could disrupt the human fetal testis. Human fetal testes were collected during the second trimester, a critical period of male sexual differentiation (development and masculinization). Testis explants were cultured for 24 h in the presence and absence of LH (10-1000 IU LH/l) and dieldrin (1 pmol and 1 nmol/l). Endocrine, immunohistological and proteome characteristics of the tissues were investigated. Exposure to dieldrin reduced LH-induced testosterone secretion (P Dieldrin altered proteins associated with cancer, apoptosis, transcription and development. Wnt-2b was reduced 3-fold and immunolocalized to Leydig and Sertoli cells. Dieldrin also reversed some LH-induced changes in protein expression, supporting the conclusion that Leydig cell function is at risk from environmental chemicals. Our findings indicate that exposure to very low, biologically relevant, concentrations of environmental chemicals could affect the fetal human Leydig cell, reducing testosterone secretion and potentially leading to subtle dysregulation of reproductive development and adult fecundity.

  7. Developmental origins of health and disease: experimental and human evidence of fetal programming for metabolic syndrome.

    Science.gov (United States)

    de Gusmão Correia, M L; Volpato, A M; Águila, M B; Mandarim-de-Lacerda, C A

    2012-07-01

    The concept of developmental origins of health and disease has been defined as the process through which the environment encountered before birth, or in infancy, shapes the long-term control of tissue physiology and homeostasis. The evidence for programming derives from a large number of experimental and epidemiological observations. Several nutritional interventions during diverse phases of pregnancy and lactation in rodents are associated with fetal and neonatal programming for metabolic syndrome. In this paper, recent experimental models and human epidemiological studies providing evidence for the fetal programming associated with the development of metabolic syndrome and related diseases are revisited.

  8. Induced pluripotent stem (iPS) cells from human fetal stem cells.

    Science.gov (United States)

    Guillot, Pascale V

    2016-02-01

    Pluripotency defines the ability of stem cells to differentiate into all the lineages of the three germ layers and self-renew indefinitely. Somatic cells can regain the developmental potential of embryonic stem cells following ectopic expression of a set of transcription factors or, in certain circumstances, via modulation of culture conditions and supplementation with small molecule, that is, induced pluripotent stem (iPS) cells. Here, we discuss the use of fetal tissues for reprogramming, focusing in particular on stem cells derived from human amniotic fluid, and the development of chemical reprogramming. We next address the advantages and disadvantages of deriving pluripotent cells from fetal tissues and the potential clinical applications.

  9. Proteolytic processing of anti-Müllerian hormone differs between human fetal testes and adult ovaries

    DEFF Research Database (Denmark)

    Mamsen, Linn; Petersen, TS; Jeppesen, JV

    2015-01-01

    and specificity of a panel of five novel high-affinity AMH monoclonal antibodies. Two recognize the mature C-terminal form of AMH, whereas three recognize the active pro-mature form of AMH in human tissue. The antibodies were tested on fetal male testicular and mesonephric tissue aged 8-19 weeks post conception...... of AMH was hardly detected in Sertoli cells, but was readily detected in GCs. This particular form was also located to the nucleus in GCs, whereas the other investigated AMH forms remained in the cytoplasm. Interestingly, the distribution of the AMH forms in the fetal serum of boys showed...

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

    Science.gov (United States)

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

    2017-01-01

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

  11. Recent advances in the prenatal interrogation of the human fetal genome.

    Science.gov (United States)

    Hui, Lisa; Bianchi, Diana W

    2013-02-01

    The amount of genetic and genomic information obtainable from the human fetus during pregnancy is accelerating at an unprecedented rate. Two themes have dominated recent technological advances in prenatal diagnosis: interrogation of the fetal genome in increasingly high resolution and the development of non-invasive methods of fetal testing using cell-free DNA in maternal plasma. These two areas of advancement have now converged with several recent reports of non-invasive assessment of the entire fetal genome from maternal blood. However, technological progress is outpacing the ability of the healthcare providers and patients to incorporate these new tests into existing clinical care, and further complicates many of the economic and ethical dilemmas in prenatal diagnosis. This review summarizes recent work in this field and discusses the integration of these new technologies into the clinic and society.

  12. GMP-grade human fetal liver-derived mesenchymal stem cells for clinical transplantation.

    Science.gov (United States)

    Larijani, Bagher; Aghayan, Hamid-Reza; Goodarzi, Parisa; Arjmand, Babak

    2015-01-01

    Stem cell therapy seems a promising avenue in regenerative medicine. Within various stem cells, mesenchymal stem cells have progressively used for cellular therapy. Because of the age-related decreasing in the frequency and differentiating capacity of adult MSCs, fetal tissues such as fetal liver, lung, pancreas, spleen, etc. have been introduced as an alternative source of MSCs for cellular therapy. On the other hand, using stem cells as advanced therapy medicinal products, must be performed in compliance with cGMP as a quality assurance system to ensure the safety, quality, and identity of cell products during translation from the basic stem cell sciences into clinical cell transplantation. In this chapter the authors have demonstrated the manufacturing of GMP-grade human fetal liver-derived mesenchymal stem cells.

  13. The Role of Placental Homeobox Genes in Human Fetal Growth Restriction

    Directory of Open Access Journals (Sweden)

    Padma Murthi

    2011-01-01

    Full Text Available Fetal growth restriction (FGR is an adverse pregnancy outcome associated with significant perinatal and paediatric morbidity and mortality, and an increased risk of chronic disease later in adult life. One of the key causes of adverse pregnancy outcome is fetal growth restriction (FGR. While a number of maternal, fetal, and environmental factors are known causes of FGR, the majority of FGR cases remain idiopathic. These idiopathic FGR pregnancies are frequently associated with placental insufficiency, possibly as a result of placental maldevelopment. Understanding the molecular mechanisms of abnormal placental development in idiopathic FGR is, therefore, of increasing importance. Here, we review our understanding of transcriptional control of normal placental development and abnormal placental development associated with human idiopathic FGR. We also assess the potential for understanding transcriptional control as a means for revealing new molecular targets for the detection, diagnosis, and clinical management of idiopathic FGR.

  14. [Morphogenesis of Human Fetal Thymus during Weeks 22-27 of Development].

    Science.gov (United States)

    Kulida, L V; Peretyatko, L P; Nazarov, S B

    2015-01-01

    Distinctive features of human fetal thymus morphogenesis in early ontogeny in the case of uncomplicated pregnancy have been characterized. A steady increase of thymus dimensions and weight occurred concomitantly to differentiation of morphofunctional zones within the organ. Cell differentiation in the subcapsular and inner cortical zones of the thymus lobes was manifested as changes in parameters of expression of T-lymphocyte antigens CD1, CD2, and CD3 and ultrastructural features of reticuloepithelial cells (REC) type I and II forming a microenvironment for lymphocytes. RECs of the medullar zone formed a glomerular syncytium with desmosomal interepithelial contacts by week 22 of fetal development. Small lymphocytes predominated among thymocytes (66%). Hassall's corpuscles, the structural correlates of morphological and functional maturity, predominated in the fetal thymuses during developmental weeks 25-27.

  15. Ectopic PDX-1 Expression Directly Reprograms Human Keratinocytes along Pancreatic Insulin-Producing Cells Fate

    Science.gov (United States)

    Chernichovski, Ellad; Nakar, Odelia; Winkler, Eyal; Mazkereth, Ram; Orenstein, Arie; Bar-Meir, Eran; Ravassard, Philippe; Meivar-Levy, Irit; Ferber, Sarah

    2011-01-01

    Background Cellular differentiation and lineage commitment have previously been considered irreversible processes. However, recent studies have indicated that differentiated adult cells can be reprogrammed to pluripotency and, in some cases, directly into alternate committed lineages. However, although pluripotent cells can be induced in numerous somatic cell sources, it was thought that inducing alternate committed lineages is primarily only possible in cells of developmentally related tissues. Here, we challenge this view and analyze whether direct adult cell reprogramming to alternate committed lineages can cross the boundaries of distinct developmental germ layers. Methodology/Principal Findings We ectopically expressed non-integrating pancreatic differentiation factors in ectoderm-derived human keratinocytes to determine whether these factors could directly induce endoderm-derived pancreatic lineage and β-cell-like function. We found that PDX-1 and to a lesser extent other pancreatic transcription factors, could rapidly and specifically activate pancreatic lineage and β-cell-like functional characteristics in ectoderm-derived human keratinocytes. Human keratinocytes transdifferentiated along the β cell lineage produced processed and secreted insulin in response to elevated glucose concentrations. Using irreversible lineage tracing for KRT-5 promoter activity, we present supporting evidence that insulin-positive cells induced by ectopic PDX-1 expression are generated in ectoderm derived keratinocytes. Conclusions/Significance These findings constitute the first demonstration of human ectoderm cells to endoderm derived pancreatic cells transdifferentiation. The study represents a proof of concept which suggests that transcription factors induced reprogramming is wider and more general developmental process than initially considered. These results expanded the arsenal of adult cells that can be used as a cell source for generating functional endocrine

  16. Discovery and Characterization of piRNAs in the Human Fetal Ovary

    Directory of Open Access Journals (Sweden)

    Zev Williams

    2015-10-01

    Full Text Available Piwi-interacting RNAs (piRNAs, a class of 26- to 32-nt non-coding RNAs (ncRNAs, function in germline development, transposon silencing, and epigenetic regulation. We performed deep sequencing and annotation of untreated and periodate-treated small RNA cDNA libraries from human fetal and adult germline and reference somatic tissues. This revealed abundant piRNAs originating from 150 piRNA-encoding genes, including some exhibiting gender-specific expression, in fetal ovary and adult testis—developmental periods coinciding with mitotic cell divisions expanding fetal germ cells prior to meiotic divisions. The absence of reads mapping uniquely to annotated piRNA genes demonstrated their paucity in fetal testis and adult ovary and absence in somatic tissues. We curated human piRNA-expressing regions and defined their precise borders and observed piRNA-guided cleavage of transcripts antisense to some piRNA-producing genes. This study provides insights into sex-specific mammalian piRNA expression and function and serves as a reference for human piRNA analysis and annotation.

  17. Expression and activation of caspase-6 in human fetal and adult tissues.

    Directory of Open Access Journals (Sweden)

    Nelly Godefroy

    Full Text Available Caspase-6 is an effector caspase that has not been investigated thoroughly despite the fact that Caspase-6 is strongly activated in Alzheimer disease brains. To understand the full physiological impact of Caspase-6 in humans, we investigated Caspase-6 expression. We performed western blot analyses to detect the pro-Caspase-6 and its active p20 subunit in fetal and adult lung, kidney, brain, spleen, muscle, stomach, colon, heart, liver, skin, and adrenals tissues. The levels were semi-quantitated by densitometry. The results show a ubiquitous expression of Caspase-6 in most fetal tissues with the lowest levels in the brain and the highest levels in the gastrointestinal system. Caspase-6 active p20 subunits were only detected in fetal stomach. Immunohistochemical analysis of a human fetal embryo showed active Caspase-6 positive apoptotic cells in the dorsal root ganglion, liver, lung, kidney, ovary, skeletal muscle and the intestine. In the adult tissues, the levels of Caspase-6 were lower than in fetal tissues but remained high in the colon, stomach, lung, kidney and liver. Immunohistological analyses revealed that active Caspase-6 was abundant in goblet cells and epithelial cells sloughing off the intestinal lining of the adult colon. These results suggest that Caspase-6 is likely important in most tissues during early development but is less involved in adult tissues. The low levels of Caspase-6 in fetal and adult brain indicate that increased expression as observed in Alzheimer Disease is a pathological condition. Lastly, the high levels of Caspase-6 in the gastrointestinal system indicate a potential specific function of Caspase-6 in these tissues.

  18. Nucleotide sequence of cloned cDNA for human pancreatic kallikrein.

    Science.gov (United States)

    Fukushima, D; Kitamura, N; Nakanishi, S

    1985-12-31

    Cloned cDNA sequences for human pancreatic kallikrein have been isolated and determined by molecular cloning and sequence analysis. The identity between human pancreatic and urinary kallikreins is indicated by the complete coincidence between the amino acid sequence deduced from the cloned cDNA sequence and that reported partially for urinary kallikrein. The active enzyme form of the human pancreatic kallikrein consists of 238 amino acids and is preceded by a signal peptide and a profragment of 24 amino acids. A sequence comparison of this with other mammalian kallikreins indicates that key amino acid residues required for both serine protease activity and kallikrein-like cleavage specificity are retained in the human sequence, and residues corresponding to some external loops of the kallikrein diverge from other kallikreins. Analyses by RNA blot hybridization, primer extension, and S1 nuclease mapping indicate that the pancreatic kallikrein mRNA is also expressed in the kidney and sublingual gland, suggesting the active synthesis of urinary kallikrein in these tissues. Furthermore, the tissue-specific regulation of the expression of the members of the human kallikrein gene family has been discussed.

  19. Insights in spatio-temporal characterization of human fetal neural stem cells.

    Science.gov (United States)

    Martín-Ibáñez, Raquel; Guardia, Inés; Pardo, Mónica; Herranz, Cristina; Zietlow, Rike; Vinh, Ngoc-Nga; Rosser, Anne; Canals, Josep M

    2017-05-01

    Primary human fetal cells have been used in clinical trials of cell replacement therapy for the treatment of neurodegenerative disorders such as Huntington's disease (HD). However, human fetal primary cells are scarce and difficult to work with and so a renewable source of cells is sought. Human fetal neural stem cells (hfNSCs) can be generated from human fetal tissue, but little is known about the differences between hfNSCs obtained from different developmental stages and brain areas. In the present work we characterized hfNSCs, grown as neurospheres, obtained from three developmental stages: 4-5, 6-7 and 8-9weeks post conception (wpc) and four brain areas: forebrain, cortex, whole ganglionic eminence (WGE) and cerebellum. We observed that, as fetal brain development proceeds, the number of neural precursors is diminished and post-mitotic cells are increased. In turn, primary cells obtained from older embryos are more sensitive to the dissociation process, their viability is diminished and they present lower proliferation ratios compared to younger embryos. However, independently of the developmental stage of derivation proliferation ratios were very low in all cases. Improvements in the expansion rates were achieved by mechanical, instead of enzymatic, dissociation of neurospheres but not by changes in the seeding densities. Regardless of the developmental stage, neurosphere cultures presented large variability in the viability and proliferation rates during the initial 3-4 passages, but stabilized achieving significant expansion rates at passage 5 to 6. This was true also for all brain regions except cerebellar derived cultures that did not expand. Interestingly, the brain region of hfNSC derivation influences the expansion potential, being forebrain, cortex and WGE derived cells the most expandable compared to cerebellar. Short term expansion partially compromised the regional identity of cortical but not WGE cultures. Nevertheless, both expanded cultures were

  20. Differential role of Hedgehog signaling in human pancreatic(patho-) physiology:An up to date review

    Institute of Scientific and Technical Information of China (English)

    Eckhard Klieser; Stefan Swierczynski; Christian Mayr; Tarkan J?ger; Johanna Schmidt; Daniel Neureiter; Tobias Kiesslich; Romana Illig

    2016-01-01

    Since the discovery of the Hedgehog(Hh)pathway in drosophila melanogaster,our knowledge of the role of Hh in embryonic development,inflammation,and cancerogenesis in humans has dramatically increased over the last decades.This is the case especially concerning the pancreas,however,real therapeutic breakthroughs are missing until now.In general,Hh signaling is essential for pancreatic organogenesis,development,and tissue maturation.In the case of acute pancreatitis,Hh has a protective role,whereas in chronic pancreatitis,Hh interacts with pancreatic stellate cells,leading to destructive parenchym fibrosis and atrophy,as well as to irregular tissue remodeling with potency of initiating cancerogenesis.In vitro and in situ analysis of Hh in pancreatic cancer revealed that the Hh pathway participates in the development of pancreatic precursor lesions and ductal adenocarcinoma including critical interactions with the tumor microenvironment.The application of specific inhibitors of components of the Hh pathway is currently subject of ongoing clinical trials(phases 1 and 2).Furthermore,a combination of Hh pathway inhibitors and established chemotherapeutic drugs could also represent a promising therapeutic approach.In this review,we give a structured survey of the role of the Hh pathway in pancreatic development,pancreatitis,pancreatic carcinogenesis and pancreatic cancer as well as an overview of current clinical trials concerning Hh pathway inhibitors and pancreas cancer.

  1. Kanglaite combined Gemcitabine inhibits growth of nude mouse subcutaneous transplantation tumor of human PC-3 pancreatic cancer cell

    Institute of Scientific and Technical Information of China (English)

    WANG Wei; JIN Jian-guang; QIN Zhao-yin

    2005-01-01

    Objective:To study the mechanisms of pancreatic cancer treatment with Kanglaite combined Gemcitabine by investigating the relationship between the apoptosis and the expression of bcl-2, Bax and VEGF in pancreatic cancer cells.Methods:Nude mouse subcutaneous transplantation tumor model of Human PC-3 pancreatic cancer was established; the expressions of bcl-2, Bax and VEGF of transplantation tumor cell were determined; the earlier apoptosis rate of pancreatic cancer cell and the gross tumor volume were determined. Results:Kanglaite combined Gemcitabine remarkably decreased the protein expression of bcl-2,raised the expression of Bax,increased the apoptosis rate of the pancreatic cancer and contract the gross tumor volume. Kanglaite greatly decreased the protein expression of VEGF of the tumor cell. Conclusion:Therapeutic efficacy of Kanglaite combined Gemcitabine is far better than separate use of the two medicines in the pancreatic cancer transplantation tumor treatment.

  2. Reactivity of Human Preformed Natural Antibodies with Various Porcine Pancreatic Cells

    Institute of Scientific and Technical Information of China (English)

    张伟杰; 熊沛; 刘绍春

    2001-01-01

    The reactivity of human preformed natural antibodies (PNAbs) with various porcine pancreatic cells and its isotypes was investigated. Eighteen serum samples from patients with insulin-dependent diabetes mellitus (IDDM) and 20 serum samples from healthy human subjects were collected. The frozen sections of the pig pancreas were incubated with these sera, and subsequently incubated with FITC-conjugated goat antihuman IgG and IgM monoclonal antibodies. The reactivity of human PNAbs with various porcine pancreatic cells was determined by indirect immunofluorescence staining technique. The results showed that 55.6 % of IDDM patients and 55.0 % of healthy human individuals contained PNAbs against porcine endocrine cells. However, the percentage of strongly reacting sera in the patient group was significantly increased as compared with that in the control group. All used sera from IDDM patients and 95 % of sera from healthy controls could react to one or more of the various pancreatic cell types, including: endocrine cells, exocrine cells, vascular endothelial cells, ductal epithelial cells and macrophages. The isotypes of PNAbs contained both IgG and IgM. In view of strongly positive reactivity of PNAbs with various porcine pancreatic cells, pretransplantly cross-matching test and graft pretreatment may be necessary for survival of islet transplants.

  3. Programmed Fetal Membrane Senescence and Exosome-Mediated Signaling: A Mechanism Associated With Timing of Human Parturition

    Directory of Open Access Journals (Sweden)

    Ramkumar Menon

    2017-08-01

    Full Text Available Human parturition is an inflammatory process that involves both fetal and maternal compartments. The precise immune cell interactions have not been well delineated in human uterine tissues during parturition, but insights into human labor initiation have been informed by studies in animal models. Unfortunately, the timing of parturition relative to fetal maturation varies among viviparous species—indicative of different phylogenetic clocks and alarms—but what is clear is that important common pathways must converge to control the birth process. Herein, we hypothesize a novel signaling mechanism initiated by human fetal membrane aging and senescence-associated inflammation. Programmed events of fetal membrane aging coincide with fetal growth and organ maturation. Mechanistically, senescence involves in telomere shortening and activation of p38 mitogen-activated signaling kinase resulting in aging-associated phenotypic transition. Senescent tissues release inflammatory signals that are propagated via exosomes to cause functional changes in maternal uterine tissues. In vitro, oxidative stress causes increased release of inflammatory mediators (senescence-associated secretory phenotype and damage-associated molecular pattern markers that can be packaged inside the exosomes. These exosomes traverse through tissues layers, reach maternal tissues to increase overall inflammatory load transitioning them from a quiescent to active state. Animal model studies have shown that fetal exosomes can travel from fetal to the maternal side. Thus, aging fetal membranes and membrane-derived exosomes cargo fetal signals to the uterus and cervix and may trigger parturition. This review highlights a novel hypothesis in human parturition research based on data from ongoing research using human fetal membrane model system.

  4. Programmed Fetal Membrane Senescence and Exosome-Mediated Signaling: A Mechanism Associated With Timing of Human Parturition.

    Science.gov (United States)

    Menon, Ramkumar; Mesiano, Sam; Taylor, Robert N

    2017-01-01

    Human parturition is an inflammatory process that involves both fetal and maternal compartments. The precise immune cell interactions have not been well delineated in human uterine tissues during parturition, but insights into human labor initiation have been informed by studies in animal models. Unfortunately, the timing of parturition relative to fetal maturation varies among viviparous species-indicative of different phylogenetic clocks and alarms-but what is clear is that important common pathways must converge to control the birth process. Herein, we hypothesize a novel signaling mechanism initiated by human fetal membrane aging and senescence-associated inflammation. Programmed events of fetal membrane aging coincide with fetal growth and organ maturation. Mechanistically, senescence involves in telomere shortening and activation of p38 mitogen-activated signaling kinase resulting in aging-associated phenotypic transition. Senescent tissues release inflammatory signals that are propagated via exosomes to cause functional changes in maternal uterine tissues. In vitro, oxidative stress causes increased release of inflammatory mediators (senescence-associated secretory phenotype and damage-associated molecular pattern markers) that can be packaged inside the exosomes. These exosomes traverse through tissues layers, reach maternal tissues to increase overall inflammatory load transitioning them from a quiescent to active state. Animal model studies have shown that fetal exosomes can travel from fetal to the maternal side. Thus, aging fetal membranes and membrane-derived exosomes cargo fetal signals to the uterus and cervix and may trigger parturition. This review highlights a novel hypothesis in human parturition research based on data from ongoing research using human fetal membrane model system.

  5. Programmed Fetal Membrane Senescence and Exosome-Mediated Signaling: A Mechanism Associated With Timing of Human Parturition

    Science.gov (United States)

    Menon, Ramkumar; Mesiano, Sam; Taylor, Robert N.

    2017-01-01

    Human parturition is an inflammatory process that involves both fetal and maternal compartments. The precise immune cell interactions have not been well delineated in human uterine tissues during parturition, but insights into human labor initiation have been informed by studies in animal models. Unfortunately, the timing of parturition relative to fetal maturation varies among viviparous species—indicative of different phylogenetic clocks and alarms—but what is clear is that important common pathways must converge to control the birth process. Herein, we hypothesize a novel signaling mechanism initiated by human fetal membrane aging and senescence-associated inflammation. Programmed events of fetal membrane aging coincide with fetal growth and organ maturation. Mechanistically, senescence involves in telomere shortening and activation of p38 mitogen-activated signaling kinase resulting in aging-associated phenotypic transition. Senescent tissues release inflammatory signals that are propagated via exosomes to cause functional changes in maternal uterine tissues. In vitro, oxidative stress causes increased release of inflammatory mediators (senescence-associated secretory phenotype and damage-associated molecular pattern markers) that can be packaged inside the exosomes. These exosomes traverse through tissues layers, reach maternal tissues to increase overall inflammatory load transitioning them from a quiescent to active state. Animal model studies have shown that fetal exosomes can travel from fetal to the maternal side. Thus, aging fetal membranes and membrane-derived exosomes cargo fetal signals to the uterus and cervix and may trigger parturition. This review highlights a novel hypothesis in human parturition research based on data from ongoing research using human fetal membrane model system. PMID:28861041

  6. Effect of mono-(2-ethylhexyl) phthalate on human and mouse fetal testis: In vitro and in vivo approaches

    Energy Technology Data Exchange (ETDEWEB)

    Muczynski, V. [Univ. Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, BP 6, 92265 Fontenay-aux-Roses (France); CEA, DSV, iRCM, SCSR, LDRG, 92265 Fontenay-aux-Roses (France); INSERM, Unité 967, F-92265, Fontenay aux Roses (France); Cravedi, J.P. [INRA, INP, Université de Toulouse, UMR1331 TOXALIM, F-31027, Toulouse (France); Lehraiki, A.; Levacher, C.; Moison, D.; Lecureuil, C.; Messiaen, S. [Univ. Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, BP 6, 92265 Fontenay-aux-Roses (France); CEA, DSV, iRCM, SCSR, LDRG, 92265 Fontenay-aux-Roses (France); INSERM, Unité 967, F-92265, Fontenay aux Roses (France); Perdu, E. [INRA, INP, Université de Toulouse, UMR1331 TOXALIM, F-31027, Toulouse (France); Frydman, R. [Service de Gynécologie-Obstétrique, Hôpital A. Béclère, Université Paris Sud F-92141 Clamart (France); Habert, R. [Univ. Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, BP 6, 92265 Fontenay-aux-Roses (France); CEA, DSV, iRCM, SCSR, LDRG, 92265 Fontenay-aux-Roses (France); INSERM, Unité 967, F-92265, Fontenay aux Roses (France); and others

    2012-05-15

    The present study was conducted to determine whether exposure to the mono-(2-ethylhexyl) phthalate (MEHP) represents a genuine threat to male human reproductive function. To this aim, we investigated the effects on human male fetal germ cells of a 10{sup −5} M exposure. This dose is slightly above the mean concentrations found in human fetal cord blood samples by biomonitoring studies. The in vitro experimental approach was further validated for phthalate toxicity assessment by comparing the effects of in vitro and in vivo exposure in mouse testes. Human fetal testes were recovered during the first trimester (7–12 weeks) of gestation and cultured in the presence or not of 10{sup −5} M MEHP for three days. Apoptosis was quantified by measuring the percentage of Caspase-3 positive germ cells. The concentration of phthalate reaching the fetal gonads was determined by radioactivity measurements, after incubations with {sup 14}C-MEHP. A 10{sup −5} M exposure significantly increased the rate of apoptosis in human male fetal germ cells. The intratesticular MEHP concentration measured corresponded to the concentration added in vitro to the culture medium. Furthermore, a comparable effect on germ cell apoptosis in mouse fetal testes was induced both in vitro and in vivo. This study suggests that this 10{sup −5} M exposure is sufficient to induce changes to the in vivo development of the human fetal male germ cells. -- Highlights: ► 10{sup −5} M of MEHP impairs germ cell development in the human fetal testis. ► Organotypic culture is a suitable approach to investigate phthalate effects in human. ► MEHP is not metabolized in the human fetal testis. ► In mice, MEHP triggers similar effects both in vivo and in vitro.

  7. Comparative studies of different cryopreservation methods for mesenchymal stem cells derived from human fetal liver.

    Science.gov (United States)

    Todorov, Plamen; Hristova, Elena; Konakchieva, Rossitza; Michova, Antoaneta; Dimitrov, Josif

    2010-03-29

    Fetal stem cells possess some intriguing characteristics, which delineate them as promising cellular therapeutics. They are less immunogenic, at lower stage of differentiation and have higher potential for repopulation and migration. Furthermore, the fetal stem cells secrete a set of cytokines and growth factors, which stimulate the regeneration of the recipient tissue. The present study indicated that the adhesive fraction of human fetal liver cells possessed the morphological characteristics of mesenchymal stem cells, as well as potential to differentiate into adipocyte and osteoblast lineages. The immunophenotypic analysis showed that the cells expressed CD13, CD73, CD90 and CD105 (typical for mesenchymal stem cells) and lacked the haematopoietic lineage markers CD34 and CD45. Addressing the issue of the low-temperature storage of the human fetal liver cells, four different methods for cryopreservation were assessed: conventional slow freezing, program freezing and two vitrification protocols. The obtained results demonstrated that the cells were cryotolerant and maintained their properties and differentiation potential after thawing. Program freezing showed to be the most efficient method for cryopreservation of the investigated cells.

  8. Relationship between pancreatic vesicular monoamine transporter 2 (VMAT2) and insulin expression in human pancreas

    OpenAIRE

    Saisho, Yoshifumi; Harris, Paul E.; Butler, Alexandra E.; Galasso, Ryan; GURLO, TATYANA; Rizza, Robert A.; Butler, Peter C.

    2008-01-01

    Vesicular monoamine transporter 2 (VMAT2) is expressed in pancreatic beta cells and has recently been proposed as a target for measurement of beta cell mass in vivo. We questioned, (1) What proportion of beta cells express VMAT2? (2) Is VMAT2 expressed by other pancreatic endocrine or non-endocrine cells? (3) Is the relationship between VMAT2 and insulin expression disturbed in type 1 (T1DM) or type 2 diabetes (T2DM)? Human pancreas (7 non-diabetics, 5 T2DM, 10 T1DM) was immunostained for ins...

  9. Ontogeny of expression of basic fibroblast growth factor and its receptors in human fetal skin

    Institute of Scientific and Technical Information of China (English)

    CHEN Wei; FU Xiao-bing; GE Shi-li; SUN Tong-zhu; SHENG Zhi-yong

    2005-01-01

    Objective : To investigate the expression characteristics of basic fibroblast growth factor (bFGF)and its receptors, flg ( FGFR1 ) and bek ( FGFR2), in fetal skin at different gestational ages underlying the relevance of these 3 proteins to skin development and the mechanisms underlying the phenotypic transition from scarless to scarforming healing.Methods: Eighteen specimens of fetal skin biopsies of human embryo were obtained from spontaneous abortions at different gestational ages of 13-32 weeks. Gene expression of bFGF, bek and flg was examined with reverse transcription-polymerase chain reaction (RT-PCR). The dynamic expression and distribution of these 3 proteins were detected with streptavidin peroxidase ( SP )immunohistochemical staining method.Results: In the early gestational fetal skin, genes of bFGF and flg were strongly expressed and more protein contents of these 2 proteins were found as compared with the genes at late gestation fetal skin (2.446 ± 0.116 and 2.066 ± 0. 152 versus 2.157 ± 0. 101 and 1.818 ± 0.086,respectively, P < 0.05). On the contrary, the levels of gene expression and protein content of bek were not differently expressed in the early gestational fetal skin versus the late ones. Protein particles of bFGF were mainly distributed in the epidermal cells and some fibroblasts. Bek was mainly located in the cell membrane and cytoplasm of epidermal cells while flg protein was principally located in the epidermal cells, endothelial cells and some fibroblasts.Conclusions: The endogenous bFGF and their receptors might be involved in the cutaneous development at fetal stage. The differently expressing levels of bFGF and flg during gestation may be related to scarless or scarforming repair during gestation.

  10. Interleukin-8 increases vascular endothelial growth factor and neuropilin expression and stimulates ERK activation in human pancreatic cancer.

    Science.gov (United States)

    Li, Min; Zhang, Yuqing; Feurino, Louis W; Wang, Hao; Fisher, William E; Brunicardi, F Charles; Chen, Changyi; Yao, Qizhi

    2008-04-01

    Interleukin-8 (IL-8) is associated with tumorigenesis by promoting angiogenesis and metastasis. Although up-regulation of IL-8 is indicated in many cancers, its function in pancreatic cancer has not been well characterized. In this study we examined the expression of IL-8 on pancreatic cancer cells and clinical tissue specimens, and investigated the effect of exogenous IL-8 on gene expression, and signaling in human pancreatic cancer cells. We found that pancreatic cancer cells expressed higher amount of IL-8 mRNA than normal human pancreatic ductal epithelium cells. IL-8 mRNA was also substantially overexpressed in 11 of 14 (79%) clinical pancreatic-adenocarcinoma samples compared with that in their surrounding normal tissues. Exogenous IL-8 up-regulated the expression of vascular endothelial growth factor(165), and neuropilin (NRP)-2 in BxPC-3 cells, one of human pancreatic cancer cell lines. IL-8 expression was inducible by hypoxia mimicking reagent cobalt chloride. In addition, IL-8 activated extracellular signal-regulated kinase (ERK)1/2 signaling pathway in BxPC-3 cells. Our studies suggest that IL-8 might be a malignant factor in human pancreatic cancer by induction of vascular endothelial growth factor and NRP-2 expression and ERK activation. Targeting IL-8 along with other antiangiogenesis therapy could be an effective treatment for this malignancy.

  11. Pancreatic tuberculosis in a human immunodeficiency virus positive patient:A case report

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Despite the increased incidence of tuberculosis related to human immunodeficiency virus (HIV) in recent decades,pancreatic tuberculosis has rarely been described.We report a case of pancreatic tuberculosis in a 39-yearold African man who presented with progressive dysphagia,vomiting,weight loss and productive cough,accompanied by localized epigastric pain and one episode of melena.HIV-1 testing was positive and lymphocyte subset profile showed CD4 count of 9/mm3.Abdominal computed tomography (CT) scan with contrast revealed a cystic mass in the body of the pancreas,significant portal and retroperitoneal cystic adenopathy,and multiple cystic lesions in the spleen and liver.CT guided cyst aspiration and node biopsy detected Mycobacterium tuberculosis.The patient responded well on antituberculosis and antiretroviral therapy.Tuberculosis rarely involves the pancreas,probably due to the presence of pancreatic enzymes which interfere with the seeding of Mycobacterium tuberculosis.Pancreatic tuberculosis is considered to be the result of dissemination of the infection from nearby lymphatic nodes.Endoscopic ultrasound or CT guided fine needle aspiration for cytology is the recommended diagnostic technique.Although the prognosis is good with antituberculosis treatment,it could be fatal without correct diagnosis and treatment.The clinician's high index of suspicion of pancreatic tuberculosis and application of FNAB to obtain pathological evidence are extremely important to a correct diagnosis,especially in young HIV positive patients.

  12. Curcumin Inhibits Tumor Growth and Angiogenesis in an Orthotopic Mouse Model of Human Pancreatic Cancer

    Directory of Open Access Journals (Sweden)

    Sabrina Bimonte

    2013-01-01

    Full Text Available Pancreatic cancer is a malignant neoplasm originating from transformed cells arising in tissues forming the pancreas. The best chemotherapeutic agent used to treat pancreatic cancer is the gemcitabine. However, gemcitabine treatment is associated with many side effects. Thus novel strategies involving less toxic agents for treatment of pancreatic cancer are necessary. Curcumin is one such agent that inhibits the proliferation and angiogenesis of a wide variety of tumor cells, through the modulation of many cell signalling pathways. In this study, we investigated whether curcumin plays antitumor effects in MIA PaCa-2 cells. In vitro studies showed that curcumin inhibits the proliferation and enhances apoptosis of MIA PaCa-2 cells. To test whether the antitumor activity of curcumin is also observed in vivo, we generated an orthotopic mouse model of pancreatic cancer by injection of MIA PaCa-2 cells in nude mice. We placed mice on diet containing curcumin at 0.6% for 6 weeks. In these treated mice tumors were smaller with respect to controls and showed a downregulation of the transcription nuclear factor NF-κB and NF-κB-regulated gene products. Overall, our data indicate that curcumin has a great potential in treatment of human pancreatic cancer through the modulation of NF-κB pathway.

  13. Involvement of Endoplasmic Reticulum Stress in Capsaicin-Induced Apoptosis of Human Pancreatic Cancer Cells

    Directory of Open Access Journals (Sweden)

    Shengzhang Lin

    2013-01-01

    Full Text Available Capsaicin, main pungent ingredient of hot chilli peppers, has been shown to have anticarcinogenic effect on various cancer cells through multiple mechanisms. In this study, we investigated the apoptotic effect of capsaicin on human pancreatic cancer cells in both in vitro and in vivo systems, as well as the possible mechanisms involved. In vitro, treatment of both the pancreatic cancer cells (PANC-1 and SW1990 with capsaicin resulted in cells growth inhibition, G0/G1 phase arrest, and apoptosis in a dose-dependent manner. Knockdown of growth arrest- and DNA damage-inducible gene 153 (GADD153, a marker of the endoplasmic-reticulum-stress- (ERS- mediated apoptosis pathway, by specific siRNA attenuated capsaicin-induced apoptosis both in PANC-1 and SW1990 cells. Moreover, in vivo studies capsaicin effectively inhibited the growth and metabolism of pancreatic cancer and prolonged the survival time of pancreatic cancer xenograft tumor-induced mice. Furthermore, capsaicin increased the expression of some key ERS markers, including glucose-regulated protein 78 (GRP78, phosphoprotein kinase-like endoplasmic reticulum kinase (phosphoPERK, and phosphoeukaryotic initiation factor-2α (phospho-eIF2α, activating transcription factor 4 (ATF4 and GADD153 in tumor tissues. In conclusion, we for the first time provide important evidence to support the involvement of ERS in the induction of apoptosis in pancreatic cancer cells by capsaicin.

  14. Targeting developmental regulators of zebrafish exocrine pancreas as a therapeutic approach in human pancreatic cancer

    Directory of Open Access Journals (Sweden)

    Nelson S. Yee

    2012-02-01

    Histone deacetylases (HDACs and RNA polymerase III (POLR3 play vital roles in fundamental cellular processes, and deregulation of these enzymes has been implicated in malignant transformation. Hdacs and Polr3 are required for exocrine pancreatic epithelial proliferation during morphogenesis in zebrafish. We aim to test the hypothesis that Hdacs and Polr3 cooperatively control exocrine pancreatic growth, and combined inhibition of HDACs and POLR3 produces enhanced growth suppression in pancreatic cancer. In zebrafish larvae, combination of a Hdac inhibitor (Trichostatin A and an inhibitor of Polr3 (ML-60218 synergistically prohibited the expansion of exocrine pancreas. In human pancreatic adenocarcinoma cells, combination of the HDAC inhibitor suberoylanilide hydroxamic acid (SAHA and ML-60218 produced augmented suppression of colony formation and proliferation, and induction of cell cycle arrest and apoptotic cell death. The enhanced cytotoxicity was associated with supra-additive upregulation of the pro-apoptotic regulator BAX and the cyclin-dependent kinase inhibitor p21CDKN1A. tRNAs have been shown to have pro-proliferative and anti-apoptotic roles, and SAHA-stimulated expression of tRNAs was reversed by ML-60218. These findings demonstrate that chemically targeting developmental regulators of exocrine pancreas can be translated into an approach with potential impact on therapeutic response in pancreatic cancer, and suggest that counteracting the pro-malignant side effect of HDAC inhibitors can enhance their anti-tumor activity.

  15. MUC1 selectively targets human pancreatic cancer in orthotopic nude mouse models.

    Directory of Open Access Journals (Sweden)

    Jeong Youp Park

    Full Text Available The goal of this study was to determine whether MUC1 antibody conjugated with a fluorophore could be used to visualize pancreatic cancer. Anti-MUC1 (CT2 antibody was conjugated with 550 nm or 650 nm fluorophores. Nude mouse were used to make subcutaneous and orthotopic models of pancreatic cancer. Western blot and flow cytometric analysis confirmed the expression of MUC1 in human pancreatic cancer cell lines including BxPC-3 and Panc-1. Immunocytochemistry with fluorophore conjugated anti-MUC1 antibody demonstrated fluorescent areas on the membrane of Panc-1 cancer cells. After injecting the conjugated anti-MUC1 antibodies via the tail vein, subcutaneously transplanted Panc-1 and BxPC-3 tumors emitted strong fluorescent signals. In the subcutaneous tumor models, the fluorescent signal from the conjugated anti-MUC1 antibody was noted around the margin of the tumor and space between the cells. The conjugated anti-MUC1 antibody bound the tumor in orthotopically-transplanted Panc-1 and BxPC-3 models enabling the tumors to be imaged. This study showed that fluorophore conjugated anti-MUC1 antibodies could visualize pancreatic tumors in vitro and in vivo and may help to improve the diagnosis and treatment of pancreatic cancer.

  16. Circadian pancreatic enzyme pattern and relationship between secretory and motor activity in fasting humans.

    Science.gov (United States)

    Keller, Jutta; Layer, Peter

    2002-08-01

    It is unknown whether nonparallel pancreatic enzyme output occurs under basal conditions in humans. We aimed to determine whether the circadian or wake-sleep cycle influences the relationship among pancreatic enzymes or between pancreatic secretory and jejunal motor activity. Using orojejunal multilumen intubation, we measured enzyme outputs and proximal jejunal motility index during consecutive daytime and nighttime periods in each of seven fasting, healthy volunteers. Enzyme outputs were correlated tightly during daytime phases of wakefulness and nighttime phases of sleep (r > 0.72, P activity was directly correlated with jejunal motility index (r > 0.50, P enzymes dominates throughout the circadian cycle. Nonparallel secretion during nocturnal phases of wakefulness may be due to merely circadian effects or to the coupling of the wake-sleep and the circadian cycle. The association between fluctuations of secretory and motor activity appears to be particularly tight during the night.

  17. Human Equilibrative Nucleoside Transporter 1 (hENT1 in Pancreatic Adenocarcinoma: Towards Individualized Treatment Decisions

    Directory of Open Access Journals (Sweden)

    Jennifer L. Spratlin

    2010-12-01

    Full Text Available Pancreatic cancer is one of the most lethal cancers, where curative surgical resections are rare and less than 5% of patients experience long-term survival. Despite numerous clinical trials, improvements in the systemic treatment of this disease have been limited. Gemcitabine, a nucleoside analogue, is still considered the standard of care chemotherapy for most patients in the advanced disease setting. To exert its cytotoxic effects, gemcitabine must enter cells via nucleoside transporters, most notably human equilibrative nucleoside transporter 1 (hENT1. Increasingly strong evidence suggests hENT1 is a prognostic biomarker in gemcitabine-treated pancreatic cancer, and may well be a predictive biomarker of gemcitabine efficacy. In this review, we synthesize the literature surrounding hENT1 in pancreatic cancer, identify the key outstanding questions, and suggest strategies to prospectively evaluate the clinical utility of hENT1 in future clinical studies.

  18. MicroRNA miR-34 inhibits human pancreatic cancer tumor-initiating cells.

    Directory of Open Access Journals (Sweden)

    Qing Ji

    Full Text Available BACKGROUND: MicroRNAs (miRNAs have been implicated in cancer initiation and progression via their ability to affect expression of genes and proteins that regulate cell proliferation and/or cell death. Transcription of the three miRNA miR-34 family members was recently found to be directly regulated by p53. Among the target proteins regulated by miR-34 are Notch pathway proteins and Bcl-2, suggesting the possibility of a role for miR-34 in the maintenance and survival of cancer stem cells. METHODOLOGY/PRINCIPAL FINDINGS: We examined the roles of miR-34 in p53-mutant human pancreatic cancer cell lines MiaPaCa2 and BxPC3, and the potential link to pancreatic cancer stem cells. Restoration of miR-34 expression in the pancreatic cancer cells by either transfection of miR-34 mimics or infection with lentiviral miR-34-MIF downregulated Bcl-2 and Notch1/2. miR-34 restoration significantly inhibited clonogenic cell growth and invasion, induced apoptosis and G1 and G2/M arrest in cell cycle, and sensitized the cells to chemotherapy and radiation. We identified that CD44+/CD133+ MiaPaCa2 cells are enriched with tumorsphere-forming and tumor-initiating cells or cancer stem/progenitor cells with high levels of Notch/Bcl-2 and loss of miR-34. More significantly, miR-34 restoration led to an 87% reduction of the tumor-initiating cell population, accompanied by significant inhibition of tumorsphere growth in vitro and tumor formation in vivo. CONCLUSIONS/SIGNIFICANCE: Our results demonstrate that miR-34 may restore, at least in part, the tumor suppressing function of the p53 in p53-deficient human pancreatic cancer cells. Our data support the view that miR-34 may be involved in pancreatic cancer stem cell self-renewal, potentially via the direct modulation of downstream targets Bcl-2 and Notch, implying that miR-34 may play an important role in pancreatic cancer stem cell self-renewal and/or cell fate determination. Restoration of miR-34 may hold significant

  19. Nanotopography Promotes Pancreatic Differentiation of Human Embryonic Stem Cells and Induced Pluripotent Stem Cells.

    Science.gov (United States)

    Kim, Jong Hyun; Kim, Hyung Woo; Cha, Kyoung Je; Han, Jiyou; Jang, Yu Jin; Kim, Dong Sung; Kim, Jong-Hoon

    2016-03-22

    Although previous studies suggest that nanotopographical features influence properties and behaviors of stem cells, only a few studies have attempted to derive clinically useful somatic cells from human pluripotent stem cells using nanopatterned surfaces. In the present study, we report that polystyrene nanopore-patterned surfaces significantly promote the pancreatic differentiation of human embryonic and induced pluripotent stem cells. We compared different diameters of nanopores and showed that 200 nm nanopore-patterned surfaces highly upregulated the expression of PDX1, a critical transcription factor for pancreatic development, leading to an approximately 3-fold increase in the percentage of differentiating PDX1(+) pancreatic progenitors compared with control flat surfaces. Furthermore, in the presence of biochemical factors, 200 nm nanopore-patterned surfaces profoundly enhanced the derivation of pancreatic endocrine cells producing insulin, glucagon, or somatostatin. We also demonstrate that nanopore-patterned surface-induced upregulation of PDX1 is associated with downregulation of TAZ, suggesting the potential role of TAZ in nanopore-patterned surface-mediated mechanotransduction. Our study suggests that appropriate cytokine treatments combined with nanotopographical stimulation could be a powerful tool for deriving a high purity of desired cells from human pluripotent stem cells.

  20. l-Methionine inhibits growth of human pancreatic cancer cells

    OpenAIRE

    BENAVIDES, MAXIMO A.; BOSLAND, MAARTEN C.; da Silva, Cássio P.; Sares, Claudia T. Gomes; de Oliveira, Alana M. Cerqueira; Kemp,Rafael; dos Reis, Rodolfo B.; Martins,Vilma R.; Sampaio,Suely V.; Bland, Kirby I.; Grizzle, William E.; José S. dos Santos

    2014-01-01

    We have previously shown that l-methionine inhibits proliferation of breast, prostate, and colon cancer cells. This study extends these findings to BXPC-3 (mutated p53) and HPAC (wild-type p53) pancreatic cancer cells and explores the reversibility of these effects. Cells were exposed to l-methionine (5 mg/ml) for 7 days or for 3 days, followed by 4 days of culture without l-methionine (recovery). Cell proliferation, apoptosis, and cell cycle effects were assessed by flow cytometry after stai...

  1. Prospective isolation of human embryonic stem cell-derived cardiovascular progenitors that integrate into human fetal heart tissue.

    Science.gov (United States)

    Ardehali, Reza; Ali, Shah R; Inlay, Matthew A; Abilez, Oscar J; Chen, Michael Q; Blauwkamp, Timothy A; Yazawa, Masayuki; Gong, Yongquan; Nusse, Roeland; Drukker, Micha; Weissman, Irving L

    2013-02-26

    A goal of regenerative medicine is to identify cardiovascular progenitors from human ES cells (hESCs) that can functionally integrate into the human heart. Previous studies to evaluate the developmental potential of candidate hESC-derived progenitors have delivered these cells into murine and porcine cardiac tissue, with inconclusive evidence regarding the capacity of these human cells to physiologically engraft in xenotransplantation assays. Further, the potential of hESC-derived cardiovascular lineage cells to functionally couple to human myocardium remains untested and unknown. Here, we have prospectively identified a population of hESC-derived ROR2(+)/CD13(+)/KDR(+)/PDGFRα(+) cells that give rise to cardiomyocytes, endothelial cells, and vascular smooth muscle cells in vitro at a clonal level. We observed rare clusters of ROR2(+) cells and diffuse expression of KDR and PDGFRα in first-trimester human fetal hearts. We then developed an in vivo transplantation model by transplanting second-trimester human fetal heart tissues s.c. into the ear pinna of a SCID mouse. ROR2(+)/CD13(+)/KDR(+)/PDGFRα(+) cells were delivered into these functioning fetal heart tissues: in contrast to traditional murine heart models for cell transplantation, we show structural and functional integration of hESC-derived cardiovascular progenitors into human heart.

  2. Combined transcriptome analysis of fetal human and mouse cerebral cortex exposed to alcohol.

    Science.gov (United States)

    Hashimoto-Torii, Kazue; Kawasawa, Yuka Imamura; Kuhn, Alexandre; Rakic, Pasko

    2011-03-08

    Fetal exposure to environmental insults increases the susceptibility to late-onset neuropsychiatric disorders. Alcohol is listed as one of such prenatal environmental risk factors and known to exert devastating teratogenetic effects on the developing brain, leading to complex neurological and psychiatric symptoms observed in fetal alcohol spectrum disorder (FASD). Here, we performed a coordinated transcriptome analysis of human and mouse fetal cerebral cortices exposed to ethanol in vitro and in vivo, respectively. Up- and down-regulated genes conserved in the human and mouse models and the biological annotation of their expression profiles included many genes/terms related to neural development, such as cell proliferation, neuronal migration and differentiation, providing a reliable connection between the two species. Our data indicate that use of the combined rodent and human model systems provides an effective strategy to reveal and analyze gene expression changes inflicted by various physical and chemical environmental exposures during prenatal development. It also can potentially provide insight into the pathogenesis of environmentally caused brain disorders in humans.

  3. Human cytomegalovirus induces a distinct innate immune response in the maternal-fetal interface.

    Science.gov (United States)

    Weisblum, Yiska; Panet, Amos; Zakay-Rones, Zichria; Vitenshtein, Alon; Haimov-Kochman, Ronit; Goldman-Wohl, Debra; Oiknine-Djian, Esther; Yamin, Rachel; Meir, Karen; Amsalem, Hagai; Imbar, Tal; Mandelboim, Ofer; Yagel, Simcha; Wolf, Dana G

    2015-11-01

    The initial interplay between human cytomegalovirus (HCMV) and innate tissue response in the human maternal-fetal interface, though crucial for determining the outcome of congenital HCMV infection, has remained unknown. We studied the innate response to HCMV within the milieu of the human decidua, the maternal aspect of the maternal-fetal interface, maintained ex vivo as an integral tissue. HCMV infection triggered a rapid and robust decidual-tissue innate immune response predominated by interferon (IFN)γ and IP-10 induction, dysregulating the decidual cytokine/chemokine environment in a distinctive fashion. The decidual-tissue response was already elicited during viral-tissue contact, and was not affected by neutralizing HCMV antibodies. Of note, IFNγ induction, reflecting immune-cell activation, was distinctive to the maternal decidua, and was not observed in concomitantly-infected placental (fetal) villi. Our studies in a clinically-relevant surrogate human model, provide a novel insight into the first-line decidual tissue response which could affect the outcome of congenital infection.

  4. Hepatitis B virus infection and replication in primarily cultured human fetal hepatocytes

    Institute of Scientific and Technical Information of China (English)

    Min Lin; Qun Chen; Li-Ye Yang; Wen-Yu Li; Xi-Biao Cao; Jiao-Ren Wu; You-Peng Peng; Mo-Rui Chen

    2007-01-01

    AIM:To investigate the infection and replication of hepatitis B virus(HBV)in primarily cultured human fetal hepatocytes(HFHs).METHODS:The human fetal hepatocytes were cultured in serum-free medium,HBV-positive serum was added into the medium to study the susceptibility of hepatocytes to HBV infection.The supernatant was collected for ELISA assay of HBsAg and HBeAg,and quantitative fluorescence PCR for HBV-DNA assay daily.Albumin and HBcAg,CK8 and CK18 expressions were detected by immunohistochemistry in cultured hepatocytes.Content of lactate dehydrogenate(LDH)was measured to find out the integrity of the cell membrane.RESULTS:A stable hepatocyte culture system was established.HBV could infect the hepatocytes and replicate,and HBcAg expression could be detected by immunohistochemistry in hepatocyte-like cells.HBV-DNA in the supernatant could be detected from d 2 to d 18 and HBsAg and HBeAg were positive on d 3-d 18 after HBV infection.HBV in medium increased from d 0 to d 6 and subsequently decreased as the cells were progressively loosing their hepatocyte phenotypes.CONCLUSION:HBV could infect human fetal hepatocytes and replicate.This in vitro model allowed a detailed Study on early events associated with human HBV entry into cells and subsequent replication.

