Sample records for human cholangiocarcinoma cell

  1. Proteasome inhibition-induces endoplasmic reticulum dysfunction and cell death of human cholangiocarcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Yucel Ustundag; Steven F Bronk; Gregory J Gores


    AIM: To determine if proteasome inhibition induces apoptosis in human cholangiocarcinoma cells, and if so, to elucidate the cellular mechanisms.METHODS: Studies were performed in the human KMCH, KMBC, and Mz-ChA-1 cholangiocarcinoma, and normal rat cell lines. MG132, a peptide aldehyde, which inhibits the chymotrypsin-like activity of the proteaosome was employed for this study. Apoptosis was assessed morphologically by 4'-6-Diamidino-2-phenylindole (DAPI) nuclear staining and fluorescence microscopy. Mitochondrial membrane potential was examined using a fluorescent unquenching assay. Ultrastructural changes during cell death were examined using transmission electron microscopy (TEM). Caspase 3/7 activity was assessed using an enzymatic-based fluorescent assay. Cytosolic-free calcium concentrations were measured using Fura-2 and digitized fluorescent microscopy.RESULTS: MG132, a proteasome inhibitor, induced apoptosis in all the cholangiocarcinoma cell lines examined. In contrast, minimal cytotoxicity was observed in normal rat cholangiocytes. Apoptosis was time- and -concentration-dependent. There was no change in the mitochondrial membrane potential between treated and untreated cells. Ultrastructural examination by transmission electron microscopy displayed the classic features of apoptosis, but in addition, there was also dramatic vacuolization of the endoplasmic reticulum (ER). Unexpectedly, no increase in caspase 3/7 activity was observed in MG132 treated cells, nor did the pancaspase inhibitor, Q-VD-OPh prevent cell death. The protein synthesis inhibitor, cycloheximide, blocked apoptosis induced by proteosome inhibitor indicating that ER dysfunction was dependent upon the formation of new proteins.CONCLUSION:Proteosome inhibition induces ER dysfunction and caspase-independent cell death selectively in human cholangiocarcinoma cells. Proteasome inhibitors warrant evaluation as anticancer agents for the treatment of human cholangiocarcinoma.

  2. Estrogens and Insulin-Like Growth Factor 1 Modulate Neoplastic Cell Growth in Human Cholangiocarcinoma (United States)

    Alvaro, Domenico; Barbaro, Barbara; Franchitto, Antonio; Onori, Paolo; Glaser, Shannon S.; Alpini, Gianfranco; Francis, Heather; Marucci, Luca; Sterpetti, Paola; Ginanni-Corradini, Stefano; Onetti Muda, Andrea; Dostal, David E.; De Santis, Adriano; Attili, Adolfo F.; Benedetti, Antonio; Gaudio, Eugenio


    We investigated the expression of estrogen receptors (ERs), insulin-like growth factor 1 (IGF-1), and IGF-1R (receptor) in human cholangiocarcinoma and cholangiocarcinoma cell lines (HuH-28, TFK-1, Mz-ChA-1), evaluating the role of estrogens and IGF-1 in the modulation of neoplastic cell growth. ER-α, ER-β, IGF-1, and IGF-1R were expressed (immunohistochemistry) in all biopsies (18 of 18) of intrahepatic cholangiocarcinoma. ER-α was expressed (Western blot) only by the HuH-28 cell line (intrahepatic cholangiocarcinoma), whereas ER-β, IGF-1, and IGF-1R were expressed in the three cell lines examined. In serum-deprived HuH-28 cells, serum readmission induced stimulation of cell proliferation that was inhibited by ER and IGF-1R antagonists. 17β-Estradiol and IGF-1 stimulated proliferation of HuH-28 cells to a similar extent to that of MCF7 (breast cancer) but greater than that of TFK-1 and Mz-ChA-1, inhibiting apoptosis and exerting additive effects. These effects of 17β-estradiol and IGF-1 were associated with enhanced protein expression of ER-α, phosphorylated (p)-ERK1/2 and pAKT but with decreased expression of ER-β. Finally, transfection of IGF-1R anti-sense oligonucleotides in HuH-28 cells markedly decreased cell proliferation. In conclusion, human intrahepatic cholangiocarcinomas express receptors for estrogens and IGF-1, which cooperate in the modulation of cell growth and apoptosis. Modulation of ER and IGF-1R could represent a strategy for the management of cholangiocarcinoma. PMID:16936263

  3. Berberine inhibits growth and induces G1 arrest and apoptosis in human cholangiocarcinoma QBC939 cells. (United States)

    He, Wei; Wang, Bin; Zhuang, Yun; Shao, Dong; Sun, Kewen; Chen, Jianping


    The chemotherapeutic approach using non-toxic natural products may be one of the strategies for the management of the cholangiocarcinoma. Here we report that in vitro treatment of human cholangiocarcinoma QBC939 cells with berberine, a naturally occurring isoquinoline alkaloid, decreased cell viability and induced cell death in a dose-dependent manner, which was associated with an increase in G1 arrest. Our western blot analysis showed that berberine-induced G1 cell cycle arrest was mediated through the increased expression of cyclin-dependent kinase inhibitors (Cdki) proteins (Cip1/p21 and Kip1/p27); a simultaneous decrease in Cdk2 and Cdk4 and cyclins D1, and reduced activity of the Cyclins-Cdk complex. In additional studies, treatment of QBC939 cells with different concentrations (10, 40, 80 μM) of berberine for 48 h resulted in a significant dose-dependent increase in apoptosis compared to the non-berberine-treated control, which was associated with an increased expression of pro-apoptotic protein Bax and decreased expression of anti-apoptotic proteins Bcl-2 and Bcl-xL. Together, this study for the first time identified berberine as a chemotherapeutic agent against human cholangiocarcinoma cells QBC939 cells in vitro. Further in vivo studies are required to determine whether berberine could be an effective chemotherapeutic agent for the management of cholangiocarcinoma.

  4. Cytotoxic activity of Thai medicinal plants against human cholangiocarcinoma, laryngeal and hepatocarcinoma cells in vitro

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    Itharat Arunporn


    Full Text Available Abstract Background Cholangiocarcinoma is a serious public health in Thailand with increasing incidence and mortality rates. The present study aimed to investigate cytotoxic activities of crude ethanol extracts of a total of 28 plants and 5 recipes used in Thai folklore medicine against human cholangiocarcinoma (CL-6, human laryngeal (Hep-2, and human hepatocarcinoma (HepG2 cell lines in vitro. Methods Cytotoxic activity of the plant extracts against the cancerous cell lines compared with normal cell line (renal epithelial cell: HRE were assessed using MTT assay. 5-fluorouracil was used as a positive control. The IC50 (concentration that inhibits cell growth by 50% and the selectivity index (SI were calculated. Results The extracts from seven plant species (Atractylodes lancea, Kaempferia galangal, Zingiber officinal, Piper chaba, Mesua ferrea, Ligusticum sinense, Mimusops elengi and one folklore recipe (Pra-Sa-Prao-Yhai exhibited promising activity against the cholangiocarcinoma CL-6 cell line with survival of less than 50% at the concentration of 50 μg/ml. Among these, the extracts from the five plants and one recipe (Atractylodes lancea, Kaempferia galangal, Zingiber officinal, Piper chaba, Mesua ferrea, and Pra-Sa-Prao-Yhai recipe showed potent cytotoxic activity with mean IC50 values of 24.09, 37.36, 34.26, 40.74, 48.23 and 44.12 μg/ml, respectively. All possessed high activity against Hep-2 cell with mean IC50 ranging from 18.93 to 32.40 μg/ml. In contrast, activity against the hepatoma cell HepG2 varied markedly; mean IC50 ranged from 9.67 to 115.47 μg/ml. The only promising extract was from Zingiber officinal (IC50 = 9.67 μg/ml. The sensitivity of all the four cells to 5-FU also varied according to cell types, particularly with CL-6 cell (IC50 = 757 micromolar. The extract from Atractylodes lancea appears to be both the most potent and most selective against cholangiocarcinoma (IC50 = 24.09 μg/ml, SI = 8.6. Conclusions The

  5. Apoptosis of human cholangiocarcinoma cells induced by ESC-3 from Crocodylus siamensis bile

    Institute of Scientific and Technical Information of China (English)

    Wei Song; Dong-Yan Shen; Jin-He Kang; Shan-Shan Li; Hui-Wang Zhan; Yan Shi; You-Xiong Xiong; Ge Liang; Qing-xi Chen


    AIM:To investigate the effects of ESC-3 isolated from crocodile bile on the growth and apoptosis induction of human cholangiocarcinoma cells.METHODS:ESC-3 was isolated from crocodile bile by Sephadex LH-20 and RP-18 reversed-phase column.3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay was conducted to determine the effects of ESC-3 on the proliferation of human cholangiocarcinoma cell lines (QBC939,Sk-ChA-1 and MZ-ChA-1).Giemsa staining,Hoechst 33258 and acridine orange/ethidium bromide staining showed the morphological changes of Mz-ChA-1 cells exposed to ESC-3 at different concentrations.Flow cytometry with regular propidium iodide (PI) staining was performed to analyze the cell cycle distribution of Mz-ChA-1 cells and to assess apoptosis by annexin v-fiuorescein isothiocyanate (VFITC)/PI staining.Rh123 staining was used to detect the alteration of mitochondrial membrane potential (△Ψm).The protein levels of Bax,Bcl-2,Cdk2,cytochrome c and caspase-3 were further confirmed by Western blotting.RESULTS:ESC-3 significantly inhibited the growth of three human cholangiocarcinoma cell lines and arrested Mz-ChA-1 cell cycle at G0/G1 phase.Mz-ChA-1 cells showed typical apoptotic morphological changes after treated with ESC-3 (10 μg/mL) for 48 h.Cell death assay indicated that Mz-ChA-1 cells underwent apoptosis in a dose-dependent manner induced by ESC-3.In addition,ESC-3 treatment could downregulate the protein level of Bcl-2 and upregulate the Bax,leading to the increase in the ratio of Bax to Bcl-2 in Mz-ChA-1 cells.Meanwhile,cytochrome c was released from the mitochondria into the cytosol,which subsequently initiated the activation of caspase-3.All these events were associated with the collapse of the mitochondrial membrane potential.CONCLUSION:ESC-3,the active ingredient of crocodile bile,induced apoptosis in Mz-ChA-1 cells through the mitochondria-dependent pathway and may be a potential chemotherapeutic drug for the treatment of

  6. Fas counterattack in cholangiocarcinoma:a mechanism for immune evasion in human hilar cholangiocarcinomas

    Institute of Scientific and Technical Information of China (English)

    Zi-Yu Li; Sheng-Quan Zou


    AIM: To investigate FasL expression in hilar cholangiocarcinoma tissues and cultured cholangiocarcinoma cells, and to assess its ability to induce apoptosis. METHODS: We studied the expression of FasL by human hilar cholangiocaroinomas tissues by immunohistochemistry, and the QBC939 cholangiocarcinomacell line by RT- PCR, immunohiatochemistry, and Western Blot. TUNEL and flow cytometry were used to detect apoptotic cells. RESJLTS: Prevalent expression of FasL was detected in 39resected hilar cholangiocarcinoma tissues. TUNEL staining disclosed a high level of cell death among lymphocytes infiltrating FasL positive areas of tumor. FasL mRNA and protein expressions in cholangiocarcinoma cells could induce Jurkat cells. CONCLUSION: Hilar cholangiocarcinomas may elude immunological surveillance by inducing, via Fas/FasL system, the apoptosis of activated lymphocytes.