  5. Metabolic gene profile in early human fetal heart development

    National Research Council Canada - National Science Library

    Iruretagoyena, J I; Davis, W; Bird, C; Olsen, J; Radue, R; Teo Broman, A; Kendziorski, C; Splinter BonDurant, S; Golos, T; Bird, I; Shah, D

    2014-01-01

    .... In order to describe normal cardiac development during late first and early second trimester in human fetuses this study used microarray and pathways analysis and created a corresponding 'normal' database...

  6. Cephalic Phase Pancreatic Polypeptide Responses to Liquid and Solid Stimuli in Humans

    OpenAIRE

    TEFF, KAREN L.

    2009-01-01

    The hormone, pancreatic polypeptide (PP) is postulated to be involved in body weight regulation. PP release is dependent on vagal activation and is a marker of vagal efferent activity. Because vagal activity plays a role in glucose homeostasis, elucidating the conditions of activation has important implications for nutrient metabolism. In humans, modified sham-feeding is known to elicit vagally-mediated hormonal responses. We present results of 3 studies in which healthy human subjects tasted...

  7. Catalytic ferrous iron in amniotic fluid as a predictive marker of human maternal-fetal disorders.

    Science.gov (United States)

    Hattori, Yuka; Mukaide, Takahiro; Jiang, Li; Kotani, Tomomi; Tsuda, Hiroyuki; Mano, Yukio; Sumigama, Seiji; Hirayama, Tasuku; Nagasawa, Hideko; Kikkawa, Fumitaka; Toyokuni, Shinya

    2015-01-01

    Amniotic fluid contains numerous biomolecules derived from fetus and mother, thus providing precious information on pregnancy. Here, we evaluated oxidative stress of human amniotic fluid and measured the concentration of catalytic Fe(II). Amniotic fluid samples were collected with consent from a total of 89 subjects in Nagoya University Hospital, under necessary medical interventions: normal pregnancy at term, normal pregnancy at the 2nd trimester, preterm delivery with maternal disorders but without fetal disorders, congenital diaphragmatic hernia, fetal growth restriction, pregnancy-induced hypertension, gestational diabetes mellitus, Down syndrome and trisomy 18. Catalytic Fe(II) and oxidative stress markers (8-hydroxy-2'-deoxyguanosine, 8-OHdG; dityrosine) were determined with RhoNox-1 and specific antibodies, respectively, using plate assays. Levels of 8-OHdG and dityrosine were higher in the 3rd trimester compared with the 2nd trimester in normal subjects, and the abnormal groups generally showed lower levels than the controls, thus suggesting that they represent fetal metabolic activities. In contrast, catalytic Fe(II) was higher in the 2nd trimester than the 3rd trimester in the normal subjects, and overall the abnormal groups showed higher levels than the controls, suggesting that high catalytic Fe(II) at late gestation reflects fetal pathologic alterations. Notably, products of H2O2 and catalytic Fe(II) remained almost constant in amniotic fluid.

  8. A high-resolution MRI study of linear growth of the human fetal skull base

    Energy Technology Data Exchange (ETDEWEB)

    Jeffery, N. [University Coll., London (United Kingdom). Dept. of Anatomy and Development Biology

    2002-04-01

    The skull base, otherwise referred to as the basicranium or cranial base, plays a key role in the process of skull development, providing both support for the brain and an architectural component of the craniofacial complex. Consequently, the fetal skull base has been the focus of numerous studies employing various methods, including sectioning, plain radiography and CT. This paper investigates high-resolution (hr) MRI as an alternative method for looking at and quantifying the fetal skull base. The evaluation tests two basic hypotheses drawn from previous studies. These suggest that the anterior segment of the midline skull base grows more rapidly than the posterior segment and that the width of the posterior cranial fossa increases disproportionately in relation to its length. I imaged 42 formalin preserved human fetuses from museum collections with hrMRI. The T2-weighted image voxels were significantly smaller than those acquired with conventional clinical MRI. Landmarks of the fetal skull base were identified on reformatted axial and sagittal images. Bivariate plots revealed that the growth rate of the anterior skull base is almost twice that of the posterior skull base and that increases in the width of the posterior cranial fossa exceed those in its length. These findings confirm those of previous investigations and show that hrMRI offers a way forward in noninvasive quantification of fetal morphology. ----------------------------------------------------------------------------.

  9. Avoidance of Maternal Cell Contamination and Overgrowth in Isolating Fetal Chorionic Villi Mesenchymal Stem Cells from Human Term Placenta.

    Science.gov (United States)

    Sardesai, Varda S; Shafiee, Abbas; Fisk, Nicholas M; Pelekanos, Rebecca A

    2017-04-01

    Human placenta is rich in mesenchymal stem/stromal cells (MSC), with their origin widely presumed fetal. Cultured placental MSCs are confounded by a high frequency of maternal cell contamination. Our recent systematic review concluded that only a small minority of placental MSC publications report fetal/maternal origin, and failed to discern a specific methodology for isolation of fetal MSC from term villi. We determined isolation conditions to yield fetal and separately maternal MSC during ex vivo expansion from human term placenta. MSCs were isolated via a range of methods in combination; selection from various chorionic regions, different commercial media, mononuclear cell digest and/or explant culture. Fetal and maternal cell identities were quantitated in gender-discordant pregnancies by XY chromosome fluorescence in situ hybridization. We first demonstrated reproducible maternal cell contamination in MSC cultures from all chorionic anatomical locations tested. Cultures in standard media rapidly became composed entirely of maternal cells despite isolation from fetal villi. To isolate pure fetal cells, we validated a novel isolation procedure comprising focal dissection from the cotyledonary core, collagenase/dispase digestion and explant culture in endothelial growth media that selected, and provided a proliferative environment, for fetal MSC. Comparison of MSC populations within the same placenta confirmed fetal to be smaller, more osteogenic and proliferative than maternal MSC. We conclude that in standard media, fetal chorionic villi-derived MSC (CV-MSC) do not grow readily, whereas maternal MSC proliferate to result in maternal overgrowth during culture. Instead, fetal CV-MSCs require isolation under specific conditions, which has implications for clinical trials using placental MSC. Stem Cells Translational Medicine 2017;6:1070-1084.

  10. Characterization of hepatic progenitors from human fetal liver using CD34 as a hepatic progenitor marker

    Institute of Scientific and Technical Information of China (English)

    Parveen Nyamath; Ayesha AM; Aejaz Habeeb; Sanjeev Khosla; Aleem A Khan; CM Habibullah

    2007-01-01

    AIM: To enrich putative hepatic progenitors from the developing human fetal liver using CD34 as a marker.METHODS: Aborted fetuses of 13-20 wk were used for the isolation of liver cells. The cells were labeled with anti CD34; a marker used for isolating progenitor population and the cells were sorted using magnetic cell sorting. The positive fractions of cells were assessed for specific hepatic markers. Further, these cells were cultured in vitro for long term investigation.RESULTS: Flow cytometric and immunocytochemical analysis for alphafetoprotein (AFP) showed that the majority of the enriched CD34 positive cells were positive for AFP. Furthermore, these enriched cells proliferated in the long term and maintained hepatic characteristics in in vitro culture.CONCLUSION: The study shows that aborted human fetal liver is a potential source for isolation of hepatic progenitors for clinical applications. The study also demonstrates that CD34 can be a good marker for the enrichment of progenitor populations.

  11. A three-dimensional study of human fetal endocervix with special reference to its epithelium

    OpenAIRE

    Barberini, F.; Makabe, S.; Motta, P M

    1998-01-01

    The development of human fetal cervix has been systematically studied by SEM, obtaining a detailed map of its fine structure, particularly concerning the differentiation and maturation of the endocervical epithelium, including its "eversion" and "squamous metaplasia", normally occurring in postnatal life, but not yet observed in detail by electron microscopy in the fetus. Cervices from spontaneous abortion at 12, 15, 18, 20, 21 and 22 weeks and from intrauterin...

  12. MCT8 expression in human fetal cerebral cortex is reduced in severe intrauterine growth restriction

    OpenAIRE

    Shiao Y Chan; Hancox, Laura A; Martín-Santos, Azucena; Loubière, Laurence S; Walter, Merlin N.M.; González, Ana-Maria; Cox, Phillip M; Logan, Ann; Christopher J. McCabe; Franklyn, Jayne A; Kilby, Mark D

    2014-01-01

    The importance of the thyroid hormone (TH) transporter, monocarboxylate transporter 8 (MCT8), to human neurodevelopment is highlighted by findings of severe global neurological impairment in subjects with MCT8 (SLC16A2) mutations. Intrauterine growth restriction (IUGR), usually due to uteroplacental failure, is associated with milder neurodevelopmental deficits, which have been partly attributed to dysregulated TH action in utero secondary to reduced circulating fetal TH concentrations and de...

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

    Science.gov (United States)

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

    2015-11-01

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

  14. Effect of placental factors on growth and function of the human fetal adrenal in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Riopel, L.; Branchaud, C.L.; Goodyer, C.G.; Zweig, M.; Lipowski, L.; Adkar, V.; Lefebvre, Y. (McGill Univ.-Montreal Children' s Hospital Research Institute, Quebec (Canada))

    1989-11-01

    Conditioned medium from human placental monolayer cultures (PM) had a marked stimulatory effect on proliferation (3H-thymidine uptake) of human fetal zone adrenal cells in primary monolayer culture, even in the absence of serum. Epidermal growth factor (EGF) and fibroblast growth factor (FGF) also significantly stimulated fetal adrenal cell growth. However, the effects of PM differed from those of EGF and FGF in several respects: (1) maximal response to PM was 2-5 times greater; (2) mitogenic effects of EGF and FGF were suppressed by adrenocorticotropic hormone (ACTH), whereas that of 50% PM was not; (3) PM inhibited ACTH-stimulated steroidogenesis (dehydroepiandrosterone sulfate and cortisol), but EGF and FGF did not. Preliminary characterization studies have indicated that approximately half of the placental growth-promoting activity is heat resistant and sensitive to bacterial proteases, and that 50-60% of the activity is lost after dialysis with membranes having a molecular weight cutoff of 3500. These findings suggest a role for the placenta in the growth and differentiated function of the human fetal adrenal gland.

  15. Expression and localization of VEGF receptors in human fetal skeletal tissues.

    Science.gov (United States)

    Marini, M; Sarchielli, E; Toce, M; Acocella, A; Bertolai, R; Ciulli, C; Orlando, C; Sgambati, E; Vannelli, G B

    2012-12-01

    During development the vertebrate skeleton is the product of deriving cells from distinct embryonic lineages. The craniofacial skeleton is formed by migrating cranial neural crest cells, whereas the axial and limb skeletons are derived from mesodermal cells. The Vascular Endothelial Growth Factors (VEGFs) / receptors (VEGFRs) system plays an important role in angiogenesis, as well as osteogenesis, during bone development, growth, and remodeling, attracting endothelial cells and osteoclasts and stimulating osteoblast differentiation. Recent evidence has shown that during development VEGFR-3 is also expressed in neural and glial precursors of forebrain and cerebellum, as well as in the eye. In this study, we found that VEGFR-1, VEGFR-2 and VEGFR-3 are expressed in human bone both in fetal and adult life. The gene expression levels were significantly higher in fetal samples especially in mandibles. In addition, higher levels of VEGFR-3 in orofacial district were confirmed by western blotting analysis. We also observed that in fetal mandibular samples VEGFRs colocalized in several osteoblasts, osteoclasts and osteoprogenitor cells. Furthermore, some cells coexpressed VEGFR-3 and ET-1, a marker of neural crest cells. The results demonstrated different expression of VEGFRs in human mandibular and femoral bones which could be correlated to their different structure, function and development during organogenesis. VEGFR-3 might represent a specific signal for ectomesenchymal lineage differentiation during early human development.

  16. Compound MMH01 possesses toxicity against human leukemia and pancreatic cancer cells.

    Science.gov (United States)

    Chen, Yu-Jen; Chou, Cheng-Jen; Chang, Tun-Tschu

    2009-04-01

    MMH01 is a compound isolated from Antrodia cinnamomea. MMH01 markedly inhibited growth of human leukemia U937 and pancreatic cancer BxPC3 cells. It resulted in distinct patterns of cell cycle distribution in U937 (G2/M, sub-G1 and polyploidy) and BxPC3 cells (G0/G1 and sub-G1). The modes of cell death in U937 cells include apoptosis and mitotic catastrophe, whereas apoptosis-associated events or necrosis in BxPC3 cells. Neither mitochondrial membrane permeabilization nor caspase dependence was noted. Proteins involving mitotic catastrophe-associated cell death such as cyclin B1 and checkpoint kinase 2 were activated in U937 cells. Only slight to moderate viability inhibition was noted to human monocytes, the normal counterpart of these myeloid leukemic cells. In conclusion, MMH01 possesses cytotoxicity against human leukemia and pancreatic cancer cells.

  17. Fetal stromal niches enhance human embryonic stem cell-derived hematopoietic differentiation and globin switch.

    Science.gov (United States)

    Lee, King Yiu; Fong, Benny Shu Pan; Tsang, Kam Sze; Lau, Tze Kin; Ng, Pak Cheung; Lam, Audrey Carmen; Chan, Kathy Yuen Yee; Wang, Chi Chiu; Kung, Hsiang Fu; Li, Chi Kong; Li, Karen

    2011-01-01

    Hematopoiesis during mammalian embryonic development has been perceived as a migratory phenomenon, from the yolk sac blood island to the aorta-gonad-mesonephros (AGM) region, fetal liver (FL), and subsequently, the fetal bone marrow. In this study, we investigated the effects of primary stromal cells from fetal hematopoietic niches and their conditioned media (CM), applied singly or in sequential orders, on induction of human embryonic stem cells, H1, H9, and H14 lines, to hematopoietic cells. Our results demonstrated that stromal support of FL, AGM + FL, and AGM + FL + fetal bone marrow significantly increased the proliferation of embryoid bodies (EB) at day 18 of hematopoietic induction in the presence of thrombopoietin, stem cell factor, and Flt-3 ligand. AGM + FL also increased hematopoietic colony-forming unit (CFU) formation. CM did not enhance EB proliferation but CM of FL and AGM + FL significantly increased the density of total CFU and early erythroid (burst-forming unit) progenitors. Increased commitment to the hematopoietic lineage was demonstrated by enhanced expressions of CD45, alpha-, beta-, and gamma-globins in CFU at day 32, compared with EB at day 18. CM of FL significantly increased these globin expressions, indicating enhanced switches from embryonic to fetal and adult erythropoiesis. Over 50% and 10% of cells derived from CFU expressed CD45 and beta-globin proteins, respectively. Expressions of hematopoietic regulatory genes (Bmi-1, β-Catenin, Hox B4, GATA-1) were increased in EB or CFU cultures supported by FL or sequential CM. Our study has provided a strategy for derivation of hematopoietic cells from embryonic stem cells under the influence of primary hematopoietic niches and CM, particularly the FL.

  18. Intracellular Immunization of Human Fetal Cord Blood Stem/Progenitor Cells with a Ribozyme Against Human Immunodeficiency Virus Type 1

    Science.gov (United States)

    Yu, Mang; Leavitt, Mark C.; Maruyama, Midori; Yamada, Osamu; Young, Dennis; Ho, Anthony D.; Wong-Staal, Flossie

    1995-01-01

    Successful treatment of human immunodeficiency virus infection may ultimately require targeting of hematopoietic stem cells. Here we used retroviral vectors carrying the ribozyme gene to transduce CD34^+ cells from human fetal cord blood. Transduction and ribozyme expression had no apparent adverse effect on cell differentiation and/or proliferation. The macrophage-like cells, differentiated from the stem/progenitor cells in vitro, expressed the ribozyme gene and resisted infection by a macrophage tropic human immunodeficiency virus type 1. These results suggest the feasibility of stem cell gene therapy for human immunodeficiency virus-infected patients.

  19. Profile of MMP and TIMP Expression in Human Pancreatic Stellate Cells: Regulation by IL-1α and TGFβ and Implications for Migration of Pancreatic Cancer Cells

    Directory of Open Access Journals (Sweden)

    Vegard Tjomsland

    2016-07-01

    Full Text Available Pancreatic ductal adenocarcinoma is characterized by a prominent fibroinflammatory stroma with both tumor-promoting and tumor-suppressive functions. The pancreatic stellate cell (PSC is the major cellular stromal component and the main producer of extracellular matrix proteins, including collagens, which are degraded by metalloproteinases (MMPs. PSCs interact with cancer cells through various factors, including transforming growth factor (TGFβ and interleukin (IL-1α. The role of TGFβ in the dual nature of tumor stroma, i.e., protumorigenic or tumor suppressive, is not clear. We aimed to investigate the roles of TGFβ and IL-1α in the regulation of MMP profiles in PSCs and the subsequent effects on cancer cell migration. Human PSCs isolated from surgically resected specimens were cultured in the presence of pancreatic cancer cell lines, as well as IL-1α or TGFβ. MMP production and activities in PSCs were quantified by gene array transcripts, mRNA measurements, fluorescence resonance energy transfer–based activity assay, and zymography. PSC-conditioned media and pancreatic cancer cells were included in a collagen matrix cell migration model. We found that production of IL-1α by pancreatic cancer cells induced alterations in MMP and tissue inhibitors of matrix metalloproteinase (TIMP profiles and activities in PSCs, upregulated expression and activation of MMP1 and MMP3, and enhanced migration of pancreatic cancer cells in the collagen matrix model. TGFβ counteracted the effects of IL-1α on PSCs, reestablished PSC MMP and TIMP profiles and activities, and inhibited migration of cancer cells. This suggests that tumor TGFβ has a role as a suppressor of stromal promotion of tumor progression through alterations in PSC MMP profiles with subsequent inhibition of pancreatic cancer cell migration.

  20. Preparation of pancreatic β-cells from human iPS cells with small molecules.

    Science.gov (United States)

    Hosoya, Masaki

    2012-01-01

    Human induced pluripotent stem (iPS) cells obtained from patients are expected to be a useful source for cell transplantation therapy, because many patients (including those with type 1 diabetes and severe type 2 diabetes) are on waiting lists for transplantation for a long time due to the shortage of donors. At present, many concerns related to clinical application of human iPS cells have been raised, but rapid development of methods for the establishment, culture, and standardization of iPS cells will lead autologous cell therapy to be realistic sooner or later. However, establishment of a method for preparing some of desired cell types is still challenging. Regarding pancreatic β-cells, there have been many reports about differentiation of these cells from human embryonic stem (ES)/iPS cells, but a protocol for clinical application has still not been established. Since there is clear proof that cell transplantation therapy is effective for diabetes based on the results of clinical islet transplantation, pancreatic β-cells prepared from human iPS cells are considered likely to be effective for reducing the burden on patients. In this article, the current status of procedures for preparing pancreatic β-cells from human ES/iPS cells, including effective use of small molecules, is summarized, and some of the problems that still need to be overcome are discussed.

  1. TGF-β1 promotes acinar to ductal metaplasia of human pancreatic acinar cells.

    Science.gov (United States)

    Liu, Jun; Akanuma, Naoki; Liu, Chengyang; Naji, Ali; Halff, Glenn A; Washburn, William K; Sun, Luzhe; Wang, Pei

    2016-08-03

    Animal studies suggest that pancreatitis-induced acinar-to-ductal metaplasia (ADM) is a key event for pancreatic ductal adenocarcinoma (PDAC) initiation. However, there has not been an adequate system to explore the mechanisms of human ADM induction. We have developed a flow cytometry-based, high resolution lineage tracing method and 3D culture system to analyse ADM in human cells. In this system, well-known mouse ADM inducers did not promote ADM in human cells. In contrast, TGF-β1 efficiently converted human acinar cells to duct-like cells (AD) in a SMAD-dependent manner, highlighting fundamental differences between the species. Functionally, AD cells gained transient proliferative capacity. Furthermore, oncogenic KRAS did not induce acinar cell proliferation, but did sustain the proliferation of AD cells, suggesting that oncogenic KRAS requires ADM-associated-changes to promote PDAC initiation. This ADM model provides a novel platform to explore the mechanisms involved in the development of human pancreatic diseases.

  2. Neuropeptide Y in the Adult and Fetal Human Pineal Gland

    Directory of Open Access Journals (Sweden)

    Morten Møller

    2014-01-01

    Full Text Available Neuropeptide Y was isolated from the porcine brain in 1982 and shown to be colocalized with noradrenaline in sympathetic nerve terminals. The peptide has been demonstrated to be present in sympathetic nerve fibers innervating the pineal gland in many mammalian species. In this investigation, we show by use of immunohistochemistry that neuropeptide Y is present in nerve fibers of the adult human pineal gland. The fibers are classical neuropeptidergic fibers endowed with large boutons en passage and primarily located in a perifollicular position with some fibers entering the pineal parenchyma inside the follicle. The distance from the immunoreactive terminals to the pinealocytes indicates a modulatory function of neuropeptide Y for pineal physiology. Some of the immunoreactive fibers might originate from neurons located in the brain and be a part of the central innervation of the pineal gland. In a series of human fetuses, neuropeptide Y-containing nerve fibers was present and could be detected as early as in the pineal of four- to five-month-old fetuses. This early innervation of the human pineal is different from most rodents, where the innervation starts postnatally.

  3. Neuropeptide Y in the adult and fetal human pineal gland.

    Science.gov (United States)

    Møller, Morten; Phansuwan-Pujito, Pansiri; Badiu, Corin

    2014-01-01

    Neuropeptide Y was isolated from the porcine brain in 1982 and shown to be colocalized with noradrenaline in sympathetic nerve terminals. The peptide has been demonstrated to be present in sympathetic nerve fibers innervating the pineal gland in many mammalian species. In this investigation, we show by use of immunohistochemistry that neuropeptide Y is present in nerve fibers of the adult human pineal gland. The fibers are classical neuropeptidergic fibers endowed with large boutons en passage and primarily located in a perifollicular position with some fibers entering the pineal parenchyma inside the follicle. The distance from the immunoreactive terminals to the pinealocytes indicates a modulatory function of neuropeptide Y for pineal physiology. Some of the immunoreactive fibers might originate from neurons located in the brain and be a part of the central innervation of the pineal gland. In a series of human fetuses, neuropeptide Y-containing nerve fibers was present and could be detected as early as in the pineal of four- to five-month-old fetuses. This early innervation of the human pineal is different from most rodents, where the innervation starts postnatally.

  4. Two Avirulent, Lentogenic Strains of Newcastle Disease Virus Are Cytotoxic for Some Human Pancreatic Tumor Lines In Vitro

    Directory of Open Access Journals (Sweden)

    Robert J Walter

    2012-09-01

    Full Text Available Context Pancreatic cancer is the fourth leading cause of cancer death in the U.S. Highly infectious Newcastle disease virus (NDV strains are known to be very cytotoxic for an array of human tumor cell types in vitro and in vivo but the effects of these and avirulent NDV strains on pancreatic neoplasms are little known. Objective Here, the direct cytolytic effects of the avirulent Hitchner-B1 (B1 and Ulster (U NDV strains on 7 human pancreatic tumor cell lines and 4 normal human cell lines were studied. Methods Cytotoxicity assays used serially diluted NDV to determine minimum cytotoxic plaque forming unit (PFU doses. Results For NDV-B1, normal human cells were killed only by relatively high doses (range: 471-3,724 PFU whereas NDV-U killed these cells at low PFU (range: 0.32-1.60 PFU. Most pancreatic cancer cell types were killed by much lower NDV-B1 doses (range: 0.40- 2.60 PFU while NDV-U killed Capan-1 and SU.86.86 cultures at very low doses (0.00041 PFU and 0.0034 PFU, respectively. Conclusions On average, 1,555 times more NDV-B1 was needed to kill normal cells than most pancreatic tumor cells and 558 times more NDV-U to kill the two most sensitive pancreatic cancer lines. These innately-targeted lentogenic viruses may have meaningful potential in treating pancreatic cancer.

  5. Effect of recombinant adenovirus vector mediated human interleukin-24 gene transfection on pancreatic carcinoma growth

    Institute of Scientific and Technical Information of China (English)

    PAN Xin-ting; ZHU Qing-yun; LI De-chun; YANG Ji-cheng; ZHANG Zi-xiang; ZHU Xing-guo; ZHAO Hua

    2008-01-01

    Background Pancreatic cancer is a highly malignant tumor affecting an ever increasing number of patients with a mean 5-year survival rate below 4%. Therefore, gene therapy for cancer has become a potential novel therapeutic modality. In this study we sought to determine the inhibitory effects of adenovirus-mediated human interleukin-24 (AdhlL-24) on pancreatic cancer.Methods Human interleukin-24 gene was cloned into replication-defective adenovirus specific for patu8988 tumor cells by virus recombination technology. Reverse transcription-polymerase chain reaction and Western blotting analysis were used to determine the expression of human interleukin-24 mRNA in patu8988 cells in vitro. Induction of apoptosis by overexpression of human interleukin-24 in patu8988 cells was determined by flow cytometry. In vivo efficacy of adenoviral delivery of human interleukin-24 was assessed in nude mice (n=10 for each group) bearing patu8988 pancreatic cancer cell lines by determining inhibition of tumor growth, endothelial growth factor and CD34 expression, and intratumoral microvessel density (MVD).Results The recombinant adenovirus vector AdVGFP/IL-24 was constructed with a packaged recombinant retrovirus titer of 1.0x1010 pfu/ml and successfully expressed of both mRNA and protein in patu8988 cells. The AdVGFP/IL-24 induced apoptosis of patu8988 tumor cells in vitro and significantly inhibited tumor growth in vivo (P <0.05). The intratumoral MVD decreased significantly in the treated tumors (P <0.05).Conclusion The recombinant adenovirus AdGFP/IL-24 can effectively express biologically active human interleukin-24, which results in inhibition of pancreatic cancer growth.

  6. Prominent periventricular fiber system related to ganglionic eminence and striatum in the human fetal cerebrum.

    Science.gov (United States)

    Vasung, L; Jovanov-Milošević, N; Pletikos, M; Mori, S; Judaš, M; Kostović, Ivica

    2011-01-01

    Periventricular pathway (PVP) system of the developing human cerebrum is situated medial to the intermediate zone in the close proximity to proliferative cell compartments. In order to elucidate chemical properties and developing trajectories of the PVP we used DTI in combination with acetylcholinesterase histochemistry, SNAP-25 immunocytochemistry and axonal cytoskeletal markers (SMI312, MAP1b) immunocytochemistry on postmortem paraformaldehyde-fixed brains of 30 human fetuses ranging in age from 10 to 38 postconceptional weeks (PCW), 2 infants (age 1-3 months) and 1 adult brain. The PVP appears in the early fetal period (10-13 PCW) as two defined fibre bundles: the corpus callosum (CC) and the fetal fronto-occipital fascicle (FOF). In the midfetal period (15-18 PCW), all four components of the PVP can be identified: (1) the CC, which at rostral levels forms a voluminous callosal plate; (2) the FOF, with SNAP-25-positive fibers; (3) the fronto-pontine pathway (FPP) which for a short distance runs within the PVP; and (4) the subcallosal fascicle of Muratoff (SFM) which contains cortico-caudate projections. The PVPs are situated medial to the internal capsule at the level of the cortico-striatal junction; they remain prominent during the late fetal and early preterm period (19-28 PCW) and represent a portion of the wider periventricular crossroad of growing associative, callosal and projection pathways. In the perinatal period, the PVPs change their topographical relationships, decrease in size and the FOF looses its SNAP-25-reactivity. In conclusion, the hitherto undescribed PVP of the human fetal cerebrum contains forerunners of adult associative and projection pathways. Its transient chemical properties and relative exuberance suggest that the PVP may exert influence on the development of cortical connectivity (intermediate targeting) and other neurogenetic events such as neuronal proliferation. The PVP's topographical position also indicates that it is a major

  7. FGF7 and cell density are required for final differentiation of pancreatic amylase-positive cells from human ES cells.

    Science.gov (United States)

    Takizawa-Shirasawa, Sakiko; Yoshie, Susumu; Yue, Fengming; Mogi, Akimi; Yokoyama, Tadayuki; Tomotsune, Daihachiro; Sasaki, Katsunori

    2013-12-01

    The major molecular signals of pancreatic exocrine development are largely unknown. We examine the role of fibroblast growth factor 7 (FGF7) in the final induction of pancreatic amylase-containing exocrine cells from induced-pancreatic progenitor cells derived from human embryonic stem (hES) cells. Our protocol consisted in three steps: Step I, differentiation of definitive endoderm (DE) by activin A treatment of hES cell colonies; Step II, differentiation of pancreatic progenitor cells by re-plating of the cells of Step I onto 24-well plates at high density and stimulation with all-trans retinoic acid; Step III, differentiation of pancreatic exocrine cells with a combination of FGF7, glucagon-like peptide 1 and nicotinamide. The expression levels of pancreatic endodermal markers such as Foxa2, Sox17 and gut tube endoderm marker HNF1β were up-regulated in both Step I and II. Moreover, in Step III, the induced cells expressed pancreatic markers such as amylase, carboxypeptidase A and chymotrypsinogen B, which were similar to those in normal human pancreas. From day 8 in Step III, cells immunohistochemically positive for amylase and for carboxypeptidase A, a pancreatic exocrine cell product, were induced by FGF7. Pancreatic progenitor Pdx1-positive cells were localized in proximity to the amylase-positive cells. In the absence of FGF7, few amylase-positive cells were identified. Thus, our three-step culture protocol for human ES cells effectively induces the differentiation of amylase- and carboxypeptidase-A-containing pancreatic exocrine cells.

  8. Human placental lactogen and unconjugated estriol concentrations in twin pregnancy: monitoring of fetal development in intrauterine growth retardation and single intrauterine fetal death.

    Science.gov (United States)

    Trapp, M; Kato, K; Bohnet, H G; Gerhard, I; Weise, H C; Leidenberger, F

    1986-11-01

    Human placental lactogen and unconjugated estriol concentrations in maternal serum were evaluated in 100 uneventful twin pregnancies, and these values were compared with those observed in 16 twin pregnancies associated with intrauterine growth retardation or single intrauterine fetal death. In pregnancies associated with intrauterine growth retardation (n = 8), human placental lactogen levels were at the lower limit of normal range for singleton pregnancies, whereas estriol levels were normal in most cases. When one of the fetuses had died before week 33 of pregnancy (n = 5), both human placental lactogen and estriol levels were low and they were almost at the levels in singleton pregnancy. When intrauterine fetal death occurred after week 36 of pregnancy (n = 3), both hormone levels remained normal until term. Thus human placental lactogen rather than estriol is a good indicator of intrauterine growth retardation in twin pregnancy. Both human placental lactogen and estriol are useful for the monitoring of the surviving fetus in the case of single intrauterine fetal death.

  9. Distinct differences in the responses of the human pancreatic β-cell line EndoC-βH1 and human islets to proinflammatory cytokines.

    Science.gov (United States)

    Oleson, Bryndon J; McGraw, Jennifer A; Broniowska, Katarzyna A; Annamalai, Mani; Chen, Jing; Bushkofsky, Justin R; Davis, Dawn B; Corbett, John A; Mathews, Clayton E

    2015-09-01

    While insulinoma cells have been developed and proven to be extremely useful in studies focused on mechanisms controlling β-cell function and viability, translating findings to human β-cells has proven difficult because of the limited access to human islets and the absence of suitable insulinoma cell lines of human origin. Recently, a human β-cell line, EndoC-βH1, has been derived from human fetal pancreatic buds. The purpose of this study was to determine whether human EndoC-βH1 cells respond to cytokines in a fashion comparable to human islets. Unlike most rodent-derived insulinoma cell lines that respond to cytokines in a manner consistent with rodent islets, EndoC-βH1 cells fail to respond to a combination of cytokines (IL-1, IFN-γ, and TNF) in a manner consistent with human islets. Nitric oxide, produced following inducible nitric oxide synthase (iNOS) expression, is a major mediator of cytokine-induced human islet cell damage. We show that EndoC-βH1 cells fail to express iNOS or produce nitric oxide in response to this combination of cytokines. Inhibitors of iNOS prevent cytokine-induced loss of human islet cell viability; however, they do not prevent cytokine-induced EndoC-βH1 cell death. Stressed human islets or human islets expressing heat shock protein 70 (HSP70) are resistant to cytokines, and, much like stressed human islets, EndoC-βH1 cells express HSP70 under basal conditions. Elevated basal expression of HSP70 in EndoC-βH1 cells is consistent with the lack of iNOS expression in response to cytokine treatment. While expressing HSP70, EndoC-βH1 cells fail to respond to endoplasmic reticulum stress activators, such as thapsigargin. These findings indicate that EndoC-βH1 cells do not faithfully recapitulate the response of human islets to cytokines. Therefore, caution should be exercised when making conclusions regarding the actions of cytokines on human islets when using this human-derived insulinoma cell line.

  10. Development of the insulin secretion mechanism in fetal and neonatal rat pancreatic B-cells: response to glucose, K+, theophylline, and carbamylcholine

    Directory of Open Access Journals (Sweden)

    A.C. Mendonça

    1998-06-01

    Full Text Available We studied the development of the insulin secretion mechanism in the pancreas of fetal (19- and 21-day-old, neonatal (3-day-old, and adult (90-day-old rats in response to stimulation with 8.3 or 16.7 mM glucose, 30 mM K+, 5 mM theophylline (Theo and 200 µM carbamylcholine (Cch. No effect of glucose or high K+ was observed on the pancreas from 19-day-old fetuses, whereas Theo and Cch significantly increased insulin secretion at this age (82 and 127% above basal levels, respectively. High K+ also failed to alter the insulin secretion in the pancreas from 21-day-old fetuses, whereas 8.3 mM and 16.7 mM glucose significantly stimulated insulin release by 41 and 54% above basal levels, respectively. Similar results were obtained with Theo and Cch. A more marked effect of glucose on insulin secretion was observed in the pancreas of 3-day-old rats, reaching 84 and 179% above basal levels with 8.3 mM and 16.7 mM glucose, respectively. At this age, both Theo and Cch increased insulin secretion to close to two-times basal levels. In islets from adult rats, 8.3 mM and 16.7 mM glucose, Theo, and Cch increased the insulin release by 104, 193, 318 and 396% above basal levels, respectively. These data indicate that pancreatic B-cells from 19-day-old fetuses were already sensitive to stimuli that use either cAMP or IP3 and DAG as second messengers, but insensitive to stimuli such as glucose and high K+ that induce membrane depolarization. The greater effect of glucose on insulin secretion during the neonatal period indicates that this period is crucial for the maturation of the glucose-sensing mechanism in B-cells.

  11. The early fetal development of human neocortical GABAergic interneurons.

    Science.gov (United States)

    Al-Jaberi, Nahidh; Lindsay, Susan; Sarma, Subrot; Bayatti, Nadhim; Clowry, Gavin J

    2015-03-01

    GABAergic interneurons are crucial to controlling the excitability and responsiveness of cortical circuitry. Their developmental origin may differ between rodents and human. We have demonstrated the expression of 12 GABAergic interneuron-associated genes in samples from human neocortex by quantitative rtPCR from 8 to 12 postconceptional weeks (PCW) and shown a significant anterior to posterior expression gradient, confirmed by in situ hybridization or immunohistochemistry for GAD1 and 2, DLX1, 2, and 5, ASCL1, OLIG2, and CALB2. Following cortical plate (CP) formation from 8 to 9 PCW, a proportion of cells were strongly stained for all these markers in the CP and presubplate. ASCL1 and DLX2 maintained high expression in the proliferative zones and showed extensive immunofluorescent double-labeling with the cell division marker Ki-67. CALB2-positive cells increased steadily in the SVZ/VZ from 10 PCW but were not double-labeled with Ki-67. Expression of GABAergic genes was generally higher in the dorsal pallium than in the ganglionic eminences, with lower expression in the intervening ventral pallium. It is widely accepted that the cortical proliferative zones may generate CALB2-positive interneurons from mid-gestation; we now show that the anterior neocortical proliferative layers especially may be a rich source of interneurons in the early neocortex.

  12. Human autologous serum as a substitute for fetal bovine serum in human Schwann cell culture.

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    Parisa Goodarzi

    2014-04-01

    Full Text Available Nowadays, cell -based and tissue engineered products have opened new horizons in treatment of incurable nervous system disorders. The number of studies on the role of Schwann cells (SC in treating nervous disorders is higher than other cell types. Different protocols have been suggested for isolation and expansion of SC which most of them have used multiple growth factors, mitogens and fetal bovine sera (FBS in culture medium. Because of potential hazards of animal-derived reagents, this study was designed to evaluate the effect of replacing FBS with human autologous serum (HAS on SC's yield and culture parameters. Samples from 10 peripheral nerve biopsies were retrieved and processed under aseptic condition. The isolated cells cultured in FBS (1st group or autologous serum (2nd group. After primary culture the cells were seeded at 10000 cell/cm2 in a 12 wells cell culture plate for each group. At 100% confluency, the cell culture parameters (count, viability, purity and culture duration of 2 groups were compared using paired t-test. The average donors' age was 35.80 (SD=13.35 and except for 1 sample the others cultured successfully. In first group, the averages of cell purity, viability and culture duration were 97% (SD=1.32, 97/33% (SD=1.22 and 11.77 (SD=2.58 days respectively. This parameters were 97.33% (SD=1.00, 97.55% (SD=1.33 and 10.33 days (SD=1.65 in second group. The difference of cell count, purity and viability were not significant between 2 groups (P>0.05. The cells of second group reached to 100% confluency in shorter period of time (P=0.03. The results of this study showed that autologous serum can be a good substitute for FBS in human SC culture. This can reduce the costs and improve the safety of cell product for clinical application.

  13. Production of embryonic and fetal-like red blood cells from human induced pluripotent stem cells.

    Science.gov (United States)

    Chang, Chan-Jung; Mitra, Koyel; Koya, Mariko; Velho, Michelle; Desprat, Romain; Lenz, Jack; Bouhassira, Eric E

    2011-01-01

    We have previously shown that human embryonic stem cells can be differentiated into embryonic and fetal type of red blood cells that sequentially express three types of hemoglobins recapitulating early human erythropoiesis. We report here that we have produced iPS from three somatic cell types: adult skin fibroblasts as well as embryonic and fetal mesenchymal stem cells. We show that regardless of the age of the donor cells, the iPS produced are fully reprogrammed into a pluripotent state that is undistinguishable from that of hESCs by low and high-throughput expression and detailed analysis of globin expression patterns by HPLC. This suggests that reprogramming with the four original Yamanaka pluripotency factors leads to complete erasure of all functionally important epigenetic marks associated with erythroid differentiation regardless of the age or the tissue type of the donor cells, at least as detected in these assays. The ability to produce large number of erythroid cells with embryonic and fetal-like characteristics is likely to have many translational applications.

  14. Potential of human fetal chorionic stem cells for the treatment of osteogenesis imperfecta.

    Science.gov (United States)

    Jones, Gemma N; Moschidou, Dafni; Abdulrazzak, Hassan; Kalirai, Bhalraj Singh; Vanleene, Maximilien; Osatis, Suchaya; Shefelbine, Sandra J; Horwood, Nicole J; Marenzana, Massimo; De Coppi, Paolo; Bassett, J H Duncan; Williams, Graham R; Fisk, Nicholas M; Guillot, Pascale V

    2014-02-01

    Osteogenesis imperfecta (OI) is a genetic bone pathology with prenatal onset, characterized by brittle bones in response to abnormal collagen composition. There is presently no cure for OI. We previously showed that human first trimester fetal blood mesenchymal stem cells (MSCs) transplanted into a murine OI model (oim mice) improved the phenotype. However, the clinical use of fetal MSC is constrained by their limited number and low availability. In contrast, human fetal early chorionic stem cells (e-CSC) can be used without ethical restrictions and isolated in high numbers from the placenta during ongoing pregnancy. Here, we show that intraperitoneal injection of e-CSC in oim neonates reduced fractures, increased bone ductility and bone volume (BV), increased the numbers of hypertrophic chondrocytes, and upregulated endogenous genes involved in endochondral and intramembranous ossification. Exogenous cells preferentially homed to long bone epiphyses, expressed osteoblast genes, and produced collagen COL1A2. Together, our data suggest that exogenous cells decrease bone brittleness and BV by directly differentiating to osteoblasts and indirectly stimulating host chondrogenesis and osteogenesis. In conclusion, the placenta is a practical source of stem cells for the treatment of OI.

  15. Production of embryonic and fetal-like red blood cells from human induced pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Chan-Jung Chang

    Full Text Available We have previously shown that human embryonic stem cells can be differentiated into embryonic and fetal type of red blood cells that sequentially express three types of hemoglobins recapitulating early human erythropoiesis. We report here that we have produced iPS from three somatic cell types: adult skin fibroblasts as well as embryonic and fetal mesenchymal stem cells. We show that regardless of the age of the donor cells, the iPS produced are fully reprogrammed into a pluripotent state that is undistinguishable from that of hESCs by low and high-throughput expression and detailed analysis of globin expression patterns by HPLC. This suggests that reprogramming with the four original Yamanaka pluripotency factors leads to complete erasure of all functionally important epigenetic marks associated with erythroid differentiation regardless of the age or the tissue type of the donor cells, at least as detected in these assays. The ability to produce large number of erythroid cells with embryonic and fetal-like characteristics is likely to have many translational applications.

  16. Microarray-bioinformatics analysis of altered genomic expression profiles between human fetal and infant myocardium

    Institute of Scientific and Technical Information of China (English)

    KONG Bo; LIU Ying-long; L(U) Xiao-dong

    2008-01-01

    Background The physiological differences between fetal and postnatal heart have been well characterized at the cellular level. However, the genetic mechanisms governing and regulating these differences have only been partially elucidated. Elucidation of the differentially expressed genes profile before and after birth has never been systematically proposed and analyzed.Methods The human oligonuclectide microarray and bioinformatics analysis approaches were applied to isolate and classify the differentially expressed genes between fetal and infant cardiac tissue samples. Quantitative real-time PCR was used to confirm the results from the microarray.Results Two hundred and forty-two differentially expressed genes were discovered and classified into 13 categories, including genes related to energy metabolism, myocyte hyperplasia, development, muscle contraction, protein synthesis and degradation, extraceUular matrix components, transcription factors, apoptosis, signal pathway molecules, organelle organization and several other biological processes. Moreover, 95 genes were identified which had not previously been reported to be expressed in the heart.Conclusions The study systematically analyzed the alteration of the gene expression profile between the human fetal and infant myocardium. A number of genes were discovered which had not been reported to be expressed in the heart. The data provided insight into the physical development mechanisms of the heart before and after birth.KONG Bo and LU Xiao-dong contributed equally to this study.

  17. The Effect of Simvastatin on Infection-Induced Inflammatory Response of Human Fetal Membranes.

    Science.gov (United States)

    Basraon, Sanmaan K; Costantine, Maged M; Saade, George; Menon, Ramkumar

    2015-07-01

    Inflammatory cytokines and matrix metalloproteinases (MMPs) contribute to preterm labor pathophysiology. The objective of this study was to test anti-inflammatory properties of simvastatin in human fetal membranes exposed to lipopolysaccharide (LPS). Normal term human fetal membrane explants (n = 11) were allocated to one of the six study groups: control, LPS only (100 ng/mL), simvastatin only (125 ng/mL), simvastatin given 6 hrs prior to LPS (S-L), simvastatin given 6 hrs post-LPS (L-S), and simvastatin and LPS given simultaneously (L+S). Explants were incubated for 24 hrs. Multiplex ELISA for cytokines: IL-1β, IL-6, IL-10, and TNF-α; soluble cytokine receptors: sIL-1R2, sIL-6R, sTNFR1, and R2; MMPs (1, 2, 7, 9, and 10); and tissue inhibitor of metalloproteinase-2 (TIMP-2) was performed on tissue culture supernatants. Pairwise comparison between different groups was conducted by least square mean estimates. Compared with controls, LPS stimulation increased cytokine production and their tissue bioavailability (measured as the molar ratio of cytokine to its soluble receptor), thus confirming membrane immune reactivity (P fetal inflammatory response associated with infection-induced preterm birth. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  18. Trans—acting factors from the human fetal liver binding to the human ε—globin gene silencer

    Institute of Scientific and Technical Information of China (English)

    YANZHIJIANG; CHUJIANG; 等

    1997-01-01

    The developmental stage-specific silencing of the human ε-globin gene during embryonic life is controlled,in part,by the silencer (-392bp- -177bp) upstream of this gene.In order to elucidate its role,the nuclear extract from the human fetal liver has been prepared and the interactions between trans-acting factors and this silencer element have been examined.By using DNaseI footprinting assay,a major protected region from -278bp to -235bp within this silencer element was identified.Furthermore,we found in gel mobility shift assay and Southwestern blotting assay that there were at least four trans-acting factors (MV≈32,28,26 and 22kD) in the nuclear extract isolated from the human fetal liver,which could specifically bind to this region.Our results suggested that these trans-acting factors might play an important role in silencing the human embryonic ε-globin gene expression at the fetal stage through the interactions with this silencer.

  19. Fetal and adult hematopoietic stem cells give rise to distinct T cell lineages in humans.

    Science.gov (United States)

    Mold, Jeff E; Venkatasubrahmanyam, Shivkumar; Burt, Trevor D; Michaëlsson, Jakob; Rivera, Jose M; Galkina, Sofiya A; Weinberg, Kenneth; Stoddart, Cheryl A; McCune, Joseph M

    2010-12-17

    Although the mammalian immune system is generally thought to develop in a linear fashion, findings in avian and murine species argue instead for the developmentally ordered appearance (or "layering") of distinct hematopoietic stem cells (HSCs) that give rise to distinct lymphocyte lineages at different stages of development. Here we provide evidence of an analogous layered immune system in humans. Our results suggest that fetal and adult T cells are distinct populations that arise from different populations of HSCs that are present at different stages of development. We also provide evidence that the fetal T cell lineage is biased toward immune tolerance. These observations offer a mechanistic explanation for the tolerogenic properties of the developing fetus and for variable degrees of immune responsiveness at birth.

  20. Pancreatic Cancer Cell Lines Can Induce Prostaglandin E2 Production from Human Blood Mononuclear Cells

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    Svitlana P. Grekova

    2011-01-01

    Full Text Available Accumulating evidence suggests an important role for cyclooxygenase-2 (COX-2 in the pathogenesis of a wide range of malignancies. The protumorigenic properties of COX-2 are generally thought to be mediated by its product, PGE2, which is shown to promote tumor spread and growth by multiple mechanisms but most importantly through modulation of the local immune response in the tumor. Pancreatic tumor cells produce various amounts of PGE2, some of them being even deficient in COX enzymes or other PGE2 synthases. Here we describe that, beside pancreatic tumor cells or stromal fibroblasts, human peripheral blood mononuclear cells can also produce PGE2 upon coculture with pancreatic cancer cells. Stimulating of cellular cPLA2 within PBMCs by secreted factors, presumably sPLA2, from tumor cells appeared crucial, while the direct contact between PBMCs and PDACs seemed to be dispensable for this effect. Our data is emphasizing the complex interactions participating in the formation of the tolerogenic immune milieu within pancreatic tumors.

  1. Generation of Rat Monoclonal Antibodies Against Human Pancreatic Ductal Adenocarcinoma Cells.

    Science.gov (United States)

    Higashi, Kiyoshi; Fujii, Nobuaki; Kushida, Masahiko; Yamada, Keita; Suzuki, Noriyuki; Saito, Koichi; Tachibana, Taro

    2016-06-01

    Pancreatic ductal adenocarcinoma is an aggressive tumor with a poor prognosis. Biomarkers that can detect the tumor in its early stages when it may be amenable to curative resection might improve prognosis. To discover novel markers expressed in primary pancreatic cancer, we generated a panel of monoclonal antibodies against pancreatic ductal adenocarcinoma cell line BxPC3 using a rat medial iliac lymph node method. The antigen recognized by 1B5A5 was expressed on the cell surface and secreted into the conditioned medium of BxPC3 cells, and characterized as glycoproteins with molecular mass between 60 and 90 kDa. A wide range of molecular weights of 1B5A5 antigen in several pancreatic cancer cell lines were observed. Immunohistochemistry using a human multiple organ tumor tissue array showed an enhanced expression of 1B5A5 antigen in pancreas, lung, stomach, breast, urinary bladder, colon, and cervix uteri cancers. Immunoprecipitation followed by proteomic analyses identified CEACAM6 as a 1B5A5 antigen. In addition, western blot analysis results indicated that the 1B5A5 epitope is located within an amino-terminal domain of CEACAM6. These results raised the possibility that our approach could lead to discovery of novel biomarkers for the early stage of cancers in a relatively short period of time.

  2. SMAD4-dependent polysome RNA recruitment in human pancreatic cancer cells.

    Science.gov (United States)

    Thornley, Jessica A; Trask, Heidi W; Ringelberg, Carol S; Ridley, Christian J A; Wang, Sinny; Sal-Lari, Richard Cowper; Moore, Jason H; Korc, Murray; Tomlinson, Craig R

    2012-10-01

    Pancreatic cancer is the fourth leading cause of cancer death in the United States because most patients are diagnosed too late in the course of the disease to be treated effectively. Thus, there is a pressing need to more clearly understand how gene expression is regulated in cancer cells and to identify new biomarkers and therapeutic targets. Translational regulation is thought to occur primarily through non-SMAD directed signaling pathways. We tested the hypothesis that SMAD4-dependent signaling does play a role in the regulation of mRNA entry into polysomes and that novel candidate genes in pancreatic cancer could be identified using polysome RNA from the human pancreatic cancer cell line BxPC3 with or without a functional SMAD4 gene. We found that (i) differentially expressed whole cell and cytoplasm RNA levels are both poor predictors of polysome RNA levels; (ii) for a majority of RNAs, differential RNA levels are regulated independently in the nucleus, cytoplasm, and polysomes; (iii) for most of the remaining polysome RNA, levels are regulated via a "tagging" of the RNAs in the nucleus for rapid entry into the polysomes; (iv) a SMAD4-dependent pathway appears to indeed play a role in regulating mRNA entry into polysomes; and (v) a gene list derived from differentially expressed polysome RNA in BxPC3 cells generated new candidate genes and cell pathways potentially related to pancreatic cancer. Copyright © 2011 Wiley Periodicals, Inc.

  3. Programmed cell death in developing human fetal CNS

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The spatial and temporal distributions of programmed cell death (PCD) in developing central nervous system (CNS) of human fetuses ranging from 12 to 39 weeks of gestation were investigated using techniques of flow cytometry and terminal transferase-mediated nick end labeling (TUNEL). The results showed that PCD did occur in every representative brain region of all fetuses examined in different stages. It was found that there were two peaks of PCD appearing at the 12th and 39th weeks respectively, which suggested that the first peak of apoptosis may be involved in the selective elimination of neurons overproduced during the early development and the second may play an important role in establishing the correct neuronal circuitry.

  4. Acute pancreatitis

    Science.gov (United States)

    ... its blood vessels. This problem is called acute pancreatitis. Acute pancreatitis affects men more often than women. Certain ... pancreatitis; Pancreas - inflammation Images Digestive system Endocrine glands Pancreatitis, acute - CT scan Pancreatitis - series References Forsmark CE. Pancreatitis. ...

  5. Activation of c-Ki-ras gene in human pancreatic cancer.

    Science.gov (United States)

    Prassolov, V S; Sakamoto, H; Nishimura, S; Terada, M; Sugimura, T

    1985-09-01

    DNA isolated from a lymph node with metastasis from pancreatic adenocarcinoma in a Japanese male patient transformed NIH3T3 cells upon transfection by the calcium-phosphate precipitation technique. Analysis of DNA from the transformant revealed the presence of an activated human c-Ki-ras gene, which is considered to be responsible for the transformation of the NIH3T3 cells.