  7. Comparative study of antitumor effects of bromelain and papain in human cholangiocarcinoma cell lines. (United States)

    Müller, Alena; Barat, Samarpita; Chen, Xi; Bui, Khac Cuong; Bozko, Przemyslaw; Malek, Nisar P; Plentz, Ruben R


    Cholangiocarcinoma (CC) worldwide is the most common biliary malignancy with poor prognostic value and new systemic treatments are desirable. Plant extracts like bromelain and papain, which are cysteine proteases from the fruit pineapple and papaya, are known to have antitumor activities. Therefore, in this study for the first time we investigated the anticancer effect of bromelain and papain in intra- and extrahepatic human CC cell lines. The effect of bromelain and papain on human CC cell growth, migration, invasion and epithelial plasticity was analyzed using cell proliferation, wound healing, invasion and apoptosis assay, as well as western blotting. Bromelain and papain lead to a decrease in the proliferation, invasion and migration of CC cells. Both plant extracts inhibited NFκB/AMPK signalling as well as their downstream signalling proteins such as p-AKT, p-ERK, p-Stat3. Additionally, MMP9 and other epithelial-mesenchymal-transition markers were partially found to be downregulated. Apoptosis was induced after bromelain and papain treatment. Interestingly, bromelain showed an overall more effective inhibition of CC as compared to papain. siRNA mediated silencing of NFκB on CC cells indicated that bromelain and papain have cytotoxic effects on human CC cell lines and bromelain and partially papain in comparison impair tumor growth by NFκB/AMPK signalling. Especially bromelain can evolve as promising, potential therapeutic option that might open new insights for the treatment of human CC.

  8. Histone deacetylase inhibitor MS-275 alone or combined with bortezomib or sorafenib exhibits strong antiproliferative action in human cholangiocarcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Viola Baradari; Michael H(o)pfner; Alexander Huether; Detlef Schuppan; Hans Scherübl


    AIM: To investigate the antiproliferative effect of the histone deacetylase (HDAC) inhibitor MS-275 on cholangiocarcinoma cells alone and in combination with conventional cytostatic drugs (gemcitabine or doxorubicin)or the novel anticancer agents sorafenib or bortezomib.METHODS: Two human bile duct adenocarcinoma cell lines (EGI-1 and TFK-1) were studied. Crystal violet staining was used for detection of cell number changes.Cytotoxicity was determined by measuring the release of the cytoplasmic enzyme lactate dehydrogenase (LDH).Apoptosis was determined by measuring the enzyme activity of caspase-3. Cell cycle status reflected by the DNA content was detected by flow cytometry.RESULTS: MS-275 treatment potently inhibited the proliferation of EGI-1 and TFK-1 cholangiocarcinoma cells by inducing apoptosis and cell cycle arrest. MS-275-induced apoptosis was characterized by activation of caspase-3, up-regulation of Bax and down-regulation of Bcl-2. Cell cycle was predominantly arrested at the G1/S checkpoint, which was associated with induction of the cyclin-dependent kinase inhibitor p21Waf/CIP1. Furthermore,additive anti-neoplastic effects were observed when MS-275 treatment was combined with gemcitabine or doxorubicin, while combination with the multikinase inhibitor sorafenib or the proteasome inhibitor bortezomib resulted in overadditive anti-neoplastic effects.CONCLUSION: The growth of human cholangiocarcinoma cells can be potently inhibited by MS-275 alone or in combination with conventional cytostatic drugs or new,targeted anticancer agents.

  9. Schisandrin B inhibits cell proliferation and induces apoptosis in human cholangiocarcinoma cells (United States)

    Yang, Xiaohui; Wang, Shuai; Mu, Yunchuan; Zheng, Yixiong


    Cholangiocarcinoma (CCA) is the second most common hepatic cancer with high resistance to current chemotherapies and extremely poor prognosis. The present study aimed to examine the effects of schisandrin B (Sch B) on CCA cells both in vitro and in vivo and to examine its underlying mechanism. We found that Sch B inhibited the viability and proliferation of CCA cells in a dose- and time-dependent manner as assessed by MTT and colony formation assays. The flow cytometric assay revealed G0/G1 phase arrest in the Sch B-treated HCCC-9810 and RBE cells. In addition, Sch B induced intrahepatic cholangiocarcinoma apoptosis as shown by the results of Annexin V/PI double staining. Rhodamine 123 staining revealed that Sch B decreased the mitochondrial membrane potential (ΔΨm) in a dose-dependent manner. Mechanistically, western blot analysis indicated that Sch B induced apoptosis by upregulating Bax, cleaved caspase-3, cleaved caspase-9 and cleaved PARP, and by downregulating cyclin D1, Bcl-2 and CDK-4. Moreover, Sch B significantly inhibited HCCC-9810 xenograft growth in athymic nude mice. In summary, these findings suggest that Sch B exhibited potent antitumor activities via the induction of CCA apoptosis and that Sch B may be a promising drug for the treatment of CCA. PMID:27499090

  10. Synergistic Anticancer Effects of Vorinostat and Epigallocatechin-3-Gallate against HuCC-T1 Human Cholangiocarcinoma Cells