  6. Cytokeratin (CK5, CK8, CK14) expression and presence of progenitor stem cells in human fetal thymuses.

    Science.gov (United States)

    Gupta, Richa; Gupta, Tulika; Kaur, Harjeet; Sehgal, Shobha; Aggarwal, Anjali; Kapoor, Kanchan; Sharma, Anshu; Sahni, Daisy; Singla, Suhalika

    2016-09-01

    The aim of the current study was to observe the expression of cytokeratins in human fetal thymuses. Specific cytokeratin markers in adult humans and mice have been well described but there has been little similar work on human fetuses. We also aimed to see whether progenitor stem cells that could be harvested to treat various immunodeficiency disorders are present in fetal thymic tissue. Thymuses obtained from 30 aborted human fetuses (12 to 31 weeks) were examined immunohistochemically to investigate changes in cytokeratin expression in the epithelial cells (TEC) at various gestational ages. Before 16 weeks of gestation, cortical (cTEC) and medullary (mTEC) TEC exhibited homogenous staining for cytokeratins CK8 and CK5. After 16 weeks there was differential staining, with cTEC positive for CK8 and mTEC for CK5 and CK14. Interestingly, both CK5 + CK8+ progenitor stem cells were present in the fetal thymic cortex at all gestational ages, with a relatively high number from 12 to 16 weeks. Cytokeratin expression in fetal thymuses was quite different from that in the adult thymus owing to the presence of undifferentiated progenitor stem cells in fetal thymic stroma along with differentiated TEC. The best time to harvest these progenitor stem cells from fetal thymic stroma in order to treat various immune deficiency disorders appears to be 12-16 weeks. Clin. Anat. 29:711-717, 2016. © 2016 Wiley Periodicals, Inc.

  7. Inner ring deiodination of thyroxine and 3,5,3'-triiodothyronine by human fetal membranes

    Energy Technology Data Exchange (ETDEWEB)

    Roti, E.; Fang, S.L.; Green, K.; Braverman, L.E.; Emerson, C.H.

    1983-12-01

    Indirect evidence, based on injection of thyroxine (T4) into the amniotic cavity of humans, and maternal thyroidectomy in the rat, suggests that fetal membranes might be capable of converting T4 to 3,3',5'-triiodothyronine (rT3) by virtue of inner ring iodothyronine deiodinase activity. The present study was undertaken to provide direct evidence that human fetal membranes contain inner ring iodothyronine deiodinase activity directed toward T4 and 3,5,3'-triiodothyronine (T3). Homogenates of human fetal membranes were incubated with 125I-labeled T4, rT3, and T3, and with stable T4. Conversion of 125(I)-T4 to 125(I)-rT3 was noted in chorion and amnion. 125I-T3 was converted to 125(I)-3,3'-diiodothyronine (T2) in chorion and amnion. 125(I)-rT3 was stable in fetal membranes under the incubation conditions employed. Time-, temperature-, pH-, and protein content-dependent conversion of stable T4 to rT3 was found in fetal membranes. Iodothyronine metabolism did not occur in the absence of dithiothreitol. These studies indicate that human fetal membranes contain an inner ring deiodinase enzyme. Because of its intimate contact with the amniotic cavity, this enzyme may generate a portion of the rT3 found in amniotic fluid.

  8. Tumor-specific gene therapy for pancreatic cancer using human neural stem cells encoding carboxylesterase.

    Science.gov (United States)

    Choi, Sung S; Yoon, Kichul; Choi, Seon-A; Yoon, Seung-Bin; Kim, Seung U; Lee, Hong J

    2016-11-15

    Advanced pancreatic cancer is one of the most lethal malignant human diseases lacking effective treatment. Its extremely low survival rate necessitates development of novel therapeutic approach. Human neural stem cells (NSCs) are known to have tumor-tropic effect. We genetically engineered them to express rabbit carboxyl esterase (F3.CE), which activates prodrug CPT-11(irinotecan) into potent metabolite SN-38. We found significant inhibition of the growth of BxPC3 human pancreatic cancer cell line in vitro by F3.CE in presence of CPT-11. Apoptosis was also markedly increased in BxPC3 cells treated with F3.CE and CPT-11. The ligand VEGF and receptor VEGF-1(Flt1) were identified to be the relevant tumor-tropic chemoattractant. We confirmed in vivo that in mice injected with BxPC3 on their skin, there was significant reduction of tumor size in those treated with both F3.CE and BxPC3 adjacent to the cancer mass. Administration of F3.CE in conjunction with CPT-11 could be a new possibility as an effective treatment regimen for patients suffering from advanced pancreatic cancer.

  9. Ontological differences in first compared to third trimester human fetal placental chorionic stem cells.

    Directory of Open Access Journals (Sweden)

    Gemma N Jones

    Full Text Available Human mesenchymal stromal/stem cells (MSC isolated from fetal tissues hold promise for use in tissue engineering applications and cell-based therapies, but their collection is restricted ethically and technically. In contrast, the placenta is a potential source of readily-obtainable stem cells throughout pregnancy. In fetal tissues, early gestational stem cells are known to have advantageous characteristics over neonatal and adult stem cells. Accordingly, we investigated whether early fetal placental chorionic stem cells (e-CSC were physiologically superior to their late gestation fetal chorionic counterparts (l-CSC. We showed that e-CSC shared a common phenotype with l-CSC, differentiating down the osteogenic, adipogenic and neurogenic pathways, and containing a subset of cells endogenously expressing NANOG, SOX2, c-MYC, and KLF4, as well as an array of genes expressed in pluripotent stem cells and primordial germ cells, including CD24, NANOG, SSEA4, SSEA3, TRA-1-60, TRA-1-81, STELLA, FRAGILIS, NANOS3, DAZL and SSEA1. However, we showed that e-CSC have characteristics of an earlier state of stemness compared to l-CSC, such as smaller size, faster kinetics, uniquely expressing OCT4A variant 1 and showing higher levels of expression of NANOG, SOX2, c-MYC and KLF4 than l-CSC. Furthermore e-CSC, but not l-CSC, formed embryoid bodies containing cells from the three germ layer lineages. Finally, we showed that e-CSC demonstrate higher tissue repair in vivo; when transplanted in the osteogenesis imperfecta mice, e-CSC, but not l-CSC increased bone quality and plasticity; and when applied to a skin wound, e-CSC, but not l-CSC, accelerated healing compared to controls. Our results provide insight into the ontogeny of the stemness phenotype during fetal development and suggest that the more primitive characteristics of early compared to late gestation fetal chorionic stem cells may be translationally advantageous.

  10. A scalable system for production of functional pancreatic progenitors from human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Thomas C Schulz

    Full Text Available Development of a human embryonic stem cell (hESC-based therapy for type 1 diabetes will require the translation of proof-of-principle concepts into a scalable, controlled, and regulated cell manufacturing process. We have previously demonstrated that hESC can be directed to differentiate into pancreatic progenitors that mature into functional glucose-responsive, insulin-secreting cells in vivo. In this study we describe hESC expansion and banking methods and a suspension-based differentiation system, which together underpin an integrated scalable manufacturing process for producing pancreatic progenitors. This system has been optimized for the CyT49 cell line. Accordingly, qualified large-scale single-cell master and working cGMP cell banks of CyT49 have been generated to provide a virtually unlimited starting resource for manufacturing. Upon thaw from these banks, we expanded CyT49 for two weeks in an adherent culture format that achieves 50-100 fold expansion per week. Undifferentiated CyT49 were then aggregated into clusters in dynamic rotational suspension culture, followed by differentiation en masse for two weeks with a four-stage protocol. Numerous scaled differentiation runs generated reproducible and defined population compositions highly enriched for pancreatic cell lineages, as shown by examining mRNA expression at each stage of differentiation and flow cytometry of the final population. Islet-like tissue containing glucose-responsive, insulin-secreting cells was generated upon implantation into mice. By four- to five-months post-engraftment, mature neo-pancreatic tissue was sufficient to protect against streptozotocin (STZ-induced hyperglycemia. In summary, we have developed a tractable manufacturing process for the generation of functional pancreatic progenitors from hESC on a scale amenable to clinical entry.

  11. LGL1 modulates proliferation, apoptosis, and migration of human fetal lung fibroblasts.

    Science.gov (United States)

    Zhang, Hui; Sweezey, Neil B; Kaplan, Feige

    2015-02-15

    Rapid growth and formation of new gas exchange units (alveogenesis) are hallmarks of the perinatal lung. Bronchopulmonary dysplasia (BPD), common in very premature infants, is characterized by premature arrest of alveogenesis. Mesenchymal cells (fibroblasts) regulate both lung branching and alveogenesis through mesenchymal-epithelial interactions. Temporal or spatial deficiency of late-gestation lung 1/cysteine-rich secretory protein LD2 (LGL1/CRISPLD2), expressed in and secreted by lung fibroblasts, can impair both lung branching and alveogenesis (LGL1 denotes late gestation lung 1 protein; LGL1 denotes the human gene; Lgl1 denotes the mouse/rat gene). Absence of Lgl1 is embryonic lethal. Lgl1 levels are dramatically reduced in oxygen toxicity rat models of BPD, and heterozygous Lgl1(+/-) mice exhibit features resembling human BPD. To explore the role of LGL1 in mesenchymal-epithelial interactions in developing lung, we developed a doxycycline (DOX)-inducible RNA-mediated LGL1 knockdown cellular model in human fetal lung fibroblasts (MRC5(LGL1KD)). We assessed the impact of LGL1 on cell proliferation, cell migration, apoptosis, and wound healing. DOX-induced MRC5(LGL1KD) suppressed cell growth and increased apoptosis of annexin V(+) staining cells and caspase 3/7 activity. LGL1-conditioned medium increased migration of fetal rat primary lung epithelial cells and human airway epithelial cells. Impaired healing by MRC5(LGL1KD) cells of a wound model was attenuated by addition of LGL1-conditioned medium. Suppression of LGL1 was associated with dysregulation of extracellular matrix genes (downregulated MMP1, ColXVα1, and ELASTIN) and proapoptosis genes (upregulated BAD, BAK, CASP2, and TNFRSF1B) and inhibition of 44/42MAPK phosphorylation. Our findings define a role for LGL1 in fibroblast expansion and migration, epithelial cell migration, and mesenchymal-epithelial signaling, key processes in fetal lung development.

  12. Recovery Rates of Human Fetal Skeletal Remains Using Varying Mesh Sizes.

    Science.gov (United States)

    Pokines, James T; De La Paz, Jade S

    2016-01-01

    Human fetal skeletal elements of different gestational ages were screened with multiple mesh sizes (6.4 mm [1/4 inch], 3.2 mm [1/8 inch], 2.0 mm, and 1.0 mm) to determine their recovery rates. All remains were previously macerated, and no significantly damaged elements were used. The 6.4 mm mesh allowed a large loss of elements (63.2% overall), including diagnostic elements, while no diagnostic elements were lost when the 1 mm mesh (0.2%) was used. When using the 3.2 mm mesh, 16.2% of the bones were lost, including some diagnostic elements (primarily tooth crowns), while 7.5% were lost using the 2.0 mm mesh. The authors recommend that the potential loss of information incurred when utilizing larger mesh sizes be taken into consideration when planning recovery methods where fetal remains may be encountered and that a minimum of 1.0 mm mesh be utilized in recovery contexts known to include fetal remains. © 2015 American Academy of Forensic Sciences.

  13. Region-specific maturation of cerebral cortex in human fetal brain: diffusion tensor imaging and histology

    Energy Technology Data Exchange (ETDEWEB)

    Trivedi, Richa; Gupta, Rakesh K.; Saksena, Sona [Sanjay Gandhi Post Graduate Institute of Medical Sciences, Department of Radiodiagnosis, Lucknow, UP (India); Husain, Nuzhat; Srivastava, Savita [CSM Medical University, Department of Pathology, Lucknow (India); Rathore, Ram K.S.; Sarma, Manoj K. [Indian Institute of Technology, Department of Mathematics and Statistics, Kanpur (India); Malik, Gyanendra K. [CSM Medical University, Department of Pediatrics, Lucknow (India); Das, Vinita [CSM Medical University, Department of Obstetrics and Gynecology, Lucknow (India); Pradhan, Mandakini [Sanjay Gandhi Postgraduate Institute of Medical Sciences, Department of Medical Genetics, Lucknow (India); Pandey, Chandra M. [Sanjay Gandhi Postgraduate Institute of Medical Sciences, Department of Biostatistics, Lucknow (India); Narayana, Ponnada A. [University of Texas Medical School at Houston, Department of Diagnostic and Interventional Imaging, Houston, TX (United States)

    2009-09-15

    In this study, diffusion tensor imaging (DTI) and glial fibrillary acidic protein (GFAP) immunohistochemical analysis in different cortical regions in fetal brains at different gestational age (GA) were performed. DTI was performed on 50 freshly aborted fetal brains with GA ranging from 12 to 42 weeks to compare age-related fractional anisotropy (FA) changes in different cerebral cortical regions that include frontal, parietal, occipital, and temporal lobes at the level of thalami. GFAP immunostaining was performed and the percentage of GFAP-positive areas was quantified. The cortical FA values in the frontal lobe peaked at around 26 weeks of GA, occipital and temporal lobes at around 20 weeks, and parietal lobe at around 23 weeks. A significant, but modest, positive correlation (r=0.31, p=0.02) was observed between cortical FA values and percentage area of GFAP expression in cortical region around the time period during which the migrational events are at its peak, i.e., GA {<=} 28 weeks for frontal cortical region and GA{<=}22 weeks for rest of the lobes. The DTI-derived FA quantification with its GFAP immunohistologic correlation in cortical regions of the various lobes of the cerebral hemispheres supports region-specific migrational and maturational events in human fetal brain. (orig.)

  14. Human embryonic and fetal mesenchymal stem cells differentiate toward three different cardiac lineages in contrast to their adult counterparts.

    Science.gov (United States)

    Ramkisoensing, Arti A; Pijnappels, Daniël A; Askar, Saïd F A; Passier, Robert; Swildens, Jim; Goumans, Marie José; Schutte, Cindy I; de Vries, Antoine A F; Scherjon, Sicco; Mummery, Christine L; Schalij, Martin J; Atsma, Douwe E

    2011-01-01

    Mesenchymal stem cells (MSCs) show unexplained differences in differentiation potential. In this study, differentiation of human (h) MSCs derived from embryonic, fetal and adult sources toward cardiomyocytes, endothelial and smooth muscle cells was investigated. Labeled hMSCs derived from embryonic stem cells (hESC-MSCs), fetal umbilical cord, bone marrow, amniotic membrane and adult bone marrow and adipose tissue were co-cultured with neonatal rat cardiomyocytes (nrCMCs) or cardiac fibroblasts (nrCFBs) for 10 days, and also cultured under angiogenic conditions. Cardiomyogenesis was assessed by human-specific immunocytological analysis, whole-cell current-clamp recordings, human-specific qRT-PCR and optical mapping. After co-culture with nrCMCs, significantly more hESC-MSCs than fetal hMSCs stained positive for α-actinin, whereas adult hMSCs stained negative. Furthermore, functional cardiomyogenic differentiation, based on action potential recordings, was shown to occur, but not in adult hMSCs. Of all sources, hESC-MSCs expressed most cardiac-specific genes. hESC-MSCs and fetal hMSCs contained significantly higher basal levels of connexin43 than adult hMSCs and co-culture with nrCMCs increased expression. After co-culture with nrCFBs, hESC-MSCs and fetal hMSCs did not express α-actinin and connexin43 expression was decreased. Conduction velocity (CV) in co-cultures of nrCMCs and hESC-MSCs was significantly higher than in co-cultures with fetal or adult hMSCs. In angiogenesis bioassays, only hESC-MSCs and fetal hMSCs were able to form capillary-like structures, which stained for smooth muscle and endothelial cell markers.Human embryonic and fetal MSCs differentiate toward three different cardiac lineages, in contrast to adult MSCs. Cardiomyogenesis is determined by stimuli from the cellular microenvironment, where connexin43 may play an important role.

  15. Growth trajectories of the human fetal brain tissues estimated from 3D reconstructed in utero MRI.

    Science.gov (United States)

    Scott, Julia A; Habas, Piotr A; Kim, Kio; Rajagopalan, Vidya; Hamzelou, Kia S; Corbett-Detig, James M; Barkovich, A James; Glenn, Orit A; Studholme, Colin

    2011-08-01

    In the latter half of gestation (20-40 gestational weeks), human brain growth accelerates in conjunction with cortical folding and the deceleration of ventricular zone progenitor cell proliferation. These processes are reflected in changes in the volume of respective fetal tissue zones. Thus far, growth trajectories of the fetal tissue zones have been extracted primarily from 2D measurements on histological sections and magnetic resonance imaging (MRI). In this study, the volumes of major fetal zones-cortical plate (CP), subplate and intermediate zone (SP+IZ), germinal matrix (GMAT), deep gray nuclei (DG), and ventricles (VENT)--are calculated from automatic segmentation of motion-corrected, 3D reconstructed MRI. We analyzed 48 T2-weighted MRI scans from 39 normally developing fetuses in utero between 20.57 and 31.14 gestational weeks (GW). The supratentorial volume (STV) increased linearly at a rate of 15.22% per week. The SP+IZ (14.75% per week) and DG (15.56% per week) volumes increased at similar rates. The CP increased at a greater relative rate (18.00% per week), while the VENT (9.18% per week) changed more slowly. Therefore, CP increased as a fraction of STV and the VENT fraction declined. The total GMAT volume slightly increased then decreased after 25 GW. We did not detect volumetric sexual dimorphisms or total hemispheric volume asymmetries, which may emerge later in gestation. Further application of the automated fetal brain segmentation to later gestational ages will bridge the gap between volumetric studies of premature brain development and normal brain development in utero. Published by Elsevier Ltd.

  16. Structural development of human brain white matter from mid-fetal to perinatal stage

    Science.gov (United States)

    Ouyang, Austin; Yu, Qiaowen; Mishra, Virendra; Chalak, Lina; Jeon, Tina; Sivarajan, Muraleedharan; Jackson, Greg; Rollins, Nancy; Liu, Shuwei; Huang, Hao

    2015-03-01

    The structures of developing human brain white matter (WM) tracts can be effectively quantified by DTI-derived metrics, including fractional anisotropy (FA), mean, axial and radial diffusivity (MD, AD and RD). However, dynamics of WM microstructure during very early developmental period from mid-fetal to perinatal stage is unknown. It is difficult to accurately measure microstructural properties of these WM tracts due to severe contamination from cerebrospinal fluid (CSF). In this study, high resolution DTI of fetal brains at mid-fetal stage (20 weeks of gestation or 20wg), 19 brains in the middle of 3rd trimester (35wg) and 17 brains around term (40wg) were acquired. We established first population-averaged DTI templates at these three time points and extracted WM skeleton. 16 major WM tracts in limbic, projection, commissural and association tract groups were traced with DTI tractography in native space. The WM skeleton in the template space was inversely transformed back to the native space for measuring core WM microstructures of each individual tract. Continuous microstructural enhancement and volumetric increase of WM tracts were found from 20wg to 40wg. The microstructural enhancement from FA measurement is decelerated in late 3rd trimester compared to mid-fetal to middle 3rd trimester, while volumetric increase of prefrontal WM tracts is accelerated. The microstructural enhancement from 35wg to 40wg is heterogeneous among different tract groups with microstructures of association tracts undergoing most dramatic change. Besides decreases of RD indicating active myelination, the decrease of AD for most WM tracts during late 3rd trimester suggests axonal packing process.

  17. High-resolution imaging diagnosis of human fetal membrane by three-dimensional optical coherence tomography

    Science.gov (United States)

    Ren, Hugang; Avila, Cecilia; Kaplan, Cynthia; Pan, Yingtian

    2011-11-01

    Microscopic chorionic pseudocyst (MCP) arising in the chorion leave of the human fetal membrane (FM) is a clinical precursor for preeclampsia which may progress to fatal medical conditions (e.g., abortion) if left untreated. To examine the utility of three-dimensional (3D) optical coherence tomography (OCT) for noninvasive delineation of the morphology of human fetal membranes and early clinical detection of MCP, 60 human FM specimens were acquired from 10 different subjects undergoing term cesarean delivery for an ex vivo feasibility study. Our results showed that OCT was able to identify the four-layer architectures of human FMs consisting of high-scattering decidua vera (DV, average thickness dDV ~ 92+/-38 μm), low-scattering chorion and trophoblast (CT, dCT ~ 150+/-67 μm), high-scattering subepithelial amnion (A, dA ~ 95+/-36 μm), and low-scattering epithelium (E, dE ~ 29+/-8 μm). Importantly, 3D OCT was able to instantaneously detect MCPs (low scattering due to edema, fluid buildup, vasodilatation) and track (staging) their thicknesses dMCP ranging from 24 to 615 μm. It was also shown that high-frequency ultrasound was able to compliment OCT for detecting more advanced thicker MCPs (e.g., dMCP>615 μm) because of its increased imaging depth.

  18. Lactobacillus rhamnosus GG and its SpaC pilus adhesin modulate inflammatory responsiveness and TLR-related gene expression in the fetal human gut

    Science.gov (United States)

    Ganguli, Kriston; Collado, Maria Carmen; Rautava, Jaana; Lu, Lei; Satokari, Reetta; von Ossowski, Ingemar; Reunanen, Justus; de Vos, Willem M.; Palva, Airi; Isolauri, Erika; Salminen, Seppo; Walker, W. Allan; Rautava, Samuli

    2015-01-01

    Background Bacterial contact in utero modulates fetal and neonatal immune responses. Maternal probiotic supplementation reduces the risk of immune-mediated disease in the infant. We investigated the immunomodulatory properties of live Lactobacillus rhamnosus GG and its SpaC pilus adhesin in human fetal intestinal models. Methods TNF-α mRNA expression was measured by qPCR in a human fetal intestinal organ culture model exposed to live L. rhamnosus GG and proinflammatory stimuli. Binding of recombinant SpaC pilus protein to intestinal epithelial cells was assessed in human fetal intestinal organ culture and the human fetal intestinal epithelial cell line H4 by immunohistochemistry and immunofluorescence, respectively. TLR-related gene expression in fetal ileal organ culture after exposure to recombinant SpaC was assessed by qPCR. Results Live L. rhamnosus GG significantly attenuates pathogen-induced TNF-α mRNA expression in the human fetal gut. Recombinant SpaC protein was found to adhere to the fetal gut and to modulate varying levels of TLR-related gene expression. Conclusion The human fetal gut is responsive to luminal microbes. L. rhamnosus GG significantly attenuates fetal intestinal inflammatory responses to pathogenic bacteria. The L. rhamnosus GG pilus adhesin SpaC binds to immature human intestinal epithelial cells and directly modulates intestinal epithelial cell innate immune gene expression. PMID:25580735

  19. Immunohistochemical localization of cyclooxygenase-1 and cyclooxygenase-2 in the human fetal and adult male reproductive tracts.

    Science.gov (United States)

    Kirschenbaum, A; Liotta, D R; Yao, S; Liu, X H; Klausner, A P; Unger, P; Shapiro, E; Leav, I; Levine, A C

    2000-09-01

    The first rate-limiting step in the conversion of arachidonic acid to PGs is catalyzed by cyclooxygenase (Cox). Two isoforms of Cox have been identified, Cox-1 (constitutively expressed) and Cox-2 (inducible form), which are the products of two different genes. In this study we describe the immunohistochemical localization of Cox-1 and -2 in the human male fetal and adult reproductive tracts. There was no Cox-1 expression in fetal samples (prostate, seminal vesicles, or ejaculatory ducts), and only minimal expression in adult tissues. There was no expression of Cox-2 in the fetal prostate. In a prepubertal prostate there was some Cox-2 expression that localized exclusively to the smooth muscle cells of the transition zone. In adult hyperplastic prostates, Cox-2 was strongly expressed in smooth muscle cells, with no expression in the luminal epithelial cells. Cox-2 was strongly expressed in epithelial cells of both fetal and adult seminal vesicles and ejaculatory ducts. The Cox-2 staining intensity in the fetal ejaculatory ducts during various times of gestation correlated with previously reported testosterone production rates by the fetal testis. These data indicate that Cox-2 is the predominant isoform expressed in the fetal male reproductive tract, and its expression may be regulated by androgens. The distinct cell type-specific expression patterns of Cox-2 in the prostate (smooth muscle) vs. the seminal vesicles and ejaculatory ducts (epithelium) may reflect the different roles of PGs in these tissues.

  20. Neurotensin is a Versatile Modulator of In Vitro Human Pancreatic Ductal Adenocarcinoma Cell (PDAC Migration

    Directory of Open Access Journals (Sweden)

    Tatjana Mijatovic

    2007-01-01

    Full Text Available Background: While the neurotensin (NT roles in pancreatic cancer growth are well documented, its effects on pancreatic cancer cell migration have not been described. Methods: The NT-induced effects on the migration process of human pancreatic ductal adenocarcinoma cells (PDACs were characterized by means of various assays including computer-assisted video-microscopy, fluorescence microscopy, ELISA-based, small GTPase pull-down and phosphorylation assays. Results: The NT-induced modifications on in vitro PDACs migration largely depended on the extra-cellular matrix environment and cell propensity to migrate collectively or individually. While NT significantly reduced the level of migration of collectively migrating PDACs on vitronectin, it significantly increased the level of individually migrating PDACs. These effects were mainly mediated through the sortilin/NTR3 receptor. Neurotensin both induced altered expression of αV and β5 integrin subunits in PDACs cultured on vitronectin resulting in modified adhesion abilities, and caused modifications to the organization of the actin cytoskeleton through the NT-mediated activation of small Rho GTPases. While the NT effects on individually migrating PDACs were mediated at least through the EGFR/ERK signaling pathways, those on collectively migrating PDACs appeared highly dependent on the PI 3-kinase pathway. Conclusion: This study strongly suggests the involvement of neurotensin in the modulation of human PDAC migration.

  1. Numb/Notch signaling pathway modulation enhances human pancreatic cancer cell radiosensitivity.

    Science.gov (United States)

    Bi, Yi-Liang; Min, Min; Shen, Wei; Liu, Yan

    2016-11-01

    The present study aims to evaluate whether repression of the Numb/Notch signaling pathway affects the radiosensitivity of human pancreatic cancer cell lines. Different doses of X-rays (0, 2, 3, 4, and 5 Gy) were applied to the PANC-1, SW1990, and MIA PaCa-2 human pancreatic cancer cell lines, and the Numb/Notch pathway inhibitor DAPT was added at different doses (0, 1, 3, and 5 μmol/l). MTT assay, colony formation assay, flow cytometry, scratch assay, and Transwell experiments were performed, and qRT-PCR and Western blot were conducted for the detection of Numb expression. Tumorigenicity assay in nude mice was carried out to verify the influence of blocker of the Numb/Notch signaling pathway on the radiosensitivity of xenograft tumors. The MTT assay, colony formation assay and flow cytometry experiments revealed that proliferation decreased as radiation dose increased. The viability of PANC-1 cells at 5 Gy, SW 1990 cells at 4 Gy and 5 Gy, and MIA PaCa-2 cells at 2-5 Gy was significantly lower than that of non-irradiated cells (all P cancer cells is associated with X-ray radiation and that inhibition of the Numb/Notch signaling pathway can enhance radiosensitivity, suggesting that inhibition of the Numb/Notch signaling pathway may serve as a potential target for clinical improvement of the radiosensitivity of pancreatic cancer.

  2. Developmental exposure to estrogen alters differentiation and epigenetic programming in a human fetal prostate xenograft model.

    Directory of Open Access Journals (Sweden)

    Camelia M Saffarini

    Full Text Available Prostate cancer is the most frequent non-cutaneous malignancy in men. There is strong evidence in rodents that neonatal estrogen exposure plays a role in the development of this disease. However, there is little information regarding the effects of estrogen in human fetal prostate tissue. This study explored early life estrogen exposure, with and without a secondary estrogen and testosterone treatment in a human fetal prostate xenograft model. Histopathological lesions, proliferation, and serum hormone levels were evaluated at 7, 30, 90, and 200-day time-points after xenografting. The expression of 40 key genes involved in prostatic glandular and stromal growth, cell-cycle progression, apoptosis, hormone receptors and tumor suppressors was evaluated using a custom PCR array. Epigenome-wide analysis of DNA methylation was performed on whole tissue, and laser capture-microdissection (LCM isolated epithelial and stromal compartments of 200-day prostate xenografts. Combined initial plus secondary estrogenic exposures had the most severe tissue changes as revealed by the presence of hyperplastic glands at day 200. Gene expression changes corresponded with the cellular events in the KEGG prostate cancer pathway, indicating that initial plus secondary exposure to estrogen altered the PI3K-Akt signaling pathway, ultimately resulting in apoptosis inhibition and an increase in cell cycle progression. DNA methylation revealed that differentially methylated CpG sites significantly predominate in the stromal compartment as a result of estrogen-treatment, thereby providing new targets for future investigation. By using human fetal prostate tissue and eliminating the need for species extrapolation, this study provides novel insights into the gene expression and epigenetic effects related to prostate carcinogenesis following early life estrogen exposure.

  3. Embelin suppresses growth of human pancreatic cancer xenografts, and pancreatic cancer cells isolated from KrasG12D mice by inhibiting Akt and Sonic hedgehog pathways.

    Directory of Open Access Journals (Sweden)

    Minzhao Huang

    Full Text Available Pancreatic cancer is a deadly disease, and therefore effective treatment and/or prevention strategies are urgently needed. The objectives of this study were to examine the molecular mechanisms by which embelin inhibited human pancreatic cancer cell growth in vitro, and xenografts in Balb C nude mice, and pancreatic cancer cell growth isolated from KrasG12D transgenic mice. XTT assays were performed to measure cell viability. AsPC-1 cells were injected subcutaneously into Balb c nude mice and treated with embelin. Cell proliferation and apoptosis were measured by Ki67 and TUNEL staining, respectively. The expression of Akt, and Sonic Hedgehog (Shh and their target gene products were measured by the immunohistochemistry, and Western blot analysis. The effects of embelin on pancreatic cancer cells isolated from 10-months old KrasG12D mice were also examined. Embelin inhibited cell viability in pancreatic cancer AsPC-1, PANC-1, MIA PaCa-2 and Hs 766T cell lines, and these inhibitory effects were blocked either by constitutively active Akt or Shh protein. Embelin-treated mice showed significant inhibition in tumor growth which was associated with reduced expression of markers of cell proliferation (Ki67, PCNA and Bcl-2 and cell cycle (cyclin D1, CDK2, and CDK6, and induction of apoptosis (activation of caspase-3 and cleavage of PARP, and increased expression of Bax. In addition, embelin inhibited the expression of markers of angiogenesis (COX-2, VEGF, VEGFR, and IL-8, and metastasis (MMP-2 and MMP-9 in tumor tissues. Antitumor activity of embelin was associated with inhibition of Akt and Shh pathways in xenografts, and pancreatic cancer cells isolated from KrasG12D mice. Furthermore, embelin also inhibited epithelial-to-mesenchymal transition (EMT by up-regulating E-cadherin and inhibiting the expression of Snail, Slug, and ZEB1. These data suggest that embelin can inhibit pancreatic cancer growth, angiogenesis and metastasis by suppressing Akt and

  4. Serum human placental lactogen levels in intra-uterine fetal growth retardation.

    Science.gov (United States)

    Zail, S S; Safro, I L

    1975-11-12

    Serum human placental lactogen (HPL) levels were measured in the last trimester of pregnancy in 16 mothers who delivered small-for-gestational-age babies. Only 3 patients had levels which were below the normal range, while 4 others had levels close to the lower limit of the normal range. The finding of a normal serum HPL level therefore does not exclude the possibility of intra-uterine fetal growth retardation. No correlation was found between serum HPL levels at 37-39 weeks and infant or placental weights in full-term normal deliveries.

  5. Hydrolysis of human milk fat globules by pancreatic lipase: role of colipase, phospholipase A2, and bile salts.

    OpenAIRE

    Bläckberg, L; Hernell, O; Olivecrona, T

    1981-01-01

    Human milk fat globules were used to explore how dietary triglycerides are hydrolyzed by pancreatic lipase. These triglycerides were hydrolyzed very slowly by lipase alone as if the surface layer of proteins and phospholipids impeded the action of the enzyme. The inhibition of lipase activity could be overcome by addition either of colipase or of pancreatic phospholipase A2. Colipase enhanced triglyceride hydrolysis in a dose-dependent manner whether bile salts were present or not. Bile salts...

  6. Knowledge Gaps in Rodent Pancreas Biology: Taking Human Pluripotent Stem Cell-Derived Pancreatic Beta Cells into Our Own Hands.

    Science.gov (United States)

    Santosa, Munirah Mohamad; Low, Blaise Su Jun; Pek, Nicole Min Qian; Teo, Adrian Kee Keong

    2015-01-01

    In the field of stem cell biology and diabetes, we and others seek to derive mature and functional human pancreatic β cells for disease modeling and cell replacement therapy. Traditionally, knowledge gathered from rodents is extended to human pancreas developmental biology research involving human pluripotent stem cells (hPSCs). While much has been learnt from rodent pancreas biology in the early steps toward Pdx1(+) pancreatic progenitors, much less is known about the transition toward Ngn3(+) pancreatic endocrine progenitors. Essentially, the later steps of pancreatic β cell development and maturation remain elusive to date. As a result, the most recent advances in the stem cell and diabetes field have relied upon combinatorial testing of numerous growth factors and chemical compounds in an arbitrary trial-and-error fashion to derive mature and functional human pancreatic β cells from hPSCs. Although this hit-or-miss approach appears to have made some headway in maturing human pancreatic β cells in vitro, its underlying biology is vaguely understood. Therefore, in this mini-review, we discuss some of these late-stage signaling pathways that are involved in human pancreatic β cell differentiation and highlight our current understanding of their relevance in rodent pancreas biology. Our efforts here unravel several novel signaling pathways that can be further studied to shed light on unexplored aspects of rodent pancreas biology. New investigations into these signaling pathways are expected to advance our knowledge in human pancreas developmental biology and to aid in the translation of stem cell biology in the context of diabetes treatments.

  7. Regulation Effect of Vascular Endothelial Growth Factor on Human Fetal Choroid Vascularization

    Institute of Scientific and Technical Information of China (English)

    JinsongZhao; YiWang; 等

    2002-01-01

    Purpose:To investigate the spatial and temporal regulation effect of vascular endothelial growth factor(VEGF) on human fetal choroids vascularization.Methods:The eyeballs of 54 human fetuses from the 9th week to the 40th week due to accidental abortion were studied by immunohistochemically stainin for the expression of VEGF and proliferation cell nuclear antigen (PCNA).Results: (1)The distribution of VEGF expression in the retinal pigment epithelium (RPE) decreased with the incrase of age,the peak of which was between the 9th and 14th week.(2)PCNA immunoreactivity was localized within choriocapillaris endothelium .The expression level decreased alone with fetus age.In this period the choriocapillaris endothelium kept proliferation,differentiation,canalization and remodeled to form the choroids vessels(3)Statistically significant correlations were shown between the expression of VEGF in the PRE and that of PCNA in choriocapillaris endothelium(r=0.933,P<0.01).Couclusin:VEGF expression in PRE was positively involved in modulating human fetal choroids vascularization .Eye Science 2000;16:11-14.

  8. Regulation Effect of Vascular Endothelial Growth Factor on Human Fetal Choroid Vascularization

    Institute of Scientific and Technical Information of China (English)

    Jinsong Zhao; Yue Song; Yi Wang; Xiaoguang Zhang

    2000-01-01

    Purpose: To investigate the spatial and temporal regulation effect of vascular endothelial growth factor(VEGF) on human fetal choroid vascularization. Methods: The eyeballs of 54 human fetuses from the 9th week to the 40th week due to accidental abortion were studied by immunohistochemically staining for the expression of VEGF and proliferation cell nuclear antigen (PCNA). Results: (1) The distribution of VEGF expression in the retinal pigment epithelium (RPE) decreased with the increase of age, the peak of which was between the 9th and 14th week. (2) PCNA immunoreactivity was localized within choriocapillaris endothelium. The expression level decreased alone with fetus age. In this period the choriocapillaris endothelium kept proliferation, differentiation, canalization and remodelled to form the choroid vessels. (3)Statistically significant correlations were shown between the expression of VEGF in the PRE and that of PCNA in choriocapillaris endothelium(r =0. 933, P < 0. 01). Conclusion: VEGF expression in RPE was positively involved in modulating human fetal choroid vascularization. Eye Science 2000; 16:11 ~ 14.

  9. Long-acting lipidated analogue of human pancreatic polypeptide is slowly released into circulation

    DEFF Research Database (Denmark)

    Bellmann-Sickert, Kathrin; Elling, Christian E; Madsen, Andreas N;

    2011-01-01

    of a human pancreatic polypeptide analogue specific for the human (h)Y(2) and hY(4) receptor with PEGs of different size and palmitic acid. Receptor specificity was demonstrated by competitive binding studies. Modifications had only a small influence on binding affinities and no influence on secondary......The main disadvantages of peptide pharmaceuticals are their rapid degradation and excretion, their low hydrophilicity, and low shelf lifes. These bottlenecks can be circumvented by acylation with fatty acids (lipidation) or polyethylene glycol (PEGylation). Here, we describe the modification...

  10. Foodborne outbreak of human brucellosis caused by ingested raw materials of fetal calf on Jeju Island.

    Science.gov (United States)

    Yoo, Jeong Rae; Heo, Sang Taek; Lee, Keun Hwa; Kim, Young Ree; Yoo, Seung Jin

    2015-02-01

    Since the first reported case of human brucellosis in 2002 in South Korea, its incidence has been increasing nationally. However, bovine brucellosis has not been present from 2005 to date on Jeju Island. Despite Jeju Island being considered a clean area for bovine brucellosis, we experienced an outbreak of human brucellosis between 2012 and 2013. Herein, we report cases with human brucellosis after ingestion of raw materials of fetal calf at a restaurant. Patients were identified by isolation of the Brucella abortus in their blood and joint tissue. Because all patients developed zoonosis by a faulty folk remedy, we emphasize the importance of educational programs to increase the awareness of zoonosis, and the need for active surveillance and detection of illegal distribution channels of the infected animal. After the outbreak, we took control of the involved restaurant and its illegal distribution channel, and there have been no further outbreaks.

  11. Fetal Mesenchymal Stromal Cells Differentiating towards Chondrocytes Acquire a Gene Expression Profile Resembling Human Growth Plate Cartilage

    NARCIS (Netherlands)

    van Gool, S.A.; Emons, J.A.M.; Leijten, Jeroen Christianus Hermanus; Decker, E.; Sticht, C.; van Houwelingen, J.C.; Goeman, J.J.; Kleijburg, C.; Scherjon, S.; Gretz, N.; Wit, J.M.; Rappold, G.; Post, Janine Nicole; Karperien, Hermanus Bernardus Johannes

    2012-01-01

    Abstract We used human fetal bone marrow-derived mesenchymal stromal cells (hfMSCs) differentiating towards chondrocytes as an alternative model for the human growth plate (GP). Our aims were to study gene expression patterns associated with chondrogenic differentiation to assess whether

  12. Correlation between Protein Expression of PTEN in Human Pancreatic Cancer and the Proliferation, Infiltration, Metastasis and Prognosis

    Institute of Scientific and Technical Information of China (English)

    TAO Jing; XIONG Jiongxin; LI Tao; YANG Zhiyong; LI Xiaohui; LI Kai; WU Heshui; WANG Chunyou

    2006-01-01

    In order to investigate the correlation between protein expression of PTEN and the proliferation, infiltration, metastasis and prognosis in pancreatic cancer, immunohistochemical SP method was used to examine the protein expression of PTEN, PCNA, MVD, MMP-2, MMP-9 and TUNEL method to detect the levels of apoptosis of pancreatic cells in 41 pancreatic head cancers from regional pancreatectomy (RP) and 10 normal pancreatic tissues. The results showed that among 41 cases of pancreatic cancers, the positive staining of PTNE (39.02 %) was significantly weaker than that in normal pancreatic tissues (P<0.05). The levels of PCNA labeling index (LI), apoptotic index(AI), microvessel density (MVD), MMP-2 LI and MMP-9 LI were decreased gradually with the increase of the expression intensity of PTEN, and there was a significant difference in the above parameters among the patients having different expression levels of PTEN (P<0.01 or P<0.05). There was a negative correlation between the expression of PTEN and PCNA LI, MVD, MMP-2 LI,MMP-9 LI, and a positive correlation between AI and the expression of PTEN. The expression intensity of PTEN was correlated with the postoperative survival of the patients with pancreatic cancer(x2=22.3400, P<0.0001, RR=2.030). It was suggested that the expression levels of PTEN protein were closely related with proliferation, infiltration and metastasis in human pancreatic cancer, and the expression of PTEN protein was one of the prognostic factors for pancreatic cancer following RP.

  13. Differentiation of human embryonic stem cells into pancreatic endoderm in patterned size-controlled clusters.

    Science.gov (United States)

    Van Hoof, Dennis; Mendelsohn, Adam D; Seerke, Rina; Desai, Tejal A; German, Michael S

    2011-05-01

    Pancreatic β-cells function optimally when clustered in islet-like structures. However, nutrient and oxygen deprivation limits the viability of cells at the core of excessively large clusters. Hence, production of functional β-cells from human embryonic stem cells (hESCs) for patients with diabetes would benefit from the growth and differentiation of these cells in size-controlled aggregates. In this study, we controlled cluster size by seeding hESCs onto glass cover slips patterned by the covalent microcontact-printing of laminin in circular patches of 120 μm in diameter. These were used as substrates to grow and differentiate hESCs first into SOX17-positive/SOX7-negative definitive endoderm, after which many clusters released and formed uniformly sized three-dimensional clusters. Both released clusters and those that remained attached differentiated into HNF1β-positive primitive gut tube-like cells with high efficiency. Further differentiation yielded pancreatic endoderm-like cells that co-expressed PDX1 and NKX6.1. Controlling aggregate size allows efficient production of uniformly-clustered pancreatic endocrine precursors for in vivo engraftment or further in vitro maturation.

  14. Depleting MEKK1 expression inhibits the ability of invasion and migration of human pancreatic cancer cells.

    Science.gov (United States)

    Su, Fuqin; Li, Hongyan; Yan, Chaoqi; Jia, Baoqing; Zhang, Yi; Chen, Xiaoguang

    2009-12-01

    Mitogen-activated protein/ERK kinase 1 (MEKK1) is a Ser/Thr protein kinase belonging to the MEKK/STE11 subgroup of the MAPKKK family and plays a key role in tumor metastasis. However, it remains unclear about its functions in pancreatic cancer. We analyzed MEKK1 expression in 41 surgically resection pancreatic cancer patient's samples by immunohistochemistry and determined its role in BxPC3 cells via RNAi experiment. The abilities of invasion, motility, and adhesion of BxPC3 cells were detected by transwell assay, wound healing assay and adhesion assay, respectively. Gelatinase activity of MMPs in cultured cells was examined by gelatin zymography. Our data showed that MEKK1 expression is positively correlated with lymphatic metastases (P BxPC3 cells were inhibited significantly (P < 0.01) when MEKK1 was depleted with a specific siRNA. We observed that the activity of MMP2 was decreased in the MEKK1 depletion cell line (P < 0.05), accompanied with decreased phosphorylated ERK1/2. Our results indicated that the depletion of MEKK1 led to a potent inhibition on the invasion and migration of human pancreatic adenocarcinoma in vitro. It suggests that MEKK1 may be a potential target for development of anti-invasion and metastasis drugs.

  15. Three-dimensional perfusion bioreactor culture supports differentiation of human fetal liver cells.

    Science.gov (United States)

    Schmelzer, Eva; Triolo, Fabio; Turner, Morris E; Thompson, Robert L; Zeilinger, Katrin; Reid, Lola M; Gridelli, Bruno; Gerlach, Jörg C

    2010-06-01

    The ability of human fetal liver cells to survive, expand, and form functional tissue in vitro is of high interest for the development of bioartificial extracorporeal liver support systems, liver cell transplantation therapies, and pharmacologic models. Conventional static two-dimensional culture models seem to be inadequate tools. We focus on dynamic three-dimensional perfusion technologies and developed a scaled-down bioreactor, providing decentralized mass exchange with integral oxygenation. Human fetal liver cells were embedded in a hyaluronan hydrogel within the capillary system to mimic an in vivo matrix and perfusion environment. Metabolic performance was monitored daily, including glucose consumption, lactate dehydrogenase activity, and secretion of alpha-fetoprotein and albumin. At culture termination cells were analyzed for proliferation and liver-specific lineage-dependent cytochrome P450 (CYP3A4/3A7) gene expression. Occurrence of hepatic differentiation in bioreactor cultures was demonstrated by a strong increase in CYP3A4/3A7 gene expression ratio, lower alpha-fetoprotein, and higher albumin secretion than in conventional Petri dish controls. Cells in bioreactors formed three-dimensional structures. Viability of cells was higher in bioreactors than in control cultures. In conclusion, the culture model implementing three-dimensionality, constant perfusion, and integral oxygenation in combination with a hyaluronan hydrogel provides superior conditions for liver cell survival and differentiation compared to conventional culture.

  16. Regeneration of insulin-producing pancreatic cells using a volatile bioactive compound and human teeth.

    Science.gov (United States)

    Okada, Mio; Imai, Toshio; Yaegaki, Ken; Ishkitiev, Nikolay; Tanaka, Tomoko

    2014-10-30

    Transplantation of insulin (INS)-secreting cells differentiated in vitro from stem cells promises a safer and easier treatment of severe diabetes mellitus. A volatile bioactive compound, hydrogen sulfide (H2S), promotes cell differentiation; human tooth-pulp stem cells undergo hepatic differentiation. The aim of this study is to develop a novel protocol using H2S to enhance pancreatic differentiation from the CD117(+) cell fraction of human tooth pulp. During the differentiation, the cells were exposed to 0.1 ng ml(-1) H2S. Immunocytochemistry, RT-PCR, determination of INS c-peptide content and flow cytometry of pancreatically related markers were carried out. Expression of WNT and the PI3K/AKT signaling pathway were also determined by PCR array. After differentiation, INS, glucagon (GCG), somatostatin (SST) and pancreatic polypeptide (PPY) were positive when examined by immunofluorescence. INS and GCG were also determined flow-cytometrically. Only the cells expressing INS increased after H2S exposure. The number of cells expressing GCG was significantly decreased. Genes involved in canonical WNT and the WNT/calcium pathways were highly expressed after H2S exposure. H2S accelerated INS synthesis and secretion by regenerated INS-producing cells from human teeth. All signaling pathway functions of the PI3K-AKT pathway were extremely activated by H2S exposure. The matured INS-producing cells originating in human teeth were increased by H2S in order to control blood-glucose level.

  17. Interdependence of Gemcitabine Treatment, Transporter Expression, and Resistance in Human Pancreatic Carcinoma Cells

    Directory of Open Access Journals (Sweden)

    Wolfgang Hagmann

    2010-09-01

    Full Text Available Gemcitabine is widely used as first-line chemotherapeutic drug in the treatment of pancreatic cancer. Our previous experimental chemotherapy studies have shown that treatment of human pancreatic carcinoma cells with 5-fluorouracil (5-FU alters the cellular transporter expression profile and that modulation of the expression of multidrug resistance protein 5 (MRP5; ABCC5 influences the chemoresistance of these tumor cells. Here, we studied the influence of acute and chronic gemcitabine treatment on the expression of relevant uptake and export transporters in pancreatic carcinoma cells by reverse transcription-polymerase chain reaction (RT-PCR, quantitative RT-PCR, and immunoblot analyses. The specific role of MRP5 in cellular gemcitabine sensitivity was studied by cytotoxicity assays using MRP5-overexpressing and MRP5-silenced cells. Exposure to gemcitabine (12 nM for 3 days did not alter the messenger RNA (mRNA expression of MRP1, MRP3, MRP5, and equilibrative nucleoside transporter 1 (ENT1, whereas high dosages of the drug (20 µM for 1 hour elicited up-regulation of these transporters in most cell lines studied. In cells with acquired gemcitabine resistance (up to 160 nM gemcitabine, the mRNA or protein expression of the gemcitabine transporters MRP5 and ENT1 was upregulated in several cell lines. Combined treatment with 5-FU and gemcitabine caused a 5- to 40-fold increase in MRP5 and ENT1 expressions. Cytotoxicity assays using either MRP5-overexpressing (HEK and PANC-1 or MRP5-silenced (PANC1/shMRP5 cells indicated that MRP5 contributes to gemcitabine resistance. Thus, our novel data not only on drug-induced alterations of transporter expression relevant for gemcitabine uptake and export but also on the link between gemcitabine sensitivity and MRP5 expression may lead to improved strategies of future chemotherapy regimens using gemcitabine in pancreatic carcinoma patients.

  18. 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.

  19. HETEROGENEITY IN THE GLYCOSYLATION PATTERN OF HUMAN PANCREATIC RIBONUCLEASE

    NARCIS (Netherlands)

    RIBO, M; BEINTEMA, JJ; OSSET, M; BRAVO, J; DELLORENS, R; CUCHILLO, CM

    1994-01-01

    Different molecular forms of ribonuclease were isolated from fresh human pancreas obtained from healthy transplant donors. The purification procedure consists of the preparation of an acetone powder, followed by (NH4)(2)SO4 precipitation and two chromatography steps (cationic exchange and reversed-p

  20. The expression of thyroid hormone transporters in the human fetal cerebral cortex during early development and in N-Tera-2 neurodifferentiation

    OpenAIRE

    Chan, S.Y.; Martín-Santos, A; Loubière, L S; González, A. M.; Stieger, B.; LOGAN, A.; McCabe, C. J.; Franklyn, J.A.; Kilby, M. D.

    2011-01-01

    Associations of neurological impairment with mutations in the thyroid hormone (TH) transporter, MCT8, and with maternal hypothyroxinaemia, suggest that THs are crucial for human fetal brain development. It has been postulated that TH transporters regulate the cellular supply of THs within the fetal brain during development. This study describes the expression of TH transporters in the human fetal cerebral cortex (7–20 weeks gestation) and during retinoic acid induced neurodifferentiation of t...

  1. Hereditary Pancreatitis

    Science.gov (United States)

    ... alcohol is a known risk factor for both acute and chronic pancreatitis. Therefore it is recommended that all HP patients ... Pancreatitis Patient Info Animated Pancreas Patient Pancreatic Cancer Chronic Pancreatitis Acute Pancreatitis Research Research Grant Application Research History Grant ...

  2. Pancreatitis - discharge

    Science.gov (United States)

    Chronic pancreatitis - discharge; Pancreatitis - chronic - discharge; Pancreatic insufficiency - discharge; Acute pancreatitis - discharge ... You were in the hospital because you have pancreatitis. This is a swelling of the pancreas. You ...

  3. Tumor-associated MUC5AC stimulates in vivo tumorigenicity of human pancreatic cancer.

    Science.gov (United States)

    Hoshi, Hirotaka; Sawada, Tetsuji; Uchida, Motoyuki; Saito, Hikaru; Iijima, Hiroko; Toda-Agetsuma, Mikako; Wada, Tsutomu; Yamazoe, Sadaaki; Tanaka, Hiroaki; Kimura, Kenjiro; Kakehashi, Anna; Wei, Min; Hirakawa, Kosei; Wanibuchi, Hideki

    2011-03-01

    MUC5AC, a high molecular weight glycoprotein, is overexpressed in the ductal region of human pancreatic cancer but is not detectable in the normal pancreas, suggesting its association with disease development. In the present study, we investigated the in vitro and in vivo effects of MUC5AC knockdown by short interfering RNA (siRNA) in the MUC5AC-overexpressing SW1990 and BxPC3 human pancreatic cancer cell lines in order to clarify its function. Significant decreases in the expression levels of MUC5AC mRNA and protein were observed in SW1990 and BxPC3 cells that had been stably transfected with a MUC5AC siRNA expression vector (SW1990/si-MUC5AC and BxPC3/si-MUC5AC cells) compared to those in cells transfected with an si-mock vector (SW1990/si-mock and BxPC3/si-mock cells). In in vitro studies, neither type of MUC5AC-knockdown cell showed any difference in cell survival, proliferation, or morphology from the si-mock cells or parental cells. However, in vivo xenograft studies demonstrated that MUC5AC knockdown significantly reduced the tumorigenicity and suppressed the tumor growth of si-MUC5AC cells compared to those of the si-mock cells. Immunohistochemical analysis revealed that CD45R/B220+ and Gr-1+ cells had infiltrated into the tumor tissue of the SW1990/si-MUC5AC cells. Furthermore, cancer-associated antigen specific antibodies were detected at high levels in the sera from the SW1990/si-MUC5AC cell-bearing mice. These results suggest that tumor-associated MUC5AC expressed on the surface of pancreatic cancer cells supports the escape of pancreatic cancer cells from immunosurveillance. The present findings highlight a new dimension of MUC5AC as a functional immunosuppressive agent and its important role in pancreatic cancer progression.