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    Tae Won Kwak


    Full Text Available The aim of this study was to investigate the effect of the combination of vorinostat and epigallocatechin-3-gallate against HuCC-T1 human cholangiocarcinoma cells. A novel chemotherapy strategy is required as cholangiocarcinomas rarely respond to conventional chemotherapeutic agents. Both vorinostat and EGCG induce apoptosis and suppress invasion, migration, and angiogenesis of tumor cells. The combination of vorinostat and EGCG showed synergistic growth inhibitory effects and induced apoptosis in tumor cells. The Bax/Bcl-2 expression ratio and caspase-3 and -7 activity increased, but poly (ADP-ribose polymerase expression decreased when compared to treatment with each agent alone. Furthermore, invasion, matrix metalloproteinase (MMP expression, and migration of tumor cells decreased following treatment with the vorinostat and EGCG combination compared to those of vorinostat or EGCG alone. Tube length and junction number of human umbilical vein endothelial cells (HUVECs decreased as well as vascular endothelial growth factor expression following vorinostat and EGCG combined treatment. These results indicate that the combination of vorinostat and EGCG had a synergistic effect on inhibiting tumor cell angiogenesis potential. We suggest that the combination of vorinostat and EGCG is a novel option for cholangiocarcinoma chemotherapy.

  11. Effect of transforming growth factor-β1 on human intrahepatic cholangiocarcinoma cell growth

    Institute of Scientific and Technical Information of China (English)

    Tetsuya Shimizu; Takashi Tajiri; Shigeki Yokomuro; Yoshiaki Mizuguchi; Yutaka Kawahigashi; Yasuo Arima; Nobuhiko Taniai; Yasuhiro Mamada; Hiroshi Yoshida; Koho Akimaru


    AIM: To elucidate the biological effects of transforming growth factor-β1 (TGF-β1) on intrahepatic cholangiocarcinoma (ICC).METHODS: We investigated the effects of TGF-β1 on human ICC cell lines (HuCCT1, MEC, and HuH-28) by monitoring the influence of TGF-β1 on tumor growth and interleukin-6 (IL-6) expression in ICC cells.RESULTS: All three human ICC cell lines produced TGF-β1 and demonstrated accelerated growth in the presence of TGF-β1 with no apoptotic effect. Studies on HuCCT1 revealed a TGF-β1-induced stimulation of the expression of TGF-β1, as well as a decrease in TGF-β1 mRNA expression induced by neutralizing anti-TGF-β1 antibody. These results indicate that TGF-β1 stimulates the production and function of TGF-β1 in an autocrine fashion. Further, IL-6 secretion was observed in all three cell lines and exhibited an inhibitory response to neutralizing anti-TGF-β1 antibody. Experiments using HuCCT1 revealed a TGF-β1-induced acceleration of IL-6 protein expression and mRNA levels. These findings demonstrate a functional interaction between TGF-β1 and IL-6. All three cell lines proliferated in the presence of IL-6. In contrast, TGF-β1 induced no growth effect in HuCCT1 in the presence of small interfering RNA against a specific cell surface receptor of IL-6 and signal transducer and activator of transcription-3.CONCLUSION: ICC cells produce TGF-β1 and confer a TGF-β1-induced growth effect in an autocrine fashion.TGF-β1 activates IL-6 production, and the functional interaction between TGF-β1 and IL-6 contributes to ICC cell growth by TGF-β1.

  12. Effects of Bile from Patient with Transduodenal Sphincteroplasty on the Growth of Human Cholangiocarcinoma Cell Line

    Institute of Scientific and Technical Information of China (English)

    吴高松; 邹声泉; 刘正人; 裘法祖


    @@ It is reported that transduodenal sphincteroplasty (TS) had a closely relationship with malignancies of the biliary tract, and cholangiocarcinoma has been suggested as a TS possible long-term complication. We used bile from patient with transduodenal sphincteroplasty (TSB) to act directly on the cholangiocacinoma cells QBC939 to see the effects of TSB on the growth of QBC939 cells. TSB was obtained from a patient who had undergone TS, and normal bile (NB) was obtained from another patient who had not undergone TS.

  13. Drug sensitivity and drug resistance profiles of human intrahepatic cholangiocarcinoma cell lines

    Institute of Scientific and Technical Information of China (English)

    Nisana Tepsiri; Liengchai Chaturat; Banchob Sripa; Wises Namwat; Sopit Wongkham; Vajarabhongsa Bhudhisawasdi; Wichittra Tassaneeyakul


    AIM: To study the effect of a number of chemotherapeutic drugs on five human intrahepatic cholangiocarcinoma (CCA) cell lines. The expressions of genes that have been proposed to influence the resistance of chemotherapeutic drugs including thymidylate synthase (TS), dihydropyrimidine dehydrogenase (DPD), glutathione-S-transferase P1 (GSTP1), multidrug resistance protein (MDR1) and multidrug resistance-associated proteins (MRPs) were also determined.METHODS: Five human CCA cell lines (KKU-100, KKU M055, KKU-M156, KKU-M214 and KKU-OCA17) weretreated with various chemotherapeutic drugs and growth inhibition was determined by 3-(4,5-dimethylthiazol-2-yl)5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay. Semi-quantitative levels of gene expression were determined by a reverse transcriptase polymerase chain reaction (RT-PCR). Results of IC50 values and the ratios of gene expression were analyzed by linear regression to predict their relationship. RESULTS: Among five CCA cell lines, KKU-M055 was the most sensitive cell line towards all chemotherapeutic drugs investigated, particularly taxane derivatives with IC50 values of 0.02-3 nmol/L, whereas KKU-100 was apparently the least sensitive cell line. When compared to other chemotherapeutic agents, doxorubicin and pirarubicin showed the lowest IC50 values (<5 μmol/L) in all five CCA cell lines. Results from RT-PCR showed that TS, MRP1, MRP3 and GSTP1 were highly expressed in these five CCA cell lines while DPD and MRP2 were only moderately expressed. It should be noted that MDR1 expression was detected only in KKU-OCA17 cell lines. A strong correlation was only found between the level of MRP3 expression and the IC50 values of etoposide, doxorubicin and pirarubicin (r = 0.86-0.98, ,P<0.05). CONCLUSION: Sensitivity to chemotherapeutic agents is not associated with the histological type of CCA. Choosing of the appropriate chemotherapeutic regimen for the treatment of CCA requires knowledge of drug