  4. Role of transcription factor CCAAT/enhancer-binding protein alpha in human fetal liver cell types in vitro.

    Science.gov (United States)

    Gerlach, Jörg C; Over, Patrick; Foka, Hubert G; Turner, Morris E; Thompson, Robert L; Gridelli, Bruno; Schmelzer, Eva

    2015-08-01

    The transcription factor CCAAT/enhancer-binding protein alpha (C/EBPα) has been shown to play an important role in liver development, cell proliferation and differentiation. It is, however, largely unknown if C/EBPα regulates cell differentiation and proliferation differently in the diverse cell types of the human liver. We investigated the role of C/EBPα in primary human fetal liver cells and liver cell subpopulations in vitro using a 3-D perfusion bioreactor as an advanced in vivo-like human organ culture model. Human fetal liver cells were investigated in vitro. C/EBPα gene expression was knocked down using siRNA or overexpressed by plasmid transfection. Cell type-specific gene expression was studied, cell populations and their proliferation were investigated, and metabolic parameters were analyzed. When C/EBPα gene expression was knocked down, we observed a significantly reduced expression of typical endothelial, hematopoietic and mesenchymal genes such as CD31, vWF, CD90, CD45 and α-smooth muscle actin in fetal cells. The intracellular expression of hepatic proteins and genes for liver-specific serum proteins α-fetoprotein and albumin were reduced, their protein secretion was increased. Fetal endothelial cell numbers were reduced and hepatoblast numbers were increased. C/EBPα overexpression in fetal cells resulted in increased endothelial numbers, but did not affect mesenchymal cell types or hepatoblasts. We demonstrated that the effects of C/EBPα are specific for the different human fetal liver cell types, using an advanced 3-D perfusion bioreactor as a human in vivo-like model. © 2014 The Japan Society of Hepatology.

  5. Human anogenital distance: an update on fetal smoke-exposure and integration of the perinatal literature on sex differences

    Science.gov (United States)

    Fowler, Paul A.; Filis, Panagiotis; Bhattacharya, Siladitya; le Bizec, Bruno; Antignac, Jean-Philippe; Morvan, Marie-Line; Drake, Amanda J.; Soffientini, Ugo; O'Shaughnessy, Peter J.

    2016-01-01

    STUDY QUESTION Do sex and maternal smoking effects on human fetal anogenital distance (AGD) persist in a larger study and how do these data integrate with the wider literature on perinatal human AGD, especially with respect to sex differences? SUMMARY ANSWER Second trimester sex differences in AGD are broadly consistent with neonatal and infant measures of AGD and maternal cigarette smoking is associated with a temporary increase in male AGD in the absence of changes in circulating testosterone. WHAT IS KNOWN ALREADY AGD is a biomarker of fetal androgen exposure, a reduced AGD in males being associated with cryptorchidism, hypospadias and reduced penile length. Normative fetal AGD data remain partial and windows of sensitivity of human fetal AGD to disruption are not known. STUDY DESIGN, SIZE, DURATION The effects of fetal sex and maternal cigarette smoking on the second trimester (11–21 weeks of gestation) human fetal AGD were studied, along with measurement of testosterone and testicular transcripts associated with apoptosis and proliferation. PARTICIPANTS/MATERIALS, SETTING METHODS AGD, measured from the centre of the anus to the posterior/caudal root of penis/clitoris (AGDapp) was determined in 56 female and 70 male morphologically normal fetuses. These data were integrated with current literature on perinatal AGD in humans. MAIN RESULTS AND THE ROLE OF CHANCE At 11–13 weeks of gestation male fetal AGDapp was 61% (P< 0.001) longer than in females, increasing to 70% at 17–21 weeks. This sexual dimorphism was independent of growth characteristics (fetal weight, length, gonad weight). We confirmed that at 14–16 weeks of gestation male fetal AGDapp was increased 28% (P < 0.05) by in utero cigarette smoke exposure. Testosterone levels were not affected by smoking. To develop normative data, our findings have been integrated with available data from in vivo ultrasound scans and neonatal studies. Inter-study variations in male/female AGD differences lead to

  6. Connexin 36 is expressed in beta and connexins 26 and 32 in acinar cells at the end of the secondary transition of mouse pancreatic development and increase during fetal and perinatal life.

    Science.gov (United States)

    Pérez-Armendariz, Elia Martha; Cruz-Miguel, Lourdes; Coronel-Cruz, Cristina; Esparza-Aguilar, Marcelino; Pinzon-Estrada, Enrique; Rancaño-Camacho, Elizabeth; Zacarias-Climaco, Gerardo; Olivares, Paola Fernández; Espinosa, Ana Maria; Becker, Ingeborg; Sáez, Juan C; Berumen, Jaime; Pérez-Palacios, Gregorio

    2012-06-01

    To identify when during fetal development connexins (Cxs) 26 (Cx26) 32 (Cx32), and 36 (Cx36) begin to be expressed, as well as to characterize their spatial distribution, real time polymerase chain reaction and immunolabeling studies were performed. Total RNA from mouse pancreases at 13 and 18 days postcoitum (dpc) and 3 days postpartum (dpp) was analyzed. In addition, pancreatic sections of mouse at 13, 14, 15, 16, 18 dpc and 3 dpp and of rat at term were double labeled with either anti-insulin or anti-α-amylase and anti-Cx26 or -Cx32 or -Cx36 antibodies and studied with confocal microscopy. From day 13 dpc, Cxs 26, 32, and 36 transcripts were identified and their levels increased with age. At 13-14 dpc, Cxs 26 and 32 were localized in few acinar cells, whereas Cx36 was distributed in small beta cell clumps. From day 14 dpc onwards, the number of labeled cells and relative immunofluorescent reactivity of all three Cxs at junctional membranes of the respective cell types increased. Cxs 26 and 32 colocalized in fetal acinar cells. In rat pancreas at term, a similar connexin distribution was found. Relative Cxs levels evaluated by immunoblotting also increased (two-fold) in pancreas homogenates from day 18 dpc to 3 dpp. The early cell specific, wide distribution, and age dependent expression of Cxs 26, 32, and 36 during fetal pancreas ontogeny suggests their possible involvement in pancreas differentiation and prenatal maturation.

  7. Heat Shock Factor-1 and Nuclear Factor-kappaB Are Systemically Activated in Human Acute Pancreatitis

    Directory of Open Access Journals (Sweden)

    Derek A O’Reilly

    2006-03-01

    Full Text Available Context :Nuclear factor-kappa B (NFkappaB is a transcription factor for a wide range of proinflammatory mediators while heat shock factor-1 (HSF-1 transcribes stress proteins that protect against cellular damage. Both are attractive therapeutic targets, undergoing investigation in other acute inflammatory conditions, such as sepsis. Objective :To evaluate the role of the transcription factors NF-kappaB and HSF-1 in human acute pancreatitis and their relationship to cytokine/chemokine production, disease severity and outcome. Patients :Twenty-four patients with acute pancreatitis and 12 healthy controls. Main outcome measures :Peripheral blood mononuclear cells were isolated. NF-kappaB and HSF-1 were measured by electrophoretic mobility shift assay. Soluble tumor necrosis factor (TNF receptor II and interleukin-8 were measured by ELISA. Acute physiology scores (APS, APACHE II scores and final Atlanta designations of severity were also determined. Results: Systemic NF-kappaB activation occurs in acute pancreatitis compared to healthy controls (P=0.004. However, there was no significant difference between those with mild and severe disease (P=0.685. Systemic activation of HSF-1 was observed in acute pancreatitis compared to healthy controls although this did not reach statistical significance (P=0.053. Activation, however, was greatest in those who had a final Atlanta designation of mild pancreatitis compared to those who had a severe attack of acute pancreatitis (P=0.036. Furthermore, HSF-1 was inversely correlated with acute physiology score (APS; r=-0.49, P=0.019 and APACHE II score (r=-0.47, P=0.026. Conclusions: Both NF-kappaB and HSF-1 are systemically activated in human acute pancreatitis. HSF-1 activation may protect against severity of pancreatitis

  8. Biotin uptake by mouse and human pancreatic beta cells/islets: a regulated, lipopolysaccharide-sensitive carrier-mediated process.

    Science.gov (United States)

    Ghosal, Abhisek; Sekar, Thillai V; Said, Hamid M

    2014-08-01

    Biotin is essential for the normal function of pancreatic beta cells. These cells obtain biotin from their surroundings via transport across their cell membrane. Little is known about the uptake mechanism involved, how it is regulated, and how it is affected by internal and external factors. We addressed these issues using the mouse-derived pancreatic beta-TC-6 cells and freshly isolated mouse and human primary pancreatic beta cells as models. The results showed biotin uptake by pancreatic beta-TC-6 cells occurs via a Na(+)-dependent, carrier-mediated process, that is sensitive to desthiobiotin, as well as to pantothenic acid and lipoate; the process is also saturable as a function of concentration (apparent Km = 22.24 ± 5.5 μM). These cells express the sodium-dependent multivitamin transporter (SMVT), whose knockdown (with doxycycline-inducible shRNA) led to a sever inhibition in biotin uptake. Similarly, uptake of biotin by mouse and human primary pancreatic islets is Na(+)-dependent and carrier-mediated, and both cell types express SMVT. Biotin uptake by pancreatic beta-TC-6 cells is also adaptively regulated (via transcriptional mechanism) by extracellular substrate level. Chronic treatment of pancreatic beta-TC-6 cells with bacterial lipopolysaccharides (LPS) leads to inhibition in biotin uptake. This inhibition is mediated via a Toll-Like receptor 4-mediated process and involves a decrease in membrane expression of SMVT. These findings show, for the first time, that pancreatic beta cells/islets take up biotin via a specific and regulated carrier-mediated process, and that the process is sensitive to the effect of LPS.

  9. Aberrant expression of zinc transporter ZIP4 (SLC39A4) significantly contributes to human pancreatic cancer pathogenesis and progression.

    Science.gov (United States)

    Li, Min; Zhang, Yuqing; Liu, Zijuan; Bharadwaj, Uddalak; Wang, Hao; Wang, Xinwen; Zhang, Sheng; Liuzzi, Juan P; Chang, Shou-Mei; Cousins, Robert J; Fisher, William E; Brunicardi, F Charles; Logsdon, Craig D; Chen, Changyi; Yao, Qizhi

    2007-11-20

    Zinc is an essential trace element and catalytic/structural component used by many metalloenzymes and transcription factors. Recent studies indicate a possible correlation of zinc levels with the cancer risk; however, the exact role of zinc and zinc transporters in cancer progression is unknown. We have observed that a zinc transporter, ZIP4 (SLC39A4), was substantially overexpressed in 16 of 17 (94%) clinical pancreatic adenocarcinoma specimens compared with the surrounding normal tissues, and ZIP4 mRNA expression was significantly higher in human pancreatic cancer cells than human pancreatic ductal epithelium (HPDE) cells. This indicates that aberrant ZIP4 up-regulation may contribute to the pancreatic cancer pathogenesis and progression. We studied the effects of ZIP4 overexpression in pancreatic cancer cell proliferation in vitro and pancreatic cancer progression in vivo. We found that forced expression of ZIP4 increased intracellular zinc levels, increased cell proliferation by 2-fold in vitro, and significantly increased tumor volume by 13-fold in the nude mice model with s.c. xenograft compared with the control cells. In the orthotopic nude mice model, overexpression of ZIP4 not only increased the primary tumor weight (7.2-fold), it also increased the peritoneal dissemination and ascites incidence. Moreover, increased cell proliferation and higher zinc content were also observed in the tumor tissues that overexpressed ZIP4. These data reveal an important outcome of aberrant ZIP4 expression in contributing to pancreatic cancer pathogenesis and progression. It may suggest a therapeutic strategy whereby ZIP4 is targeted to control pancreatic cancer growth.

  10. Pancreatic beta cells and islets take up thiamin by a regulated carrier-mediated process: studies using mice and human pancreatic preparations

    Science.gov (United States)

    Mee, Lisa; Nabokina, Svetlana M.; Sekar, V. Thillai; Subramanian, Veedamali S.; Maedler, Kathrin; Said, Hamid M.

    2009-01-01

    Thiamin is essential for the normal function of the endocrine pancreas, but very little is known about uptake mechanism(s) and regulation by beta cells. We addressed these issues using mouse-derived pancreatic beta-TC-6 cells, and freshly isolated primary mouse and human pancreatic islets. Results showed that thiamin uptake by beta-TC-6 cells involves a pH (but not Na+)-dependent carrier-mediated process that is saturable at both the nanomolar (apparent Km = 37.17 ± 9.9 nM) and micromolar (apparent Km = 3.26 ± 0.86 μM) ranges, cis-inhibited by thiamin structural analogs, and trans-stimulated by unlabeled thiamin. Involvement of carrier-mediated process was also confirmed in primary mouse and human pancreatic islets. Both THTR-1 and THTR-2 were found to be expressed in these mouse and human pancreatic preparations. Maintaining beta-TC-6 cells in the presence of a high level of thiamin led to a significant (P < 0.01) decrease in thiamin uptake, which was associated with a significant downregulation in level of expression of THTR-1 and THTR-2 at the protein and mRNA levels and a decrease in transcriptional (promoter) activity. Modulators of intracellular Ca2+/calmodulin- and protein-tyrosine kinase-mediated pathways also altered thiamin uptake. Finally, confocal imaging of live beta-TC-6 cells showed that clinical mutants of THTR-1 have mixed expression phenotypes and all led to impairment in thiamin uptake. These studies demonstrate for the first time that thiamin uptake by the endocrine pancreas is carrier mediated and is adaptively regulated by the prevailing vitamin level via transcriptional mechanisms. Furthermore, clinical mutants of THTR-1 impair thiamin uptake via different mechanisms. PMID:19423748

  11. Fetal reprogramming and senescence in hypoplastic left heart syndrome and in human pluripotent stem cells during cardiac differentiation.

    Science.gov (United States)

    Gaber, Naila; Gagliardi, Mark; Patel, Pranali; Kinnear, Caroline; Zhang, Cindy; Chitayat, David; Shannon, Patrick; Jaeggi, Edgar; Tabori, Uri; Keller, Gordon; Mital, Seema

    2013-09-01

    Hypoplastic left heart syndrome (HLHS) is a severe cardiac malformation characterized by left ventricle (LV) hypoplasia and abnormal LV perfusion and oxygenation. We studied hypoxia-associated injury in fetal HLHS and human pluripotent stem cells during cardiac differentiation to assess the effect of microenvironmental perturbations on fetal cardiac reprogramming. We studied LV myocardial samples from 32 HLHS and 17 structurally normal midgestation fetuses. Compared with controls, the LV in fetal HLHS samples had higher nuclear expression of hypoxia-inducible factor-1α but lower angiogenic growth factor expression, higher expression of oncogenes and transforming growth factor (TGF)-β1, more DNA damage and senescence with cell cycle arrest, fewer cardiac progenitors, myocytes and endothelial lineages, and increased myofibroblast population (P cells (SMCs) had less DNA damage compared with endothelial cells and myocytes. We recapitulated the fetal phenotype by subjecting human pluripotent stem cells to hypoxia during cardiac differentiation. DNA damage was prevented by treatment with a TGF-β1 inhibitor (P cells). The hypoplastic LV in fetal HLHS samples demonstrates hypoxia-inducible factor-1α up-regulation, oncogene-associated cellular senescence, TGF-β1-associated fibrosis and impaired vasculogenesis. The phenotype is recapitulated by subjecting human pluripotent stem cells to hypoxia during cardiac differentiation and rescued by inhibition of TGF-β1. This finding suggests that hypoxia may reprogram the immature heart and affect differentiation and development.

  12. Temporal and spatial distribution of mast cells and steroidogenic enzymes in the human fetal adrenal.

    Science.gov (United States)

    Naccache, Alexandre; Louiset, Estelle; Duparc, Céline; Laquerrière, Annie; Patrier, Sophie; Renouf, Sylvie; Gomez-Sanchez, Celso E; Mukai, Kuniaki; Lefebvre, Hervé; Castanet, Mireille

    2016-10-15

    Mast cells are present in the human adult adrenal with a potential role in the regulation of aldosterone secretion in both normal cortex and adrenocortical adenomas. We have investigated the human developing adrenal gland for the presence of mast cells in parallel with steroidogenic enzymes profile and serotonin signaling pathway. RT-QPCR and immunohistochemical studies were performed on adrenals at 16-41 weeks of gestation (WG). Tryptase-immunopositive mast cells were found from 18 WG in the adrenal subcapsular layer, close to 3βHSD- and CYP11B2-immunoreactive cells, firstly detected at 18 and 24 WG, respectively. Tryptophan hydroxylase and serotonin receptor type 4 expression increased at 30 WG before the CYP11B2 expression surge. In addition, HDL and LDL cholesterol receptors were expressed in the subcapsular zone from 24 WG. Altogether, our findings suggest the implication of mast cells and serotonin in the establishment of the mineralocorticoid synthesizing pathway during fetal adrenal development.

  13. Weak functional connectivity in the human fetal brain prior to preterm birth

    Science.gov (United States)

    Thomason, Moriah E.; Scheinost, Dustin; Manning, Janessa H.; Grove, Lauren E.; Hect, Jasmine; Marshall, Narcis; Hernandez-Andrade, Edgar; Berman, Susan; Pappas, Athina; Yeo, Lami; Hassan, Sonia S.; Constable, R. Todd; Ment, Laura R.; Romero, Roberto

    2017-01-01

    It has been suggested that neurological problems more frequent in those born preterm are expressed prior to birth, but owing to technical limitations, this has been difficult to test in humans. We applied novel fetal resting-state functional MRI to measure brain function in 32 human fetuses in utero and found that systems-level neural functional connectivity was diminished in fetuses that would subsequently be born preterm. Neural connectivity was reduced in a left-hemisphere pre-language region, and the degree to which connectivity of this left language region extended to right-hemisphere homologs was positively associated with the time elapsed between fMRI assessment and delivery. These results provide the first evidence that altered functional connectivity in the preterm brain is identifiable before birth. They suggest that neurodevelopmental disorders associated with preterm birth may result from neurological insults that begin in utero. PMID:28067865

  14. Weak functional connectivity in the human fetal brain prior to preterm birth.

    Science.gov (United States)

    Thomason, Moriah E; Scheinost, Dustin; Manning, Janessa H; Grove, Lauren E; Hect, Jasmine; Marshall, Narcis; Hernandez-Andrade, Edgar; Berman, Susan; Pappas, Athina; Yeo, Lami; Hassan, Sonia S; Constable, R Todd; Ment, Laura R; Romero, Roberto

    2017-01-09

    It has been suggested that neurological problems more frequent in those born preterm are expressed prior to birth, but owing to technical limitations, this has been difficult to test in humans. We applied novel fetal resting-state functional MRI to measure brain function in 32 human fetuses in utero and found that systems-level neural functional connectivity was diminished in fetuses that would subsequently be born preterm. Neural connectivity was reduced in a left-hemisphere pre-language region, and the degree to which connectivity of this left language region extended to right-hemisphere homologs was positively associated with the time elapsed between fMRI assessment and delivery. These results provide the first evidence that altered functional connectivity in the preterm brain is identifiable before birth. They suggest that neurodevelopmental disorders associated with preterm birth may result from neurological insults that begin in utero.

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

    Directory of Open Access Journals (Sweden)

    Ya'an Kang

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

  16. Characterization of pancreatic stem cells derived from adult human pancreas ducts by fluorescence activated cell sorting

    Institute of Scientific and Technical Information of China (English)

    Han-Tso Lin; Shih-Hwa Chiou; Chung-Lan Kao; Yi-Ming Shyr; Chien-Jen Hsu; Yih-Wen Tarng; Larry L-T Ho; Ching-Fai Kwok; Hung-Hai Ku

    2006-01-01

    AIM: To isolate putative pancreatic stem cells (PSCs)from human adult tissues of pancreas duct using serumfree, conditioned medium. The characterization of surface phenotype of these PSCs was analyzed by flow cytometry. The potential for pancreatic lineage and the capability of β-cell differentiation in these PSCs were evaluated as well.METHODS: By using serum-free medium supplemented with essential growth factors, we attempted to isolate the putative PSCs which has been reported to express nestin and pdx-1. The MatrigelTM was employed to evaluate the differential capacity of isolated cells. Dithizone staining, insulin content/secretion measurement, and immunohistochemistry staining were used to monitor the differentiation. Fluorescence activated cell sorting (FACS)was used to detect the phenotypic markers of putative PSCs.RESULTS: A monolayer of spindle-like cells was cultivated. The putative PSCs expressed pdx-1 and nestin.They were also able to differentiate into insulin-, glucagon-, and somatostatin-positive cells. The spectrum of phenotypic markers in PSCs was investigated; a similarity was revealed when using human bone marrow-derived stem cells as the comparative experiment, such as CD29,CD44, CD49, CD50, CD51, CD62E, PDGFR-α, CD73 (SH2),CD81, CD105(SH3).CONCLUSION: In this study, we successfully isolated PSCs from adult human pancreatic duct by using serumfree medium. These PSCs not only expressed nestin and pdx-1 but also exhibited markers attributable to mesenchymal stem cells. Although work is needed to elucidate the role of these cells, the application of these PSCs might be therapeutic strategies for diabetes mellitus.

  17. Effects of apigenin on cell proliferation of human pancreatic carcinoma cell line BxPC-3 in vitro

    Institute of Scientific and Technical Information of China (English)

    Jiancang Ma; Qiang Li; Jun Zhao; Ying Guo; Qinghua Su; Zongzheng Ji

    2007-01-01

    Objective: To observe the effects of apigenin on cell proliferation of human pancreatic carcinoma cell line BxPC-3 in vitro.Methods :The inhibitive effects of apigenin at different concentrations (0 μmol/L, 100 μmol/L, 200 μmol/L, and 400 μmol/L)on human pancreatic carcinoma cell line BxPC-3 were detected by MTT assays, transmission electron microscope, agarose gel electrophoresis and flow cytometry. The immunohistochemistry was used to detect the expression of Bcl-2 and Bax gene. Results:Apigenin at different concentrations could inhibit the proliferation of human pancreatic carcinoma cell lines BxPC-3, and the inhibitive effect was dose-dependent. The cell cycle of pancreatic carcinoma cells was arrested at G2/M phase. The results of immunohistochemistry showed that the density of apigenin increased, and the expression of Bcl-2 gene was reduced gradually. At the same time the expression of Bax gene was enhanced. Conclusion: Apigenin could inhibit the proliferation of human pancreatic carcinoma cell lines BxPC-3 in vitro. The effect of apoptosis was accompanied with the expression of Bcl-2 decrease and Bax increase.

  18. Neurotransmitters act as paracrine signals to regulate insulin secretion from the human pancreatic islet.

    Science.gov (United States)

    Rodriguez-Diaz, Rayner; Menegaz, Danusa; Caicedo, Alejandro

    2014-08-15

    In this symposium review we discuss the role of neurotransmitters as paracrine signals that regulate pancreatic islet function. A large number of neurotransmitters and their receptors has been identified in the islet, but relatively little is known about their involvement in islet biology. Interestingly, neurotransmitters initially thought to be present in autonomic axons innervating the islet are also present in endocrine cells of the human islet. These neurotransmitters can thus be released as paracrine signals to help control hormone release. Here we propose that the role of neurotransmitters may extend beyond controlling endocrine cell function to work as signals modulating vascular flow and immune responses within the islet.

  19. Impact of Global Fxr Deficiency on Experimental Acute Pancreatitis and Genetic Variation in the FXR Locus in Human Acute Pancreatitis

    NARCIS (Netherlands)

    Nijmeijer, Rian M.; Schaap, Frank G.; Smits, Alexander J. J.; Kremer, Andreas E.; Akkermans, Louis M. A.; Kroese, Alfons B. A.; Rijkers, Ger. T.; Schipper, Marguerite E. I.; Verheem, Andre; Wijmenga, Cisca; Gooszen, Hein G.; van Erpecum, Karel J.

    2014-01-01

    Background: Infectious complications often occur in acute pancreatitis, related to impaired intestinal barrier function, with prolonged disease course and even mortality as a result. The bile salt nuclear receptor farnesoid X receptor (FXR), which is expressed in the ileum, liver and other organs

  20. Impact of Global Fxr Deficiency on Experimental Acute Pancreatitis and Genetic Variation in the FXR Locus in Human Acute Pancreatitis

    NARCIS (Netherlands)

    Nijmeijer, R.M.; Schaap, F.G.; Smits, A.J.A.; Kremer, A.E.; Akkermans, L.M.; Kroese, A.B.A.; Rijkers, G.T.; Schipper, M.E.; Verheem, A.; Wijmenga, C.; Gooszen, H.G.; Erpecum, K.J. van

    2014-01-01

    BACKGROUND: Infectious complications often occur in acute pancreatitis, related to impaired intestinal barrier function, with prolonged disease course and even mortality as a result. The bile salt nuclear receptor farnesoid X receptor (FXR), which is expressed in the ileum, liver and other organs

  1. Impact of Global Fxr Deficiency on Experimental Acute Pancreatitis and Genetic Variation in the FXR Locus in Human Acute Pancreatitis

    NARCIS (Netherlands)

    Nijmeijer, Rian M; Schaap, Frank G; Smits, Alexander J J; Kremer, Andreas E; Akkermans, Louis M A; Kroese, A.B.; Rijkers, Ger T; Schipper, Marguerite E I; Verheem, André; Wijmenga, Cisca; Gooszen, Hein G; van Erpecum, Karel J

    2014-01-01

    BACKGROUND: Infectious complications often occur in acute pancreatitis, related to impaired intestinal barrier function, with prolonged disease course and even mortality as a result. The bile salt nuclear receptor farnesoid X receptor (FXR), which is expressed in the ileum, liver and other organs in

  2. Impact of Global Fxr Deficiency on Experimental Acute Pancreatitis and Genetic Variation in the FXR Locus in Human Acute Pancreatitis

    NARCIS (Netherlands)

    Nijmeijer, Rian M.; Schaap, Frank G.; Smits, Alexander J. J.; Kremer, Andreas E.; Akkermans, Louis M. A.; Kroese, Alfons B. A.; Rijkers, Ger. T.; Schipper, Marguerite E. I.; Verheem, Andre; Wijmenga, Cisca; Gooszen, Hein G.; van Erpecum, Karel J.

    2014-01-01

    Background: Infectious complications often occur in acute pancreatitis, related to impaired intestinal barrier function, with prolonged disease course and even mortality as a result. The bile salt nuclear receptor farnesoid X receptor (FXR), which is expressed in the ileum, liver and other organs in

  3. Investigation on Hepatopoietin and Other Novel Genes from Human Fetal Liver

    Institute of Scientific and Technical Information of China (English)

    He Fuchu; Zhang Chenggang; Li Yong; Lu Chengrong; Zhang Lingqiang

    2007-01-01

    The aim of this study is to discover the molecular mechanism of the 22-week gestated human fetal liver( HFL ) which rarely displays both hematopoietic and hepatic functions. Based on large-scale cDNA library sequencing and bioinformatic analysis, the largest gene expression profile of human fetal liver in the world was successfully established. A set of gene clusters functionally related to the liver development, hepatocarcinogenesis and hematopoiesis have been identified. This is for the first time that we could panoramically understand the molecular mechanism of the dual functions of human fetal liver. Moreover, 201 unrecorded human homologous genes and 609 novel genes have been identified and annotated, which accounting for more than 7% of the known human genes in 2001. In the recent human genome annotation map (human genome build 35.1 ), 45 genes were nominated based on this study.In addition, we have characterized a set of gene families represented by hepatopoietin (HPO), Semaphorin,LSECtin and ARFGAP. Two distinctive novel pathways,"extracellular HPO→ HPO receptor→ EGF receptor→Raf→ MEK→ MAPK" for autocrine and "intracellular HPO→ JAB1→c-JUN (AP-1 )" for intracrine of HPO, an unusual cytokine functioned in the regeneration of liver,has been reported for the first time, which have shed new lights on the study of the signal transduction of the entire HPO family. We have also demonstrated that HPO could act as a FAD thioloxidase and that only its intracrine pathway is dependent on the enzymatic activity. It is also known for the first time that the enzyme activity is critically important for the cytokine HPO. Regarding the regulation of the gene expression of HPO, it was demonstrated that HPO promoter includes a negative regulatory element and a core promoter (comprises an initiator and its flanking three tandem IFE elements).Furthermore, two novel members of Semaphorin family,SEMA6C and SEMA6D, were cloned and shown to be able to determine the

  4. Cyclin C stimulates β-cell proliferation in rat and human pancreatic β-cells

    Science.gov (United States)

    Jiménez-Palomares, Margarita; López-Acosta, José Francisco; Villa-Pérez, Pablo; Moreno-Amador, José Luis; Muñoz-Barrera, Jennifer; Fernández-Luis, Sara; Heras-Pozas, Blanca; Perdomo, Germán; Bernal-Mizrachi, Ernesto

    2015-01-01

    Activation of pancreatic β-cell proliferation has been proposed as an approach to replace reduced functional β-cell mass in diabetes. Quiescent fibroblasts exit from G0 (quiescence) to G1 through pRb phosphorylation mediated by cyclin C/cdk3 complexes. Overexpression of cyclin D1, D2, D3, or cyclin E induces pancreatic β-cell proliferation. We hypothesized that cyclin C overexpression would induce β-cell proliferation through G0 exit, thus being a potential therapeutic target to recover functional β-cell mass. We used isolated rat and human islets transduced with adenovirus expressing cyclin C. We measured multiple markers of proliferation: [3H]thymidine incorporation, BrdU incorporation and staining, and Ki67 staining. Furthermore, we detected β-cell death by TUNEL, β-cell differentiation by RT-PCR, and β-cell function by glucose-stimulated insulin secretion. Interestingly, we have found that cyclin C increases rat and human β-cell proliferation. This augmented proliferation did not induce β-cell death, dedifferentiation, or dysfunction in rat or human islets. Our results indicate that cyclin C is a potential target for inducing β-cell regeneration. PMID:25564474

  5. Purinergic receptors and calcium signalling in human pancreatic duct cell lines

    DEFF Research Database (Denmark)

    Hansen, Mette R; Krabbe, Simon; Novak, Ivana

    2008-01-01

    Purinergic receptors regulate various processes including epithelial transport. There are several studies on P2 receptors in pancreatic ducts of various species, but relatively little is known about these receptors in human tissue. The aim of this study was to identify purinergic receptors in human......ATP, commonly used to stimulate P2X7 receptors, elicited non-oscillatory and transient Ca(2+) responses. Ivermectin, a potentiator of P2X4 receptors, increased Ca(2+) signals evoked by ATP. The single cell Ca(2+) measurements indicated functional expression of P2Y2 and other P2Y receptors, and notably...... expression of P2X4 and P2X7 receptors. Expression of P2Y2, P2X4 and P2X7 receptors was confirmed by immunocytochemistry. This fingerprint of P2 receptors in human pancreatic duct models forms the basis for studying effect of nucleotides on ion and fluid secretion, as well as on Ca(2+) and tissue homeostasis...

  6. Efficient gene delivery and silencing of mouse and human pancreatic islets

    Directory of Open Access Journals (Sweden)

    Moerman Ericka

    2010-03-01

    Full Text Available Abstract Background In view of the importance of beta cells in glucose homeostasis and the profound repercussions of beta cell pathology on human health, the acquisition of tools to study pancreatic islet function is essential for the design of alternative novel therapies for diabetes. One promising approach toward this goal involves the modification of gene expression profile of beta cells. Results This study describes a new method of gene and siRNA delivery into human pancreatic islets by microporation technology. We demonstrated that mild islet distention with accutase greatly enhanced the transfection efficiency without compromising in vitro function (secretion, apoptosis and viability. As an example, the recently identified gene involved in type 2 diabetes, ZnT8, can be over-expressed or silenced by RNA interference using this technology. Microporation can also be used on rodent islets. Conclusions Taken together, our results demonstrate that microporation technology can be used to modify gene expression in whole rodent and human islets without altering their in vitro function and will be key to the elucidation of the factors responsible for proper islet function.

  7. Plasticity of adult human pancreatic duct cells by neurogenin3-mediated reprogramming.

    Directory of Open Access Journals (Sweden)

    Nathalie Swales

    Full Text Available AIMS/HYPOTHESIS: Duct cells isolated from adult human pancreas can be reprogrammed to express islet beta cell genes by adenoviral transduction of the developmental transcription factor neurogenin3 (Ngn3. In this study we aimed to fully characterize the extent of this reprogramming and intended to improve it. METHODS: The extent of the Ngn3-mediated duct-to-endocrine cell reprogramming was measured employing genome wide mRNA profiling. By modulation of the Delta-Notch signaling or addition of pancreatic endocrine transcription factors Myt1, MafA and Pdx1 we intended to improve the reprogramming. RESULTS: Ngn3 stimulates duct cells to express a focused set of genes that are characteristic for islet endocrine cells and/or neural tissues. This neuro-endocrine shift however, is incomplete with less than 10% of full duct-to-endocrine reprogramming achieved. Transduction of exogenous Ngn3 activates endogenous Ngn3 suggesting auto-activation of this gene. Furthermore, pancreatic endocrine reprogramming of human duct cells can be moderately enhanced by inhibition of Delta-Notch signaling as well as by co-expressing the transcription factor Myt1, but not MafA and Pdx1. CONCLUSIONS/INTERPRETATION: The results provide further insight into the plasticity of adult human duct cells and suggest measurable routes to enhance Ngn3-mediated in vitro reprogramming protocols for regenerative beta cell therapy in diabetes.

  8. Study on the immune responses against pancreatic cancer induced by mucin 4 and human telomerase reverse transcriptase mRNA co-transfected dendritic cells in vitro

    Institute of Scientific and Technical Information of China (English)

    陈江

    2014-01-01

    Objective To investigate the anti-tumor immune response induced by human pancreatic cancer mucin 4mRNA and human telomerase reverse transcriptase(hTERT)mRNA cotransfected dendritic cells(DC),and to provide the experimental evidences for the treatment of pancreatic cancer with multi-epitope loaded DC vaccine.Methods DC were isolated from peripheral DC.

  9. Differential response of the epithelium and interstitium in developing human fetal lung explants to hyperoxia.

    Science.gov (United States)

    Bustani, Porus; Hodge, Rachel; Tellabati, Ananth; Li, Juan; Pandya, Hitesh; Kotecha, Sailesh

    2006-03-01

    Hyperoxia is closely linked with the development of chronic lung disease of prematurity (CLD), but the exact mechanisms whereby hyperoxia alters the lung architecture in the developing lung remain largely unknown. We developed a fetal human lung organ culture model to investigate (a) the morphologic changes induced by hyperoxia and (b) whether hyperoxia resulted in differential cellular responses in the epithelium and interstitium. The effects of hyperoxia on lung morphometry were analyzed using computer-assisted image analysis. The lung architecture remained largely unchanged in normoxia lasting as long as 4 d. In contrast, hyperoxic culture of pseudoglandular fetal lungs resulted in significant dilatation of airways, thinning of the epithelium, and regression of the interstitium including the pulmonary vasculature. Although there were no significant differences in Ki67 between normoxic and hyperoxic lungs, activated caspase-3 was significantly increased in interstitial cells, but not epithelial cells, under hyperoxic conditions. These changes show that exposure of pseudoglandular lungs to hyperoxia modulates the lung architecture to resemble saccular lungs.

  10. Immunomodulatory properties of human adult and fetal multipotent mesenchymal stem cells.

    Science.gov (United States)

    Chen, Pei-Min; Yen, Men-Luh; Liu, Ko-Jiunn; Sytwu, Huey-Kang; Yen, B-Linju

    2011-07-18

    In recent years, a large number of studies have contributed to our understanding of the immunomodulatory mechanisms used by multipotent mesenchymal stem cells (MSCs). Initially isolated from the bone marrow (BM), MSCs have been found in many tissues but the strong immunomodulatory properties are best studied in BM MSCs. The immunomodulatory effects of BM MSCs are wide, extending to T lymphocytes and dendritic cells, and are therapeutically useful for treatment of immune-related diseases including graft-versus-host disease as well as possibly autoimmune diseases. However, BM MSCs are very rare cells and require an invasive procedure for procurement. Recently, MSCs have also been found in fetal-stage embryo-proper and extra-embryonic tissues, and these human fetal MSCs (F-MSCs) have a higher proliferative profile, and are capable of multilineage differentiation as well as exert strong immunomodulatory effects. As such, these F-MSCs can be viewed as alternative sources of MSCs. We review here the current understanding of the mechanisms behind the immunomodulatory properties of BM MSCs and F-MSCs. An increase in our understanding of MSC suppressor mechanisms will offer insights for prevalent clinical use of these versatile adult stem cells in the near future.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2016-03-15

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

  12. Glycosylation of human fetal mucins: a similar repertoire of O-glycans along the intestinal tract.

    Science.gov (United States)

    Robbe-Masselot, Catherine; Maes, Emmanuel; Rousset, Monique; Michalski, Jean-Claude; Capon, Calliope

    2009-05-01

    Intestinal mucins are very high molecular weight glycoproteins secreted by goblet cells lining the crypt and the surface of the colonic mucosa. Profound alterations of mucin O-glycans are observed in diseases such as cancer and inflammation, modifying the function of the cell and its antigenic and adhesive properties. Based on immunohistochemical studies, certain cancer- and inflammation- associated glycans have been defined as oncofetal antigens. However, little or no chemical analysis has allowed the structural elucidation of O-glycans expressed on human fetal mucins. In this paper, mucins were isolated from different regions of the normal human intestine (ileum, right, transverse and left colon) of eight fetuses with A, B or O blood group. After alkaline borohydride treatment, the released oligosaccharides were investigated by nanoESI Q-TOF MS/MS (electrospray ionization quadrupole time-of-flight tandem mass spectrometry). More than 117 different glycans were identified, mainly based on core 2 structures. Some core 1, 3 and 4 oligosaccharides were also found. Most of the structures were acidic with NeuAc residues mainly alpha2-6 linked to the N-acetylgalactosaminitol and sulphate residues 3-linked to galactose or 6-linked to GlcNAc. In contrast to adult human intestinal mucins, Sda/Cad determinants were not expressed on fetal mucin O-glycans and the presence of an acidic gradient along the intestinal tract was not observed. Similar patterns of glycosylation were found in each part of the intestine and the level of expression of the major oligosaccharides was in the same order of magnitude. This study could help determining new oncofetal antigens, which can be exploited for the diagnosis or the treatment of intestinal diseases.

  13. Evaluation of human platelet lysate versus fetal bovine serum for culture of mesenchymal stromal cells.

    Science.gov (United States)

    Hemeda, Hatim; Giebel, Bernd; Wagner, Wolfgang

    2014-02-01

    Culture media for therapeutic cell preparations-such as mesenchymal stromal cells (MSCs)-usually comprise serum additives. Traditionally, fetal bovine serum is supplemented in basic research and in most clinical trials. Within the past years, many laboratories adapted their culture conditions to human platelet lysate (hPL), which further stimulates proliferation and expansion of MSCs. Particularly with regard to clinical application, human alternatives for fetal bovine serum are clearly to be preferred. hPL is generated from human platelet units by disruption of the platelet membrane, which is commonly performed by repeated freeze and thaw cycles. Such culture supplements are notoriously ill-defined, and many parameters contribute to batch-to-batch variation in hPL such as different amounts of plasma, a broad range of growth factors and donor-specific effects. The plasma components of hPL necessitate addition of anticoagulants such as heparins to prevent gelatinization of hPL medium, and their concentration must be standardized. Labels for description of hPL-such as "xenogen-free," "animal-free" and "serum free"-are not used consistently in the literature and may be misleading if not critically assessed. Further analysis of the precise composition of relevant growth factors, attachment factors, microRNAs and exosomes will pave the way for optimized and defined culture conditions. The use of hPL has several advantages and disadvantages: they must be taken into account because the choice of cell culture additive has major impact on cell preparations. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  14. MCT8 expression in human fetal cerebral cortex is reduced in severe intrauterine growth restriction.

    Science.gov (United States)

    Chan, Shiao Y; Hancox, Laura A; Martín-Santos, Azucena; Loubière, Laurence S; Walter, Merlin N M; González, Ana-Maria; Cox, Phillip M; Logan, Ann; McCabe, Christopher J; Franklyn, Jayne A; Kilby, Mark D

    2014-02-01

    The importance of the thyroid hormone (TH) transporter, monocarboxylate transporter 8 (MCT8), to human neurodevelopment is highlighted by findings of severe global neurological impairment in subjects with MCT8 (SLC16A2) mutations. Intrauterine growth restriction (IUGR), usually due to uteroplacental failure, is associated with milder neurodevelopmental deficits, which have been partly attributed to dysregulated TH action in utero secondary to reduced circulating fetal TH concentrations and decreased cerebral thyroid hormone receptor expression. We postulate that altered MCT8 expression is implicated in this pathophysiology; therefore, in this study, we sought to quantify changes in cortical MCT8 expression with IUGR. First, MCT8 immunohistochemistry was performed on occipital and parietal cerebral cortex sections obtained from appropriately grown for gestational age (AGA) human fetuses between 19 weeks of gestation and term. Secondly, MCT8 immunostaining in the occipital cortex of stillborn IUGR human fetuses at 24-28 weeks of gestation was objectively compared with that in the occipital cortex of gestationally matched AGA fetuses. Fetuses demonstrated widespread MCT8 expression in neurons within the cortical plate and subplate, in the ventricular and subventricular zones, in the epithelium of the choroid plexus and ependyma, and in microvessel wall. When complicated by IUGR, fetuses showed a significant fivefold reduction in the percentage area of cortical plate immunostained for MCT8 compared with AGA fetuses (PMCT8 expression was negatively correlated with the severity of IUGR indicated by the brain:liver weight ratios (r(2)=0.28; PMCT8 expression in the IUGR fetal brain could further compromise TH-dependent brain development.

  15. Human pancreatic β-cell G1/S molecule cell cycle atlas.

    Science.gov (United States)

    Fiaschi-Taesch, Nathalie M; Kleinberger, Jeffrey W; Salim, Fatimah G; Troxell, Ronnie; Wills, Rachel; Tanwir, Mansoor; Casinelli, Gabriella; Cox, Amy E; Takane, Karen K; Scott, Donald K; Stewart, Andrew F

    2013-07-01

    Expansion of pancreatic β-cells is a key goal of diabetes research, yet induction of adult human β-cell replication has proven frustratingly difficult. In part, this reflects a lack of understanding of cell cycle control in the human β-cell. Here, we provide a comprehensive immunocytochemical "atlas" of G1/S control molecules in the human β-cell. This atlas reveals that the majority of these molecules, previously known to be present in islets, are actually present in the β-cell. More importantly, and in contrast to anticipated results, the human β-cell G1/S atlas reveals that almost all of the critical G1/S cell cycle control molecules are located in the cytoplasm of the quiescent human β-cell. Indeed, the only nuclear G1/S molecules are the cell cycle inhibitors, pRb, p57, and variably, p21: none of the cyclins or cdks necessary to drive human β-cell proliferation are present in the nuclear compartment. This observation may provide an explanation for the refractoriness of human β-cells to proliferation. Thus, in addition to known obstacles to human β-cell proliferation, restriction of G1/S molecules to the cytoplasm of the human β-cell represents an unanticipated obstacle to therapeutic human β-cell expansion.

  16. [Chronic pancreatitis, acute pancreatitis].

    Science.gov (United States)

    Mabuchi, T; Katada, N; Nishimura, D; Hoshino, H; Shimizu, F; Suzuki, R; Sano, H; Kato, K

    1998-11-01

    MRCP has been recognized as a safe and noninvasive diagnostic method. In the present study we evaluated the usefulness of MRCP in diagnosis of chronic and acute pancreatitis. Two-dimensional fast asymmetric spin-echo (FASE) MRCP was performed in 40 patients with chronic pancreatitis and 13 with acute pancreatitis. In 29 patients (72.5%) with chronic pancreatitis and 9 (66.7%) with acute pancreatitis, main pancreatic duct (MPD) was visualized entirely. MRCP could demonstrate the characteristic findings of chronic pancreatitis such as dilatation and irregularity of MPD in most cases. In acute pancreatitis, MRCP indicated that MPD was normal in diameter, but irregular in configuration compared with that of the control group. MRCP may facilitate the diagnosis of chronic and acute pancreatitis.

  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. Varicella zoster virus infection of human fetal lung cells alters mitochondrial morphology.

    Science.gov (United States)

    Keller, Amy C; Badani, Hussain; McClatchey, P Mason; Baird, Nicholas L; Bowlin, Jacqueline L; Bouchard, Ron; Perng, Guey-Chuen; Reusch, Jane E B; Kaufer, Benedikt B; Gilden, Don; Shahzad, Aamir; Kennedy, Peter G E; Cohrs, Randall J

    2016-10-01

    Varicella zoster virus (VZV) is a ubiquitous alphaherpesvirus that establishes latency in ganglionic neurons throughout the neuraxis after primary infection. Here, we show that VZV infection induces a time-dependent significant change in mitochondrial morphology, an important indicator of cellular health, since mitochondria are involved in essential cellular functions. VZV immediate-early protein 63 (IE63) was detected in mitochondria-rich cellular fractions extracted from infected human fetal lung fibroblasts (HFL) by Western blotting. IE63 interacted with cytochrome c oxidase in bacterial 2-hybrid analyses. Confocal microscopy of VZV-infected HFL cells at multiple times after infection revealed the presence of IE63 in the nucleus, mitochondria, and cytoplasm. Our data provide the first evidence that VZV infection induces alterations in mitochondrial morphology, including fragmentation, which may be involved in cellular damage and/or death during virus infection.

  19. Anatomical relationships between testis and epididymis during the fetal period in humans (10-36 weeks postconception)

    NARCIS (Netherlands)

    Favorito, LA; Sampaio, FJB

    1998-01-01

    Objective: To determine the anatomy of the epididymis and its relationship with the testis during the fetal period in normal individuals. Methods: We studied bilaterally 146 testes and epididymides taken from 73 normal fresh human fetuses ranging in age from 10 to 36 weeks postconception. The epidid

  20. FA1 immunoreactivity in endocrine tumours and during development of the human fetal pancreas; negative correlation with glucagon expression

    DEFF Research Database (Denmark)

    Tornehave, D; Jensen, Charlotte Harken; Teisner, B

    1996-01-01

    Fetal antigen 1 (FA1) is a glycoprotein containing six epidermal growth factor (EGF)-like repeats. It is closely similar to the protein translated from the human delta-like (dlk) cDNA and probably constitutes a proteolytically processed form of dlk. dlk is homologous to the Drosophila homeotic pr...

  1. Celecoxib Inhibits Proliferation and Induces Apoptosis via Cyclooxygen-ase-2 Pathway in Human Pancreatic Carcinoma Cells

    Institute of Scientific and Technical Information of China (English)

    WU Gaosong; YI Jilin; DI Fang; ZOU Shengquan; LI Xingrui

    2005-01-01

    In order to evaluate the effects and mechanisms of celecoxib in inhibiting proliferation and inducing apoptosis on human pancreatic carcinoma cells, the anti-proliferative effect was measured by using methabenzthiazuron (MTT) assay. Cell cycle and apoptosis were analyzed by using flow cytometry (FCM), and the PGE2 levels in the supernatant of cultured pancreatic carcinoma cells were quantitated by enzyme-linked immunoabsordent assay (ELISA). Our results showed that celecoxib suppressed the production of PGE2 and inhibited the growth of JF-305 cells, and the anti-proliferative effect of celecoxib could be abolished by addition of PGE2. FCM revealed that celecoxib could inhibit proliferation and induce apoptosis by G1-S cell cycle arrest. It was concluded that cyclooxygenase-2 specific inhibitor celecoxib could inhibit proliferation and induced apoptosis of human pancreatic carcinoma cells via suppression of PGE2 production in vitro.

  2. A Convenient and Efficient Method to Enrich and Maintain Highly Proliferative Human Fetal Liver Stem Cells.

    Science.gov (United States)

    Guo, Xuan; Wang, Shu; Dou, Ya-ling; Guo, Xiang-fei; Chen, Zhao-li; Wang, Xin-wei; Shen, Zhi-qiang; Qiu, Zhi-gang; Jin, Min; Li, Jun-wen

    2015-06-01

    Pluripotent human hepatic stem cells have broad research and clinical applications, which are, however, restricted by both limited resources and technical difficulties with respect to isolation of stem cells from the adult or fetal liver. In this study, we developed a convenient and efficient method involving a two-step in situ collagenase perfusion, gravity sedimentation, and Percoll density gradient centrifugation to enrich and maintain highly proliferative human fetal liver stem cells (hFLSCs). Using this method, the isolated hFLSCs entered into the exponential growth phase within 10 days and maintained sufficient proliferative activity to permit subculture for at least 20 passages without differentiation. Immunocytochemistry, immunofluorescence, and flow cytometry results showed that these cells expressed stem cell markers, such as c-kit, CD44, epithelial cell adhesion molecule (EpCAM), oval cell marker-6 (OV-6), epithelial marker cytokeratin 18 (CK18), biliary ductal marker CK19, and alpha-fetoprotein (AFP). Gene expression analysis showed that these cells had stable mRNA expression of c-Kit, EpCAM, neural cell adhesion molecule (NCAM), CK19, CK18, AFP, and claudin 3 (CLDN-3) throughout each passage while maintaining low levels of ALB, but with complete absence of cytochrome P450 3A4 (C3A4), phosphoenolpyruvate carboxykinase (PEPCK), telomeric repeat binding factor (TRF), and connexin 26 (CX26) expression. When grown in appropriate medium, these isolated liver stem cells could differentiate into hepatocytes, cholangiocytes, osteoblasts, adipocytes, or endothelial cells. Thus, we have demonstrated a more economical and efficient method to isolate hFLSCs than magnetic-activated cell sorting (MACS). This novel approach may provide an excellent tool to isolate highly proliferative hFLSCs for tissue engineering and regenerative therapies.

  3. Protein phosphatase 1 (PP-1)-dependent inhibition of insulin secretion by leptin in INS-1 pancreatic β-cells and human pancreatic islets.

    Science.gov (United States)

    Kuehnen, Peter; Laubner, Katharina; Raile, Klemens; Schöfl, Christof; Jakob, Franz; Pilz, Ingo; Päth, Günter; Seufert, Jochen

    2011-05-01

    Leptin inhibits insulin secretion from pancreatic β-cells, and in turn, insulin stimulates leptin biosynthesis and secretion from adipose tissue. Dysfunction of this adipoinsular feedback loop has been proposed to be involved in the development of hyperinsulinemia and type 2 diabetes mellitus. At the molecular level, leptin acts through various pathways, which in combination confer inhibitory effects on insulin biosynthesis and secretion. The aim of this study was to identify molecular mechanisms of leptin action on insulin secretion in pancreatic β-cells. To identify novel leptin-regulated genes, we performed subtraction PCR in INS-1 β-cells. Regulated expression of identified genes was confirmed by RT-PCR and Northern and Western blotting. Furthermore, functional impact on β-cell function was characterized by insulin-secretion assays, intracellular Ca²(+) concentration measurements, and enzyme activity assays. PP-1α, the catalytic subunit of protein phosphatase 1 (PP-1), was identified as a novel gene down-regulated by leptin in INS-1 pancreatic β-cells. Expression of PP-1α was verified in human pancreatic sections. PP-1α mRNA and protein expression is down-regulated by leptin, which culminates in reduction of PP-1 enzyme activity in β-cells. In addition, glucose-induced insulin secretion was inhibited by nuclear inhibitor of PP-1 and calyculin A, which was in part mediated by a reduction of PP-1-dependent calcium influx into INS-1 β-cells. These results identify a novel molecular pathway by which leptin confers inhibitory action on insulin secretion, and impaired PP-1 inhibition by leptin may be involved in dysfunction of the adipoinsular axis during the development of hyperinsulinemia and type 2 diabetes mellitus.