  14. Impact of Salinomycin on human cholangiocarcinoma: induction of apoptosis and impairment of tumor cell proliferation in vitro

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    Lieke Thorsten


    Full Text Available Abstract Background Cholangiocarcinoma (CC is a primary liver cancer with increasing incidence worldwide. Despite all efforts made in past years, prognosis remains to be poor. At least in part, this might be explained by a pronounced resistance of CC cells to undergo apoptosis. Thus, new therapeutic strategies are imperatively required. In this study we investigated the effect of Salinomycin, a polyether ionophore antibiotic, on CC cells as an appropriate agent to treat CC. Salinomycin was quite recently identified to induce apoptosis in cancer stem cells and to overcome apoptosis-resistance in several leukemia-cells and other cancer cell lines of different origin. Methods To delineate the effects of Salinomycin on CC, we established an in vitro cell culture model using three different human CC cell lines. After treatment apoptosis as well as migration and proliferation behavior was assessed and additional cell cycle analyses were performed by flowcytometry. Results By demonstrating Annexin V and TUNEL positivity of human CC cells, we provide evidence that Salinomycin reveals the capacity to break apoptosis-resistance in CC cells. Furthermore, we are able to demonstrate that the non-apoptotic cell fraction is characterized by sustainable impaired migration and proliferation. Cell cycle analyses revealed G2-phase accumulation of human CC cells after treatment with Salinomycin. Even though apoptosis is induced in two of three cell lines of CC cells, one cell line remained unaffected in regard of apoptosis but revealed as the other CC cells decreased proliferation and migration. Conclusion In this study, we are able to demonstrate that Salinomycin is an effective agent against previously resistant CC cells and might be a potential candidate for the treatment of CC in the future.

  15. Upregulation of human telomerase reverse transcriptase mRNA expression by in vitro transfection of hepatitis B virus X gene into human hepatocarcinoma and cholangiocarcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Zhen-Liang Qu; Sheng-Quan Zou; Nai-Qiang Cui; Xian-Zhong Wu; Ming-Fang Qin; Di Kong; Zhen-Li Zhou


    AIM: To study the changes of human telomerase reverse transcriptase (hTERT) mRNA expression in human hepatocarcinoma cell lines (HepG2) and cholangiocarcinoma cell lines (QBC939) after HBx gene transfection and to illustrate the significance of transcriptional regulation of hTERT gene by HBx gene in the carcinogenesis.METHODS: HepG2 and QBC939 cell lines were cultured and co-transfected with eukaryotic expression vector containing the HBx coding region and cloning vector containing enhanced green fluorescent protein (EGFP) coding sequence using lipid-mediated gene transduction technique. Thirty-six hours after transfection, EGFP expression in cells was used as the indicator of successful transfection. Flow cytometry was performed to determine the transfection efficiency.Cells were harvested and total RNA was extracted using TRIzol() reagent. The expression of hTERT mRNA in HepG2and QBC939 cell lines was assayed by reverse transcriptionpolymerase chain reaction. The expression of HBx protein in both cell lines was detected by immunocytochemical staining and Western blotting.RESULTS: Flow cytometry showed that the transfection efficiency was 46.4% in HepG2 cells and 29.6% in QBC939cells for both HBx gene expression vector and blank vector. The expression of hTERT mRNA was meaningfully increased in HepG2 and QBC939 cell lines when transfected with HBx gene expression vector compared to those transfected with OPTI-MEM medium and blank vector.Immunocytochemical staining and Western blotting revealed HBx protein expression in HepG2 and QBC939cells only when transfected with HBx gene.CONCLUSION: HBx gene transfection can upregulate the transcriptional expression of hTERT mRNA. The transactivation of hTERT gene by HBx gene is a newfound mechanism for pathogenesis of hepatocarcinomas and cholangiocarcinomas after HBV infection.

  16. Suppressing Effects of Down-regulating DNMT1 and DNMT3b Expression on the Growth of Human Cholangiocarcinoma Cell Line

    Institute of Scientific and Technical Information of China (English)