  4. Human placenta metabolizes fatty acids: implications for fetal fatty acid oxidation disorders and maternal liver diseases.

    Science.gov (United States)

    Shekhawat, Prem; Bennett, Michael J; Sadovsky, Yoel; Nelson, D Michael; Rakheja, Dinesh; Strauss, Arnold W

    2003-06-01

    The role of fat metabolism during human pregnancy and in placental growth and function is poorly understood. Mitochondrial fatty acid oxidation disorders in an affected fetus are associated with maternal diseases of pregnancy, including preeclampsia, acute fatty liver of pregnancy, and the hemolysis, elevated liver enzymes, and low platelets syndrome called HELLP. We have investigated the developmental expression and activity of six fatty acid beta-oxidation enzymes at various gestational-age human placentas. Placental specimens exhibited abundant expression of all six enzymes, as assessed by immunohistochemical and immunoblot analyses, with greater staining in syncytiotrophoblasts compared with other placental cell types. beta-Oxidation enzyme activities in placental tissues were higher early in gestation and lower near term. Trophoblast cells in culture oxidized tritium-labeled palmitate and myristate in substantial amounts, indicating that the human placenta utilizes fatty acids as a significant metabolic fuel. Thus human placenta derives energy from fatty acid oxidation, providing a potential explanation for the association of fetal fatty acid oxidation disorders with maternal liver diseases in pregnancy.

  5. The roadmap of WT1 protein expression in the human fetal heart.

    Science.gov (United States)

    Duim, Sjoerd N; Smits, Anke M; Kruithof, Boudewijn P T; Goumans, Marie-José

    2016-01-01

    The transcription factor Wilms' Tumor-1 (WT1) is essential for cardiac development. Deletion of Wt1 in mice results in disturbed epicardial and myocardial formation and lack of cardiac vasculature, causing embryonic lethality. Little is known about the role of WT1 in the human fetal heart. Therefore, as a first step, we analyzed the expression pattern of WT1 protein during human cardiac development from week 4 till week 20. WT1 expression was apparent in epicardial, endothelial and endocardial cells in a spatiotemporal manner. The expression of WT1 follows a pattern starting at the epicardium and extending towards the lumen of the heart, with differences in timing and expression levels between the atria and ventricles. The expression of WT1 in cardiac arterial endothelial cells reduces in time, whereas WT1 expression in the endothelial cells of cardiac veins and capillaries remains present at all stages studied. This study provides for the first time a detailed description of the expression of WT1 protein during human cardiac development, which indicates an important role for WT1 also in human cardiogenesis. Copyright © 2015 Elsevier Ltd. All rights reserved.

  6. Killing effect of adenoviral mediated cytosine deaminase gene on human pancreatic cancer cell line PaTu 8988

    Institute of Scientific and Technical Information of China (English)

    PAN Xue; LI Zhao-shen; XU Guo-ming; CUI Long; ZHANG Su-zhen; GONG Yan-fang; TU Zhen-xing

    2001-01-01

    Objective: To evaluate the in vitro killing effects of cytosine deaminase gene mediated by adenovirus vector on human pancreatic carcinoma. Methods: Cytosine Deaminase (CD) gene was cloned into pAdTrack-CMV-CD, pAdTrack-CMV-CD and pAdEasy-1 were recombined in bacteria, and the products containing green fluorescent protein (GFP)were propagated in 293 cells and purified by cesium chloride gradient centrifugation. Human pancreatic carcinoma cell line 8988 were infected with this virus, then 5-FC was added; XTT assay was used to estimate the relative numbers of viable cells. Results: The positive clones were confirmed by using endonuclease digestion, and the titer of the virus containing CD gene was 2 × 1011 pfu/ml. It was found that 5-FC possessed significant cytotoxic activities for CD gene transfected 8988cell line, but had little effects on non-transfected pancreatic carcinoma cells. Conclusion: CD gene mediated by adenovirus has a high infectivity and is efficient for killing cultured pancreatic carcinoma cells, indicating suicide gene may be effective for pancreatic cancer in furure.

  7. Evaluation of Traditional Indian Antidiabetic Medicinal Plants for Human Pancreatic Amylase Inhibitory Effect In Vitro

    Directory of Open Access Journals (Sweden)

    Sudha Ponnusamy

    2011-01-01

    Full Text Available Pancreatic α-amylase inhibitors offer an effective strategy to lower the levels of post prandial hyperglycemia via control of starch breakdown. Eleven Ayurvedic Indian medicinal plants with known hypoglycemic properties were subjected to sequential solvent extraction and tested for α-amylase inhibition, in order to assess and evaluate their inhibitory potential on pancreatic α-amylase. Analysis of 91 extracts, showed that 10 exhibited strong Human Pancreatic Amylase (HPA inhibitory potential. Of these, 6 extracts showed concentration dependent inhibition with IC50 values, namely, cold and hot water extracts from Ficus bengalensis bark (4.4 and 125 μgmL-1, Syzygium cumini seeds (42.1 and 4.1 μgmL-1, isopropanol extracts of Cinnamomum verum leaves (1.0 μgmL-1 and Curcuma longa rhizome (0.16 μgmL-1. The other 4 extracts exhibited concentration independent inhibition, namely, methanol extract of Bixa orellana leaves (49 μgmL-1, isopropanol extract from Murraya koenigii leaves (127 μgmL-1, acetone extracts from C. longa rhizome (7.4 μgmL-1 and Tribulus terrestris seeds (511 μgmL-1. Thus, the probable mechanism of action of the above fractions is due to their inhibitory action on HPA, thereby reducing the rate of starch hydrolysis leading to lowered glucose levels. Phytochemical analysis revealed the presence of alkaloids, proteins, tannins, cardiac glycosides, flavonoids, saponins and steroids as probable inhibitory compounds.

  8. Transgenic expression of the human growth hormone minigene promotes pancreatic β-cell proliferation.

    Science.gov (United States)

    Baan, Mieke; Kibbe, Carly R; Bushkofsky, Justin R; Harris, Ted W; Sherman, Dawn S; Davis, Dawn Belt

    2015-10-01

    Transgenic mouse models are designed to study the role of specific proteins. To increase transgene expression the human growth hormone (hGH) minigene, including introns, has been included in many transgenic constructs. Until recently, it was thought that the hGH gene was not spliced, transcribed, and translated to produce functional hGH protein. We generated a transgenic mouse with the transcription factor Forkhead box M1 (FoxM1) followed by the hGH minigene, under control of the mouse insulin promoter (MIP) to target expression specifically in the pancreatic β-cell. Expression of FoxM1 in isolated pancreatic islets in vitro stimulates β-cell proliferation. We aimed to investigate the effect of FoxM1 on β-cell mass in a mouse model for diabetes mellitus. However, we found inadvertent coexpression of hGH protein from a spliced, bicistronic mRNA. MIP-FoxM1-hGH mice had lower blood glucose and higher pancreatic insulin content, due to increased β-cell proliferation. hGH signals through the murine prolactin receptor, and expression of its downstream targets tryptophan hydroxylase-1 (Tph1), tryptophan hydroxylase-2 (Tph2), and cytokine-inducible SH2 containing protein (Cish) was increased. Conversely, transcriptional targets of FoxM1 were not upregulated. Our data suggest that the phenotype of MIP-FoxM1-hGH mice is due primarily to hGH activity and that the FoxM1 protein remains largely inactive. Over the past decades, multiple transgenic mouse strains were generated that make use of the hGH minigene to increase transgene expression. Our work suggests that each will need to be carefully screened for inadvertent hGH production and critically evaluated for the use of proper controls.

  9. Cells Lines vs. Animals Studies for Developing New Therapeutic Strategies in Human Pancreatic Cancer?

    Directory of Open Access Journals (Sweden)

    Raffaele Pezzilli

    2006-05-01

    Full Text Available In the past, most of the knowledge gained regarding the physiology and the pathology of the pancreas has been evaluated in experimental studies on animals especially on rats/mice. This approach has been criticized in recent years because most of the data obtained from animals cannot be fully applied to humans [1]. A new approach to the physiology and pathology of the pancreas comes from studying its molecular biology, and the results obtained seem to be more reliable than those obtained in animals. An example of this assumption comes from studies on pancreatic ductal adenocarcinoma: this cancer seems to result from a progressive accumulation of mutations in genes such as K-ras, CDKN2A, p53, BRCA2, p164ink, and SMAD4 [2]; in particular, the SMAD4 mutations which result in the constitutive activation of transforming growth factor b1 signalling, are generally considered to be responsible for the desmoplastic response, which includes upregulated expression of the extracellular matrix, and type I collagen [3, 4, 5, 6]. Grzesiak et al. [7, 8] have also shown that a2b1 integrin-mediated adhesion on type I collagen promotes a malignant phenotype in FG pancreatic cells, as defined by increased proliferation and haptokinetic cell migration, downregulated expression and localization of E-cadherin and b-catenin in cell-cell contacts, increased phosphorylation of GSK3b and PKB/Akt, and downregulated expression of PTHrP, IL-6, and IL-8 as compared to fibronectin, type IV collagen, laminin, or vitronectin. These results are in agreement with previous studies demonstrating that type I collagen downregulates E-cadherin expression in Panc-1, BxPC-3, and PaTu8988s pancreatic cancer cells, resulting in increased proliferation and migration compared to fibronectin.

  10. Adenoviral vectors stimulate glucagon transcription in human mesenchymal stem cells expressing pancreatic transcription factors.

    Directory of Open Access Journals (Sweden)

    Arnaud Zaldumbide

    Full Text Available Viral gene carriers are being widely used as gene transfer systems in (transdifferentiation and reprogramming strategies. Forced expression of key regulators of pancreatic differentiation in stem cells, liver cells, pancreatic duct cells, or cells from the exocrine pancreas, can lead to the initiation of endocrine pancreatic differentiation. While several viral vector systems have been employed in such studies, the results reported with adenovirus vectors have been the most promising in vitro and in vivo. In this study, we examined whether the viral vector system itself could impact the differentiation capacity of human bone-marrow derived mesenchymal stem cells (hMSCs toward the endocrine lineage. Lentivirus-mediated expression of Pdx-1, Ngn-3, and Maf-A alone or in combination does not lead to robust expression of any of the endocrine hormones (i.e. insulin, glucagon and somatostatin in hMSCs. Remarkably, subsequent transduction of these genetically modified cells with an irrelevant early region 1 (E1-deleted adenoviral vector potentiates the differentiation stimulus and promotes glucagon gene expression in hMSCs by affecting the chromatin structure. This adenovirus stimulation was observed upon infection with an E1-deleted adenovirus vector, but not after exposure to helper-dependent adenovirus vectors, pointing at the involvement of genes retained in the E1-deleted adenovirus vector in this phenomenon. Lentivirus mediated expression of the adenovirus E4-ORF3 mimics the adenovirus effect. From these data we conclude that E1-deleted adenoviral vectors are not inert gene-transfer vectors and contribute to the modulation of the cellular differentiation pathways.

  11. An investigation of the endocrine-disruptive effects of bisphenol a in human and rat fetal testes.

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    Millissia Ben Maamar

    Full Text Available Few studies have been undertaken to assess the possible effects of bisphenol A (BPA on the reproductive hormone balance in animals or humans with often contradictory results. We investigated possible direct endocrine disruption by BPA of the fetal testes of 2 rat strains (14.5-17.5 days post-coitum and humans (8-12 gestational weeks and under different culture conditions. BPA concentrations of 10(-8M and 10(-5M for 72 h reduced testosterone production by the Sprague-Dawley fetal rat testes, while only 10-5M suppressed it in the Wistar strain. The suppressive effects at 10-5M were seen as early as 24h and 48 h in both strains. BPA at 10(-7-10(-5M for 72 h suppressed the levels of fetal rat Leydig cell insulin-like factor 3 (INSL3. BPA exposure at 10(-8M, 10(-7M, and 10(-5M for 72 h inhibited testosterone production in fetal human testes. For the lowest doses, the effects observed occurred only when no gonadotrophin was added to the culture media and were associated with a poorly preserved testicular morphology. We concluded that (i BPA can display anti-androgenic effects both in rat and human fetal testes; (ii it is essential to ascertain that the divergent effects of endocrine disruptors between species in vitro do not result from the culture conditions used, and/or the rodent strain selected; (iii the optimization of each in vitro assay for a given species should be a major objective rather than the search of an hypothetical trans-species consensual model-system, as the organization of the testis is intrinsically different between mammalian species; (iv due to the uncertainty existing on the internal exposure of the human fetal testis to BPA, and the insufficient number of epidemiological studies on the endocrine disruptive effects of BPA, caution should be taken in the extrapolation of our present results to the human reproductive health after fetal exposure to BPA.

  12. Monocytic cells derived from human embryonic stem cells and fetal liver share common differentiation pathways and homeostatic functions.

    Science.gov (United States)

    Klimchenko, Olena; Di Stefano, Antonio; Geoerger, Birgit; Hamidi, Sofiane; Opolon, Paule; Robert, Thomas; Routhier, Mélanie; El-Benna, Jamel; Delezoide, Anne-Lise; Boukour, Siham; Lescure, Bernadette; Solary, Eric; Vainchenker, William; Norol, Françoise

    2011-03-17

    The early emergence of macrophages and their large pattern of tissue distribution during development suggest that they may play a critical role in the initial steps of embryogenesis. In the present study, we show that monocytic cells derived from human embryonic stem cells (hESCs) and from fetal liver follow a differentiation pathway different to that of adult cells, leading to specific functions. Embryonic and fetal monocytic cells differentiated from a CD14(low)CD16(-) precursor to form CD14(high)CD16(+) cells without producing the CD14(high)CD16(-) cell population that predominates in adult peripheral blood. Both demonstrated an enhanced expression of genes encoding tissue-degrading enzymes, chemokines, and scavenger receptors, as was previously reported for M2 macrophages. Compared with adult blood monocytes, embryonic and fetal monocytic cells secreted high amounts of proteins acting on tissue remodeling and angiogenesis, and most of them expressed the Tie2 receptor. Furthermore, they promoted vascular remodeling in xenotransplanted human tumors. These findings suggest that the regulation of human fetal and embryonic monocytic cell differentiation leads to the generation of cells endowed mainly with anti-inflammatory and remodeling functions. Trophic and immunosuppressive functions of M2-polarized macrophages link fetus and tumor development, and hESCs offer a valuable experimental model for in vitro studies of mechanisms sustaining these processes.

  13. Human fetal striatum-derived neural stem (NS) cells differentiate to mature neurons in vitro and in vivo.

    Science.gov (United States)

    Monni, Emanuela; Cusulin, Carlo; Cavallaro, Maurizio; Lindvall, Olle; Kokaia, Zaal

    2014-01-01

    Clonogenic neural stem (NS) cell lines grown in adherent cultures have previously been established from embryonic stem cells and fetal and adult CNS in rodents and from human fetal brain and spinal cord. Here we describe the isolation of a new cell line from human fetal striatum (hNS cells). These cells showed properties of NS cells in vitro such as monolayer growth, high proliferation rate and expression of radial glia markers. The hNS cells expressed an early neuronal marker while being in the proliferative state. Under appropriate conditions, the hNS cells were efficiently differentiated to neurons, and after 4 weeks about 50% of the cells were βIII tubulin positive. They also expressed the mature neuronal marker NeuN and markers of neuronal subtypes, GABA, calbindin, and DARPP32. After intrastriatal implantation into newborn rats, the hNS cells survived and many of them migrated outside the transplant core into the surrounding tissue. A high percentage of cells in the grafts expressed the neuroblast marker DCX, indicating their neurogenic potential, and some of the cells differentiated to NeuN+ mature neurons. The human fetal striatum-derived NS cell line described here should be a useful tool for studies on cell replacement strategies in models of the striatal neuronal loss occurring in Huntington's disease and stroke.

  14. Increased oxidative stress in human fetal membranes overlying the cervix from term non-labouring and post labour deliveries.

    Science.gov (United States)

    Chai, M; Barker, G; Menon, R; Lappas, M

    2012-08-01

    Enzymatic breakdown of the collagen-rich extracellular matrix (ECM) that connects the amnion and chorion layers of the fetal membranes is one of the key events leading to rupture of membranes. Oxidant stress caused by increased formation of reactive oxygen species and/or reduced antioxidant capacity may predispose to membrane rupture, a major cause of preterm birth. The aim of this study was to determine the effect of human labour and supracervical (SC) apposition on antioxidant enzymes and 8-isoprostane (a marker of lipid peroxidation). To determine the effect of human labour on oxidative stress status, fetal membranes from the SC site (SCS) were collected from women at term Caesarean section (no labour), and from the site of membrane rupture (SOR) after spontaneous labour onset and delivery (post labour). To determine the effect of SC apposition on oxidative stress status, amnion was collected from the SCS and a distal site (DS) in women at term Caesarean section in the absence of labour. The release of 8-isoprostane was significantly higher in amnion from the SCS compared to DS, and in fetal membranes from the SOR compared to the SCS. Glutathione peroxidase (GPx) and superoxide dismutase (SOD) activity were lower in amnion from the SC compared to DS. SOD gene expression and enzyme activity were lower in fetal membranes after labour. There was no difference in expression or activity in catalase, GPx and glutathione reductase (GSR) between no labour and post labour fetal membranes. In primary amnion cells, SOD supplementation significantly augmented IL-1β induced MMP-9 expression and activity. In summary, non-labouring SC fetal membranes are characterised by reduced antioxidant enzyme activity when compared to distal membranes, and, as such, may be more susceptible to oxidative damage and thus membrane rupture. Copyright © 2012 Elsevier Ltd. All rights reserved.

  15. /sup 125/I-human epidermal growth factor specific binding to placentas and fetal membranes from varoius pregnancy states

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    Hofmann, G.E.; Siddiqi, T.A.; Rao, Ch. V.; Carman, F.R.

    1988-01-01

    Specific binding of /sup 125/I-human epidermal growth factor (hEGF) to homogenates of term human placentas and fetal membranes from normal and appropriate for gestational age (N = 20), intrauterine growth retarded (N = 9), twin (N = 11), White class AB diabetic (N = 12), and large for gestational age (N = 13) pregnancies was measured. In all pregnancy states, placentas bound approximately four times more /sup 125/I-hEGF than did fetal membranes (P<0.0001). There was no significant differnce in /sup 125/I-hEGF binding to fetal membranes from the various pregnancy states (P<0.05). /sup 125/I-hEGF specific binding to placentas from intrauterine growth retarded or twin pregnancies was significantly greater compared with placentas from normal and appropriate for gestational age pregnancies (P<0.05). The binding to placentas from pregnancies complicated by White class AB diabetes or large for gestational age infants, on the other hand, was not significantly different from that to placentas from normal and appropriate for gestational age pregnancies. /sup 125/I-hEGF specific binding did not differ between placentas from intrauterine growth retarded or twin pregnancies (P<0.05). Placental and fetal membrane /sup 125/I-hEGF binding did not vary with fetal sex, maternal race, placental weight, or gestational age between 37 to 42 weeks (P<0.05). Placental but not fetal membrane /sup 125/I-hEGF binding increased with increasing infant weight when appropriate for gestational age and large for gestational age infants were included (P<0.05, r = 0.38, N = 32) but not for intrauterine growth retarded, appropriate for gestational age, or large for gestational age infants alone.

  16. Characterization of hepatic progenitors from human fetal liver during second trimester

    Institute of Scientific and Technical Information of China (English)

    Mekala Subba Rao; Aleem Ahmed Khan; Nyamath Parveen; Mohammed Aejaz Habeeb; Chittor Mohammed Habibullah; Gopal Pande

    2008-01-01

    AIM: To enrich hepatic progenitors using epithelial cell adhesion molecule (EpCAH) as a marker from human fetal liver and investigate the expression of human leukocyte antigen (HLA) and their markers associated with hepatic progenitor cells.METHODS: EpCAM +ve cells were isolated usingmagnetic cell sorting (MACS) from human fetuses (n =10) at 15-25 wk gestation.Expression of markers for hepatic progenitors such as albumin,alpha-fetoprotein (AFP),CD29 (integrin β1),CD49f (integrin α6) and CD90 (Thy 1) was studied by using flow cytometry,immunocytochemistry and RT-PCR; HLA class Ⅰ (A,B,C) and class Ⅱ (DR) expression was studied by flow cytometry only.RESULTS: FACS analysis indicated that EpCAM +ve cells were positive for CD29,CD49f,CD90,CD34,HLA class Ⅰ,albumin and AFP but negative for HLA class Ⅱ (DR) and CD45.RT PCR showed that EpCAM +ve cells expressed liver epithelial markers (CK18),biliary specific marker (CK19) and hepatic markers (albumin,AFP).On immunocytochemical staining,EpCAH +ve cells were shown positive signals for CK18 and albumin.CONCLUSION: Our study suggests that these EpCAM +ve cells can be used as hepatic progenitors for cell transplantation with a minimum risk of alloreactivity and these cells may serve as a potential source for enrichment of hepatic progenitor.

  17. Multilineage potential of stable human mesenchymal stem cell line derived from fetal marrow.

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    Atsushi Nagai

    Full Text Available Human bone marrow contains two major cell types, hematopoietic stem cells (HSCs and mesenchymal stem cells (MSCs. MSCs possess self-renewal capacity and pluripotency defined by their ability to differentiate into osteoblasts, chondrocytes, adipocytes and muscle cells. MSCs are also known to differentiate into neurons and glial cells in vitro, and in vivo following transplantation into the brain of animal models of neurological disorders including ischemia and intracerebral hemorrhage (ICH stroke. In order to obtain sufficient number and homogeneous population of human MSCs, we have clonally isolated permanent and stable human MSC lines by transfecting primary cell cultures of fetal human bone marrow MSCs with a retroviral vector encoding v-myc gene. One of the cell lines, HM3.B10 (B10, was found to differentiate into neural cell types including neural stem cells, neurons, astrocytes and oligodendrocytes in vitro as shown by expression of genetic markers for neural stem cells (nestin and Musashi1, neurons (neurofilament protein, synapsin and MAP2, astrocytes (glial fibrillary acidic protein, GFAP and oligodendrocytes (myelin basic protein, MBP as determined by RT-PCR assay. In addition, B10 cells were found to differentiate into neural cell types as shown by immunocytochical demonstration of nestin (for neural stem cells, neurofilament protein and beta-tubulin III (neurons GFAP (astrocytes, and galactocerebroside (oligodendrocytes. Following brain transplantation in mouse ICH stroke model, B10 human MSCs integrate into host brain, survive, differentiate into neurons and astrocytes and induce behavioral improvement in the ICH animals. B10 human MSC cell line is not only a useful tool for the studies of organogenesis and specifically for the neurogenesis, but also provides a valuable source of cells for cell therapy studies in animal models of stroke and other neurological disorders.

  18. Validation of In utero Tractography of Human Fetal Commissural and Internal Capsule Fibers with Histological Structure Tensor Analysis

    OpenAIRE

    Mitter, Christian; Jakab, András; Peter C Brugger; Ricken, Gerda; Gruber, Gerlinde M.; Bettelheim, Dieter; Scharrer, Anke; Langs, Georg; Hainfellner, Johannes A.; Prayer, Daniela; Kasprian, Gregor

    2015-01-01

    Diffusion tensor imaging (DTI) and tractography offer the unique possibility to visualize the developing white matter macroanatomy of the human fetal brain in vivo and in utero and are currently under investigation for their potential use in the diagnosis of developmental pathologies of the human central nervous system. However, in order to establish in utero DTI as a clinical imaging tool, an independent comparison between macroscopic imaging and microscopic histology data in the same subjec...

  19. Validation of In utero Tractography of Human Fetal Commissural and Internal Capsule Fibers with Histological Structure Tensor Analysis.

    Science.gov (United States)

    Mitter, Christian; Jakab, András; Brugger, Peter C; Ricken, Gerda; Gruber, Gerlinde M; Bettelheim, Dieter; Scharrer, Anke; Langs, Georg; Hainfellner, Johannes A; Prayer, Daniela; Kasprian, Gregor

    2015-01-01

    Diffusion tensor imaging (DTI) and tractography offer the unique possibility to visualize the developing white matter macroanatomy of the human fetal brain in vivo and in utero and are currently under investigation for their potential use in the diagnosis of developmental pathologies of the human central nervous system. However, in order to establish in utero DTI as a clinical imaging tool, an independent comparison between macroscopic imaging and microscopic histology data in the same subject is needed. The present study aimed to cross-validate normal as well as abnormal in utero tractography results of commissural and internal capsule fibers in human fetal brains using postmortem histological structure tensor (ST) analysis. In utero tractography findings from two structurally unremarkable and five abnormal fetal brains were compared to the results of postmortem ST analysis applied to digitalized whole hemisphere sections of the same subjects. An approach to perform ST-based deterministic tractography in histological sections was implemented to overcome limitations in correlating in utero tractography to postmortem histology data. ST analysis and histology-based tractography of fetal brain sections enabled the direct assessment of the anisotropic organization and main fiber orientation of fetal telencephalic layers on a micro- and macroscopic scale, and validated in utero tractography results of corpus callosum and internal capsule fiber tracts. Cross-validation of abnormal in utero tractography results could be achieved in four subjects with agenesis of the corpus callosum (ACC) and in two cases with malformations of internal capsule fibers. In addition, potential limitations of current DTI-based in utero tractography could be demonstrated in several brain regions. Combining the three-dimensional nature of DTI-based in utero tractography with the microscopic resolution provided by histological ST analysis may ultimately facilitate a more complete morphologic

  20. Validation of In utero Tractography of Human Fetal Commissural and Internal Capsule Fibers with Histological Structure Tensor Analysis

    Science.gov (United States)

    Mitter, Christian; Jakab, András; Brugger, Peter C.; Ricken, Gerda; Gruber, Gerlinde M.; Bettelheim, Dieter; Scharrer, Anke; Langs, Georg; Hainfellner, Johannes A.; Prayer, Daniela; Kasprian, Gregor

    2015-01-01

    Diffusion tensor imaging (DTI) and tractography offer the unique possibility to visualize the developing white matter macroanatomy of the human fetal brain in vivo and in utero and are currently under investigation for their potential use in the diagnosis of developmental pathologies of the human central nervous system. However, in order to establish in utero DTI as a clinical imaging tool, an independent comparison between macroscopic imaging and microscopic histology data in the same subject is needed. The present study aimed to cross-validate normal as well as abnormal in utero tractography results of commissural and internal capsule fibers in human fetal brains using postmortem histological structure tensor (ST) analysis. In utero tractography findings from two structurally unremarkable and five abnormal fetal brains were compared to the results of postmortem ST analysis applied to digitalized whole hemisphere sections of the same subjects. An approach to perform ST-based deterministic tractography in histological sections was implemented to overcome limitations in correlating in utero tractography to postmortem histology data. ST analysis and histology-based tractography of fetal brain sections enabled the direct assessment of the anisotropic organization and main fiber orientation of fetal telencephalic layers on a micro- and macroscopic scale, and validated in utero tractography results of corpus callosum and internal capsule fiber tracts. Cross-validation of abnormal in utero tractography results could be achieved in four subjects with agenesis of the corpus callosum (ACC) and in two cases with malformations of internal capsule fibers. In addition, potential limitations of current DTI-based in utero tractography could be demonstrated in several brain regions. Combining the three-dimensional nature of DTI-based in utero tractography with the microscopic resolution provided by histological ST analysis may ultimately facilitate a more complete morphologic

  1. Validation of in utero tractography of human fetal commissural and internal capsule fibers with histological structure tensor analysis

    Directory of Open Access Journals (Sweden)

    Christian eMitter

    2015-12-01

    Full Text Available Diffusion tensor imaging (DTI and tractography offer the unique possibility to visualize the developing white matter macroanatomy of the human fetal brain in vivo and in utero and are currently under investigation for their potential use in the diagnosis of developmental pathologies of the human central nervous system. However, in order to establish in utero DTI as a clinical imaging tool, an independent comparison between macroscopic imaging and microscopic histology data in the same subject is needed. The present study aimed to cross-validate normal as well as abnormal in utero tractography results of commissural and internal capsule fibers in human fetal brains using postmortem histological structure tensor (ST analysis. In utero tractography findings from two structurally unremarkable and five abnormal fetal brains were compared to the results of postmortem ST analysis applied to digitalized whole hemisphere sections of the same subjects. An approach to perform ST-based deterministic tractography in histological sections was implemented to overcome limitations in correlating in utero tractography to postmortem histology data. ST analysis and histology-based tractography of fetal brain sections enabled the direct assessment of the anisotropic organization and main fiber orientation of fetal telencephalic layers on a micro- and macroscopic scale, and validated in utero tractography results of corpus callosum and internal capsule fiber tracts. Cross-validation of abnormal in utero tractography results could be achieved in four subjects with agenesis of the corpus callosum and in two cases with malformations of internal capsule fibers. In addition, potential limitations of current DTI-based in utero tractography could be demonstrated in several brain regions. Combining the three-dimensional nature of DTI-based in utero tractography with the microscopic resolution provided by histological ST analysis may ultimately facilitate a more complete

  2. Effects of disulfiram on apoptosis in PANC-1 human pancreatic cancer cell line

    Science.gov (United States)

    Dastjerdi, M. Nikbakht; Babazadeh, Z.; Rabbani, M.; Gharagozloo, M.; Esmaeili, A.; Narimani, M.

    2014-01-01

    Pancreatic carcinoma is currently considered as a rapidly progressive and fatal disease, and is typically diagnosed late in its natural course. It is characterized by a poor diagnosis and lack of response to conventional therapy. Recent studies have suggested that disulfiram (DSF), a member of the dithiocarbamate family, may have antitumor activity. This study aimed to evaluate the in vitro effect of DSF on apoptosis in human pancreatic cancerous cell line (PANC-1). PANC-1 cells were cultured and treated with DSF at doses of 5, 10, 13 μM for 24 h and apoptosis was measured. Methylation specific PCR (MS-PCR) and real-time quantitative PCR were carried out to detect the methylation pattern and to estimate the mRNA expression levels of RASSF1A, p21 and Bax. MS-PCR analysis demonstrated that no unmethylated band was apeared in PANC-1 cell line after DSF treatments. The real-time quantitative PCR results showed no significant mRNA expression for RASSF1A (p>0.05); whereas p21 and Bax expression were significantly (p<0.01) enhanced after treatment with DSF. The results of the current study indicated that DSF can induce appoptosis in PANC-1 through p21 and Bax pathway but not through RASSF1A. PMID:25657800

  3. THE AUTOCRINE REGULATORY EFFECT OF VASOACTIVE INTESTINAL PEPTIDE ON THE GROWTH OF HUMAN PANCREATIC CARCINOMA CELLS

    Institute of Scientific and Technical Information of China (English)

    陈元方; 陈潜; 陆国钧; 范振符; 钟守先

    1994-01-01

    In the present study, the effets of VIP on the growth of two human pancreatic carcinoma cell lines PU-PAH-1 and PANC-1 were determined using tritiated thymidine incorporation, VIP receptors, intracellular cAMP and polyamings were investlsa.ted, The results indicated that VIP at a concentcation of 10-8mol/L to 10-7 mol/L can significantly Stimulate the growth of PU-PAN-1 cells but not PANC-1 cells, This effect is dose-dependent and abolished by VIP receptor antagonist, [4-CI-Phe6 , Leu7] VIP, suggesting VIP receptors in PU-PAN-1 cells maymediate this effect. VIP can markedly elevate the levels of intracellular cAMP and polyammes in PU-PAN-1 cells,indicating that the growth-promoting effect stimulated by VIP may be via a rapid increase in the biosynth~es of cAMP and polyamines. In addition, the VIP-antibody ir2Libited the growth of PU-PAN-1 cells in serum-free culture medium. The results above suggested that VIP has an autoctine regulatory effect on this pancreatic carcinoma cell line(PU-PAN-1).

  4. Direct long-term effects of L-asparaginase on rat and human pancreatic islets

    DEFF Research Database (Denmark)

    Clausen, Niels; Nielsen, Jens Høiriis

    1989-01-01

    L-Asparaginase, an effective agent in the treatment of acute lymphoblastic leukemia, may induce a diabetic state. The pathogenesis of the diabetogenic effect was studied in cultured pancreatic islets. Mean serum concentrations in three children with acute lymphoblastic leukemia were 2.4 U/mL (range...... 1.4-4.5) before and 31.5 U/mL (range 18.6-51.8) immediately after an intravenous injection of 1000 U/kg L-asparaginase. Glucose-induced insulin release from pancreatic islets of rat and man was measured after 3 and 7 days of culture in media with or without clinically relevant concentrations...... of Escherichia coli L-asparaginase (0.01-100 U/mL). After culture, the remaining insulin, glucagon, and DNA in the islets were determined. After 7 days of culture of adult rat or human islets, both the accumulation of insulin in the medium and the content of insulin and glucagon in the islets were significantly...

  5. Fetal functional imaging portrays heterogeneous development of emerging human brain networks.

    Science.gov (United States)

    Jakab, András; Schwartz, Ernst; Kasprian, Gregor; Gruber, Gerlinde M; Prayer, Daniela; Schöpf, Veronika; Langs, Georg

    2014-01-01

    The functional connectivity architecture of the adult human brain enables complex cognitive processes, and exhibits a remarkably complex structure shared across individuals. We are only beginning to understand its heterogeneous structure, ranging from a strongly hierarchical organization in sensorimotor areas to widely distributed networks in areas such as the parieto-frontal cortex. Our study relied on the functional magnetic resonance imaging (fMRI) data of 32 fetuses with no detectable morphological abnormalities. After adapting functional magnetic resonance acquisition, motion correction, and nuisance signal reduction procedures of resting-state functional data analysis to fetuses, we extracted neural activity information for major cortical and subcortical structures. Resting fMRI networks were observed for increasing regional functional connectivity from 21st to 38th gestational weeks (GWs) with a network-based statistical inference approach. The overall connectivity network, short range, and interhemispheric connections showed sigmoid expansion curve peaking at the 26-29 GW. In contrast, long-range connections exhibited linear increase with no periods of peaking development. Region-specific increase of functional signal synchrony followed a sequence of occipital (peak: 24.8 GW), temporal (peak: 26 GW), frontal (peak: 26.4 GW), and parietal expansion (peak: 27.5 GW). We successfully adapted functional neuroimaging and image post-processing approaches to correlate macroscopical scale activations in the fetal brain with gestational age. This in vivo study reflects the fact that the mid-fetal period hosts events that cause the architecture of the brain circuitry to mature, which presumably manifests in increasing strength of intra- and interhemispheric functional macro connectivity.

  6. Fetal functional imaging portrays heterogeneous development of emerging human brain networks

    Science.gov (United States)

    Jakab, András; Schwartz, Ernst; Kasprian, Gregor; Gruber, Gerlinde M.; Prayer, Daniela; Schöpf, Veronika; Langs, Georg

    2014-01-01

    The functional connectivity architecture of the adult human brain enables complex cognitive processes, and exhibits a remarkably complex structure shared across individuals. We are only beginning to understand its heterogeneous structure, ranging from a strongly hierarchical organization in sensorimotor areas to widely distributed networks in areas such as the parieto-frontal cortex. Our study relied on the functional magnetic resonance imaging (fMRI) data of 32 fetuses with no detectable morphological abnormalities. After adapting functional magnetic resonance acquisition, motion correction, and nuisance signal reduction procedures of resting-state functional data analysis to fetuses, we extracted neural activity information for major cortical and subcortical structures. Resting fMRI networks were observed for increasing regional functional connectivity from 21st to 38th gestational weeks (GWs) with a network-based statistical inference approach. The overall connectivity network, short range, and interhemispheric connections showed sigmoid expansion curve peaking at the 26–29 GW. In contrast, long-range connections exhibited linear increase with no periods of peaking development. Region-specific increase of functional signal synchrony followed a sequence of occipital (peak: 24.8 GW), temporal (peak: 26 GW), frontal (peak: 26.4 GW), and parietal expansion (peak: 27.5 GW). We successfully adapted functional neuroimaging and image post-processing approaches to correlate macroscopical scale activations in the fetal brain with gestational age. This in vivo study reflects the fact that the mid-fetal period hosts events that cause the architecture of the brain circuitry to mature, which presumably manifests in increasing strength of intra- and interhemispheric functional macro connectivity. PMID:25374531

  7. IMMUNE MODULATORY EFFECTS of HUMAN CHORIONIC GONADOTROPIN on DENDRITIC CELLS SUPPORTING FETAL SURVIVAL in MURINE PREGNANCY

    Directory of Open Access Journals (Sweden)

    Dominique Dauven

    2016-11-01

    Full Text Available Dendritic cells (DCs are critically involved in the determination of immunity versus tolerance. Hence, DCs are key regulators of immune responses either favoring or disfavoring fetal survival. Several factors were proposed to modulate DC phenotype and function during preg-nancy. Here, we studied whether the pregnancy hormone human Chorionic Gonadotropin (hCG is involved in DC regulation.In vitro, bone-marrow-derived DCs (BMDCs were stimulated in the presence or absence of urine-purified (uhCG or recombinant hCG (rhCG preparations. Subsequently, BMDC matu-ration was assessed. Cytokine secretion of activated BMDCs and their capability to enforce TH1, TH2, TH17 or Treg cell differentiation was determined after rhCG treatment. Moreover, the in vivo potential of hCG-modulated BMDCs to influence pregnancy outcome, Treg cell number and local cytokine expression was evaluated after adoptive transfer in a murine abor-tion-prone model before and after conception. Both hCG preparations impaired the maturation process of BMDCs. rhCG treatment did nei-ther alter cytokine secretion by BMDCs nor their ability to drive TH1, TH2 or TH17 differen-tiation. rhCG-treated BMDCs augmented the number of Treg cells within the T cell popula-tion. Adoptive transfer of rhCG-treated BMDCs after conception did not influence pregnancy outcome. However, transfer of hCG-treated BMDCs prior to mating had a protective effect on pregnancy. This positive effect was accompanied by increased Treg cell numbers and decidual IL-10 and TGF-β expression. Our results unveil the importance of hCG in retaining DCs in a tolerogenic state, thereby promoting Treg cell increment and supporting fetal survival.

  8. Immune Modulatory Effects of Human Chorionic Gonadotropin on Dendritic Cells Supporting Fetal Survival in Murine Pregnancy

    Science.gov (United States)

    Dauven, Dominique; Ehrentraut, Stefanie; Langwisch, Stefanie; Zenclussen, Ana Claudia; Schumacher, Anne

    2016-01-01

    Dendritic cells (DCs) are critically involved in the determination of immunity vs. tolerance. Hence, DCs are key regulators of immune responses either favoring or disfavoring fetal survival. Several factors were proposed to modulate DC phenotype and function during pregnancy. Here, we studied whether the pregnancy hormone human chorionic gonadotropin (hCG) is involved in DC regulation. In vitro, bone marrow-derived DCs (BMDCs) were stimulated in the presence or absence of urine-purified or recombinant hCG (rhCG) preparations. Subsequently, BMDC maturation was assessed. Cytokine secretion of activated BMDCs and their capability to enforce TH1, TH2, TH17, or Treg cell differentiation was determined after rhCG treatment. Moreover, the in vivo potential of hCG-modulated BMDCs to influence pregnancy outcome, Treg cell number, and local cytokine expression was evaluated after adoptive transfer in a murine abortion-prone model before and after conception. Both hCG preparations impaired the maturation process of BMDCs. rhCG treatment did neither alter cytokine secretion by BMDCs nor their ability to drive TH1, TH2, or TH17 differentiation. rhCG-treated BMDCs augmented the number of Treg cells within the T cell population. Adoptive transfer of rhCG-treated BMDCs after conception did not influence pregnancy outcome. However, transfer of hCG-treated BMDCs prior to mating had a protective effect on pregnancy. This positive effect was accompanied by increased Treg cell numbers and decidual IL-10 and TGF-β expression. Our results unveil the importance of hCG in retaining DCs in a tolerogenic state, thereby promoting Treg cell increment and supporting fetal survival. PMID:27895621

  9. Induction of human pancreatic beta cell replication by inhibitors of dual specificity tyrosine regulated kinase

    Science.gov (United States)

    Wang, Peng; Alvarez-Perez, Juan-Carlos; Felsenfeld, Dan P.; Liu, Hongtao; Sivendran, Sharmila; Bender, Aaron; Kumar, Anil; Sanchez, Roberto; Scott, Donald K.; Garcia-Ocaña, Adolfo; Stewart, Andrew F.

    2015-01-01

    Types 1 and 2 diabetes affect some 380 million people worldwide. Both result ultimately from a deficiency of functional pancreatic insulin-producing beta cells. Beta cells proliferate in humans during a brief temporal window beginning around the time of birth, with peak beta cell labeling indices achieving approximately 2% in first year of life1-4. In embryonic life and after early childhood, beta cell replication rates are very low. While beta cell expansion seems an obvious therapeutic approach to beta cell deficiency, adult human beta cells have proven recalcitrant to such efforts1-8. Hence, there remains an urgent need for diabetes therapeutic agents that can induce regeneration and expansion of adult human beta cells in vivo or ex vivo. Here, we report the results of a high-throughput small molecule screen (HTS) revealing a novel class of human beta cell mitogenic compounds, analogues of the small molecule, harmine. We also define dual specificity tyrosine-regulated kinase-1a (DYRK1A) as the likely target of harmine, and the Nuclear Factors of activated T-cells (NFAT) family of transcription factors as likely mediators of human beta cell proliferation as well as beta cell differentiation. These observations suggest that harmine analogues (“harmalogs”) may have unique therapeutic promise for human diabetes therapy. Enhancing potency and beta cell specificity are important future challenges. PMID:25751815

  10. The novel mTORC1/2 dual inhibitor INK-128 suppresses survival and proliferation of primary and transformed human pancreatic cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Lou, Hai-zhou [Department of Medical Oncology, Sir Run Run Shaw Hospital, College of Medicine, Zhejiang University, Hangzhou 310016 (China); Weng, Xiao-chuan [Department of Anesthesiology, Hangzhou Xia-sha Hospital, Hangzhou 310018 (China); Pan, Hong-ming; Pan, Qin [Department of Medical Oncology, Sir Run Run Shaw Hospital, College of Medicine, Zhejiang University, Hangzhou 310016 (China); Sun, Peng [Department of Medical Oncology, Sun Yat-Sen University Cancer Center, Guangzhou 510060 (China); Liu, Li-li [Department of Medical Oncology, Sir Run Run Shaw Hospital, College of Medicine, Zhejiang University, Hangzhou 310016 (China); Chen, Bin, E-mail: chenbinhangzhou126@126.com [Department of Hepatopancreatobiliary Surgery, First People’s Hospital of Hangzhou, Hangzhou 310006 (China)

    2014-07-25

    Highlights: • INK-128 inhibits the survival and growth of human pancreatic cancer cells. • INK-128 induced pancreatic cancer cell apoptosis and necrosis simultaneously. • INK-128 blocks mTORC1/2 activation simultaneously in pancreatic cancer cells. • INK-128 down-regulates cyclin D1 and causes pancreatic cancer cell cycle arrest. • INK-128 significantly increases sensitivity of pancreatic cancer cells to gemcitabine. - Abstract: Pancreatic cancer has one of worst prognosis among all human malignancies around the world, the development of novel and more efficient anti-cancer agents against this disease is urgent. In the current study, we tested the potential effect of INK-128, a novel mammalian target of rapamycin (mTOR) complex 1 and 2 (mTORC1/2) dual inhibitor, against pancreatic cancer cells in vitro. Our results demonstrated that INK-128 concentration- and time-dependently inhibited the survival and growth of pancreatic cancer cells (both primary cells and transformed cells). INK-128 induced pancreatic cancer cell apoptosis and necrosis simultaneously. Further, INK-128 dramatically inhibited phosphorylation of 4E-binding protein 1 (4E-BP1), ribosomal S6 kinase 1 (S6K1) and Akt at Ser 473 in pancreatic cancer cells. Meanwhile, it downregulated cyclin D1 expression and caused cell cycle arrest. Finally, we found that a low concentration of INK-128 significantly increased the sensitivity of pancreatic cancer cells to gemcitabine. Together, our in vitro results suggest that INK-128 might be further investigated as a novel anti-cancer agent or chemo-adjuvant for pancreatic cancer treatment.

  11. Calcium sensing receptor suppresses human pancreatic tumorigenesis through a novel NCX1/Ca(2+)/β-catenin signaling pathway.

    Science.gov (United States)

    Tang, Bo; Chow, Jimmy Y C; Dong, Tobias Xiao; Yang, Shi-Ming; Lu, De-Sheng; Carethers, John M; Dong, Hui

    2016-07-10

    The calcium sensing receptor (CaSR) is functionally expressed in normal human pancreases, but its pathological role in pancreatic tumorigenesis is currently unknown. We sought to investigate the role of CaSR in pancreatic cancer (PC) and the underlying molecular mechanisms. We revealed that the expression of CaSR was consistently downregulated in the primary cancer tissues from PC patients, which was correlated with tumor size, differentiation and poor survival of the patients. CaSR activation markedly suppressed pancreatic tumorigenesis in vitro and in vivo likely through the Ca(2+) entry mode of Na(+)/Ca(2+) exchanger 1 (NCX1) to induce Ca(2+) entry into PC cells. Moreover, NCX1-mediated Ca(2+) entry resulted in Ca(2+)-dependent inhibition of β-catenin signaling in PC cells, eventually leading to the inhibition of pancreatic tumorigenesis. Collectively, we demonstrate for the first time that CaSR exerts a suppressive function in pancreatic tumorigenesis through a novel NCX1/Ca(2+)/β-catenin signaling pathway. Targeting this specific signaling pathway could be a potential therapeutic strategy for PC.

  12. LLL12 inhibits endogenous and exogenous interleukin-6-induced STAT3 phosphorylation in human pancreatic cancer cells.

    Science.gov (United States)

    Liu, Aiguo; Liu, Yan; Li, Pui-Kai; Li, Chenglong; Lin, Jiayuh

    2011-06-01

    Pancreatic cancer is one of the most serious types of cancer, with a five-year survival rate at only 6%. There is a critical need to develop more effective treatments for pancreatic cancer. Growing evidence shows that chronic inflammation plays a crucial role in tumor initiation and progression. Here we demonstrated that the endogenous expression of the inflammatory cytokine interleukin-6 (IL-6) correlates with signal transducer and activator of transcription 3 (STAT3) phosphorylation in human pancreatic cancer cells. Inhibition of the endogenous IL-6/STAT3 pathway reduces cell viability. Exogenous IL-6 induces STAT3 phosphorylation, but differently induces phosphorylation of STAT3 upstream kinases, Janus kinase 1(JAK1), JAK2, and tyrosine kinase 2 (TYK2). Interestingly, LLL12, a nonpeptide, cell-permeable small molecule, selectively blocked exogenous IL-6-induced STAT3 phosphorylation and nuclear translocation in both PANC-1 and ASPC-1 pancreatic cancer cell lines independently of the phosphorylation of JAK1, JAK2, and TYK2. These results suggest that the inhibition of endogenous and exogenous IL-6-mediated STAT3 signaling may be a potential therapeutic approach for pancreatic cancer.

  13. A developmental stage-specific switch from DAZL to BOLL occurs during fetal oogenesis in humans, but not mice.

    Directory of Open Access Journals (Sweden)

    Jing He

    Full Text Available The Deleted in Azoospermia gene family encodes three germ cell-specific RNA-binding proteins (DAZ, DAZL and BOLL that are essential for gametogenesis in diverse species. Targeted disruption of Boll in mice causes male-specific spermiogenic defects, but females are apparently fertile. Overexpression of human BOLL promotes the derivation of germ cell-like cells from genetically female (XX, but not male (XY human ES cells however, suggesting a functional role for BOLL in regulating female gametogenesis in humans. Whether BOLL is expressed during oogenesis in mammals also remains unclear. We have therefore investigated the expression of BOLL during fetal oogenesis in humans and mice. We demonstrate that BOLL protein is expressed in the germ cells of the human fetal ovary, at a later developmental stage than, and almost mutually-exclusive to, the expression of DAZL. Strikingly, BOLL is downregulated, and DAZL re-expressed, as primordial follicles form, revealing BOLL expression to be restricted to a narrow window during fetal oogenesis. By quantifying the extent of co-expression of DAZL and BOLL with markers of meiosis, we show that this window likely corresponds to the later stages of meiotic prophase I. Finally, we demonstrate that Boll is also transiently expressed during oogenesis in the fetal mouse ovary, but is simultaneously co-expressed within the same germ cells as Dazl. These data reveal significant similarities and differences between the expression of BOLL homologues during oogenesis in humans and mice, and raise questions as to the validity of the Boll(-/- mouse as a model for understanding BOLL function during human oogenesis.

  14. Genes expressed in specific areas of the human fetal cerebral cortex display distinct patterns of evolution.

    Directory of Open Access Journals (Sweden)

    Nelle Lambert

    Full Text Available The developmental mechanisms through which the cerebral cortex increased in size and complexity during primate evolution are essentially unknown. To uncover genetic networks active in the developing cerebral cortex, we combined three-dimensional reconstruction of human fetal brains at midgestation and whole genome expression profiling. This novel approach enabled transcriptional characterization of neurons from accurately defined cortical regions containing presumptive Broca and Wernicke language areas, as well as surrounding associative areas. We identified hundreds of genes displaying differential expression between the two regions, but no significant difference in gene expression between left and right hemispheres. Validation by qRTPCR and in situ hybridization confirmed the robustness of our approach and revealed novel patterns of area- and layer-specific expression throughout the developing cortex. Genes differentially expressed between cortical areas were significantly associated with fast-evolving non-coding sequences harboring human-specific substitutions that could lead to divergence in their repertoires of transcription factor binding sites. Strikingly, while some of these sequences were accelerated in the human lineage only, many others were accelerated in chimpanzee and/or mouse lineages, indicating that genes important for cortical development may be particularly prone to changes in transcriptional regulation across mammals. Genes differentially expressed between cortical regions were also enriched for transcriptional targets of FoxP2, a key gene for the acquisition of language abilities in humans. Our findings point to a subset of genes with a unique combination of cortical areal expression and evolutionary patterns, suggesting that they play important roles in the transcriptional network underlying human-specific neural traits.

  15. Conditionally immortalized human pancreatic stellate cell lines demonstrate enhanced proliferation and migration in response to IGF-I

    Energy Technology Data Exchange (ETDEWEB)

    Rosendahl, Ann H., E-mail: ann.rosendahl@med.lu.se [Lund University, Department of Clinical Sciences Lund, Division of Surgery, Lund (Sweden); Lund University and Skåne University Hospital, Department of Clinical Sciences Lund, Division of Oncology and Pathology, Lund (Sweden); Gundewar, Chinmay; Said Hilmersson, Katarzyna [Lund University, Department of Clinical Sciences Lund, Division of Surgery, Lund (Sweden); Ni, Lan; Saleem, Moin A. [University of Bristol, School of Clinical Sciences, Children' s Renal Unit and Academic Renal Unit, Bristol (United Kingdom); Andersson, Roland [Lund University, Department of Clinical Sciences Lund, Division of Surgery, Lund (Sweden)

    2015-01-15

    Pancreatic stellate cells (PSCs) play a key role in the dense desmoplastic stroma associated with pancreatic ductal adenocarcinoma. Studies on human PSCs have been minimal due to difficulty in maintaining primary PSC in culture. We have generated the first conditionally immortalized human non-tumor (NPSC) and tumor-derived (TPSC) pancreatic stellate cells via transformation with the temperature-sensitive SV40 large T antigen and human telomerase (hTERT). These cells proliferate at 33°C. After transfer to 37°C, the SV40LT is switched off and the cells regain their primary PSC phenotype and growth characteristics. NPSC contained cytoplasmic vitamin A-storing lipid droplets, while both NPSC and TPSC expressed the characteristic markers αSMA, vimentin, desmin and GFAP. Proteome array analysis revealed that of the 55 evaluated proteins, 27 (49%) were upregulated ≥3-fold in TPSC compared to NPSC, including uPA, pentraxin-3, endoglin and endothelin-1. Two insulin-like growth factor binding proteins (IGFBPs) were inversely expressed. Although discordant IGFBP-2 and IGFBP-3 levels, IGF-I was found to stimulate proliferation of both NPSC and TPSC. Both basal and IGF-I stimulated motility was significantly enhanced in TPSC compared to NPSC. In conclusion, these cells provide a unique resource that will facilitate further study of the active stroma compartment associated with pancreatic cancer. - Highlights: • Generation of human conditionally immortalized human pancreatic stellate cell lines. • Temperature-sensitive SV40LT allows switch to primary PSC phenotype characteristics. • Proteome profiling revealed distinct expression patterns between TPSC and NPSC. • Enhanced IGF-I-stimulated proliferation and motility by TPSC compared to NPSC.

  16. Expression of nestin in human fetal lung%巢蛋白在人胚胎肺中的表达

    Institute of Scientific and Technical Information of China (English)

    董丽萍; 邹鹰; 黄河; 张喆; 魏楚蓉; 伍赶球

    2011-01-01

    目的:研究不同时期人胚胎肺中巢蛋白的表达变化,探讨巢蛋白在肺发育过程中的作用.方法:取3~8月人胚胎肺组织,常规石蜡切片,采用免疫组织化学法检测巢蛋白在胚胎肺中的表达及分布.结果:各胎龄段(3~8月)肺均有巢蛋白的表达,巢蛋白阳性细胞主要分布于肺内支气管旁和血管中,而在支气管上皮中未发现阳性细胞.结论:正常人胚胎肺中有巢蛋白的表达,随着胎龄的增加,巢蛋白表达量下降.%Objective To study the changes of expression of nestin in fetal lung and to explore the function of nestin during lung development in human. Methods Fetal lung tissues from 3-8 months' embryos were dissected and fixed for paraffin section. Immunohistochemical staining was proceeded to examine the expression level and distribution of nestin in fetal lung. Results Nestin was highly expressed strongly throughout lung in human fetal of 3 - 8 months' embryos. Nestin-positive cells distributed in pulmonary, entobronchus and blood vessels, but it was not found in bronchial epithelium. Conclusion Nestin widely expresses in normal human fetal lung. The level of expression decreases with the increase of gestational age.

  17. Fate and Development of Human Vomeronasal Organ - A Microscopic Fetal Study.

    Science.gov (United States)

    Vasuki, A K Manicka; Fenn, T K Aleyemma; Devi, M Nirmala; Hebzibah, T Deborah Joy; Jamuna, M; Sundaram, K Kalyana

    2016-03-01

    The existence of Vomeronasal organ in human is a controversial subject. Presence of Vomeronasal organ and its structure was not reported in standard text books. The presence of Vomeronasal organ in fetal life is doubtful. Hence identification of the organ by histological examination was planned. A study was conducted on resected specimens of nasal septum obtained from 45 spontaneously aborted fetuses from Obstetrics and Gynaecology department, PSG Institute of Medical Sciences and Research, Coimbatore, after ethical clearance. The histological structure of Vomeronasal organ was observed from 11 weeks old fetus. The epithelial lining of the organ, presence of cilia, presence of lamina propria, acini and the blood vessel and the types of cells were observed. The organ was lined by pseudostratified columnar epithelium. The organ showed Lamina propria with serous acini from 18 weeks fetus. Vomeronasal duct opening into the nasal cavity and three types of cells were observed in 28 weeks fetus. Knowledge about the persistence of Vomeronasal organ in fetuses and its structure need to be known. The organ may be found as a putative pit posterior to anterior nasal spine. The organ may be damaged in nasal septal surgeries and nasal endoscopic procedures. The organ may not be seen on gross examination in all human fetuses and cadavers.

  18. Differential Responses of Human Fetal Brain Neural Stem Cells to Zika Virus Infection

    Directory of Open Access Journals (Sweden)

    Erica L. McGrath

    2017-03-01

    Full Text Available Zika virus (ZIKV infection causes microcephaly in a subset of infants born to infected pregnant mothers. It is unknown whether human individual differences contribute to differential susceptibility of ZIKV-related neuropathology. Here, we use an Asian-lineage ZIKV strain, isolated from the 2015 Mexican outbreak (Mex1-7, to infect primary human neural stem cells (hNSCs originally derived from three individual fetal brains. All three strains of hNSCs exhibited similar rates of Mex1-7 infection and reduced proliferation. However, Mex1-7 decreased neuronal differentiation in only two of the three stem cell strains. Correspondingly, ZIKA-mediated transcriptome alterations were similar in these two strains but significantly different from that of the third strain with no ZIKV-induced neuronal reduction. This study thus confirms that an Asian-lineage ZIKV strain infects primary hNSCs and demonstrates a cell-strain-dependent response of hNSCs to ZIKV infection.

  19. Morphometric Development of Sphincter of Oddi in Human Fetuses During Fetal Period: Microscopic Study

    Directory of Open Access Journals (Sweden)

    E. Hilal Evcil

    2012-09-01

    Full Text Available Objective: In this study, morphometric developments of the sphincter of Oddi in human fetuses were observed. Material and Methods: We observed 113 human fetuses consisting of 67 male and 46 female subjects, whose ages varied between 14 to 40 weeks who showed no signs of any pathology or anomaly externally. The common external measurements of fetuses were carried out, followed by abdominal dissection to determine where the sphincters of Oddi were localized within the duodenum and pancreas. Histological specimens of tissue samples were gathered from the inner wall of the duodenum where it was assumed that the sphincters of Oddi had been localized. The parameters of total external diameters, lumen diameters, wall thickness, diameters of ductus choledochus and ductus pancreaticus, and the distance between these two structures, which are also known as the origins of the sphincter of Oddi, were measured by using a light microscope. The standard deviations of the measurements were calculated for each gestational week and trimester. Results: The calculations suggested that there were statistically significant correlations between gestational age and all of the other parameters with the exception of the ductus choledochus (p0.05.Conclusion: The data we collected in our study were considered as useful for the evaluation of the development of the sphincter of Oddi area and fetal stage.

  20. Species-Related Differences in the Proteome of Rat and Human Pancreatic Beta Cells

    Directory of Open Access Journals (Sweden)

    G. A. Martens

    2015-01-01

    Full Text Available The core proteomes of human and rat pancreatic beta cells were compared by label-free LC-MS/MS: this resulted in quantification of relative molar abundances of 707 proteins belonging to functional pathways of intermediary metabolism, protein synthesis, and cytoskeleton. Relative molar abundances were conserved both within and between pathways enabling the selection of a housekeeping network for geometric normalization and the analysis of potentially relevant differential expressions. Human beta cells differed from rat beta cells in their lower level of enzymes involved in glucose sensing (MDH1, PC, and ACLY and upregulation of lysosomal enzymes. Human cells also expressed more heat shock proteins and radical scavenging systems: apart from SOD2, they expressed high levels of H2O2-scavenger peroxiredoxin 3 (PRDX3, confirmed by microarray, Western blotting, and microscopy. Besides conferring lower susceptibility to oxidative stress to human cells PRDX3 might also play a role in physiological redox regulation as, in rat, its expression was restricted to a beta cell subset with higher metabolic glucose responsiveness. In conclusion, although their core proteomic architecture is conserved, human and rat beta cells differ in their molar expression of key enzymes involved in glucose sensing and redox control.

  1. The human fetal lymphocyte lineage: identification by CD27 and LIN28B expression in B cell progenitors

    Science.gov (United States)

    McWilliams, Laurie; Su, Kuei-Ying; Liang, Xiaoe; Liao, Dongmei; Floyd, Serina; Amos, Joshua; Moody, M. Anthony; Kelsoe, Garnett; Kuraoka, Masayuki

    2013-01-01

    CD27, a member of the TNFR superfamily, is used to identify human memory B cells. Nonetheless, CD27+ B cells are present in patients with HIGM1 syndrome who are unable to generate GCs or memory B cells. CD27+IgD+ fetal B cells are present in umbilical cord blood, and CD27 may also be a marker of the human B1-like B cells. To define the origin of naïve CD27+IgD+ human B cells, we studied B cell development in both fetal and adult tissues. In human FL, most CD19+ cells coexpressed CD10, a marker of human developing B cells. Some CD19+CD10+ B cells expressed CD27, and these fetal CD27+ cells were present in the pro-B, pre-B, and immature/transitional B cell compartments. Lower frequencies of phenotypically identical cells were also identified in adult BM. CD27+ pro-B, pre-B, and immature/transitional B cells expressed recombination activating gene-1, terminal deoxynucleotidyl transferase and Vpre-B mRNA comparably to their CD27− counterparts. CD27+ and CD27− developing B cells showed similar Ig heavy chain gene usage with low levels of mutations, suggesting that CD27+ developing B cells are distinct from mutated memory B cells. Despite these similarities, CD27+ developing B cells differed from CD27− developing B cells by their increased expression of LIN28B, a transcription factor associated with the fetal lymphoid lineages of mice. Furthermore, CD27+ pro-B cells efficiently generated IgM+IgD+ immature/transitional B cells in vitro. Our observations suggest that CD27 expression during B cell development identifies a physiologic state or lineage for human B cell development distinct from the memory B cell compartment. PMID:23901121

  2. Adult Human Biliary Tree Stem Cells Differentiate to β-Pancreatic Islet Cells by Treatment with a Recombinant Human Pdx1 Peptide.

    Directory of Open Access Journals (Sweden)

    Vincenzo Cardinale

    Full Text Available Generation of β-pancreatic cells represents a major goal in research. The aim of this study was to explore a protein-based strategy to induce differentiation of human biliary tree stem cells (hBTSCs towards β-pancreatic cells. A plasmid containing the sequence of the human pancreatic and duodenal homeobox 1 (PDX1 has been expressed in E. coli. Epithelial-Cell-Adhesion-Molecule positive hBTSCs or mature human hepatocyte cell line, HepG2, were grown in medium to which Pdx1 peptide was added. Differentiation toward pancreatic islet cells were evaluated by the expression of the β-cell transcription factors, Pdx1 and musculoapo-neurotic fibrosarcoma oncogene homolog A, and of the pancreatic hormones, insulin, glucagon, and somatostatin, investigated by real time polymerase chain reaction, western blot, light microscopy and immunofluorescence. C-peptide secretion in response to high glucose was also measured. Results indicated how purified Pdx1 protein corresponding to the primary structure of the human Pdx1 by mass spectroscopy was efficiently produced in bacteria, and transduced into hBTSCs. Pdx1 exposure triggered the expression of both intermediate and mature stage β-cell differentiation markers only in hBTSCs but not in HepG2 cell line. Furthermore, hBTSCs exposed to Pdx1 showed up-regulation of insulin, glucagon and somatostatin genes and formation of 3-dimensional islet-like structures intensely positive for insulin and glucagon. Finally, Pdx1-induced islet-like structures exhibited glucose-regulated C-peptide secretion. In conclusion, the human Pdx1 is highly effective in triggering hBTSC differentiation toward functional β-pancreatic cells.

  3. Expression, biosynthesis and release of preadipocyte factor-1/ delta-like protein/fetal antigen-1 in pancreatic beta-cells

    DEFF Research Database (Denmark)

    Friedrichsen, B N; Carlsson, C; Møldrup, Annette

    2003-01-01

    Preadipocyte factor-1 (Pref-1)/delta-like protein/fetal antigen-1 (FA1) is a member of the epidermal growth factor-like family. It is widely expressed in embryonic tissues, whereas in adults it is confined to the adrenal gland, the anterior pituitary, the endocrine pancreas, the testis and the ov......Preadipocyte factor-1 (Pref-1)/delta-like protein/fetal antigen-1 (FA1) is a member of the epidermal growth factor-like family. It is widely expressed in embryonic tissues, whereas in adults it is confined to the adrenal gland, the anterior pituitary, the endocrine pancreas, the testis...

  4. Lipid-Conjugation of Endogenous Neuropeptides: Improved Biotherapy against Human Pancreatic Cancer.

    Science.gov (United States)

    Gopalakrishnan, Gopakumar; Lepetre, Sinda; Maksimenko, Andrei; Mura, Simona; Desmaële, Didier; Couvreur, Patrick

    2015-05-01

    Neuropeptides are small neuronal signaling molecules that act as neuromodulators for a variety of neural functions including analgesia, reproduction, social behavior, learning, and memory. One of the endogenous neuropeptides-Met-Enkephalin (Met-Enk), has been shown to display an inhibitory effect on cell proliferation and differentiation. Here, a novel lipid-modification approach is shown to create a small library of neuropeptides that will allow increased bioavailability and plasma stability after systemic administration. It is demonstrated, on an experimental model of human pancreatic adenocarcinoma, that lipid conjugation of Met-Enk enhances its tumor suppression efficacy compared to its nonlipidated counterparts, both in vitro and in vivo. More strikingly, the in vivo studies show that a combination therapy with a reduced concentration of Gemcitabine has suppressed the tumor growth considerably even three weeks after the last treatment.

  5. A nuclear-directed human pancreatic ribonuclease (PE5) targets the metabolic phenotype of cancer cells.

    Science.gov (United States)

    Vert, Anna; Castro, Jessica; Ribó, Marc; Benito, Antoni; Vilanova, Maria

    2016-04-05

    Ribonucleases represent a new class of antitumor RNA-damaging drugs. However, many wild-type members of the vertebrate secreted ribonuclease family are not cytotoxic because they are not able to evade the cytosolic ribonuclease inhibitor. We previously engineered the human pancreatic ribonuclease to direct it to the cell nucleus where the inhibitor is not present. The best characterized variant is PE5 that kills cancer cells through apoptosis mediated by the p21(WAF1/CIP1) induction and the inactivation of JNK. Here, we have used microarray-derived transcriptional profiling to identify PE5 regulated genes on the NCI/ADR-RES ovarian cancer cell line. RT-qPCR analyses have confirmed the expression microarray findings. The results show that PE5 cause pleiotropic effects. Among them, it is remarkable the down-regulation of multiple genes that code for enzymes involved in deregulated metabolic pathways in cancer cells.

  6. Phenotypic and functional analysis of human fetal liver hematopoietic stem cells in culture.

    Science.gov (United States)

    Rollini, Pierre; Faes-Van't Hull, Eveline; Kaiser, Stefan; Kapp, Ursula; Leyvraz, Serge

    2007-04-01

    Steady-state hematopoiesis and hematopoietic transplantation rely on the unique potential of stem cells to undergo both self-renewal and multilineage differentiation. Fetal liver (FL) represents a promising alternative source of hematopoietic stem cells (HSCs), but limited by the total cell number obtained in a typical harvest. We reported that human FL nonobese diabetic/severe combined immunodeficient (NOD/SCID) repopulating cells (SRCs) could be expanded under simple stroma-free culture conditions. Here, we sought to further characterize FL HSC/SRCs phenotypically and functionally before and following culture. Unexpanded or cultured FL cell suspensions were separated into various subpopulations. These were tested for long-term culture potential and for in vivo repopulating function following transplantation into NOD/SCID mice. We found that upon culture of human FL cells, a tight association between classical stem cell phenotypes, such as CD34(+) /CD38(-) and/or side population, and NOD/SCID repopulating function was lost, as observed with other sources. Although SRC activity before and following culture consistently correlated with the presence of a CD34(+) cell population, we provide evidence that, contrary to umbilical cord blood and adult sources, stem cells present in both CD34(+) and CD34(-) FL populations can sustain long-term hematopoietic cultures. Furthermore, upon additional culture, CD34-depleted cell suspensions, devoid of SRCs, regenerated a population of CD34(+) cells possessing SRC function. Our studies suggest that compared to neonatal and adult sources, the phenotypical characteristics of putative human FL HSCs may be less strictly defined, and reinforce the accumulated evidence that human FL represents a unique, valuable alternative and highly proliferative source of HSCs for clinical applications.

  7. Effect of gemcitabine on the expression of apoptosis-related genes in human pancreatic cancer cells

    Institute of Scientific and Technical Information of China (English)

    Pei-Hong Jiang; Yoshiharu Motoo; Norio Sawabu; Toshinari Minamoto

    2006-01-01

    AIM: To investigate the expression of genes involved in the gemcitabine-induced cytotoxicity in human pancreatic cancer cells.METHODS: A human pancreatic cancer cell line,PANC-1, was cultured. 1×104 PANC-1 cells were plated in 96-well microtiter plates. After being incubated for 24 h,gemcitabine was added to the medium at concentrations ranging 2.5 -1 000 mg/L. The AlamarBlue dye method was used for cell growth analysis. DNA fragmentation was quantitatively assayed using a DNA fragmentation enzyme-linked immunosorbent assay (ELISA) kit. PAP and TP53INP1 mRNA expression was determined using the reverse transcription-polymerase chain reaction with semi-quantitative analysis. The expression of GSK-3β and phospho-GSK-3β proteins was examined with Western blot analysis.RESULTS: The IC50 for the drug after a 48-h exposure to gemcitabine was 16 mg/L. The growth of PANC-1 cells was inhibited by gemcitabine in a concentrationdependent manner (P< 0.0001) and the cell growth was also inhibited throughout the time course (P<0.0001).The DNA fragmentation rate in the gemcitabine-treated group at 48 h was 44.7 %, whereas it was 25.3 % in the untreated group. The PAP mRNA expression was decreased after being treated with gemcitabine, whereas the TP53INP1 mRNA was increased by the gemcitabine treatment. Western blot analysis showed that phosphoGSK-3βser9 was induced by the gemcitabine treatment.CONCLUSION: Gemcitabine suppresses PANC-1cell proliferation and induces apoptosis. Apoptosis is considered to be associated with the inhibition of PAP and GSK-3β, and the activation of TP53INP1 and posphoGSK-33ser9 .

  8. Protection of Human Pancreatic Islets from Lipotoxicity by Modulation of the Translocon.

    Directory of Open Access Journals (Sweden)

    R Cassel

    Full Text Available Type 2 diabetes is characterized by peripheral insulin resistance and pancreatic beta cell dysfunction. Elevated free fatty acids (FFAs may impair beta cell function and mass (lipotoxicity. Altered calcium homeostasis may be involved in defective insulin release. The endoplasmic reticulum (ER is the major intracellular calcium store. Lipotoxicity induces ER stress and in parallel an ER calcium depletion through unknown ER calcium leak channels. The main purposes of this study is first to identify one of these channels and secondly, to check the opportunity to restore beta cells function (i.e., insulin secretion after pharmacological inhibition of ER calcium store depletion. We investigated the functionality of translocon, an ER calcium leak channel and its involvement on FFAs-induced alterations in MIN6B1 cells and in human pancreatic islets. We evidenced that translocon acts as a functional ER calcium leak channel in human beta cells using anisomycin and puromycin (antibiotics, respectively blocker and opener of this channel. Puromycin induced a significant ER calcium release, inhibited by anisomycin pretreatment. Palmitate treatment was used as FFA model to induce a mild lipotoxic effect: ER calcium content was reduced, ER stress but not apoptosis were induced and glucose induced insulin secretion was decreased in our beta cells. Interestingly, translocon inhibition by chronic anisomycin treatment prevented dysfunctions induced by palmitate, avoiding reticular calcium depletion, ER stress and restoring insulin secretion. Our results provide for the first time compelling evidence that translocon actively participates to the palmitate-induced ER calcium leak and insulin secretion decrease in beta cells. Its inhibition reduces these lipotoxic effects. Taken together, our data indicate that TLC may be a new potential target for the treatment of type 2 diabetes.

  9. Fetal Circulation

    Science.gov (United States)

    ... Peripheral Artery Disease Venous Thromboembolism Aortic Aneurysm More Fetal Circulation Updated:Oct 18,2016 click to enlarge The ... fetal heart. These two bypass pathways in the fetal circulation make it possible for most fetuses to survive ...

  10. Renewal and preliminary study of expressed sequence tags database on human fetal liver aged 22 wk of gestation

    Institute of Scientific and Technical Information of China (English)

    CHEN TingGui; WU SongFeng; ZHOU GangQiao; ZHU YunPing; HE FuChu

    2008-01-01

    With the developments of international human transcriptome data and our ESTs of human fetal liver aged 22 weeks (wk) of gestation (HFL22w), the former research must be renewed. In this work, the EST data were firstly clustered by blasting against the ESTs of HFL22w, UniGene, DOTS, MGC and Twin-scan-predicted human transcriptome. Then, after EST assembly and gene identification, the known genes were classified by GO (gene ontology), and the unknown genes were predicted by Pfam and ScanProsite to clarify their functions. In the end, the relations of 5 tissues including fetal liver, adult liver, bone marrow, thymus and lymph node that possess hemopoiesis or can indicate fetal liver char-acteristics were analyzed by hierarchical clustering. The results show that: (i) By comparing the 5 newest human transcriptome databases, we can largely reduce the probability that the ESTS belonging to unconnected parts of one gene were probably divided into different clusters, so it is recommended to blast against the newest databases when clustering EST data; (ii) some previous unknown ESTs had been identified as function-known genes, and 1379 genes were identified as fully new sequences pos-sessed in our lab; (iii) through GO classification, we got a rough understanding of HFL22w, and ob-tained 6 cell migration genes and 6 hemopoiesis genes; (iv) prediction of gene function had enabled us to obtain 277 profiles, among them, there are 5 categories distributed in more than 10 genes; (v) five tissue relations analyzed by hierarchical clustering are related to their functions; (vi) We have built the world's largest EST database on HFL22w. Renewal and preliminary analysis of EST database on HFL22w will help to understand hemopoiesis and cell migration mechanism, and promote future re-search on human fetal liver.

  11. Ex vivo culture of human fetal gonads: manipulation of meiosis signalling by retinoic acid treatment disrupts testis development.

    Science.gov (United States)

    Jørgensen, A; Nielsen, J E; Perlman, S; Lundvall, L; Mitchell, R T; Juul, A; Rajpert-De Meyts, E

    2015-10-01

    What are the effects of experimentally manipulating meiosis signalling by addition of retinoic acid (RA) in cultured human fetal gonads? RA-treatment accelerated meiotic entry in cultured fetal ovary samples, while addition of RA resulted in a dysgenetic gonadal phenotype in fetal testis cultures. One of the first manifestations of sex differentiation is the initiation of meiosis in fetal ovaries. In contrast, meiotic entry is actively prevented in the fetal testis at this developmental time-point. It has previously been shown that RA-treatment mediates initiation of meiosis in human fetal ovary ex vivo. This was a controlled ex vivo study of human fetal gonads treated with RA in 'hanging-drop' tissue cultures. The applied experimental set-up preserves germ cell-somatic niche interactions and the investigated outcomes included tissue integrity and morphology, cell proliferation and survival and the expression of markers of meiosis and sex differentiation. Tissue from 24 first trimester human fetuses was included in this study, all from elective terminations at gestational week (GW) 7-12. Gonads were cultured for 2 weeks with and without addition of 1 µM RA. Samples were subsequently formalin-fixed and investigated by immunohistochemistry and cell counting. Proteins investigated and quantified included; octamer-binding transcription factor 4 (OCT4), transcription factor AP-2 gamma (AP2γ) (embryonic germ cell markers), SRY (sex determining region Y)-box 9 (SOX9), anti-Müllerian hormone (AMH) (immature Sertoli cell markers), COUP transcription factor 2 (COUP-TFII) (marker of interstitial cells), forkhead box L2 (FOXL2) (granulosa cell marker), H2A histone family, member X (γH2AX) (meiosis marker), doublesex and mab-3 related transcription factor 1 (DMRT1) (meiosis regulator), cleaved poly ADP ribose polymerase (PARP), cleaved Caspase 3 (apoptosis markers) and Ki-67 antigen (Ki-67) (proliferation marker). Also, proliferation was determined using a 5'-bromo-2

  12. Structural Basis for Accelerated Cleavage of Bovine Pancreatic Trypsin Inhibitor (BPTI) by Human Mesotrypsin

    Energy Technology Data Exchange (ETDEWEB)

    Salameh,M.; Soares, A.; Hockla, A.; Radisky, E.

    2008-01-01

    Human mesotrypsin is an isoform of trypsin that displays unusual resistance to polypeptide trypsin inhibitors and has been observed to cleave several such inhibitors as substrates. Whereas substitution of arginine for the highly conserved glycine 193 in the trypsin active site has been implicated as a critical factor in the inhibitor resistance of mesotrypsin, how this substitution leads to accelerated inhibitor cleavage is not clear. Bovine pancreatic trypsin inhibitor (BPTI) forms an extremely stable and cleavage-resistant complex with trypsin, and thus provides a rigorous challenge of mesotrypsin catalytic activity toward polypeptide inhibitors. Here, we report kinetic constants for mesotrypsin and the highly homologous (but inhibitor sensitive) human cationic trypsin, describing inhibition by, and cleavage of BPTI, as well as crystal structures of the mesotrypsin-BPTI and human cationic trypsin-BPTI complexes. We find that mesotrypsin cleaves BPTI with a rate constant accelerated 350-fold over that of human cationic trypsin and 150,000-fold over that of bovine trypsin. From the crystal structures, we see that small conformational adjustments limited to several side chains enable mesotrypsin-BPTI complex formation, surmounting the predicted steric clash introduced by Arg-193. Our results show that the mesotrypsin-BPTI interface favors catalysis through (a) electrostatic repulsion between the closely spaced mesotrypsin Arg-193 and BPTI Arg-17, and (b) elimination of two hydrogen bonds between the enzyme and the amine leaving group portion of BPTI. Our model predicts that these deleterious interactions accelerate leaving group dissociation and deacylation.

  13. Highly efficient differentiation of human ES cells and iPS cells into mature pancreatic insulin-producing cells

    Institute of Scientific and Technical Information of China (English)

    Donghui Zhang; Wei Jiang; Meng Liu; Xin Sui; Xiaolei Yin; Song Chen; Yan Shi; Hongkui Deng

    2009-01-01

    Human pluripotent stem cells represent a potentially unlimited source of functional pancreatic endocrine lineage cells. Here we report a highly efficient approach to induce human embryonic stem (ES) cells and induced pluripotent stem (iPS) cells to differentiate into mature insulin-producing cells in a chemical-defined culture system. The differentiated human ES cells obtained by this approach comprised nearly 25% insulin-positive cells as assayed by flow cytometry analysis, which released insulin/C-peptide in response to glucose stimuli in a manner comparable to that of adult human islets. Most of these insulin-producing cells co-expressed mature β cell-specific markers such as NKX6-1 and PDX1, indicating a similar gene expression pattern to adult islet β cells in vivo. In this study, we also demonstrated that EGF facilitates the expansion of PDX1-positive pancreatic progenitors. Moreover, our protocol also succeeded in efficiently inducing human iPS cells to differentiate into insulin-producing cells. Therefore, this work not only provides a new model to study the mechanism of human pancreatic specialization and maturation in vitro, but also enhances the possibility of utilizing patient-specific iPS cells for the treatment of diabetes.

  14. Plasmacytoid Dendritic Cell Activation and IFN-α Production Are Prominent Features of Murine Autoimmune Pancreatitis and Human IgG4-Related Autoimmune Pancreatitis.

    Science.gov (United States)

    Arai, Yasuyuki; Yamashita, Kouhei; Kuriyama, Katsutoshi; Shiokawa, Masahiro; Kodama, Yuzo; Sakurai, Toshiharu; Mizugishi, Kiyomi; Uchida, Kazushige; Kadowaki, Norimitsu; Takaori-Kondo, Akifumi; Kudo, Masatoshi; Okazaki, Kazuichi; Strober, Warren; Chiba, Tsutomu; Watanabe, Tomohiro

    2015-10-01

    The abnormal immune response accompanying IgG4-related autoimmune pancreatitis (AIP) is presently unclear. In this study, we examined the role of plasmacytoid dendritic cell (pDC) activation and IFN-α production in this disease as well as in a murine model of AIP (MRL/Mp mice treated with polyinosinic-polycytidylic acid). We found that the development of AIP in treated MRL/Mp mice occurred in parallel with pancreatic accumulation of pDCs producing IFN-α, and with pDC depletion and IFN-α-blocking studies, we showed that such accumulation was necessary for AIP induction. In addition, we found that the pancreas of treated MRL/Mp mice contained neutrophil extracellular traps (NETs) shown previously to stimulate pDCs to produce IFN-α. Consistent with these findings, we found that patients with IgG4-related AIP also exhibited pancreatic tissue localization of IFN-α-expressing pDCs and had significantly higher serum IFN-α levels than healthy controls. In addition, the inflamed pancreas of these patients but not controls also contained NETs that were shown to be capable of pDC activation. More importantly, patient pDCs cultured in the presence of NETs produced greatly increased levels of IFN-α and induced control B cells to produce IgG4 (but not IgG1) as compared with control pDCs. These data suggest that pDC activation and production of IFN-α is a major cause of murine AIP; in addition, the increased pDC production of IFN-α and its relation to IgG4 production observed in IgG4-related AIP suggest that this mechanism also plays a role in the human disease.

  15. Evaluation of a gene-directed enzyme-product therapy (GDEPT in human pancreatic tumor cells and their use as in vivo models for pancreatic cancer.

    Directory of Open Access Journals (Sweden)

    Juraj Hlavaty

    Full Text Available BACKGROUND: Gene-directed enzyme prodrug therapy (GDEPT is a two-step treatment protocol for solid tumors that involves the transfer of a gene encoding a prodrug-activating enzyme followed by administration of the inactive prodrug that is subsequently activated by the enzyme to its tumor toxic form. However, the establishment of such novel treatment regimes to combat pancreatic cancer requires defined and robust animal model systems. METHODS: Here, we comprehensively compared six human pancreatic cancer cell lines (PaCa-44, PANC-1, MIA PaCa-2, Hs-766T, Capan-2, and BxPc-3 in subcutaneous and orthotopical mouse models as well as in their susceptibility to different GDEPTs. RESULTS: Tumor uptake was 83% to 100% in the subcutaneous model and 60% to 100% in the orthotopical mouse model, except for Hs-766T cells, which did not grow orthotopically. Pathohistological analyses of the orthotopical models revealed an infiltrative growth of almost all tumors into the pancreas; however, the different cell lines gave rise to tumors with different morphological characteristics. All of the resultant tumors were positive for MUC-1 staining indicating their origin from glandular or ductal epithelium, but revealed scattered pan-cytokeratin staining. Transfer of the cytochrome P450 and cytosine deaminase suicide gene, respectively, into the pancreatic cancer cell lines using retroviral vector technology revealed high level infectibility of these cell lines and allowed the analysis of the sensitivity of these cells to the chemotherapeutic drugs ifosfamide and 5-fluorocytosine, respectively. CONCLUSION: These data qualify the cell lines as part of valuable in vitro and in vivo models for the use in defined preclinical studies for pancreas tumor therapy.

  16. Dimethylenastron suppresses human pancreatic cancer cell migration and invasion in vitro via allosteric inhibition of mitotic kinesin Eg5

    Institute of Scientific and Technical Information of China (English)

    Xiao-dong SUN; LIU Jun ZHOU; Xing-juan SHl; Xiao-ou SUN; You-guang LUO; Xiao-jing WU; Chang-fu YAO; Hai-yang YU; Deng-wen; LI Min

    2011-01-01

    The mitotic kinesin Eg5 plays a critical role in bipolar spindle assembly,and its inhibitors have shown impressive anticancer activity in preclinical studies.This study was undertaken to investigate the effect of dimethylenastron,a specific inhibitor of Eg5,on the migration and invasion of pancreatic cancer cells.Methods:Human pancreatic cancer cell lines PANC1,EPP85,BxPC3,CFPAC1,and AsPAC1 were used.Eg5 expression was examined using immunofluorescence microscopy.Cell migration and invasion were analyzed with wound healing and transwell assays.Cell pro-liferation was examined using sulforhodamine B and MTT assays.The binding of dimethylenastron to Eg5 was analyzed with a molecular modeling study,and the ADP release rate was examined with the MANT-ADP reagent.Results:Eg5 expression was 9-16-fold up-regulated in the 5 pancreatic cancer cell lines.Treatment of PANC1 pancreatic cancer cells with dimethylenastron (3 and 10 μmol/L) for 24 h suppressed the migratory ability of the cancer cells in a concentration-dependent manner.The invasion ability of the cancer cells was also reduced by the treatment.However,treatment of PANC1 cells with dimeth-ylenastron (3 and 10 μmol/L) for 24 h had no detectable effect on their proliferation,which was inhibited when the cancer cells were treated with the drug for 72 h.Molecular modeling study showed that dimethylenastron could allosterically inhibit the motor domain ATPase of Eg5 by decreasing the rate of ADP release.Conclusion:Dimethylenastron inhibits the migration and invasion of PANC1 pancreatic cancer cells,independent of suppressing the cell proliferation.The findings provide a novel insight into the mechanisms of targeting Eg5 for pancreatic cancer chemotherapy.

  17. Human telomerase activity, telomerase and telomeric template expression in hepatic stem cells and in livers from fetal and postnatal donors.

    Science.gov (United States)

    Schmelzer, Eva; Reid, Lola M

    2009-10-01

    Although telomerase activity has been analyzed in various normal and malignant tissues, including liver, it is still unknown to what extent telomerase can be associated with specific maturational lineage stages. We assessed human telomerase activity, protein and gene expression for the telomerase reverse transcriptase, as well as expression of the telomeric template RNA hTER in hepatic stem cells and in various developmental stages of the liver from fetal to adult. In addition, the effect of growth factors on telomerase activity was analyzed in hepatic stem cells in vitro. Telomerase was found to be highly active in fetal liver cells and was significantly higher than in hepatic stem cells, correlating with gene and protein expression levels. Activity in postnatal livers from all donor ages varied considerably and did not correlate with age or gene expression levels. The hter expression could be detected throughout the development. A short stimulation by growth factors of cultured hepatic stem cells did not increase telomerase activity. Telomerase is considerably active in fetal liver and variably in postnatal livers. Although telomerase protein is present at varying levels in liver cells of all donor ages, gene expression is solely associated with fetal liver cells.

  18. Reactivating Fetal Hemoglobin Expression in Human Adult Erythroblasts Through BCL11A Knockdown Using Targeted Endonucleases

    Directory of Open Access Journals (Sweden)

    Carmen F Bjurström

    2016-01-01

    Full Text Available We examined the efficiency, specificity, and mutational signatures of zinc finger nucleases (ZFNs, transcriptional activator-like effector nucleases (TALENs, and clustered regularly interspaced short palindromic repeat (CRISPR/Cas9 systems designed to target the gene encoding the transcriptional repressor BCL11A, in human K562 cells and human CD34+ progenitor cells. ZFNs and TALENs were delivered as in vitro transcribed mRNA through electroporation; CRISPR/Cas9 was codelivered by Cas9 mRNA with plasmid-encoded guideRNA (gRNA (pU6.g1 or in vitro transcribed gRNA (gR.1. Analyses of efficacy revealed that for these specific reagents and the delivery methods used, the ZFNs gave rise to more allelic disruption in the targeted locus compared to the TALENs and CRISPR/Cas9, which was associated with increased levels of fetal hemoglobin in erythroid cells produced in vitro from nuclease-treated CD34+ cells. Genome-wide analysis to evaluate the specificity of the nucleases revealed high specificity of this specific ZFN to the target site, while specific TALENs and CRISPRs evaluated showed off-target cleavage activity. ZFN gene-edited CD34+ cells had the capacity to engraft in NOD-PrkdcSCID-IL2Rγnull mice, while retaining multi-lineage potential, in contrast to TALEN gene-edited CD34+ cells. CRISPR engraftment levels mirrored the increased relative plasmid-mediated toxicity of pU6.g1/Cas9 in hematopoietic stem/progenitor cells (HSPCs, highlighting the value for the further improvements of CRISPR/Cas9 delivery in primary human HSPCs.

  19. Adult, embryonic and fetal hemoglobin are expressed in human glioblastoma cells.

    Science.gov (United States)

    Emara, Marwan; Turner, A Robert; Allalunis-Turner, Joan

    2014-02-01

    Hemoglobin is a hemoprotein, produced mainly in erythrocytes circulating in the blood. However, non-erythroid hemoglobins have been previously reported in other cell types including human and rodent neurons of embryonic and adult brain, but not astrocytes and oligodendrocytes. Human glioblastoma multiforme (GBM) is the most aggressive tumor among gliomas. However, despite extensive basic and clinical research studies on GBM cells, little is known about glial defence mechanisms that allow these cells to survive and resist various types of treatment. We have shown previously that the newest members of vertebrate globin family, neuroglobin (Ngb) and cytoglobin (Cygb), are expressed in human GBM cells. In this study, we sought to determine whether hemoglobin is also expressed in GBM cells. Conventional RT-PCR, DNA sequencing, western blot analysis, mass spectrometry and fluorescence microscopy were used to investigate globin expression in GBM cell lines (M006x, M059J, M059K, M010b, U87R and U87T) that have unique characteristics in terms of tumor invasion and response to radiotherapy and hypoxia. The data showed that α, β, γ, δ, ζ and ε globins are expressed in all tested GBM cell lines. To our knowledge, we are the first to report expression of fetal, embryonic and adult hemoglobin in GBM cells under normal physiological conditions that may suggest an undefined function of those expressed hemoglobins. Together with our previous reports on globins (Ngb and Cygb) expression in GBM cells, the expression of different hemoglobins may constitute a part of series of active defence mechanisms supporting these cells to resist various types of treatments including chemotherapy and radiotherapy.

  20. Fetal calf serum heat inactivation and lipopolysaccharide contamination influence the human T lymphoblast proteome and phosphoproteome

    Directory of Open Access Journals (Sweden)

    Rahman Hazir

    2011-11-01

    Full Text Available Abstract Background The effects of fetal calf serum (FCS heat inactivation and bacterial lipopolysaccharide (LPS contamination on cell physiology have been studied, but their effect on the proteome of cultured cells has yet to be described. This study was undertaken to investigate the effects of heat inactivation of FCS and LPS contamination on the human T lymphoblast proteome. Human T lymphoblastic leukaemia (CCRF-CEM cells were grown in FCS, either non-heated, or heat inactivated, having low ( Results A total of four proteins (EIF3M, PRS7, PSB4, and SNAPA were up-regulated when CCRF-CEM cells were grown in media supplemented with heat inactivated FCS (HE as compared to cells grown in media with non-heated FCS (NHE. Six proteins (TCPD, ACTA, NACA, TCTP, ACTB, and ICLN displayed a differential phosphorylation pattern between the NHE and HE groups. Compared to the low concentration LPS group, regular levels of LPS resulted in the up-regulation of three proteins (SYBF, QCR1, and SUCB1. Conclusion The present study provides new information regarding the effect of FCS heat inactivation and change in FCS-LPS concentration on cellular protein expression, and post-translational modification in human T lymphoblasts. Both heat inactivation and LPS contamination of FCS were shown to modulate the expression and phosphorylation of proteins involved in basic cellular functions, such as protein synthesis, cytoskeleton stability, oxidative stress regulation and apoptosis. Hence, the study emphasizes the need to consider both heat inactivation and LPS contamination of FCS as factors that can influence the T lymphoblast proteome.

  1. Development and morphogenesis of human wrist joint during embryonic and early fetal period.

    Science.gov (United States)

    Hita-Contreras, Fidel; Martínez-Amat, Antonio; Ortiz, Raúl; Caba, Octavio; Alvarez, Pablo; Prados, José C; Lomas-Vega, Rafael; Aránega, Antonia; Sánchez-Montesinos, Indalecio; Mérida-Velasco, Juan A

    2012-06-01

    The development of the human wrist joint has been studied widely, with the main focus on carpal chondrogenesis, ligaments and triangular fibrocartilage. However, there are some discrepancies concerning the origin and morphogenetic time-table of these structures, including nerves, muscles and vascular elements. For this study we used serial sections of 57 human embryonic (n = 30) and fetal (n = 27) specimens from O'Rahilly stages 17-23 and 9-14 weeks, respectively. The following phases in carpal morphogenesis have been established: undifferentiated mesenchyme (stage 17), condensated mesenchyme (stages 18 and 19), pre-chondrogenic (stages 19 and 20) and chondrogenic (stages 21 and over). Carpal chondrification and osteogenic processes are similar, starting with capitate and hamate (stage 19) and ending with pisiform (stage 22). In week 14, a vascular bud penetrates into the lunate cartilaginous mold, early sign of the osteogenic process that will be completed after birth. In stage 18, median, ulnar and radial nerves and thenar eminence appear in the hand plate. In stage 21, there are indications of the interosseous muscles, and in stage 22 flexor digitorum superficialis, flexor digitorum profundus and lumbrical muscles, transverse carpal ligament and collateral ligaments emerge. In stage 23, the articular disc, radiocarpal and ulnocarpal ligaments and deep palmar arterial arch become visible. Radiate carpal and interosseous ligaments appear in week 9, and in week 10, dorsal radiocarpal ligament and articular capsule are evident. Finally, synovial membrane is observed in week 13. We have performed a complete analysis of the morphogenesis of the structures of the human wrist joint. Our results present new data on nervous and arterial elements and provide the basis for further investigations on anatomical pathology, comparative morphology and evolutionary anthropology. © 2012 The Authors. Journal of Anatomy © 2012 Anatomical Society.

  2. Nickel nanowires induced and reactive oxygen species mediated apoptosis in human pancreatic adenocarcinoma cells

    Directory of Open Access Journals (Sweden)

    Kleve MG

    2011-07-01

    Full Text Available Md. Zakir Hossain1, Maurice G Kleve21Applied Biosciences (Bionanotechnology Research, Department of Applied Science, 2Molecular Biotechnology and Microscopy Laboratory, Department of Biology, University of Arkansas at Little Rock, Little Rock, Arkansas, USABackground: The ability to evade apoptosis is one of the key properties of cancer. The apoptogenic effect of nickel nanowires (Ni NWs on cancer cell lines has never been adequately addressed. Due to the unique physicochemical characteristics of Ni NWs, we envision the development of a novel anticancer therapeutics specifically for pancreatic cancer. Thus, we investigated whether Ni NWs induce ROS-mediated apoptosis in human pancreatic adenocarcinoma (Panc-1 cells. Methods: In this study Ni NWs were fabricated using the electrodeposition method. Synthesized Ni NWs were physically characterized by energy dispersive X-ray analysis, UV-Vis spectroscopy of NanoDrop 2000 (UV-Vis, magnetization study, scanning electron microscopy, and transmission electron microscopy. Assessment of morphological apoptotic characteristics by phase contrast microscopy (PCM, Ni-NWs-induced apoptosis staining with ethidium bromide (EB and acridine orange (AO followed by fluorescence microscopy (FM was performed. For molecular biological and biochemical characterization, Panc-1 cell culture and cytotoxic effect of Ni NWs were determined by using 3-(4, 5-dimethylthiazol-2-yl-2, 5-diphenyl tetrazolium bromide (MTT assay. Quantitative apoptosis was analyzed by flow cytometry staining with propidium iodide through cell cycle arrest and generation of ROS using 2', 7'-dichlorofluorescein diacetate fluorescence intensity. In all experiments, Panc-1 cancer cells without any treatment were used as the negative controls.Results: The intracellular uptake of Ni NWs through endocytosis by Panc-1 cells was observed by PCM. EB and AO staining of FM and MTT assay qualitatively and quantitatively confirmed the extent of apoptosis. Flow

  3. MicroRNA expression in alpha and beta cells of human pancreatic islets.

    Directory of Open Access Journals (Sweden)

    Dagmar Klein

    Full Text Available microRNAs (miRNAs play an important role in pancreatic development and adult β-cell physiology. Our hypothesis is based on the assumption that each islet cell type has a specific pattern of miRNA expression. We sought to determine the profile of miRNA expression in α-and β-cells, the main components of pancreatic islets, because this analysis may lead to a better understanding of islet gene regulatory pathways. Highly enriched (>98% subsets of human α-and β-cells were obtained by flow cytometric sorting after intracellular staining with c-peptide and glucagon antibody. The method of sorting based on intracellular staining is possible because miRNAs are stable after fixation. MiRNA expression levels were determined by quantitative high throughput PCR-based miRNA array platform screening. Most of the miRNAs were preferentially expressed in β-cells. From the total of 667 miRNAs screened, the Significant Analysis of Microarray identified 141 miRNAs, of which only 7 were expressed more in α-cells (α-miRNAs and 134 were expressed more in β-cells (β-miRNAs. Bioinformatic analysis identified potential targets of β-miRNAs analyzing the Beta Cell Gene Atlas, described in the T1Dbase, the web platform, supporting the type 1 diabetes (T1D community. cMaf, a transcription factor regulating glucagon expression expressed selectively in α-cells (TFα is targeted by β-miRNAs; miR-200c, miR-125b and miR-182. Min6 cells treated with inhibitors of these miRNAs show an increased expression of cMaf RNA. Conversely, over expression of miR-200c, miR-125b or miR-182 in the mouse alpha cell line αTC6 decreases the level of cMAF mRNA and protein. MiR-200c also inhibits the expression of Zfpm2, a TFα that inhibits the PI3K signaling pathway, at both RNA and protein levels.In conclusion, we identified miRNAs differentially expressed in pancreatic α- and β-cells and their potential transcription factor targets that could add new insights into different

  4. Selection of polymers for application in scaffolds applicable for human pancreatic islet transplantation.

    Science.gov (United States)

    Smink, Alexandra M; de Haan, Bart J; Paredes-Juarez, Genaro A; Wolters, Anouk H G; Kuipers, Jeroen; Giepmans, Ben N G; Schwab, Leendert; Engelse, Marten A; van Apeldoorn, Aart A; de Koning, Eelco; Faas, Marijke M; de Vos, Paul

    2016-05-13

    The liver is currently the site for transplantation of islets in humans. This is not optimal for islets, but alternative sites in humans are not available. Polymeric scaffolds in surgically accessible areas are a solution. As human donors are rare, the polymers should not interfere with functional survival of human-islets. We applied a novel platform to test the adequacy of polymers for application in scaffolds for human-islet transplantation. Viability, functionality, and immune parameters were included to test poly(D,L-lactide-co-ε-caprolactone) (PDLLCL), poly(ethylene oxide terephthalate)/polybutylene terephthalate (PEOT/PBT) block copolymer, and polysulfone. The type of polymer influenced the functional survival of human islets. In islets cultured on PDLLCL the glucagon-producing α-cells and insulin-producing β-cells contained more hormone granules than in islets in contact with PEOT/PBT or polysulfone. This was studied with ultrastructural analysis by electron microscopy (nanotomy) during 7 d of culture. PDLLCL was also associated with statistically significant lower release of double-stranded DNA (dsDNA, a so called danger-associate molecular pattern (DAMP)) from islets on PDLLCL when compared to the other polymers. DAMPs support undesired immune responses. Hydrophilicity of the polymers did not influence dsDNA release. Islets on PDLLCL also showed less cellular outgrowth. These outgrowing cells were mainly fibroblast and some β-cells undergoing epithelial to mesenchymal cell transition. None of the polymers influenced the glucose-stimulated insulin secretion. As PDLLCL was associated with less release of DAMPs, it is a promising candidate for creating a scaffold for human islets. Our study demonstrates that for sensitive, rare cadaveric donor tissue such as pancreatic islets it might be necessary to first select materials that do not influence functionality before proposing the biomaterial for in vivo application. Our presented platform may facilitate

  5. Nicotine Alters the Proteome of Two Human Pancreatic Duct Cell Lines

    Science.gov (United States)

    Paulo, Joao A

    2015-01-01

    Context Cigarette smoking is a known risk factor of pancreatic disease. Nicotine - a major cigarette tobacco component - can traffic through the circulatory system and may induce fibrosis and metastasis, hallmarks of chronic pancreatitis and pancreatic adenocarcinoma, respectively. However, at the biomolecular level, particularly in pancreatic research, the effects of nicotine remain unresolved. Methods The effects of nicotine on the proteomes of two pancreatic duct cell lines–an immortalized normal cell line (HPNE) and a cancer cell line (PanC1)- were investigated using mass spectrometry-based proteomics. For each cell line, the global proteomesof cells exposed to nicotine for 24 hrswere compared with untreated cells in triplicate using 6-plex tandem mass tag-based isobaric labeling techniques. Results Over 5,000 proteins were detectedper cell line. Of these, over 900 proteins were differentially abundant with statistical significance (corrected p-value <0.01) upon nicotine treatment, 57 of which were so in both cell lines. Amyloid precursor protein, previously observed to increase expression in pancreatic stellate cells when exposed to nicotine, was also up-regulated in both cell lines.In general, the two cell lines varied in the classes of proteins altered by nicotine treatment, supporting published evidence that nicotine may play different roles in the initiation and progression of pancreatic disease. Conclusions Understanding the underlying mechanisms associating nicotine with pancreatic function is paramount to intervention aiming to retard, arrest, or ameliorate pancreatic disease. PMID:25262714

  6. Effects of extremely low frequency electromagnetic fields on human fetal scleral fibroblasts.

    Science.gov (United States)

    Zhu, Huang; Wang, Jie; Cui, Jiefeng; Fan, Xianqun

    2016-06-01

    This study investigated the effects of extremely low frequency electromagnetic fields (ELF-EMFs) on human fetal scleral fibroblasts (HFSFs). HFSFs were subjected to 50 Hz artificial ELF-EMFs generated by Helmholtz coils with 0.1, 0.2, 0.5, and 1.0 mT field intensities for 6 to 48 h. The viability and factors involved in scleral structuring of HFSFs were determined. The growth rate of HFSFs significantly decreased after only 24 h of exposure to ELF-EMFs (0.2 mT). The messenger RNA (mRNA) expression of collagen type I (COL1A1) decreased and expression of matrix metalloproteinase-2 (MMP-2) increased significantly. There was a decrease in tissue inhibitor of MMP-2 mRNA levels between treated and control cells only at the 1.0 mT intensity level. Transforming growth factor beta-2 mRNA increased in exposed cells, and, simultaneously, fibroblast growth factor-2 mRNA levels decreased. The protein expressions of COL1A1 and MMP-2 were also significantly altered subsequent to exposure (p effects on HFSFs and could cause abnormality in scleral collagen.

  7. The 2008 National Institute of Child Health and Human Development workshop report on electronic fetal monitoring: update on definitions, interpretation, and research guidelines

    National Research Council Canada - National Science Library

    Macones, George A; Hankins, Gary D V; Spong, Catherine Y; Hauth, John; Moore, Thomas

    2008-01-01

    In April 2008, the Eunice Kennedy Shriver National Institute of Child Health and Human Development, the American College of Obstetricians and Gynecologists, and the Society for Maternal-Fetal Medicine...

  8. Observation of the human fetal corpses with maxillofacial malformations. 1. CT and MRI examinations of the fetal cleft lip and/or palate

    Energy Technology Data Exchange (ETDEWEB)

    Saito, Chikara; Nakano, Yoko; Shigematsu, Shiro [Tokyo Dental Coll., Chiba (Japan)] (and others)

    1999-06-01

    Of the various types of congenital malformations, the cleft lip and/or palate is one of the most frequent. Observation of human fetal corpses exhibiting cleft lip and palate is very important to research on its onset of its mechanism and development. In recent years, some of researchers have performed clinical studies on prenatal diagnosis and surgical treatment for the entirey. However, there have hardly been any reports on detailed observations of the maxillofacial structure of a fetus with cleft lip and palate. We seized an opportunity of observing the maxillofacial structure of fetuses with cleft lip and/or palate using three-dimensional CT (3D-CT) and MR imaging as non-disjunctive methods. In the present study, nine fetal corpses having cleft lip and/or palate were examined. The results were as follows: CT and MRI were useful for non-invasive observation of the maxillofacial structure, including soft tissues. Because the osseous tissues of young fetus tissue is not fully mature, observation of bone structures was slightly difficult. When corpses were immersed in formalin for a long time, osseous tissue was decalcified, thus making it difficult to obtain clear images. We could observe the details of the maxillofacial structures such as the alveolar process, the hard palate, the maxillary sinus, the nasal cavity, the nasal bone, and the vomer, in some of the cases. 3D-CT and MR findings observed in the fetuses with cleft lip and/or palate should provide some basement of the imaging diagnosis of congenital disorder. (author)

  9. Human pancreas development.

    Science.gov (United States)

    Jennings, Rachel E; Berry, Andrew A; Strutt, James P; Gerrard, David T; Hanley, Neil A

    2015-09-15

    A wealth of data and comprehensive reviews exist on pancreas development in mammals, primarily mice, and other vertebrates. By contrast, human pancreatic development has been less comprehensively reviewed. Here, we draw together those studies conducted directly in human embryonic and fetal tissue to provide an overview of what is known about human pancreatic development. We discuss the relevance of this work to manufacturing insulin-secreting β-cells from pluripotent stem cells and to different aspects of diabetes, especially permanent neonatal diabetes, and its underlying causes.