    Shi ZUO; Jian LUO; Minfeng LIU; Lining XU; Jingqing DONG; Wei GUO; Shengquan ZOU


    Hypermethylation in the promoter region is an important epigenetic mechanism for the transcriptional repression of a number of cancer-associated genes, and over-expression and/or increased activity of DNA methyltransferases are considered to be the main cause of promoter hypermethylation. In order to explore the roles of two methyltransferase members (DNMT1 and DNMT3b) in the cholangiocarcinoma tumorigenesis, antisense eukaryotic expression plasmid of DNMT1 and DNMT3b gene was constructed respectively, and were co-transfected into the human cholangiocarcinoma cell line QBC-939 to observe their biological effects on the cell growth and proliferation ability, apoptosis, cell cycle alteration, and the tumorigenesis ability in the subcutaneous tissue of nude mouse. The results demonstrated that co-transfection with antisense eukaryotic expression plasmid of DNMT1 and DNMT3b gene and single transfection with antisense eukaryotic expression plasmid of DNMT1 gene can suppress the growth and proliferation of QBC-939, block the cell cycle at G1 phase, increase the apoptosis rate, minimize the tumor size in the subcutaneous tissue of nude mouse. The suppressing biological effect of co-transfection is stronger than single transfection with antisense DNMT1. Meanwhile, single transfection with antisense eukaryotic expression plasmid of DNMT3b gene has no effects on the biological characteristics of QBC-939. This study suggests that DNMT1 gene plays a key role in DNA methylation and DNMT3b gene may act as an accessory to support its function in inactivation of tumor suppressor genes. Combination DNMT1 and DNMT3b will inhere their biological effects and have the synergistic effect on suppressing the growth of human cholangiocarcinoma cell line QBC-939.

  17. Sensitivity of Human Intrahepatic Cholangiocarcinoma Subtypes to Chemotherapeutics and Molecular Targeted Agents: A Study on Primary Cell Cultures (United States)

    Fraveto, Alice; Cardinale, Vincenzo; Bragazzi, Maria Consiglia; Giuliante, Felice; De Rose, Agostino Maria; Grazi, Gian Luca; Napoletano, Chiara; Semeraro, Rossella; Lustri, Anna Maria; Costantini, Daniele; Nevi, Lorenzo; Di Matteo, Sabina; Renzi, Anastasia; Carpino, Guido; Gaudio, Eugenio; Alvaro, Domenico


    We investigated the sensitivity of intrahepatic cholangiocarcinoma (IHCCA) subtypes to chemotherapeutics and molecular targeted agents. Primary cultures of mucin- and mixed-IHCCA were prepared from surgical specimens (N. 18 IHCCA patients) and evaluated for cell proliferation (MTS assay) and apoptosis (Caspase 3) after incubation (72 hours) with increasing concentrations of different drugs. In vivo, subcutaneous human tumor xenografts were evaluated. Primary cultures of mucin- and mixed-IHCCA were characterized by a different pattern of expression of cancer stem cell markers, and by a different drug sensitivity. Gemcitabine and the Gemcitabine-Cisplatin combination were more active in inhibiting cell proliferation in mixed-IHCCA while Cisplatin or Abraxane were more effective against mucin-IHCCA, where Abraxane also enhances apoptosis. 5-Fluoracil showed a slight inhibitory effect on cell proliferation that was more significant in mixed- than mucin-IHCCA primary cultures and, induced apoptosis only in mucin-IHCCA. Among Hg inhibitors, LY2940680 and Vismodegib showed slight effects on proliferation of both IHCCA subtypes. The tyrosine kinase inhibitors, Imatinib Mesylate and Sorafenib showed significant inhibitory effects on proliferation of both mucin- and mixed-IHCCA. The MEK 1/2 inhibitor, Selumetinib, inhibited proliferation of only mucin-IHCCA while the aminopeptidase-N inhibitor, Bestatin was more active against mixed-IHCCA. The c-erbB2 blocking antibody was more active against mixed-IHCCA while, the Wnt inhibitor, LGK974, similarly inhibited proliferation of mucin- and mixed-IHCCA. Either mucin- or mixed-IHCCA showed high sensitivity to nanomolar concentrations of the dual PI3-kinase/mTOR inhibitor, NVP-BEZ235. In vivo, in subcutaneous xenografts, either NVP-BEZ235 or Abraxane, blocked tumor growth. In conclusion, mucin- and mixed-IHCCA are characterized by a different drug sensitivity. Cisplatin, Abraxane and the MEK 1/2 inhibitor, Selumetinib were more

  18. Sensitivity of Human Intrahepatic Cholangiocarcinoma Subtypes to Chemotherapeutics and Molecular Targeted Agents: A Study on Primary Cell Cultures.

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    Alice Fraveto

    Full Text Available We investigated the sensitivity of intrahepatic cholangiocarcinoma (IHCCA subtypes to chemotherapeutics and molecular targeted agents. Primary cultures of mucin- and mixed-IHCCA were prepared from surgical specimens (N. 18 IHCCA patients and evaluated for cell proliferation (MTS assay and apoptosis (Caspase 3 after incubation (72 hours with increasing concentrations of different drugs. In vivo, subcutaneous human tumor xenografts were evaluated. Primary cultures of mucin- and mixed-IHCCA were characterized by a different pattern of expression of cancer stem cell markers, and by a different drug sensitivity. Gemcitabine and the Gemcitabine-Cisplatin combination were more active in inhibiting cell proliferation in mixed-IHCCA while Cisplatin or Abraxane were more effective against mucin-IHCCA, where Abraxane also enhances apoptosis. 5-Fluoracil showed a slight inhibitory effect on cell proliferation that was more significant in mixed- than mucin-IHCCA primary cultures and, induced apoptosis only in mucin-IHCCA. Among Hg inhibitors, LY2940680 and Vismodegib showed slight effects on proliferation of both IHCCA subtypes. The tyrosine kinase inhibitors, Imatinib Mesylate and Sorafenib showed significant inhibitory effects on proliferation of both mucin- and mixed-IHCCA. The MEK 1/2 inhibitor, Selumetinib, inhibited proliferation of only mucin-IHCCA while the aminopeptidase-N inhibitor, Bestatin was more active against mixed-IHCCA. The c-erbB2 blocking antibody was more active against mixed-IHCCA while, the Wnt inhibitor, LGK974, similarly inhibited proliferation of mucin- and mixed-IHCCA. Either mucin- or mixed-IHCCA showed high sensitivity to nanomolar concentrations of the dual PI3-kinase/mTOR inhibitor, NVP-BEZ235. In vivo, in subcutaneous xenografts, either NVP-BEZ235 or Abraxane, blocked tumor growth. In conclusion, mucin- and mixed-IHCCA are characterized by a different drug sensitivity. Cisplatin, Abraxane and the MEK 1/2 inhibitor, Selumetinib

  19. Cholangiocarcinoma cell line TK may be useful for the pharmacokinetic study of the chemotherapeutic agent gemcitabine. (United States)

    Kamada, Minori; Akiyoshi, Kohei; Akiyama, Nobutake; Funamizu, Naotake; Watanabe, Michiko; Fujioka, Kouki; Ikeda, Kei-Ichi; Manome, Yoshinobu


    Cholangiocarcinoma is a disease with a poor prognosis. A human cholangiocarcinoma cell line, TK, was previously established to enable further understanding of the disease. We conducted this investigation to determine whether or not the TK line is useful for pharmacokinetic study of the chemotherapeutic agent gemcitabine (GEM). Along with the BXPC3 human pancreatic adenocarcinoma cell line, the sensitivity to and effects on the TK cell line of GEM were compared. The influence of deoxycytidine kinase (dCK) transduction was also comparatively investigated. The effects of GEM in terms of drug sensitivity of the TK cell line, cell cycle and levels of transcripts of key enzymes were comparable to the BXPC3 cell line. Responses to the drug were similar in both cell lines. In contrast to pancreatic carcinoma, cell lines for research on cholangiocarcinoma have been limited. This study suggests the application of the TK cell line to the pharmacokinetic study of the chemosensitization of therapeutic drugs, such as GEM.

  20. Xanthohumol inhibits STAT3 activation pathway leading to growth suppression and apoptosis induction in human cholangiocarcinoma cells. (United States)

    Dokduang, Hasaya; Yongvanit, Puangrat; Namwat, Nisana; Pairojkul, Chawalit; Sangkhamanon, Sakkarn; Yageta, Mika Sakurai; Murakami, Yoshinori; Loilome, Watcharin


    STAT3 plays a significant role in the development of cholangiocarcinoma (CCA) associated with the liver fluke (Opisthorchis viverrini; Ov). Xanthohumol (XN), a prenylated flavonoid extracted from hops, has known anticancer activity and could potentially target STAT3. The present study determined the effect of XN on STAT3, as well as ascertained its usefulness against CCA. The CCA cell proliferation at 20 µM and 50 µM of XN was shown to inhibited, while 20 µM partially inhibited IL-6-induced STAT3 activation. At 50 µM, the inhibition was complete. The reduction in STAT3 activity at 20 and 50 µM was associated with a significant reduction of CCA cell growth and apoptosis. We also found that the administration of 50 µM XN orally in drinking water to nude mice inoculated with CCA led to a reduction in tumor growth in comparison with controls. In addition, apoptosis of cancer cells increased although there was no visible toxicity. The present study shows that XN can inhibit STAT3 activation both in vivo and in vitro due to suppression of the Akt-NFκB signaling pathway. XN should be considered as a possible therapeutic agent against CCA.

  1. A polysaccharide from Pinellia ternata inhibits cell proliferation and metastasis in human cholangiocarcinoma cells by targeting of Cdc42 and 67kDa Laminin Receptor (LR). (United States)

    Li, Yong; Li, Dajiang; Chen, Jian; Wang, Shuguang


    In this study, we isolated and purified a polysaccharide (PTPA) from the tubers of Pinellia ternate. We aimed to evaluate the cytotoxic effects of PTPA on human cholangiocarcinoma (CCA) cell lines and to identify the underlying molecular mechanism. PTPA at the dose from 25 to 200μg/mL showed significant inhibitory effect on the proliferation of four cancer cell lines (SNU-245, CL-6, Sk-ChA-1 and MZ-ChA-1), among which Sk-ChA-1 was a most sensitive cell line to PTPA treatment via induction of apoptosis. Interestingly, RNA interference of Sk-ChA-1 cells with 67LR or Cdc42-targeted shRNAs resulted a similar potency in decreasing cell viability and causing apoptotic death. Moreover, PTPA (100μg/mL) or 67LR or Cdc42 special shRNAs increased the ratio of pro-apoptotic Bax to anti-apoptotic Bcl-2, induced the activation of caspase-9 and caspase-3, but not caspsase-8, and inhibited the expression of 67LR or Cdc42 protein in Sk-ChA-1 cells. Taken together, the inhibitory effect of PTPA on the cell growth of Sk-ChA-1 cells was at least in part mediated via the activation of the intrinsic mitochondrial apoptotic pathway and the downregulation of 67LR or Cdc42 protein expression. Thus, PTPA may be developed as a promising candidate for chemopreventive agent in the prevention and treatment of human CCA.