  10. Expression profiles of miRNAs in human pancreatic cancer cell lines%胰腺癌细胞系中miRNAs的表达分析

    Institute of Scientific and Technical Information of China (English)

    Shineng Zhang; Haijun Zuo; Zhong Yu; Fengting Huang; Wa Zhong

    2009-01-01

    Objective:To analyze initially the differences of miRNAs expression profiles in human pancreatic cencer cell lines by microarray technique.Methods:A total of 743 probes were designed according to lhe known miRNAs sequences of human,mice,and rats.miRNAs microarray was manufactured and its credibility was verified.Total RNAs were extracted and miRNAs were separated from human pancreatic cancer cell lines(SW1990,Capan-2.BxPC-3,Aspc-1,and Panc 1)and immortal human pancreatic duct epithelial cell line H6C7.They were labeled with T4 RNA ligase.then were hybridized with microarray.Through array scan and analysis,miRNAs expression profiles in pancreatic cancer were obtained.The results were verified by Northern blotting and RT-PCR.Results:A totsl of 63 miRNAs related to pancreatic cancer were found to be differentially expressed in 5 pancreatic cancer cell lines.including 25 down-regulated and 38 up-regulated miRNAs.Expres- sions of mir-21 and let-7 were also confirmed.Conclusion:The results suggested that miRNAs expression profiles could befound in pancreatic cancer cells.

  11. CpG sites with continuously increasing or decreasing methylation from early to late human fetal brain development.

    Science.gov (United States)

    Schneider, Eberhard; Dittrich, Marcus; Böck, Julia; Nanda, Indrajit; Müller, Tobias; Seidmann, Larissa; Tralau, Tim; Galetzka, Danuta; El Hajj, Nady; Haaf, Thomas

    2016-10-30

    Normal human brain development is dependent on highly dynamic epigenetic processes for spatial and temporal gene regulation. Recent work identified wide-spread changes in DNA methylation during fetal brain development. We profiled CpG methylation in frontal cortex of 27 fetuses from gestational weeks 12-42, using Illumina 450K methylation arrays. Sites showing genome-wide significant correlation with gestational age were compared to a publicly available data set from gestational weeks 3-26. Altogether, we identified 2016 matching developmentally regulated differentially methylated positions (m-dDMPs): 1767m-dDMPs were hypermethylated and 1149 hypomethylated during fetal development. M-dDMPs are underrepresented in CpG islands and gene promoters, and enriched in gene bodies. They appear to cluster in certain chromosome regions. M-dDMPs are significantly enriched in autism-associated genes and CpGs. Our results promote the idea that reduced methylation dynamics during fetal brain development may predispose to autism. In addition, m-dDMPs are enriched in genes with human-specific brain expression patterns and/or histone modifications. Collectively, we defined a subset of dDMPs exhibiting constant methylation changes from early to late pregnancy. The same epigenetic mechanisms involving methylation changes in cis-regulatory regions may have been adopted for human brain evolution and ontogeny. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  12. Wnt/β-Catenin Stimulation and Laminins Support Cardiovascular Cell Progenitor Expansion from Human Fetal Cardiac Mesenchymal Stromal Cells.

    Science.gov (United States)

    Månsson-Broberg, Agneta; Rodin, Sergey; Bulatovic, Ivana; Ibarra, Cristián; Löfling, Marie; Genead, Rami; Wärdell, Eva; Felldin, Ulrika; Granath, Carl; Alici, Evren; Le Blanc, Katarina; Smith, C I Edvard; Salašová, Alena; Westgren, Magnus; Sundström, Erik; Uhlén, Per; Arenas, Ernest; Sylvén, Christer; Tryggvason, Karl; Corbascio, Matthias; Simonson, Oscar E; Österholm, Cecilia; Grinnemo, Karl-Henrik

    2016-04-12

    The intrinsic regenerative capacity of human fetal cardiac mesenchymal stromal cells (MSCs) has not been fully characterized. Here we demonstrate that we can expand cells with characteristics of cardiovascular progenitor cells from the MSC population of human fetal hearts. Cells cultured on cardiac muscle laminin (LN)-based substrata in combination with stimulation of the canonical Wnt/β-catenin pathway showed increased gene expression of ISL1, OCT4, KDR, and NKX2.5. The majority of cells stained positive for PDGFR-α, ISL1, and NKX2.5, and subpopulations also expressed the progenitor markers TBX18, KDR, c-KIT, and SSEA-1. Upon culture of the cardiac MSCs in differentiation media and on relevant LNs, portions of the cells differentiated into spontaneously beating cardiomyocytes, and endothelial and smooth muscle-like cells. Our protocol for large-scale culture of human fetal cardiac MSCs enables future exploration of the regenerative functions of these cells in the context of myocardial injury in vitro and in vivo.

  13. Wnt/β-Catenin Stimulation and Laminins Support Cardiovascular Cell Progenitor Expansion from Human Fetal Cardiac Mesenchymal Stromal Cells

    Directory of Open Access Journals (Sweden)

    Agneta Månsson-Broberg

    2016-04-01

    Full Text Available The intrinsic regenerative capacity of human fetal cardiac mesenchymal stromal cells (MSCs has not been fully characterized. Here we demonstrate that we can expand cells with characteristics of cardiovascular progenitor cells from the MSC population of human fetal hearts. Cells cultured on cardiac muscle laminin (LN-based substrata in combination with stimulation of the canonical Wnt/β-catenin pathway showed increased gene expression of ISL1, OCT4, KDR, and NKX2.5. The majority of cells stained positive for PDGFR-α, ISL1, and NKX2.5, and subpopulations also expressed the progenitor markers TBX18, KDR, c-KIT, and SSEA-1. Upon culture of the cardiac MSCs in differentiation media and on relevant LNs, portions of the cells differentiated into spontaneously beating cardiomyocytes, and endothelial and smooth muscle-like cells. Our protocol for large-scale culture of human fetal cardiac MSCs enables future exploration of the regenerative functions of these cells in the context of myocardial injury in vitro and in vivo.

  14. Pasireotide and octreotide antiproliferative effects and sst2 trafficking in human pancreatic neuroendocrine tumor cultures.

    Science.gov (United States)

    Mohamed, Amira; Blanchard, Marie-Pierre; Albertelli, Manuela; Barbieri, Federica; Brue, Thierry; Niccoli, Patricia; Delpero, Jean-Robert; Monges, Genevieve; Garcia, Stephane; Ferone, Diego; Florio, Tullio; Enjalbert, Alain; Moutardier, Vincent; Schonbrunn, Agnes; Gerard, Corinne; Barlier, Anne; Saveanu, Alexandru

    2014-10-01

    Gastroenteropancreatic neuroendocrine tumors (GEP-NETs) raise difficult therapeutic problems despite the emergence of targeted therapies. Somatostatin analogs (SSA) remain pivotal therapeutic drugs. However, the tachyphylaxis and the limited antitumoral effects observed with the classical somatostatin 2 (sst2) agonists (octreotide and lanreotide) led to the development of new SSA, such as the pan sst receptor agonist pasireotide. Our aim was to compare the effects of pasireotide and octreotide on cell survival, chromogranin A (CgA) secretion, and sst2 phosphorylation/trafficking in pancreatic NET (pNET) primary cells from 15 tumors. We established and characterized the primary cultures of human pancreatic tumors (pNETs) as powerful preclinical models for understanding the biological effects of SSA. At clinically relevant concentrations (1-10 nM), pasireotide was at least as efficient as octreotide in inhibiting CgA secretion and cell viability through caspase-dependent apoptosis during short treatments, irrespective of the expression levels of the different sst receptors or the WHO grade of the parental tumor. Interestingly, unlike octreotide, which induces a rapid and persistent partial internalization of sst2 associated with its phosphorylation on Ser341/343, pasireotide did not phosphorylate sst2 and induced a rapid and transient internalization of the receptor followed by a persistent recycling at the cell surface. These results provide the first evidence, to our knowledge, of striking differences in the dynamics of sst2 trafficking in pNET cells treated with the two SSAs, but with similar efficiency in the control of CgA secretion and cell viability.

  15. Structure of Human Pancreatic Lipase-Related Protein 2 with the Lid in an Open Conformation

    Energy Technology Data Exchange (ETDEWEB)

    Eydoux, Cecilia; Spinelli, Silvia; Davis, Tara L.; Walker, John R.; Seitova, Alma; Dhe-Paganon, Sirano; De Caro, Alain; Cambillau, Christian; Carriere, Frederic (CNRS-UMR); (Toronto)

    2008-10-02

    Access to the active site of pancreatic lipase (PL) is controlled by a surface loop, the lid, which normally undergoes conformational changes only upon addition of lipids or amphiphiles. Structures of PL with their lids in the open and functional conformation have required cocrystallization with amphiphiles. Here we report two crystal structures of wild-type and unglycosylated human pancreatic lipase-related protein 2 (HPLRP2) with the lid in an open conformation in the absence of amphiphiles. These structures solved independently are strikingly similar, with some residues of the lid being poorly defined in the electron-density map. The open conformation of the lid is however different from that previously observed in classical liganded PL, suggesting different kinetic properties for HPLRP2. Here we show that the HPLRP2 is directly inhibited by E600, does not present interfacial activation, and acts preferentially on substrates forming monomers or small aggregates (micelles) dispersed in solution like monoglycerides, phospholipids and galactolipids, whereas classical PL displays reverse properties and a high specificity for unsoluble substrates like triglycerides and diglycerides forming oil-in-water interfaces. These biochemical properties imply that the lid of HPLRP2 is likely to spontaneously adopt in solution the open conformation observed in the crystal structure. This open conformation generates a large cavity capable of accommodating the digalactose polar head of galactolipids, similar to that previously observed in the active site of the guinea pig PLRP2, but absent from the classical PL. Most of the structural and kinetic properties of HPLRP2 were found to be different from those of rat PLRP2, the structure of which was previously obtained with the lid in a closed conformation. Our findings illustrate the essential role of the lid in determining the substrate specificity and the mechanism of action of lipases.

  16. A three-dimensional study of human fetal endocervix with special reference to its epithelium.

    Science.gov (United States)

    Barberini, F; Makabe, S; Motta, P M

    1998-07-01

    The development of human fetal cervix has been systematically studied by SEM, obtaining a detailed map of its fine structure, particularly concerning the differentiation and maturation of the endocervical epithelium, including its "eversion" and "squamous metaplasia", normally occurring in postnatal life, but not yet observed in detail by electron microscopy in the fetus. Cervices from spontaneous abortion at 12, 15, 18, 20, 21 and 22 weeks and from intrauterine fetal death (hydrocephalus) at 31 weeks of development have been examined. At 12-15 weeks, as the canalization of the cervix proceeded, the endocervical epithelium consisted of high polyhedral cells, with regularly flattened or concave apices exhibiting scarce microvilli and often single primary cilia. Some narrow intercellular infoldings probably corresponded to primordial tubular glands. At the 18th week the epithelium was made up of a mosaic of flat or slightly raised polygonal cells, whose apical surface showed thin microplicae. At the 20th week a pseudostratified epithelium with many apically convex cells lined the cervical canal and the tubular glands. At 21 and 22 weeks "plicae palmatae" developed, covered by cells, often showing a smooth central area surrounded by microvilli, provided with a primary cilium and swollen by secretory material. This also formed rounded masses on the epithelium. In the lower part of the endocervix some very elongated cells showed short microplicae resulting from fusion of microvilli. At the 31st week secretion increased and its products spreading from the bottom of the glands contacted isolated ciliated cells at their openings and diffusely covered the surface epithelium. Most of the ectocervix exhibited squamous elements, with well-developed labyrinthine microplicae. These cells could overlap each other and also desquamate. The zone of the portio vaginalis around the os of the cervical canal appeared infolded and hypertrophic. Here, an indented squamo-columnar junction

  17. TGFβ Pathway Inhibition Redifferentiates Human Pancreatic Islet β Cells Expanded In Vitro.

    Directory of Open Access Journals (Sweden)

    Ginat Toren-Haritan

    Full Text Available In-vitro expansion of insulin-producing cells from adult human pancreatic islets could provide an abundant cell source for diabetes therapy. However, proliferation of β-cell-derived (BCD cells is associated with loss of phenotype and epithelial-mesenchymal transition (EMT. Nevertheless, BCD cells maintain open chromatin structure at β-cell genes, suggesting that they could be readily redifferentiated. The transforming growth factor β (TGFβ pathway has been implicated in EMT in a range of cell types. Here we show that human islet cell expansion in vitro involves upregulation of the TGFβ pathway. Blocking TGFβ pathway activation using short hairpin RNA (shRNA against TGFβ Receptor 1 (TGFBR1, ALK5 transcripts inhibits BCD cell proliferation and dedifferentiation. Treatment of expanded BCD cells with ALK5 shRNA results in their redifferentiation, as judged by expression of β-cell genes and decreased cell proliferation. These effects, which are reproducible in cells from multiple human donors, are mediated, at least in part, by AKT-FOXO1 signaling. ALK5 inhibition synergizes with a soluble factor cocktail to promote BCD cell redifferentiation. The combined treatment may offer a therapeutically applicable way for generating an abundant source of functional insulin-producing cells following ex-vivo expansion.

  18. Characterization of the human pancreatic islet proteome by two-dimensional LC/MS/MS.

    Science.gov (United States)

    Metz, Thomas O; Jacobs, Jon M; Gritsenko, Marina A; Fontès, Ghislaine; Qian, Wei-Jun; Camp, David G; Poitout, Vincent; Smith, Richard D

    2006-12-01

    The pancreatic beta-cell plays a central role in the maintenance of glucose homeostasis and in the pathogenesis of both type 1 and type 2 diabetes mellitus. Elucidation of the insulin secretory defects observed in diabetes first requires a better understanding of the complex mechanisms regulating insulin secretion, which are only partly understood. While there have been reports detailing proteomic analyses of islet cell lines or isolated rodent islets, the information gained is not always applicable to humans. Therefore, definition of the human islet proteome could contribute to a better understanding of islet biology and lead to more effective treatment strategies. We have applied a two-dimensional LC-MS/MS-based analysis to the characterization of the human islet proteome, resulting in the confident identification of 29,021 different tryptic peptides covering 3365 proteins (> or =2 unique peptide identifications per protein). As expected, the three major islet hormones (insulin, glucagon, and somatostatin) were detected, as well as various beta-cell enriched secretory products, ion channels, and transcription factors. In addition, significant proteome coverage of metabolic enzymes and cellular pathways was observed, including the integrin signaling cascade and the MAP kinase, NF-kappa beta, and JAK/STAT pathways. The resulting peptide reference library provides a resource for future higher throughput and quantitative studies of islet biology.

  19. Downregulation of tight junction-associated MARVEL protein marvelD3 during epithelial-mesenchymal transition in human pancreatic cancer cells.

    Science.gov (United States)

    Kojima, Takashi; Takasawa, Akira; Kyuno, Daisuke; Ito, Tatsuya; Yamaguchi, Hiroshi; Hirata, Koichi; Tsujiwaki, Mitsuhiro; Murata, Masaki; Tanaka, Satoshi; Sawada, Norimasa

    2011-10-01

    The novel tight junction protein marvelD3 contains a conserved MARVEL (MAL and related proteins for vesicle trafficking and membrane link) domain like occludin and tricellulin. However, little is yet known about the detailed role and regulation of marvelD3 in normal epithelial cells and cancer cells, including pancreatic cancer. In the present study, we investigated marvelD3 expression in well and poorly differentiated human pancreatic cancer cell lines and normal pancreatic duct epithelial cells in which the hTERT gene was introduced into human pancreatic duct epithelial cells in primary culture, and the changes of marvelD3 during Snail-induced epithelial-mesenchymal transition (EMT) under hypoxia, TGF-β treatment and knockdown of FOXA2 in well differentiated pancreatic cancer HPAC cells. MarvelD3 was transcriptionally downregulated in poorly differentiated pancreatic cancer cells and during Snail-induced EMT of pancreatic cancer cells in which Snail was highly expressed and the fence function downregulated, whereas it was maintained in well differentiated human pancreatic cancer cells and normal pancreatic duct epithelial cells. Depletion of marvelD3 by siRNAs in HPAC cells resulted in downregulation of barrier functions indicated as a decrease in transepithelial electric resistance and an increase of permeability to fluorescent dextran tracers, whereas it did not affect fence function of tight junctions. In conclusion, marvelD3 is transcriptionally downregulated in Snail-induced EMT during the progression for the pancreatic cancer. Copyright © 2011 Elsevier Inc. All rights reserved.

  20. Is there a place for human fetal-derived stem cells for cell replacement therapy in Huntington's disease?

    Science.gov (United States)

    Precious, Sophie V; Zietlow, Rike; Dunnett, Stephen B; Kelly, Claire M; Rosser, Anne E

    2017-06-01

    Huntington's disease (HD) is a neurodegenerative disease that offers an excellent paradigm for cell replacement therapy because of the associated relatively focal cell loss in the striatum. The predominant cells lost in this condition are striatal medium spiny neurons (MSNs). Transplantation of developing MSNs taken from the fetal brain has provided proof of concept that donor MSNs can survive, integrate and bring about a degree of functional recovery in both pre-clinical studies and in a limited number of clinical trials. The scarcity of human fetal tissue, and the logistics of coordinating collection and dissection of tissue with neurosurgical procedures makes the use of fetal tissue for this purpose both complex and limiting. Alternative donor cell sources which are expandable in culture prior to transplantation are currently being sought. Two potential donor cell sources which have received most attention recently are embryonic stem (ES) cells and adult induced pluripotent stem (iPS) cells, both of which can be directed to MSN-like fates, although achieving a genuine MSN fate has proven to be difficult. All potential donor sources have challenges in terms of their clinical application for regenerative medicine, and thus it is important to continue exploring a wide variety of expandable cells. In this review we discuss two less well-reported potential donor cell sources; embryonic germ (EG) cells and fetal neural precursors (FNPs), both are which are fetal-derived and have some properties that could make them useful for regenerative medicine applications. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  1. Human fetal chromaffin cells: a potential tool for cell pain therapy.

    Science.gov (United States)

    Jozan, Suzanne; Aziza, Jacqueline; Châtelin, Sophie; Evra, Corinne; Courtade-Saïdi, Monique; Parant, Olivier; Sol, Jean Christophe; Zhou, Huafang; Lazorthes, Yves

    2007-06-01

    Transplantation of adrenal medulla cells has been proposed in the treatment of various conditions. Indeed, these cells possess a bipotentiality: neural and neuroendocrine, which could be exploited for brain repair or pain therapy. In a previous study, we characterized these human cells in vitro over 7-10 gestational weeks (GW) [Zhou, H., Aziza, J., Sol, J.C., Courtade-Saidi, M., Chatelin, S., Evra, C., Parant, O., Lazorthes, Y., and Jozan, S., 2006. Cell therapy of pain: Characterization of human fetal chromaffin cells at early adrenal medulla development. Exp. Neurol. 198, 370-381]. We report here our results on the extension to 23 GW. This developmental period can be split into three stages. During the first stage (7-10 GW), we observed in situ that extra-adrenal surrounding cells display the same morphology and phenotype as the intra-adrenal chromaffin cells. We also found that the intra-adrenal chromaffin cells could be committed in vitro towards an adrenergic phenotype using differentiating agents. During the second stage (11 to 15-16 GW), two types of cells (Type 1 and Type 2 cells) were identified morphologically both inside and outside the gland. Interestingly, we noted that the Type 2 cells stem from the Type 1 cells. However, during this developmental period only the intra-adrenal Type 2 cells will evolve towards an adrenergic phenotype. In the third stage (17-23 GW), we observed the ultimate location of the medulla gland. Both the in situ results and the in vitro experiments indicate that particular procedures need to be implemented prior transplantation of chromaffin cells. First, in order to obtain a large number of immature chromaffin cells, they must be isolated from the intra and extra-adrenal gland and should then be committed towards an adrenergic phenotype in vitro for subsequent use in pain therapy. This strategy is under investigation in our laboratory.

  2. Asymmetry of Radial and Symmetry of Tangential Neuronal Migration Pathways in Developing Human Fetal Brains.

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    Miyazaki, Yuta; Song, Jae W; Takahashi, Emi

    2016-01-01

    The radial and tangential neural migration pathways are two major neuronal migration streams in humans that are critical during corticogenesis. Corticogenesis is a complex process of neuronal proliferation that is followed by neuronal migration and the formation of axonal connections. Existing histological assessments of these two neuronal migration pathways have limitations inherent to microscopic studies and are confined to small anatomic regions of interest (ROIs). Thus, little evidence is available about their three-dimensional (3-D) fiber pathways and development throughout the entire brain. In this study, we imaged and analyzed radial and tangential migration pathways in the whole human brain using high-angular resolution diffusion MR imaging (HARDI) tractography. We imaged ten fixed, postmortem fetal (17 gestational weeks (GW), 18 GW, 19 GW, three 20 GW, three 21 GW and 22 GW) and eight in vivo newborn (two 30 GW, 34 GW, 35 GW and four 40 GW) brains with no neurological/pathological conditions. We statistically compared the volume of the left and right radial and tangential migration pathways, and the volume of the radial migration pathways of the anterior and posterior regions of the brain. In specimens 22 GW or younger, the volume of radial migration pathways of the left hemisphere was significantly larger than that of the right hemisphere. The volume of posterior radial migration pathways was also larger when compared to the anterior pathways in specimens 22 GW or younger. In contrast, no significant differences were observed in the radial migration pathways of brains older than 22 GW. Moreover, our study did not identify any significant differences in volumetric laterality in the tangential migration pathways. These results suggest that these two neuronal migration pathways develop and regress differently, and radial neuronal migration varies regionally based on hemispheric and anterior-posterior laterality, potentially explaining regional differences in

  3. Effect of prolonged exposure to sublethal concentrations of DDT and DDE on protein expression in human pancreatic beta cells.

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    Pavlikova, Nela; Smetana, Pavel; Halada, Petr; Kovar, Jan

    2015-10-01

    Pollution of the environment represents one of less explored potential reasons for the worldwide epidemic of type 2 diabetes. One of the most prevalent organochlorine pollutants remains the pesticide DDT and its degradation product DDE. Despite some epidemiologic correlations between levels of DDT and DDE in human organism and the prevalence of diabetes, there is almost no information about the exact targets of these compounds inside pancreatic beta cells. To detect functional areas of pancreatic beta cells that could be affected by exposure to DDT and DDE, we analyzed changes in protein expression in the NES2Y human pancreatic beta cell line exposed to three sublethal concentrations (0.1 μM, 1 μM, 10 μM) of DDT and DDE for 1 month. Protein separation and identification was achieved using high-resolution 2D-electrophoresis, computer analysis and mass spectrometry. With these techniques, four proteins were found downregulated after exposure to 10 μM DDT: three cytoskeletal proteins (cytokeratin 8, cytokeratin 18 and actin) and one protein involved in glycolysis (alpha-enolase). Two proteins were downregulated after exposure to 10 μM DDE: cytokeratin 18 and heterogenous nuclear ribonucleoprotein H1 (HNRH1). These changes correlate with previously described effects of other stress conditions (e.g. exposure to palmitate, hyperglycemia, imidazoline derivative, and cytokines) on protein expression in pancreatic beta cells. We conclude that cytoskeletal proteins and their processing, glucose metabolism, and mRNA processing may represent targets affected by exposure to conditions hostile to pancreatic beta cells, including exposure to DDT and DDE.

  4. Establishment and characterization of a new human pancreatic adenocarcinoma cell line with high metastatic potential to the lung

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    Schwarzenbach Heidi

    2010-06-01

    Full Text Available Abstract Background Pancreatic cancer is still associated with devastating prognosis. Real progress in treatment options has still not been achieved. Therefore new models are urgently needed to investigate this deadly disease. As a part of this process we have established and characterized a new human pancreatic cancer cell line. Methods The newly established pancreatic cancer cell line PaCa 5061 was characterized for its morphology, growth rate, chromosomal analysis and mutational analysis of the K-ras, EGFR and p53 genes. Gene-amplification and RNA expression profiles were obtained using an Affymetrix microarray, and overexpression was validated by IHC analysis. Tumorigenicity and spontaneous metastasis formation of PaCa 5061 cells were analyzed in pfp-/-/rag2-/- mice. Sensitivity towards chemotherapy was analysed by MTT assay. Results PaCa 5061 cells grew as an adhering monolayer with a doubling time ranging from 30 to 48 hours. M-FISH analyses showed a hypertriploid complex karyotype with multiple numerical and unbalanced structural aberrations. Numerous genes were overexpressed, some of which have previously been implicated in pancreatic adenocarcinoma (GATA6, IGFBP3, IGFBP6, while others were detected for the first time (MEMO1, RIOK3. Specifically highly overexpressed genes (fold change > 10 were identified as EGFR, MUC4, CEACAM1, CEACAM5 and CEACAM6. Subcutaneous transplantation of PaCa 5061 into pfp-/-/rag2-/- mice resulted in formation of primary tumors and spontaneous lung metastasis. Conclusion The established PaCa 5061 cell line and its injection into pfp-/-/rag2-/- mice can be used as a new model for studying various aspects of the biology of human pancreatic cancer and potential treatment approaches for the disease.

  5. Fetal liver blood flow distribution: role in human developmental strategy to prioritize fat deposition versus brain development.

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    Keith M Godfrey

    Full Text Available Among primates, human neonates have the largest brains but also the highest proportion of body fat. If placental nutrient supply is limited, the fetus faces a dilemma: should resources be allocated to brain growth, or to fat deposition for use as a potential postnatal energy reserve? We hypothesised that resolving this dilemma operates at the level of umbilical blood distribution entering the fetal liver. In 381 uncomplicated pregnancies in third trimester, we measured blood flow perfusing the fetal liver, or bypassing it via the ductus venosus to supply the brain and heart using ultrasound techniques. Across the range of fetal growth and independent of the mother's adiposity and parity, greater liver blood flow was associated with greater offspring fat mass measured by dual-energy X-ray absorptiometry, both in the infant at birth (r = 0.43, P<0.001 and at age 4 years (r = 0.16, P = 0.02. In contrast, smaller placentas less able to meet fetal demand for essential nutrients were associated with a brain-sparing flow pattern (r = 0.17, p = 0.02. This flow pattern was also associated with a higher degree of shunting through ductus venosus (P = 0.04. We propose that humans evolved a developmental strategy to prioritize nutrient allocation for prenatal fat deposition when the supply of conditionally essential nutrients requiring hepatic inter-conversion is limited, switching resource allocation to favour the brain if the supply of essential nutrients is limited. Facilitated placental transfer mechanisms for glucose and other nutrients evolved in environments less affluent than those now prevalent in developed populations, and we propose that in circumstances of maternal adiposity and nutrient excess these mechanisms now also lead to prenatal fat deposition. Prenatal developmental influences play important roles in the human propensity to deposit fat.

  6. Adult stromal cells derived from human adipose tissue provoke pancreatic cancer cell death both in vitro and in vivo.

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    Beatrice Cousin

    Full Text Available BACKGROUND: Normal tissue homeostasis is maintained by dynamic interactions between epithelial cells and their microenvironment. Disrupting this homeostasis can induce aberrant cell proliferation, adhesion, function and migration that might promote malignant behavior. Indeed, aberrant stromal-epithelial interactions contribute to pancreatic ductal adenocarcinoma (PDAC spread and metastasis, and this raises the possibility that novel stroma-targeted therapies represent additional approaches for combating this malignant disease. The aim of the present study was to determine the effect of human stromal cells derived from adipose tissue (ADSC on pancreatic tumor cell proliferation. PRINCIPAL FINDINGS: Co-culturing pancreatic tumor cells with ADSC and ADSC-conditioned medium sampled from different donors inhibited cancer cell viability and proliferation. ADSC-mediated inhibitory effect was further extended to other epithelial cancer-derived cell lines (liver, colon, prostate. ADSC conditioned medium induced cancer cell necrosis following G1-phase arrest, without evidence of apoptosis. In vivo, a single intra-tumoral injection of ADSC in a model of pancreatic adenocarcinoma induced a strong and long-lasting inhibition of tumor growth. CONCLUSION: These data indicate that ADSC strongly inhibit PDAC proliferation, both in vitro and in vivo and induce tumor cell death by altering cell cycle progression. Therefore, ADSC may constitute a potential cell-based therapeutic alternative for the treatment of PDAC for which no effective cure is available.

  7. Targeting pancreatic progenitor cells in human embryonic stem cell differentiation for the identification of novel cell surface markers.

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    Fishman, Bettina; Segev, Hanna; Kopper, Oded; Nissenbaum, Jonathan; Schulman, Margarita; Benvenisty, Nissim; Itskovitz-Eldor, Joseph; Kitsberg, Danny

    2012-09-01

    New sources of beta cells are needed in order to develop cell therapies for patients with diabetes. An alternative to forced expansion of post-mitotic beta cells is the induction of differentiation of stem-cell derived progenitor cells that have a natural self-expansion capacity into insulin-producing cells. In order to learn more about these progenitor cells at different stages along the differentiation process in which they become progressively more committed to the final beta cell fate, we took the approach of identifying, isolating and characterizing stage specific progenitor cells. We generated human embryonic stem cell (HESC) clones harboring BAC GFP reporter constructs of SOX17, a definitive endoderm marker, and PDX1, a pancreatic marker, and identified subpopulations of GFP expressing cells. Using this approach, we isolated a highly enriched population of pancreatic progenitor cells from hESCs and examined their gene expression with an emphasis on the expression of stage-specific cell surface markers. We were able to identify novel molecules that are involved in the pancreatic differentiation process, as well as stage-specific cell markers that may serve to define (alone or in combination with other markers) a specific pancreatic progenitor cell. These findings may help in optimizing conditions for ultimately generating and isolating beta cells for transplantation therapy.

  8. CXCL12 chemokine expression suppresses human pancreatic cancer growth and metastasis.

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    Ishan Roy

    Full Text Available Pancreatic ductal adenocarcinoma is an unsolved health problem with nearly 75% of patients diagnosed with advanced disease and an overall 5-year survival rate near 5%. Despite the strong link between mortality and malignancy, the mechanisms behind pancreatic cancer dissemination and metastasis are poorly understood. Correlative pathological and cell culture analyses suggest the chemokine receptor CXCR4 plays a biological role in pancreatic cancer progression. In vivo roles for the CXCR4 ligand CXCL12 in pancreatic cancer malignancy were investigated. CXCR4 and CXCR7 were consistently expressed in normal and cancerous pancreatic ductal epithelium, established cell lines, and patient-derived primary cancer cells. Relative to healthy exocrine ducts, CXCL12 expression was pathologically repressed in pancreatic cancer tissue specimens and patient-derived cell lines. To test the functional consequences of CXCL12 silencing, pancreatic cancer cell lines stably expressingthe chemokine were engineered. Consistent with a role for CXCL12 as a tumor suppressor, cells producing the chemokine wereincreasingly adherent and migration deficient in vitro and poorly metastatic in vivo, compared to control cells. Further, CXCL12 reintroduction significantly reduced tumor growth in vitro, with significantly smaller tumors in vivo, leading to a pronounced survival advantage in a preclinical model. Together, these data demonstrate a functional tumor suppressive role for the normal expression of CXCL12 in pancreatic ducts, regulating both tumor growth andcellulardissemination to metastatic sites.

  9. Examining the effects of hyperglycemia on pancreatic endocrine function in humans

    DEFF Research Database (Denmark)

    Solomon, Thomas P J; Knudsen, Sine H; Karstoft, Kristian;

    2012-01-01

    Investigating the impact of hyperglycemia on pancreatic endocrine function promotes our understanding of the pathophysiology of hyperglycemia-related disease.......Investigating the impact of hyperglycemia on pancreatic endocrine function promotes our understanding of the pathophysiology of hyperglycemia-related disease....

  10. Disabled-1 alternative splicing in human fetal retina and neural tumors.

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    Sachin Katyal

    Full Text Available BACKGROUND: The Reelin-Dab1 signaling pathway plays a critical role in the positioning of migrating neurons, dendrite formation and lamination in the developing central nervous system. We have previously identified two alternatively spliced forms of Dab1 in the developing chick retina: an early form, Dab1-E, expressed in retinal progenitor cells, and a late form, Dab1 or Dab1-L, expressed in amacrine and ganglion cells. Compared to Dab1-L, Dab1-E lacks two exons that encode two Src family kinase (SFK phosphorylation sites. PRINCIPAL FINDINGS: Both Dab1-L and Dab1-E-like transcripts were identified in human fetal retina. Expression of human Dab1-L in primary chick retinal cultures resulted in Reelin-mediated induction of SFK phosphorylation and formation of neurite-like processes. In contrast, human Dab1-E-expressing cells retained an undifferentiated morphology. The human Dab1 gene is located within a common fragile site, and it has been postulated that it may function as a tumor suppressor. Analysis of Dab1 splice forms in retinoblastoma and neuroblastoma tumor cells revealed relative enrichment of Dab1-L-like (includes exons 7 and 8 and Dab1-E-like (excludes exons 7 and 8 transcripts in retinoblastoma and neuroblastoma, respectively. Treatment of retinoblastoma cell line RB522A with Reelin resulted in increased tyrosine phosphorylation of Dab1. As Nova2 has previously been implicated in the exclusion of exons 9B and 9C in Dab1, we examined the expression of this splicing factor in neuroblastoma and retinoblastoma cell lines. Nova2 was only detected in neuroblastoma cells, suggesting a correlation between Nova2 expression and increased levels of Dab1-E-like splice forms in neuroblastoma. CONCLUSIONS: These results indicate that alternative splicing of Dab1 is conserved in avian and mammalian species, with Dab1-L driving SFK phosphorylation in both species. Dab1-E- and Dab-L-like isoforms are also expressed in childhood neural tumors, with

  11. Disabled-1 Alternative Splicing in Human Fetal Retina and Neural Tumors

    Science.gov (United States)

    Katyal, Sachin; Glubrecht, Darryl D.; Li, Lei; Gao, Zhihua; Godbout, Roseline

    2011-01-01

    Background The Reelin-Dab1 signaling pathway plays a critical role in the positioning of migrating neurons, dendrite formation and lamination in the developing central nervous system. We have previously identified two alternatively spliced forms of Dab1 in the developing chick retina: an early form, Dab1-E, expressed in retinal progenitor cells, and a late form, Dab1 or Dab1-L, expressed in amacrine and ganglion cells. Compared to Dab1-L, Dab1-E lacks two exons that encode two Src family kinase (SFK) phosphorylation sites. Principal Findings Both Dab1-L and Dab1-E-like transcripts were identified in human fetal retina. Expression of human Dab1-L in primary chick retinal cultures resulted in Reelin-mediated induction of SFK phosphorylation and formation of neurite-like processes. In contrast, human Dab1-E-expressing cells retained an undifferentiated morphology. The human Dab1 gene is located within a common fragile site, and it has been postulated that it may function as a tumor suppressor. Analysis of Dab1 splice forms in retinoblastoma and neuroblastoma tumor cells revealed relative enrichment of Dab1-L-like (includes exons 7 and 8) and Dab1-E-like (excludes exons 7 and 8) transcripts in retinoblastoma and neuroblastoma, respectively. Treatment of retinoblastoma cell line RB522A with Reelin resulted in increased tyrosine phosphorylation of Dab1. As Nova2 has previously been implicated in the exclusion of exons 9B and 9C in Dab1, we examined the expression of this splicing factor in neuroblastoma and retinoblastoma cell lines. Nova2 was only detected in neuroblastoma cells, suggesting a correlation between Nova2 expression and increased levels of Dab1-E-like splice forms in neuroblastoma. Conclusions These results indicate that alternative splicing of Dab1 is conserved in avian and mammalian species, with Dab1-L driving SFK phosphorylation in both species. Dab1-E- and Dab-L-like isoforms are also expressed in childhood neural tumors, with preferential enrichment

  12. Glucosyl epi-cyclophellitol allows mechanism-based inactivation and structural analysis of human pancreatic α-amylase.

    Science.gov (United States)

    Caner, Sami; Zhang, Xiaohua; Jiang, Jianbing; Chen, Hong-Ming; Nguyen, Nham T; Overkleeft, Hermen; Brayer, Gary D; Withers, Stephen G

    2016-04-01

    As part of a search for selective, mechanism-based covalent inhibitors of human pancreatic α-amylase we describe the chemoenzymatic synthesis of the disaccharide analog α-glucosyl epi-cyclophellitol, demonstrate its stoichiometric reaction with human pancreatic α-amylase and evaluate the time dependence of its inhibition. X-ray crystallographic analysis of the covalent derivative so formed confirms its reaction at the active site with formation of a covalent bond to the catalytic nucleophile D197. The structure illuminates the interactions with the active site and confirms OH4' on the nonreducing end sugar as a good site for attachment of fluorescent tags in generating probes for localization and quantitation of amylase in vivo.

  13. Acquisition of innate-like microbial reactivity in mucosal tissues during human fetal MAIT-cell development

    Science.gov (United States)

    Leeansyah, Edwin; Loh, Liyen; Nixon, Douglas F.; Sandberg, Johan K.

    2014-01-01

    Innate-like, evolutionarily conserved MR1-restricted mucosa-associated invariant T (MAIT) cells represent a large antimicrobial T-cell subset in humans. Here, we investigate the development of these cells in second trimester human fetal tissues. MAIT cells are rare and immature in the fetal thymus, spleen and mesenteric lymph nodes. In contrast, mature IL-18Rα+ CD8αα MAIT cells are enriched in the fetal small intestine, liver and lung. Independently of localization, MAIT cells express CD127 and Ki67 in vivo and readily proliferate in response to Escherichia coli in vitro. Maturation is accompanied by the gradual post-thymic acquisition of the PLZF transcription factor and the ability to produce IFNγ and IL-22 in response to bacteria in mucosa. Thus, MAIT cells acquire innate-like antimicrobial responsiveness in mucosa before exposure to environmental microbes and the commensal microflora. Establishment of this arm of immunity before birth may help protect the newborn from a range of pathogenic microbes.

  14. Fetal Research

    Science.gov (United States)

    Hansen, John T.; Sladek, John R.

    1989-11-01

    This article reviews some of the significant contributions of fetal research and fetal tissue research over the past 20 years. The benefits of fetal research include the development of vaccines, advances in prenatal diagnosis, detection of malformations, assessment of safe and effective medications, and the development of in utero surgical therapies. Fetal tissue research benefits vaccine development, assessment of risk factors and toxicity levels in drug production, development of cell lines, and provides a source of fetal cells for ongoing transplantation trials. Together, fetal research and fetal tissue research offer tremendous potential for the treatment of the fetus, neonate, and adult.

  15. In vitro differentiation potential of human haematopoietic CD34(+) cells towards pancreatic β-cells.

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    Sunitha, Manne Mudhu; Srikanth, Lokanathan; Santhosh Kumar, Pasupuleti; Chandrasekhar, Chodimella; Sarma, Potukuchi Venkata Gurunadha Krishna

    2016-10-01

    Haematopoietic stem cells (HSCs) possess multipotent ability to differentiate into various types of cells on providing appropriate niche. In the present study, the differentiating potential of human HSCs into β-cells of islets of langerhans was explored. Human HSCs were apheretically isolated from a donor and cultured. Phenotypic characterization of CD34 glycoprotein in the growing monolayer HSCs was confirmed by immunocytochemistry and flow cytometry techniques. HSCs were induced by selection with beta cell differentiating medium (BDM), which consists of epidermal growth factor (EGF), fibroblast growth factor (FGF), transferrin, Triiodo-l-Tyronine, nicotinamide and activin A. Distinct morphological changes of differentiated cells were observed on staining with dithizone (DTZ) and expression of PDX1, insulin and synaptophysin was confirmed by immunocytochemistry. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis revealed distinct expression of specific β-cell markers, pancreatic and duodenal homeobox-1 (PDX1), glucose transporter-2 (GLUT-2), synaptophysin (SYP) and insulin (INS) in these differentiated cells compared to HSCs. Further, these cells exhibited elevated expression of INS gene at 10 mM glucose upon inducing with different glucose concentrations. The prominent feature of the obtained β-cells was the presence of glucose sensors, which was determined by glucokinase activity and high glucokinase activity compared with CD34(+) stem cells. These findings illustrate the differentiation of CD34(+) HSCs into β-cells of islets of langerhans.

  16. Human pancreatic cancer tumors are nutrient poor and tumor cells actively scavenge extracellular protein.

    Science.gov (United States)

    Kamphorst, Jurre J; Nofal, Michel; Commisso, Cosimo; Hackett, Sean R; Lu, Wenyun; Grabocka, Elda; Vander Heiden, Matthew G; Miller, George; Drebin, Jeffrey A; Bar-Sagi, Dafna; Thompson, Craig B; Rabinowitz, Joshua D

    2015-02-01

    Glucose and amino acids are key nutrients supporting cell growth. Amino acids are imported as monomers, but an alternative route induced by oncogenic KRAS involves uptake of extracellular proteins via macropinocytosis and subsequent lysosomal degradation of these proteins as a source of amino acids. In this study, we examined the metabolism of pancreatic ductal adenocarcinoma (PDAC), a poorly vascularized lethal KRAS-driven malignancy. Metabolomic comparisons of human PDAC and benign adjacent tissue revealed that tumor tissue was low in glucose, upper glycolytic intermediates, creatine phosphate, and the amino acids glutamine and serine, two major metabolic substrates. Surprisingly, PDAC accumulated essential amino acids. Such accumulation could arise from extracellular proteins being degraded through macropinocytosis in quantities necessary to meet glutamine requirements, which in turn produces excess of most other amino acids. Consistent with this hypothesis, active macropinocytosis is observed in primary human PDAC specimens. Moreover, in the presence of physiologic albumin, we found that cultured murine PDAC cells grow indefinitely in media lacking single essential amino acids and replicate once in the absence of free amino acids. Growth under these conditions was characterized by simultaneous glutamine depletion and essential amino acid accumulation. Overall, our findings argue that the scavenging of extracellular proteins is an important mode of nutrient uptake in PDAC.

  17. Human immunodeficiency virus type 1 efficiently binds to human fetal astrocytes and induces neuroinflammatory responses independent of infection

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    Potash Mary

    2007-05-01

    Full Text Available Abstract Background HIV-1 infects human astrocytes in vitro and in vivo but the frequency of infected cells is low and its biological significance is unknown. In studies in vitro, recombinant gp120 alone can induce profound effects on astrocyte biology, suggesting that HIV-1 interaction with astrocytes and its functional consequences extend beyond the limited levels of infection in these cells. Here we determined the relative efficiencies of HIV-1 binding and infection in human fetal astrocytes (HFA, mainly at the single cell level, using HIV-1 tagged with green fluorescence protein (GFP-Vpr fusion proteins, termed HIV-GFP, to detect virus binding and HIV-1 expressing Rev and NefGFP fusion proteins to detect productive infection. Results Essentially all HFA in a population bound HIV-GFP specifically and independently of CCR5 and CXCR4. The dynamics of this binding at 37°C resembled binding of an HIV fusion mutant to CD4-positive cells, indicating that most of HIV-GFP arrested infection of HFA at the stage of virus-cell fusion. Despite extensive binding, only about 1% of HFA were detectably infected by HIV-RevGFP or HIV-NefGFP, but this proportion increased to the majority of HFA when the viruses were pseudotyped with vesicular stomatitis virus envelope glycoprotein G, confirming that HFA impose a restriction upon HIV-1 entry. Exposure of HFA to HIV-1 through its native proteins rapidly induced synthesis of interleukin-6 and interleukin-8 with increased mRNA detected within 3 h and increased protein detected within 18 h of exposure. Conclusion Our results indicate that HIV-1 binding to human astrocytes, although extensive, is not generally followed by virus entry and replication. Astrocytes respond to HIV-1 binding by rapidly increased cytokine production suggesting a role of this virus-brain cell interaction in HIV-1 neuropathogenesis.

  18. Fetal development of the elastic-fiber-mediated enthesis in the human middle ear.

    Science.gov (United States)

    Takanashi, Yoshitaka; Shibata, Shunichi; Katori, Yukio; Murakami, Gen; Abe, Shinichi; Rodríguez-Vázquez, Jose Francisco; Kawase, Tetsuaki

    2013-10-01

    In the human middle ear, the annular ligament of the incudostapedial joint and the insertions of the tensor tympani and stapedius muscles contain abundant elastic fibers; i.e., the elastic-fiber-mediated entheses. Hyaluronan also coexists with the elastic fibers. In the present study using immunohistochemistry, we demonstrated the distribution of elastin not only in the incudostapedial joint but also in the other two joints of the middle ear in adults and fetuses. In adults, the expression of elastin did not extend out of the annular ligament composed of mature elastic fibers but clearly overlapped with it. Electron microscopic observations of the annular ligament demonstrated a few microfibrils along the elastic fibers. Thus, in contrast to the vocal cord, the middle ear entheses seemed not to contain elaunin and oxytalan fibers. In mid-term fetuses (at approximately 15-16 weeks of gestation) before opening of the external acoustic meatus, the incudostapedial joint showed abundant elastic fibers, but the incudomalleolar and stapediovestibular joints did not. At this stage, hyaluronan was not colocalized, but distributed diffusely in loose mesenchymal tissues surrounding the ear ossicles. Therefore, fetal development of elastin and elastic fibers in the middle ear entheses is unlikely to require acoustic oscillation. In late-stage fetuses (25-30 weeks), whose ear ossicles were almost the same size as those in adults, we observed bundling and branching of elastic fibers. However, hyaluronan expression was not as strong as in adults. Colocalization between elastic fibers and hyaluronan appeared to be a result of postnatal maturation of the entheses.

  19. MORPHOLOGICAL FEATURES AND MORPHOMETRIC PARAMETERS OF HUMAN FETAL VERMIFORM APPENDIX AT DIFFERENT GESTATIONAL

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    Reshma.Mohammad

    2013-09-01

    Full Text Available Background:Vermiform appendix is a vestigial organ of variable position in the abdomen. Its location, size andshape are subject to alterations with the race of the population and limited information is available on devel-opmental morphology and morphometry of fetal appendix.Materials and Methods:In the present study 60appendix specimens from aborted human fetuses of 17-40 weeks gestational age and both sexes were studiedby dissection method for age related morphological features and morphometric parameters. The morphologi-cal parameters observed include its location in relation to abdominal region, caecum and ileum, clock position,position of base in relation to caecal wall and direction of tip of appendix. The morphometric parameters oflength, diameter and distance between ileo-caecal orifice and appendicular orifice were measured.Results:The location of appendix in relation to abdominal region presented higher incidence of sub-hepatic position inless than 30 weeks fetuses and right iliac fossa location in more than 30 weeks fetuses.Discussion: in comparisionwith the literature available on adult vermiform appendix the observations in the present study arein favor ofinfluence of developmental processes on the localization of appendix including its base, ileo-caecalorifice,direction of tip, distance from McBurney’s point.Conclusion:Results of this work suggests variability in local-ization of appendix during prenatal development and the influence of gestational age, sex, size, growth ofcaecum and gut on its final position and was different from that of adults. There is increase in themorphomet-ric parameters of appendix with increase in gestational age. Both morphological and morphometric param-eters were different between sexes.

  20. The Metastatic Potential and Chemoresistance of Human Pancreatic Cancer Stem Cells.

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    Vikash J Bhagwandin

    Full Text Available Cancer stem cells (CSCs typically have the capacity to evade chemotherapy and may be the principal source of metastases. CSCs for human pancreatic ductal carcinoma (PDAC have been identified, but neither the metastatic potential nor the chemoresistance of these cells has been adequately evaluated. We have addressed these issues by examining side-population (SP cells isolated from the Panc-1 and BxPC3 lines of human PDAC cells, the oncogenotypes of which differ. SP cells could be isolated from monolayers of Panc-1, but only from spheroids of BxPC3. Using orthotopic xenografts into the severely immunocompromised NSG mouse, we found that SP cells isolated from both cell lines produced tumors that were highly metastatic, in contrast to previous experience with PDAC cell lines. SP cells derived from both cell lines expressed the ABCG2 transporter, which was demonstrably responsible for the SP phenotype. SP cells gave rise to non-SP (NSP cells in vitro and in vivo, a transition that was apparently due to posttranslational inhibition of the ABCG2 transporter. Twenty-two other lines of PDAC cells also expressed ABCG2. The sensitivity of PDAC SP cells to the vinca alkaloid vincristine could be greatly increased by verapamil, a general inhibitor of transporters. In contrast, verapamil had no effect on the killing of PDAC cells by gemcitabine, the current first-line therapeutic for PDAC. We conclude that the isolation of SP cells can be a convenient and effective tool for the study of PDAC CSCs; that CSCs may be the principal progenitors of metastasis by human PDAC; that the ABCG2 transporter is responsible for the SP phenotype in human PDAC cells, and may be a ubiquitous source of drug-resistance in PDAC, but does not confer resistance to gemcitabine; and that inhibition of ABCG2 might offer a useful adjunct in a therapeutic attack on the CSCs of PDAC.

  1. The Metastatic Potential and Chemoresistance of Human Pancreatic Cancer Stem Cells.

    Science.gov (United States)

    Bhagwandin, Vikash J; Bishop, J Michael; Wright, Woodring E; Shay, Jerry W

    2016-01-01

    Cancer stem cells (CSCs) typically have the capacity to evade chemotherapy and may be the principal source of metastases. CSCs for human pancreatic ductal carcinoma (PDAC) have been identified, but neither the metastatic potential nor the chemoresistance of these cells has been adequately evaluated. We have addressed these issues by examining side-population (SP) cells isolated from the Panc-1 and BxPC3 lines of human PDAC cells, the oncogenotypes of which differ. SP cells could be isolated from monolayers of Panc-1, but only from spheroids of BxPC3. Using orthotopic xenografts into the severely immunocompromised NSG mouse, we found that SP cells isolated from both cell lines produced tumors that were highly metastatic, in contrast to previous experience with PDAC cell lines. SP cells derived from both cell lines expressed the ABCG2 transporter, which was demonstrably responsible for the SP phenotype. SP cells gave rise to non-SP (NSP) cells in vitro and in vivo, a transition that was apparently due to posttranslational inhibition of the ABCG2 transporter. Twenty-two other lines of PDAC cells also expressed ABCG2. The sensitivity of PDAC SP cells to the vinca alkaloid vincristine could be greatly increased by verapamil, a general inhibitor of transporters. In contrast, verapamil had no effect on the killing of PDAC cells by gemcitabine, the current first-line therapeutic for PDAC. We conclude that the isolation of SP cells can be a convenient and effective tool for the study of PDAC CSCs; that CSCs may be the principal progenitors of metastasis by human PDAC; that the ABCG2 transporter is responsible for the SP phenotype in human PDAC cells, and may be a ubiquitous source of drug-resistance in PDAC, but does not confer resistance to gemcitabine; and that inhibition of ABCG2 might offer a useful adjunct in a therapeutic attack on the CSCs of PDAC.

  2. Severe Acute Pancreatitis in Pregnancy

    Directory of Open Access Journals (Sweden)

    Bahiyah Abdullah

    2015-01-01

    Full Text Available This is a case of a pregnant lady at 8 weeks of gestation, who presented with acute abdomen. She was initially diagnosed with ruptured ectopic pregnancy and ruptured corpus luteal cyst as the differential diagnosis. However she then, was finally diagnosed as acute hemorrhagic pancreatitis with spontaneous complete miscarriage. This is followed by review of literature on this topic. Acute pancreatitis in pregnancy is not uncommon. The emphasis on high index of suspicion of acute pancreatitis in women who presented with acute abdomen in pregnancy is highlighted. Early diagnosis and good supportive care by multidisciplinary team are crucial to ensure good maternal and fetal outcomes.

  3. Liposomal insulin promoter-thymidine kinase gene therapy followed by ganciclovir effectively ablates human pancreatic cancer in mice.

    Science.gov (United States)

    Wu, James X; Liu, Shi-He; Nemunaitis, John J; Brunicardi, F Charles

    2015-04-10

    PDX1 is overexpressed in pancreatic cancer, and activates the insulin promoter (IP). Adenoviral IP-thymidine kinase and ganciclovir (TK/GCV) suppresses human pancreatic ductal carcinoma (PDAC) in mice, but repeated doses carry significant toxicity. We hypothesized that multiple cycles of liposomal IP-TK/GCV ablate human PDAC in SCID mice with minimal toxicity compared to adenoviral IP-TK/GCV. SCID mice with intraperitoneal human pancreatic cancer PANC-1 tumor implants were given a single cycle of 35 µg iv L-IP-TK, or four cycles of 1, 10, 20, 30, or 35 µg iv L-IP-TK (n = 20 per group), followed by intraperitoneal GCV. Insulin and glucose levels were monitored in mice treated with four cycles of 35 µg iv L-IP-TK. We found that four cycles of 10-35 µg L-IP-TK/GCV ablated more PANC-1 tumor volume compared to a single cycle with 35 µg. Mice that received four cycles of 10 µg L-IP-TK demonstrated the longest survival (P SCID mice with minimal toxicity, suggesting non-viral vectors are superior to adenoviral vectors for IP-gene therapy.

  4. Loss of canonical Smad4 signaling promotes KRAS driven malignant transformation of human pancreatic duct epithelial cells and metastasis.

    Directory of Open Access Journals (Sweden)

    Lisa Leung

    Full Text Available Pancreatic ductal adenocarcinoma (PDAC is the fourth most common cause of cancer death in North America. Activating KRAS mutations and Smad4 loss occur in approximately 90% and 55% of PDAC, respectively. While their roles in the early stages of PDAC development have been confirmed in genetically modified mouse models, their roles in the multistep malignant transformation of human pancreatic duct cells have not been directly demonstrated. Here, we report that Smad4 represents a barrier in KRAS-mediated malignant transformation of the near normal immortalized human pancreatic duct epithelial (HPDE cell line model. Marked Smad4 downregulation by shRNA in KRAS (G12V expressing HPDE cells failed to cause tumorigenic transformation. However, KRAS-mediated malignant transformation occurred in a new HPDE-TGF-β resistant (TβR cell line that completely lacks Smad4 protein expression and is resistant to the mito-inhibitory activity of TGF-β. This transformation resulted in tumor formation and development of metastatic phenotype when the cells were implanted orthotopically into the mouse pancreas. Smad4 restoration re-established TGF-β sensitivity, markedly increased tumor latency by promoting apoptosis, and decreased metastatic potential. These results directly establish the critical combination of the KRAS oncogene and complete Smad4 inactivation in the multi-stage malignant transformation and metastatic progression of normal human HPDE cells.

  5. The protective effect of ursodeoxycholic acid in an in vitro model of the human fetal heart occurs via targeting cardiac fibroblasts.

    Science.gov (United States)

    Schultz, Francisca; Hasan, Alveera; Alvarez-Laviada, Anita; Miragoli, Michele; Bhogal, Navneet; Wells, Sarah; Poulet, Claire; Chambers, Jenny; Williamson, Catherine; Gorelik, Julia

    2016-01-01

    Bile acids are elevated in the blood of women with intrahepatic cholestasis of pregnancy (ICP) and this may lead to fetal arrhythmia, fetal hypoxia and potentially fetal death in utero. The bile acid taurocholic acid (TC) causes abnormal calcium dynamics and contraction in neonatal rat cardiomyocytes. Ursodeoxycholic acid (UDCA), a drug clinically used to treat ICP, prevents adverse effects of TC. During development, the fetus is in a state of relative hypoxia. Although this is essential for the development of the heart and vasculature, resident fibroblasts can transiently differentiate into myofibroblasts and form gap junctions with cardiomyocytes in vitro, resulting in cardiomyocyte depolarization. We expanded on previously published work using an in vitro hypoxia model to investigate the differentiation of human fetal fibroblasts into myofibroblasts. Recent evidence shows that potassium channels are involved in maintaining the membrane potential of ventricular fibroblasts and that ATP-dependent potassium (KATP) channel subunits are expressed in cultured fibroblasts. KATP channels are a valuable target as they are thought to have a cardioprotective role during ischaemic and hypoxic conditions. We investigated whether UDCA could modulate fibroblast membrane potential. We established the isolation and culture of human fetal cardiomyocytes and fibroblasts to investigate the effect of hypoxia, TC and UDCA on human fetal cardiac cells. UDCA hyperpolarized myofibroblasts and prevented TC-induced depolarisation, possibly through the activation of KATP channels that are expressed in cultured fibroblasts. Also, similar to the rat model, UDCA can counteract TC-induced calcium abnormalities in human fetal cultures of cardiomyocytes and myofibroblasts. Under normoxic conditions, we found a higher number of myofibroblasts in cultures derived from human fetal hearts compared to cells isolated from neonatal rat hearts, indicating a possible increased number of myofibroblasts

  6. Pancreatic Cancer

    Science.gov (United States)

    ... hormones that help control blood sugar levels. Pancreatic cancer usually begins in the cells that produce the juices. Some risk factors for developing pancreatic cancer include Smoking Long-term diabetes Chronic pancreatitis Certain ...

  7. Pancreatic pseudocysts

    African Journals Online (AJOL)

    at formulating a classification that can be used to dictate treatment strategy ... standing of the natural history of pancreatic fluid collec- ... ring, protein plug, or stone, the ongoing pancreatic secretion ... exocrine function in chronic pancreatitis.

  8. LIN28 is selectively expressed by primordial and pre-meiotic germ cells in the human fetal ovary.

    Science.gov (United States)

    Childs, Andrew J; Kinnell, Hazel L; He, Jing; Anderson, Richard A

    2012-09-01

    Germ cell development requires timely transition from primordial germ cell (PGC) self-renewal to meiotic differentiation. This is associated with widespread changes in gene expression, including downregulation of stem cell-associated genes, such as OCT4 and KIT, and upregulation of markers of germ cell differentiation and meiosis, such as VASA, STRA8, and SYCP3. The stem cell-expressed RNA-binding protein Lin28 has recently been demonstrated to be essential for PGC specification in mice, and LIN28 is expressed in human germ cell tumors with phenotypic similarities to human fetal germ cells. We have therefore examined the expression of LIN28 during normal germ cell development in the human fetal ovary, from the PGC stage, through meiosis to the initiation of follicle formation. LIN28 transcript levels were highest when the gonad contained only PGCs, and decreased significantly with increasing gestation, coincident with the onset of germ cell differentiation. Immunohistochemistry revealed LIN28 protein expression to be germ cell-specific at all stages examined. All PGCs expressed LIN28, but at later gestations expression was restricted to a subpopulation of germ cells, which we demonstrate to be primordial and premeiotic germ cells based on immunofluorescent colocalization of LIN28 and OCT4, and absence of overlap with the meiosis marker SYCP3. We also demonstrate the expression of the LIN28 target precursor pri-microRNA transcripts pri-LET7a/f/d and pri-LET-7g in the human fetal ovary, and that expression of these is highest at the PGC stage, mirroring that of LIN28. The spatial and temporal restriction of LIN28 expression and coincident peaks of expression of LIN28 and target pri-microRNAs suggest important roles for this protein in the maintenance of the germline stem cell state and the regulation of microRNA activity in the developing human ovary.

  9. Bone morphogenetic protein-4 and 7 and receptors regulate vascular endothelial growth factor and receptors in human fetal leptomeninges.

    Science.gov (United States)

    Johnson, Mahlon D; Reeder, Jay E; O'Connell, Mary

    2015-10-08

    Bone morphogenetic proteins (BMP) 4 and 7 have important roles in neuronal differentiation and cortical development in the murine brain. However BMP4 and BMP7 expression and functions in the developing human brain are unknown. In this study, frozen tissue human fetal leptomeninges, formalin-fixed tissue and primary fetal leptomeningeal cell cultures were studied. By western blot, BMP4, BMP7 and BMPRIa were demonstrated in 15, 17 20, 23 week (wk) human leptomeninges. BMP receptor II was detected at 15 and 17 wks. Immunohistochemically, BMP4 immunoreactivity was also found in 20 to 39 wk human leptomeninges. BMP4 significantly reduced basal DNA synthesis at 22 wks. BMP7 100 and 300 ng/ml stimulated basal DNA synthesis in the 15, 17 and 22 wk leptomeninges. BMP4 and BMP7 increased phosphorylation of SMAD-1, 5, 8 in most cells and had no effect on phosphorylation of p-38MAPK, or p44/42MAPK. BMP4 and BMP7 produced a decrease in VEGF RNA expression in 2 of 4 leptomeninges. BMP4 and BMP7 increased VEGFR1 RNA in 2 or 3 of 4 leptomeningeal cultures respectively. BMP4 produced a decrease in VEGFR2 RNA in 2 of 4 and BMP7 in 3 of 4 while BMP7 reduced VEGFR2 protein in the leptomeninges. The findings show, for the first time, BMP4, BMP7 and receptors are expressed and active in the human fetal leptomeninges. They suggest these BMPs influence vascular development in this tissue by regulating VEGF and its receptors. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  10. Expansion of tumor-infiltrating lymphocytes (TIL) from human pancreatic tumors.

    Science.gov (United States)

    Hall, MacLean; Liu, Hao; Malafa, Mokenge; Centeno, Barbara; Hodul, Pamela J; Pimiento, José; Pilon-Thomas, Shari; Sarnaik, Amod A

    2016-01-01

    We evaluated whether tumor infiltrating lymphocytes (TIL) could be expanded from surgically resected tumors from pancreatic cancer patients. Tumors were resected from pancreatic cancer patients. Tumors were minced into fragments and cultured in media containing high dose interleukin-2 (IL-2) for up to 6 weeks. T cell phenotype, activation markers, and reactivity were measured. TIL expansion was measured in 19 patient samples. The majority of these TIL were CD4(+) T cells and were highly activated. Purified CD8(+) T cells produced IFN-γ in response to HLA-matched pancreatic tumor targets. PD-1 blockade and 4-1BB stimulation were demonstrated as effective strategies to improve effective TIL yield, including the production of tumor-reactive pancreatic TIL. TIL expanded from pancreatic tumors are functional and able to respond to pancreatic tumor associated antigens. PD-1 blockade, 41BB stimulation, and CD8(+) T cell enrichment are effective strategies to improve TIL yield and tumor reactivity. These results support the development of adoptive cell therapy strategies using TIL for the treatment of pancreatic cancer.

  11. Protocadherin 11X/Y a human-specific gene pair: an immunohistochemical survey of fetal and adult brains.

    Science.gov (United States)

    Priddle, Thomas H; Crow, Tim J

    2013-08-01

    Protocadherins 11X and 11Y are cell adhesion molecules of the δ1-protocadherin family. Pcdh11X is present throughout the mammalian radiation; however, 6 million years ago (MYA), a reduplicative translocation of the Xq21.3 block onto what is now human Yp11 created the Homo sapiens-specific PCDH11Y. Therefore, modern human females express PCDH11X whereas males express both PCDH11X and PCDH11Y. PCDH11X/Y has been subject to accelerated evolution resulting in human-specific changes to both proteins, most notably 2 cysteine substitutions in the PCDH11X ectodomain that may alter binding characteristics. The PCDH11X/Y gene pair is postulated to be critical to aspects of human brain evolution related to the neural correlates of language. Therefore, we raised antibodies to investigate the temporal and spatial expression of PCDH11X/Y in cortical and sub-cortical areas of the human fetal brain between 12 and 34 postconceptional weeks. We then used the antibodies to determine if this expression was consistent in a series of adult brains. PCDH11X/Y immunoreactivity was detectable at all developmental stages. Strong expression was detected in the fetal neocortex, ganglionic eminences, cerebellum, and inferior olive. In the adult brain, the cerebral cortex, hippocampal formation, and cerebellum were strongly immunoreactive, with expression also detectable in the brainstem.

  12. Characterization of human fetal cord blood steroid profiles in relation to fetal sex and mode of delivery using temperature-dependent inclusion chromatography and principal component analysis (PCA).

    Science.gov (United States)

    Clifton, Vicki L; Bisits, Andrew; Zarzycki, Paweł K

    2007-08-15

    In the present work, human male and female fetal cord blood samples were purified, selectively extracted and separated to examine a fraction of steroids ranging from polar estetrol to relatively non-polar progesterone using solid phase extraction based on C-18 tubes and beta-cyclodextrin driven temperature dependent inclusion chromatography. Resulting UV diode array chromatographic patterns revealed the presence of 27 peaks. Chromatographic patterns of UV detected steroids were analyzed using principal components analysis which revealed differences between male/female and labour/not-in-labour clusters. Quantitative analysis of nine identified steroids including: estetrol, 17beta-estradiol, estrone, estriol, cortisol, cortisone, progesterone, 20 alpha-hydroxyprogesterone and 17 alpha-hydroxyprogesterone were not significantly different between males and females. Significant differences between male and female fetuses were related to as yet unidentified compounds. Four peaks were significantly different with labour which corresponded with cortisol, cortisone and two unidentified compounds. This protocol may distinguish significant differences between clinical groups that are not readily identifiable using univariate measurements of single steroids or different low molecular mass biomarkers. Moreover, we have provided new evidence that despite the absence of testosterone there are number of steroids and low molecular mass compounds that differ between male and female fetuses.

  13. In the absence of nutrients, pancreatic-biliary secretions in the jejunum do not exert feedback control of human pancreatic or gastric function.

    Science.gov (United States)

    Krawisz, B R; Miller, L J; DiMagno, E P; Go, V L

    1980-01-01

    Feedback inhibition of basal pancreatic enzyme secretion by luminal pancreatic enzymes appears to be an important regulator of pancreatic secretion in some laboratory animals. To determine whether pancreatic enzymes in the jejunum influence pancreatic or gastric functions in healthy man, we intubated six subjects with a gastric sump tube and a four-lumen duodenal tube which provided (1) a duodenal perfusion site, (2) a duodenal aspiration site, (3) an inflatable balloon immediately distal to the aspiration site, and (4) a jejunal perfusion site immediately beyond the balloon. In this way, the gastroduodenal segment could be functionally separated from the remainder of the intestine. The jejunum was exposed to normal saline, active pancreatic-biliary secretions, or pancreatic-biliary secretions in which the enzymes had been inactivated by heat. Ten minutes after initiation of each jejunal perfusion, normal saline was instilled into the stomach. No differences in trypsin secretion, gastric acid secretion, or gastric emptying occurred with the different jejunal perfusates. We therefore conclude that normal man, in the absence of intraluminal nutrients, does not exhibit a jejunal pancreatic enzyme-dependent feedback control mechanism for pancreatic enzyme or gastric secretion. However, our study does not exclude the possibility of a duodenal feedback regulatory mechanism.

  14. Human fetal exposure to triclosan and triclocarban in an urban population from Brooklyn, New York.

    Science.gov (United States)

    Pycke, Benny F G; Geer, Laura A; Dalloul, Mudar; Abulafia, Ovadia; Jenck, Alizee M; Halden, Rolf U

    2014-01-01

    Triclosan (TCS) and triclocarban (TCC) are antimicrobial agents formulated in a wide variety of consumer products (including soaps, toothpaste, medical devices, plastics, and fabrics) that are regulated by the U.S. Food and Drug Administration (FDA) and U.S. Environmental Protection Agency. In late 2014, the FDA will consider regulating the use of both chemicals, which are under scrutiny regarding lack of effectiveness, potential for endocrine disruption, and potential contribution to bacterial resistance to antibiotics. Here, we report on body burdens of TCS and TCC resulting from real-world exposures during pregnancy. Using liquid chromatography tandem mass spectrometry, we determined the concentrations of TCS, TCC, and its human metabolites (2'-hydroxy-TCC and 3'-hydroxy-TCC) as well as the manufacturing byproduct (3'-chloro-TCC) as total concentrations (Σ-) after conjugate hydrolysis in maternal urine and cord blood plasma from a cohort of 181 expecting mother/infant pairs in an urban multiethnic population from Brooklyn, NY recruited in 2007-09. TCS was detected in 100% of urine and 51% of cord blood samples after conjugate hydrolysis. The interquartile range (IQR) of detected TCS concentrations in urine was highly similar to the IQR reported previously for the age-matched population of the National Health and Nutrition Examination Survey (NHANES) from 2003 to 2004, but typically higher than the IQR reported previously for the general population (detection frequency = 74.6%). Urinary levels of TCC are reported here for the first time from real-world exposures during pregnancy, showing a median concentration of 0.21 μg/L. Urinary concentrations of TCC correlated well with its phase-I metabolite ∑-2'-hydroxy-TCC (r = 0.49) and the manufacturing byproduct ∑-3'-chloro-TCC C (r = 0.79), and ∑-2'-hydroxy-TCC correlated strongly with ∑-3'-hydroxy-TCC (r = 0.99). This human biomonitoring study presents the first body burden data for TCC from exposures

  15. Optimization of The Electroporation Conditions for Transfection of Human Factor IX into The Goat Fetal Fibroblasts

    Directory of Open Access Journals (Sweden)

    Amir Amiri Yekta

    2013-01-01

    Full Text Available Objective: Electroporation is the most common method used for the transfection of DNA. Although this method has been attributed for various cell using different buffer compositions, the effects of DNA concentration on the transfection efficiency has not yet been studied. Here the correlation between the efficiency of electroporation reaction and increments of DNA concentration has been investigated. Following this investigation, a study was set out to produce a transgenic goat which is capable of secreting recombinant human coagulation factor IX in its milk.Materials and Methods: In this experimental study, a linearized recombinant vector (pBC1 entailing human coagulation factor IX (5.7 kb cDNA was introduced into goat fetal fibroblast cells (three sub passages via electroporation in four separate experiments (while other variables were kept constant, beginning with 10 μg DNA per pulse in the first experiment and increments of 10 μg/pulse for the next three reactions. Thereafter, polymerase chain reaction (PCR-positive cell culture plates were diluted by several factors for further analysis of the transfected wells. Subsequently, positive cells were isolated for fluorescence in situ hybridization. Logistic regression model was used for data analyzing. Significance was defined as p< 0.05.Results: The results showed no significant difference among three first concentrations except for group 1 (10 μg/pulse and group 3 (30 μg/pulse, but there was a significant difference between these groups and the fourth group (p<0.05, as this group (40 μg/pulse statistically showed the highest rate of transfection. As the fluorescence in situ hybridization (FISH results indicated the transgene was integrated in a single position in PCR positive cells.Conclusion: Increasing amount of using the vector 40μg/pulse efficiently increased the number of transfected cells. Besides of voltage and buffer strength which had been previously shown to play a critical role

  16. Histo-blood group antigens in human fetal thymus and in thymomas

    DEFF Research Database (Denmark)

    Engel, P; Dabelsteen, Erik; Francis, D

    1996-01-01

    -y, Le-x and sialyl-Le-x) of the ABO-histo-blood group system was investigated in 19 normal fetal thymuses (gestational age 16 to 39 weeks) and in 19 thymomas in order to study possible tumor-associated changes in the glycosylation pattern. The material was investigated by immunochemical stainings...... of formalin-fixed paraffin-imbedded tissue using monoclonal antibodies with defined specificity. In fetal thymus the epithelial cells of the medulla and the Hassal's bodies strongly expressed elongated carbohydrate structures (Le-y, Le-x and sialyl-Le-x). In a few cases the cortical epithelial cells weakly...

  17. Acute Pancreatitis and Pregnancy

    Science.gov (United States)

    ... Pancreatitis Acute Pancreatitis and Pregnancy Acute Pancreatitis and Pregnancy Timothy Gardner, MD Acute pancreatitis is defined as ... pancreatitis in pregnancy. Reasons for Acute Pancreatitis and Pregnancy While acute pancreatitis is responsible for almost 1 ...

  18. Crystal structures of human pancreatic alpha-amylase in complex with carbohydrate and proteinaceous inhibitors.

    Science.gov (United States)

    Nahoum, V; Roux, G; Anton, V; Rougé, P; Puigserver, A; Bischoff, H; Henrissat, B; Payan, F

    2000-01-01

    Crystal structures of human pancreatic alpha-amylase (HPA) in complex with naturally occurring inhibitors have been solved. The tetrasaccharide acarbose and a pseudo-pentasaccharide of the trestatin family produced identical continuous electron densities corresponding to a pentasaccharide species, spanning the -3 to +2 subsites of the enzyme, presumably resulting from transglycosylation. Binding of the acarviosine core linked to a glucose residue at subsites -1 to +2 appears to be a critical part of the interaction process between alpha-amylases and trestatin-derived inhibitors. Two crystal forms, obtained at different values of pH, for the complex of HPA with the protein inhibitor from Phaseolus vulgaris (alpha-amylase inhibitor) have been solved. The flexible loop typical of the mammalian alpha-amylases was shown to exist in two different conformations, suggesting that loop closure is pH-sensitive. Structural information is provided for the important inhibitor residue, Arg-74, which has not been observed previously in structural analyses. PMID:10657258

  19. Epidrug-induced upregulation of functional somatostatin type 2 receptors in human pancreatic neuroendocrine tumor cells.

    Science.gov (United States)

    Veenstra, Marije J; van Koetsveld, Peter M; Dogan, Fadime; Farrell, William E; Feelders, Richard A; Lamberts, Steven W J; de Herder, Wouter W; Vitale, Giovanni; Hofland, Leo J

    2016-05-19

    Somatostatin receptors are a pivotal target for treatment of pancreatic neuroendocrine tumors (pNET), either with somatostatin analogues (SSA) or radiolabeled SSA. The highest affinity target for the most commonly used SSA is the somatostatin receptor type 2 (sst2). An important factor that may complicate treatment efficacy, is the variable number of receptors expressed on pNETs. Gene expression is subject to complex regulation, in which epigenetics has a central role. In this study we explored the possible role of epigenetic modifications in the variations in sst2 expression levels in two human pNET cell lines, BON-1 and QGP-1. We found upregulation of sst2 mRNA after treatment with the epidrugs 5-aza-2'-deoxycytidine (5-aza-dC) and valproic acid (VPA), an increased uptake of radiolabeled octreotide, as well as increased sensitivity to the SSA octreotide in functional cAMP inhibition. At epigenetic level we observed low methylation levels of the sst2 gene promoter region irrespective of expression. Activating histone mark H3K9Ac can be regulated with epidrug treatment, with an angle of effect corresponding to the effect on mRNA expression. Repressive histone mark H3K27me3 is not regulated by either 5-aza-dC or VPA. We conclude that epidrug treatment, in particular with combined 5-aza-dC and VPA treatment, might hold promise for improving and adding to current SSA treatment strategies of patients with pNETs.

  20. The nuclear transport capacity of a human-pancreatic ribonuclease variant is critical for its cytotoxicity.

    Science.gov (United States)

    Tubert, Pere; Rodríguez, Montserrat; Ribó, Marc; Benito, Antoni; Vilanova, Maria

    2011-10-01

    We have previously described a human pancreatic-ribonuclease variant, named PE5, which carries a non-contiguous extended bipartite nuclear localization signal. This signal comprises residues from at least three regions of the protein. We postulated that the introduction of this signal in the ribonuclease provides it with cytotoxic activity because although the variant poorly evades the ribonuclease inhibitor in vitro, it is routed to the nucleus, which is devoid of the inhibitor. In this work, we have investigated the relationship between the cytotoxicity produced by PE5 and its ability to reach the nucleus. First, we show that this enzyme, when incubated with HeLa cells, specifically cleaves nuclear RNA while it leaves cytoplasmic RNA unaffected. On the other hand, we have created new variants in which the residues of the nuclear localization signal that are important for the nuclear transport have been replaced. As expected, the individual changes produce a significant decrease in the cytotoxicity of the resulting variants. We conclude that the nuclear transport of PE5 is critical for its cytotoxicity. Therefore, routing a ribonuclease to the nucleus is an alternative strategy to endow it with cytotoxic activity.

  1. Constitutive expression of human pancreatic lipase-related protein 1 in Pichia pastoris.

    Science.gov (United States)

    Aloulou, Ahmed; Grandval, Philippe; De Caro, Josiane; De Caro, Alain; Carrière, Frédéric

    2006-06-01

    High-level constitutive expression of the human pancreatic lipase-related protein 1 (HPLRP1) was achieved using the methylotrophic yeast Pichia pastoris. The HPLRP1 cDNA, including its original leader sequence, was subcloned into the pGAPZB vector and further integrated into the genome of P. pastoris X-33 under the control of the glyceraldehyde 3-phosphate dehydrogenase (GAP) constitutive promoter. A major protein with a molecular mass of 50 kDa was found to be secreted into the culture medium and was identified using anti-HPLRP1 polyclonal antibodies as HPLRP1 recombinant protein. The level of expression reached 100-120 mg of HPLRP1 per liter of culture medium after 40 h, as attested by specific and quantitative enzyme-linked immunosorbent assay. A single cation-exchange chromatography sufficed to obtain a highly purified recombinant HPLRP1 after direct batch adsorption onto S-Sepharose of the HPLRP1 present in the culture medium, at pH 5.5. N-terminal sequencing and mass spectrometry analysis were carried out to monitor the production of the mature protein and to confirm that its signal peptide was properly processed.

  2. Autoimmunity against INS-IGF2 protein expressed in human pancreatic islets.

    Science.gov (United States)

    Kanatsuna, Norio; Taneera, Jalal; Vaziri-Sani, Fariba; Wierup, Nils; Larsson, Helena Elding; Delli, Ahmed; Skärstrand, Hanna; Balhuizen, Alexander; Bennet, Hedvig; Steiner, Donald F; Törn, Carina; Fex, Malin; Lernmark, Åke

    2013-10-04

    Insulin is a major autoantigen in islet autoimmunity and progression to type 1 diabetes. It has been suggested that the insulin B-chain may be critical to insulin autoimmunity in type 1 diabetes. INS-IGF2 consists of the preproinsulin signal peptide, the insulin B-chain, and eight amino acids of the C-peptide in addition to 138 amino acids from the IGF2 gene. We aimed to determine the expression of INS-IGF2 in human pancreatic islets and autoantibodies in newly diagnosed children with type 1 diabetes and controls. INS-IGF2, expressed primarily in beta cells, showed higher levels of expression in islets from normal compared with donors with either type 2 diabetes (p = 0.006) or high HbA1c levels (p INS-IGF2 autoantibody levels were increased in newly diagnosed patients with type 1 diabetes (n = 304) compared with healthy controls (n = 355; p INS-IGF2 revealed that more patients than controls had doubly reactive insulin-INS-IGF2 autoantibodies. These data suggest that INS-IGF2, which contains the preproinsulin signal peptide, the B-chain, and eight amino acids of the C-peptide may be an autoantigen in type 1 diabetes. INS-IGF2 and insulin may share autoantibody-binding sites, thus complicating the notion that insulin is the primary autoantigen in type 1 diabetes.

  3. Functional proteomics screen enables enrichment of distinct cell types from human pancreatic islets.

    Directory of Open Access Journals (Sweden)

    Revital Sharivkin

    Full Text Available The current world-wide epidemic of diabetes has prompted attempts to generate new sources of insulin-producing cells for cell replacement therapy. An inherent challenge in many of these strategies is the lack of cell-surface markers permitting isolation and characterization of specific cell types from differentiating stem cell populations. Here we introduce an iterative proteomics procedure allowing tag-free isolation of cell types based on their function. Our method detects and associates specific cell-surface markers with particular cell functionality by coupling cell capture on antibody arrays with immunofluorescent labeling. Using this approach in an iterative manner, we discovered marker combinations capable of enriching for discrete pancreatic cell subtypes from human islets of Langerhans: insulin-producing beta cells (CD9high/CD56+, glucagon-producing alpha cells (CD9-/CD56+ and trypsin-producing acinar cells (CD9-/CD56-. This strategy may assist future beta cell research and the development of diagnostic tools for diabetes. It can also be applied more generally for function-based purification of desired cell types from other limited and heterogeneous biological samples.

  4. Structural features for the mechanism of antitumor action of a dimeric human pancreatic ribonuclease variant

    Science.gov (United States)

    Merlino, Antonello; Avella, Giovanna; Di Gaetano, Sonia; Arciello, Angela; Piccoli, Renata; Mazzarella, Lelio; Sica, Filomena

    2009-01-01

    A specialized class of RNases shows a high cytotoxicity toward tumor cell lines, which is critically dependent on their ability to reach the cytosol and to evade the action of the ribonuclease inhibitor (RI). The cytotoxicity and antitumor activity of bovine seminal ribonuclease (BSRNase), which exists in the native state as an equilibrium mixture of a swapped and an unswapped dimer, are peculiar properties of the swapped form. A dimeric variant (HHP2-RNase) of human pancreatic RNase, in which the enzyme has been engineered to reproduce the sequence of BSRNase helix-II (Gln28→Leu, Arg31→Cys, Arg32→Cys, and Asn34→Lys) and to eliminate a negative charge on the surface (Glu111→Gly), is also extremely cytotoxic. Surprisingly, this activity is associated also to the unswapped form of the protein. The crystal structure reveals that on this molecule the hinge regions, which are highly disordered in the unswapped form of BSRNase, adopt a very well-defined conformation in both subunits. The results suggest that the two hinge peptides and the two Leu28 side chains may provide an anchorage to a transient noncovalent dimer, which maintains Cys31 and Cys32 of the two subunits in proximity, thus stabilizing a quaternary structure, similar to that found for the noncovalent swapped dimer of BSRNase, that allows the molecule to escape RI and/or to enhance the formation of the interchain disulfides. PMID:19177350

  5. Comparison of hematopoietic supportive capacity between human fetal and adult bone marrow mesenchymal stem cells in vitro.

    Science.gov (United States)

    Liu, Meng; Yang, Shao-Guang; Xing, Wen; Lu, Shi-Hong; Zhao, Qin-Jun; Ren, Hong-Ying; Chi, Ying; Ma, Feng-Xia; Han, Zhong-Chao

    2011-08-01

    Hematopoietic stem cells (HSC) shift from fetal liver and spleen to bone marrow at neonatal stages and this movement may be due to inductive signals from different microenvironments. Mesenchymal stem cells (MSC) are the precursors of stromal cells in bone marrow microenvironments such as osteoblasts and endothelial cells. Some researchers speculated that fetal bone marrow before birth might be not perfectly suit HSC growth. However, it is still lack of direct evidence to prove this hypothesis. This study was aimed to compare the hematopoietic supportive capacity between human fetal and adult bone marrow MSC in vitro. Adult bone marrow MSC (ABM-MSC) were isolated from three healthy donors and fetal bone marrow MSC (FBM-MSC) were isolated from three fetuses between gestations of 19 to 20 weeks. After irradiation, MSC were co-cultured with CD34(+) cells isolated from umbilical cord blood in long-term culture-initiating cell (LTC-IC) assay. The colony number of colony forming cells (CFC) was counted and the phenotypic changes of co-cultured CD34(+) cells were analyzed by flow cytometry. Cytokine expressions in both kinds of MSC were detected by reverse transcription polymerase chain reaction (RT-PCR). The results showed that ABM-MSC had a stronger hematopoietic supportive capacity than FBM-MSC. Both of them enhanced the differentiation of CD34(+) cells into myeloid lineages. Cytokines were expressed differently in ABM-MSC and FBM-MSC. It is concluded that ABM-MSC possess more potential application in some treatments than FBM-MSC, especially in hematopoietic reconstitution.

  6. Neurosensory Differentiation and Innervation Patterning in the Human Fetal Vestibular End Organs between the Gestational Weeks 8–12

    Science.gov (United States)

    Johnson Chacko, Lejo; Pechriggl, Elisabeth J.; Fritsch, Helga; Rask-Andersen, Helge; Blumer, Michael J. F.; Schrott-Fischer, Anneliese; Glueckert, Rudolf

    2016-01-01

    Balance orientation depends on the precise operation of the vestibular end organs and the vestibular ganglion neurons. Previous research on the assemblage of the neuronal network in the developing fetal vestibular organ has been limited to data from animal models. Insights into the molecular expression profiles and signaling moieties involved in embryological development of the human fetal inner ear have been limited. We present an investigation of the cells of the vestibular end organs with specific focus on the hair cell differentiation and innervation pattern using an uninterrupted series of unique specimens from gestational weeks 8–12. Nerve fibers positive for peripherin innervate the entire fetal crista and utricle. While in rodents only the peripheral regions of the cristae and the extra-striolar region of the statolithic organs are stained. At week 9, transcription factors PAX2 and PAX8 were observed in the hair cells whereas PAX6 was observed for the first time among the supporting cells of the cristae and the satellite glial cells of the vestibular ganglia. Glutamine synthetase, a regulator of the neurotransmitter glutamate, is strongly expressed among satellite glia cells, transitional zones of the utricle and supporting cells in the sensory epithelium. At gestational week 11, electron microscopic examination reveals bouton contacts at hair cells and first signs of the formation of a protocalyx at type I hair cells. Our study provides first-hand insight into the fetal development of the vestibular end organs as well as their pattern of innervation by means of immunohistochemical and EM techniques, with the aim of contributing toward our understanding of balance development. PMID:27895556

  7. Neurosensory Differentiation and Innervation Patterning in the Human Fetal Vestibular End Organs between the Gestational Weeks 8-12.

    Science.gov (United States)

    Johnson Chacko, Lejo; Pechriggl, Elisabeth J; Fritsch, Helga; Rask-Andersen, Helge; Blumer, Michael J F; Schrott-Fischer, Anneliese; Glueckert, Rudolf

    2016-01-01

    Balance orientation depends on the precise operation of the vestibular end organs and the vestibular ganglion neurons. Previous research on the assemblage of the neuronal network in the developing fetal vestibular organ has been limited to data from animal models. Insights into the molecular expression profiles and signaling moieties involved in embryological development of the human fetal inner ear have been limited. We present an investigation of the cells of the vestibular end organs with specific focus on the hair cell differentiation and innervation pattern using an uninterrupted series of unique specimens from gestational weeks 8-12. Nerve fibers positive for peripherin innervate the entire fetal crista and utricle. While in rodents only the peripheral regions of the cristae and the extra-striolar region of the statolithic organs are stained. At week 9, transcription factors PAX2 and PAX8 were observed in the hair cells whereas PAX6 was observed for the first time among the supporting cells of the cristae and the satellite glial cells of the vestibular ganglia. Glutamine synthetase, a regulator of the neurotransmitter glutamate, is strongly expressed among satellite glia cells, transitional zones of the utricle and supporting cells in the sensory epithelium. At gestational week 11, electron microscopic examination reveals bouton contacts at hair cells and first signs of the formation of a protocalyx at type I hair cells. Our study provides first-hand insight into the fetal development of the vestibular end organs as well as their pattern of innervation by means of immunohistochemical and EM techniques, with the aim of contributing toward our understanding of balance development.

  8. Impact of exposure to low concentrations of nitric oxide on protein profile in murine and human pancreatic islet cells.

    Science.gov (United States)

    Tapia-Limonchi, Rafael; Díaz, Irene; Cahuana, Gladys M; Bautista, Mario; Martín, Franz; Soria, Bernat; Tejedo, Juan R; Bedoya, Francisco J

    2014-01-01

    Homeostatic levels of nitric oxide (NO) protect efficiently against apoptotic death in both human and rodent pancreatic β cells, but the protein profile of this action remains to be determined. We have applied a 2 dimensional LC-MS-MALDI-TOF/TOF-based analysis to study the impact of protective NO in rat insulin-producing RINm5F cell line and in mouse and human pancreatic islets (HPI) exposed to serum deprivation condition. 24 proteins in RINm5F and 22 in HPI were identified to undergo changes in at least one experimental condition. These include stress response mitochondrial proteins (UQCRC2, VDAC1, ATP5C1, ATP5A1) in RINm5F cells and stress response endoplasmic reticulum proteins (HSPA5, PDIA6, VCP, GANAB) in HPI. In addition, metabolic and structural proteins, oxidoreductases and chaperones related with protein metabolism are also regulated by NO treatment. Network analysis of differentially expressed proteins shows their interaction in glucocorticoid receptor and NRF2-mediated oxidative stress response pathways and eNOS signaling. The results indicate that exposure to exogenous NO counteracts the impact of serum deprivation on pancreatic β cell proteome. Species differences in the proteins involved are apparent.

  9. Impact of exposure to low concentrations of nitric oxide on protein profile in murine and human pancreatic islet cells

    Science.gov (United States)

    Tapia-Limonchi, Rafael; Díaz, Irene; Cahuana, Gladys M; Bautista, Mario; Martín, Franz; Soria, Bernat; Tejedo, Juan R; Bedoya, Francisco J

    2014-01-01

    Homeostatic levels of nitric oxide (NO) protect efficiently against apoptotic death in both human and rodent pancreatic β cells, but the protein profile of this action remains to be determined. We have applied a 2 dimensional LC-MS-MALDI-TOF/TOF-based analysis to study the impact of protective NO in rat insulin-producing RINm5F cell line and in mouse and human pancreatic islets (HPI) exposed to serum deprivation condition. 24 proteins in RINm5F and 22 in HPI were identified to undergo changes in at least one experimental condition. These include stress response mitochondrial proteins (UQCRC2, VDAC1, ATP5C1, ATP5A1) in RINm5F cells and stress response endoplasmic reticulum proteins (HSPA5, PDIA6, VCP, GANAB) in HPI. In addition, metabolic and structural proteins, oxidoreductases and chaperones related with protein metabolism are also regulated by NO treatment. Network analysis of differentially expressed proteins shows their interaction in glucocorticoid receptor and NRF2-mediated oxidative stress response pathways and eNOS signaling. The results indicate that exposure to exogenous NO counteracts the impact of serum deprivation on pancreatic β cell proteome. Species differences in the proteins involved are apparent. PMID:25658244

  10. Expression of lysophosphatidic acid and its receptor in human pancreatic cancer and its clinical evaluation of diagnosis and therapy

    Institute of Scientific and Technical Information of China (English)

    WANG Shao-kai; TAO Chen-jie; WANG Wei-dong; L(U)Guang-mei; GONG Yong-ling

    2011-01-01

    Lysophosphatidic acid(LPA) is a naturally occurring phospholipid with diverse effects in various cells, ranging from immediate morphological alteration to long lasting cellular function changes, such as induction of stimulation of cell proliferation, survival, drug resistance and motility. Like many other biomediators, LPA interacts with cells through specific cell surface receptors(G protein-coupled receptors). LPA1/Edg-2,LPA2/Edg-4 and LPA3/Edg-7, named as Edg/LP subfamily, are the three most common lysophosphatidic acid receptors. LPA plays a critical role as a general growth, survival and pro-angiogenic factor in the regulation of pathophysiological processes in vivo and in vitro. Recent literatures suggest that abnormalities in LPA metabolism and function in pancreatic cancer patients may contribute to the initiation and progression of the disease. Thus, LPA might be a potential target for clinical pancreatic cancer diagnosis and therapy. Herein we review the expression of LPA and its receptors in the carcinogenesis of human malignancies, with focus on human pancreatic cancer, and also clinical diagnosis and treatment has been evaluated.

  11. Suppression of FAT/CD36 mRNA by human growth hormone in pancreatic β-cells

    DEFF Research Database (Denmark)

    Dalgaard, Louise Torp; Thams, Peter Grevsen; Gaarn, Louise Winkel;

    2011-01-01

    of this study was to examine the effect of human growth hormone (hGH) on mRNAs of fatty acid transport and binding proteins expressed in pancreatic β-cells, and to examine this in relation to β-cell survival after exposure to fatty acids. hGH decreased mRNA levels of FAT/CD36, whereas mRNAs of GPR40, FASN, FABP......Fatty acid-induced damage in pancreatic β-cells is assumed to play an important role in the development of type 2 diabetes. Lactogens (prolactin, placental lactogen and growth hormone) improve β-cell survival via STAT5 activation but the molecular targets are incompletely characterized. The aim...

  12. Suppression of FAT/CD36 mRNA by human growth hormone in pancreatic ß-cells

    DEFF Research Database (Denmark)

    Dalgaard, Louise Torp; Thams, Peter Grevsen; Gaarn, Louise Winkel;

    2011-01-01

    of this study was to examine the effect of human growth hormone (hGH) on mRNAs of fatty acid transport and binding proteins expressed in pancreatic ß-cells, and to examine this in relation to ß-cell survival after exposure to fatty acids. hGH decreased mRNA levels of FAT/CD36, whereas mRNAs of GPR40, FASN, FABP......Fatty acid-induced damage in pancreatic ß-cells is assumed to play an important role in the development of type 2 diabetes. Lactogens (prolactin, placental lactogen and growth hormone) improve ß-cell survival via STAT5 activation but the molecular targets are incompletely characterized. The aim...

  13. Expression profiling of wild type and β-catenin gene disrupted human BxPC-3 pancreatic adenocarcinoma cells

    Directory of Open Access Journals (Sweden)

    Petter Angell Olsen

    2015-06-01

    Full Text Available To study the role of WNT/β-catenin signaling in pancreatic adenocarcinoma, human BxPC-3 cell lines deficient of the central canonical WNT signaling protein β-catenin were established by using zinc-finger nuclease mediated targeted genomic disruption of the β-catenin gene (CTNNB1. Comparison of the global transcription levels in wild type cells with two β-catenin gene disrupted clones identified 85 transcripts that were the most differentially regulated. Gene ontology (GO term enrichment analysis of these transcripts identified “cell adhesion” as the most significantly enriched GO term. Here we describe the data from the transcription profiling analysis published in the article “Implications of Targeted Genomic Disruption of β-Catenin in BxPC-3 Pancreatic Adenocarcinoma Cells” [1]. Data have been deposited to the Gene Expression Omnibus (GEO database repository with the dataset identifier GSE63072.

  14. Evidence that P12, a specific variant of P16{sup INK4A}, plays a suppressive role in human pancreatic carcinogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Poi, Ming J. [Department of Pharmacy, The Arthur G. James Cancer Hospital and Richard J. Solove Research Institute, The Ohio State University, Columbus, OH 43210 (United States); Knobloch, Thomas J. [Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210 (United States); Division of Environmental Health Sciences, College of Public Health, The Ohio State University, Columbus, OH 43210 (United States); Yuan, Chunhua [Campus Chemical Instrument Center, The Ohio State University, Columbus, OH 43210 (United States); Tsai, Ming-Daw [Genomics Research Center and Institute of Biological Chemistry, Academia Sinica, Taiwan (China); Weghorst, Christopher M. [Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210 (United States); Division of Environmental Health Sciences, College of Public Health, The Ohio State University, Columbus, OH 43210 (United States); Li, Junan, E-mail: li.225@osu.edu [Comprehensive Cancer Center, The Ohio State University, Columbus, OH 43210 (United States); Division of Environmental Health Sciences, College of Public Health, The Ohio State University, Columbus, OH 43210 (United States)

    2013-06-28

    Highlights: •P12, a variant of P16{sup INK4A}, inhibits the proliferation of pancreatic cancer cells. •P12 is distinct from P16 in function and structure. •Genetic alterations of p12 are prevalent in human pancreatic carcinoma. •P12 represents a potential pancreas-specific tumor suppressor. -- Abstract: The INK4a-ARF locus plays a central role in the development of pancreatic tumors as evidenced by the fact that up to 98% of pancreatic tumor specimens harbored genetic alterations at the INK4a-ARF locus. Interestingly, in addition to the well-known P16{sup INK4A} (P16) and P14ARF tumor suppressors, the INK4a-ARF locus in pancreas encodes another protein, P12, whose structure, function, and contributions to pancreatic carcinogenesis remain to be elucidated. In the current study, we demonstrated that over-expression of p12 in human pancreatic cancer cells led to cell arrest at the G1 phase and such cell cycle arrest was related to down-regulation of a number of oncogenes, such as c-Jun, Fos, and SEI1. Furthermore, unlike P16, P12 did not retain any cyclin-dependent kinase 4 (CDK4)-inhibitory activity. Instead, P12 exhibited a transactivating activity not found in P16. We also examined the genetic status of p12 in a cohort of 40 pancreatic tumor specimens and found that p12 alteration was prevalent in pancreatic tumors with an incidence of 70% (28/40). These results support that P12 is a tumor suppressive protein distinct from P16, and its genetic inactivation is associated with pancreatic carcinogenesis.

  15. Fetal calcium regulates branching morphogenesis in the developing human and mouse lung: involvement of voltage-gated calcium channels.

    Science.gov (United States)

    Brennan, Sarah C; Finney, Brenda A; Lazarou, Maria; Rosser, Anne E; Scherf, Caroline; Adriaensen, Dirk; Kemp, Paul J; Riccardi, Daniela

    2013-01-01

    Airway branching morphogenesis in utero is essential for optimal postnatal lung function. In the fetus, branching morphogenesis occurs during the pseudoglandular stage (weeks 9-17 of human gestation, embryonic days (E)11.5-16.5 in mouse) in a hypercalcaemic environment (~1.7 in the fetus vs. ~1.1-1.3 mM for an adult). Previously we have shown that fetal hypercalcemia exerts an inhibitory brake on branching morphogenesis via the calcium-sensing receptor. In addition, earlier studies have shown that nifedipine, a selective blocker of L-type voltage-gated Ca(2+) channels (VGCC), inhibits fetal lung growth, suggesting a role for VGCC in lung development. The aim of this work was to investigate the expression of VGCC in the pseudoglandular human and mouse lung, and their role in branching morphogenesis. Expression of L-type (CaV1.2 and CaV1.3), P/Q type (CaV2.1), N-type (CaV2.2), R-type (CaV2.3), and T-type (CaV3.2 and CaV3.3) VGCC was investigated in paraffin sections from week 9 human fetal lungs and E12.5 mouse embryos. Here we show, for the first time, that Cav1.2 and Cav1.3 are expressed in both the smooth muscle and epithelium of the developing human and mouse lung. Additionally, Cav2.3 was expressed in the lung epithelium of both species. Incubating E12.5 mouse lung rudiments in the presence of nifedipine doubled the amount of branching, an effect which was partly mimicked by the Cav2.3 inhibitor, SNX-482. Direct measurements of changes in epithelial cell membrane potential, using the voltage-sensitive fluorescent dye DiSBAC2(3), demonstrated that cyclic depolarisations occur within the developing epithelium and coincide with rhythmic occlusions of the lumen, driven by the naturally occurring airway peristalsis. We conclude that VGCC are expressed and functional in the fetal human and mouse lung, where they play a role in branching morphogenesis. Furthermore, rhythmic epithelial depolarisations evoked by airway peristalsis would allow for branching to match

  16. Fetal calcium regulates branching morphogenesis in the developing human and mouse lung: involvement of voltage-gated calcium channels.

    Directory of Open Access Journals (Sweden)

    Sarah C Brennan

    Full Text Available Airway branching morphogenesis in utero is essential for optimal postnatal lung function. In the fetus, branching morphogenesis occurs during the pseudoglandular stage (weeks 9-17 of human gestation, embryonic days (E11.5-16.5 in mouse in a hypercalcaemic environment (~1.7 in the fetus vs. ~1.1-1.3 mM for an adult. Previously we have shown that fetal hypercalcemia exerts an inhibitory brake on branching morphogenesis via the calcium-sensing receptor. In addition, earlier studies have shown that nifedipine, a selective blocker of L-type voltage-gated Ca(2+ channels (VGCC, inhibits fetal lung growth, suggesting a role for VGCC in lung development. The aim of this work was to investigate the expression of VGCC in the pseudoglandular human and mouse lung, and their role in branching morphogenesis. Expression of L-type (CaV1.2 and CaV1.3, P/Q type (CaV2.1, N-type (CaV2.2, R-type (CaV2.3, and T-type (CaV3.2 and CaV3.3 VGCC was investigated in paraffin sections from week 9 human fetal lungs and E12.5 mouse embryos. Here we show, for the first time, that Cav1.2 and Cav1.3 are expressed in both the smooth muscle and epithelium of the developing human and mouse lung. Additionally, Cav2.3 was expressed in the lung epithelium of both species. Incubating E12.5 mouse lung rudiments in the presence of nifedipine doubled the amount of branching, an effect which was partly mimicked by the Cav2.3 inhibitor, SNX-482. Direct measurements of changes in epithelial cell membrane potential, using the voltage-sensitive fluorescent dye DiSBAC2(3, demonstrated that cyclic depolarisations occur within the developing epithelium and coincide with rhythmic occlusions of the lumen, driven by the naturally occurring airway peristalsis. We conclude that VGCC are expressed and functional in the fetal human and mouse lung, where they play a role in branching morphogenesis. Furthermore, rhythmic epithelial depolarisations evoked by airway peristalsis would allow for branching to

  17. Echovirus 6 Infects Human Exocrine and Endocrine Pancreatic Cells and Induces Pro-Inflammatory Innate Immune Response

    Directory of Open Access Journals (Sweden)

    Luis Sarmiento

    2017-01-01

    Full Text Available Human enteroviruses (HEV, especially coxsackievirus serotype B (CVB and echovirus (E, have been associated with diseases of both the exocrine and endocrine pancreas, but so far evidence on HEV infection in human pancreas has been reported only in islets and ductal cells. This study aimed to investigate the capability of echovirus strains to infect human exocrine and endocrine pancreatic cells. Infection of explanted human islets and exocrine cells with seven field strains of E6 caused cytopathic effect, virus titer increase and production of HEV protein VP1 in both cell types. Virus particles were found in islets and acinar cells infected with E6. No cytopathic effect or infectious progeny production was observed in exocrine cells exposed to the beta cell-tropic strains of E16 and E30. Endocrine cells responded to E6, E16 and E30 by upregulating the transcription of interferon-induced with helicase C domain 1 (IF1H1, 2'-5'-oligoadenylate synthetase 1 (OAS1, interferon-β (IFN-β, chemokine (C–X–C motif ligand 10 (CXCL10 and chemokine (C–C motif ligand 5 (CCL5. Echovirus 6, but not E16 or E30, led to increased transcription of these genes in exocrine cells. These data demonstrate for the first time that human exocrine cells represent a target for E6 infection and suggest that certain HEV serotypes can replicate in human pancreatic exocrine cells, while the pancreatic endocrine cells are permissive to a wider range of HEV.

  18. Fetal antigen 2: an amniotic protein identified as the aminopropeptide of the alpha 1 chain of human procollagen type I

    DEFF Research Database (Denmark)

    Teisner, B; Rasmussen, H B; Højrup, P

    1992-01-01

    -PAGE analysis gave an M(r) = 27 kDa under reducing and non-reducing conditions for both forms, whereas the exact M(r) determined by mass spectrometry was 14,343 +/- 3 Da. FA2 was N-terminally blocked and after tryptic digestion the amino acid composition and sequences of the peptides showed identity...... with the aminopropeptide of the alpha 1 chain of human procollagen type I as determined by nucleotide sequences. After oxidative procedures normally employed for radio-iodination (iodogen and chloramine-T), FA2 lost its immunoreactivity. An antigen which cross-reacted with polyclonal rabbit anti-human FA2 was demonstrated...... in fetal calf serum. Gel filtration with analysis of fractions by inhibition ELISA showed that the bovine homologue was present in the same molecular forms as those in human amniotic fluid, and immunohistochemical analysis with anti-human FA2 showed that its distribution in bovine skin was identical...

  19. Solanine Induces Mitochondria-Mediated Apoptosis in Human Pancreatic Cancer Cells

    OpenAIRE

    Hongwei Sun; Chongqing Lv; Longlong Yang; Yingxiu Wang; Qingshun Zhang; Suhui Yu; Hongru Kong; Meng Wang; Jianming Xie; Chunwu Zhang; Mengtao Zhou

    2014-01-01

    Steroid alkaloids have been suggested as potential anticancer compounds. However, the underlying mechanisms of how steroid alkaloids inhibit the tumor growth are largely unknown. Here, we reported that solanine, a substance of steroid alkaloids, has a positive effect on the inhibition of pancreatic cancer cell growth in vitro and in vivo. In pancreatic cancer cells and nu/nu nude mice mod