  2. XIAP antagonist embelin inhibited proliferation of cholangiocarcinoma cells.

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    Cody J Wehrkamp

    Full Text Available Cholangiocarcinoma cells are dependent on antiapoptotic signaling for survival and resistance to death stimuli. Recent mechanistic studies have revealed that increased cellular expression of the E3 ubiquitin-protein ligase X-linked inhibitor of apoptosis (XIAP impairs TRAIL- and chemotherapy-induced cytotoxicity, promoting survival of cholangiocarcinoma cells. This study was undertaken to determine if pharmacologic antagonism of XIAP protein was sufficient to sensitize cholangiocarcinoma cells to cell death. We employed malignant cholangiocarcinoma cell lines and used embelin to antagonize XIAP protein. Embelin treatment resulted in decreased XIAP protein levels by 8 hours of treatment with maximal effect at 16 hours in KMCH and Mz-ChA-1 cells. Assessment of nuclear morphology demonstrated a concentration-dependent increase in nuclear staining. Interestingly, embelin induced nuclear morphology changes as a single agent, independent of the addition of TNF-related apoptosis inducing ligand (TRAIL. However, caspase activity assays revealed that increasing embelin concentrations resulted in slight inhibition of caspase activity, not activation. In addition, the use of a pan-caspase inhibitor did not prevent nuclear morphology changes. Finally, embelin treatment of cholangiocarcinoma cells did not induce DNA fragmentation or PARP cleavage. Apoptosis does not appear to contribute to the effects of embelin on cholangiocarcinoma cells. Instead, embelin caused inhibition of cell proliferation and cell cycle analysis indicated that embelin increased the number of cells in S and G2/M phase. Our results demonstrate that embelin decreased proliferation in cholangiocarcinoma cell lines. Embelin treatment resulted in decreased XIAP protein expression, but did not induce or enhance apoptosis. Thus, in cholangiocarcinoma cells the mechanism of action of embelin may not be dependent on apoptosis.

  3. High mobility group A1 enhances tumorigenicity of human cholangiocarcinoma and confers resistance to therapy

    DEFF Research Database (Denmark)

    Quintavalle, Cristina; Burmeister, Katharina; Piscuoglio, Salvatore


    High mobility group A1 (HMGA1) protein has been described to play an important role in numerous types of human carcinoma. By the modulation of several target genes HMGA1 promotes proliferation and epithelial-mesenchymal transition of tumor cells. However, its role in cholangiocarcinoma (CCA) has ...

  4. Proteomic Studies of Cholangiocarcinoma and Hepatocellular Carcinoma Cell Secretomes

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    Chantragan Srisomsap


    Full Text Available Cholangiocarcinoma (CCA and hepatocellular carcinoma (HCC occur with relatively high incidence in Thailand. The secretome, proteins secreted from cancer cells, are potentially useful as biomarkers of the diseases. Proteomic analysis was performed on the secreted proteins of cholangiocarcinoma (HuCCA-1 and hepatocellular carcinoma (HCC-S102, HepG2, SK-Hep-1, and Alexander cell lines. The secretomes of the five cancer cell lines were analyzed by SDS-PAGE combined with LC/MS/MS. Sixty-eight proteins were found to be expressed only in HuCCA-1. Examples include neutrophil gelatinase-associated lipocalin (lipocalin 2, laminin 5 beta 3, cathepsin D precursor, desmoplakin, annexin IV variant, and annexin A5. Immunoblotting was used to confirm the presence of lipocalin 2 in conditioned media and cell lysate of 5 cell lines. The results showed that lipocalin 2 was a secreted protein which is expressed only in the conditioned media of the cholangiocarcinoma cell line. Study of lipocalin 2 expression in different types of cancer and normal tissues from cholangiocarcinoma patients showed that lipocalin 2 was expressed only in the cancer tissues. We suggest that lipocalin 2 may be a potential biomarker for cholangiocarcinoma.

  5. Identification of CXCL5/ENA-78 as a factor involved in the interaction between cholangiocarcinoma cells and cancer-associated fibroblasts. (United States)

    Okabe, Hirohisa; Beppu, Toru; Ueda, Mitsuharu; Hayashi, Hiromitsu; Ishiko, Takatoshi; Masuda, Toshiro; Otao, Ryu; Horlad, Hasita; Mima, Kosuke; Miyake, Keisuke; Iwatsuki, Masaaki; Baba, Yoshifumi; Takamori, Hiroshi; Jono, Hirofumi; Shinriki, Satoru; Ando, Yukio; Baba, Hideo


    Knowledge of tumor-stromal interactions is essential for understanding tumor development. We focused on the interaction between cholangiocarcinoma and cancer-associated fibroblasts (CAFs) in intrahepatic cholangiocarcinoma and reported their positive interaction in vitro and in vivo. The aim of this study is to identify the key protein involved in the interaction between cholangiocarcinoma cells and CAFs and its role on cholangiocarcinoma progression. Using the conditioning medium from cholangiocarcinoma cells, hepatic stellate cells and coculture of them, Protein-Chip analysis with SELDI-TOF-MS showed that the peak of an 8,360-Da protein remarkably increased in the coculture medium. This protein was identified as CXCL5/ENA78, epithelial cell-derived neutrophil-activating peptide-78, by q-TOF/MS/MS analysis. Two cholangiocarcinoma cell lines, HuCCT1 and RBE, produced CXCL5 that promoted their invasion and migration in an autocrine fashion. These effects of CXCL5 significantly decreased by inhibition of CXC-receptor 2, which is the receptor for CXCL5. In addition, IL-1β produced by hepatic stellate cells induced the expression of CXCL5 in cholangiocarcinoma cells. In human tissue samples, a significant correlation was observed between CAFs and CXCL5 produced by cholangiocarcinoma cells in intrahepatic cholangiocarcinoma (p = 0.0044). Furthermore, the high-CXCL5-expression group exhibited poor overall survival after curative hepatic resection (p = 0.027). The presence of tumor-infiltrating neutrophils expressing CD66b was associated with CXCL5 expression in tumor cells (p < 0.0001). These data suggest that CXCL5 is important for the interaction between cholangiocarcinoma and CAFs, and inhibition of tumor-stromal interactions may be a useful therapeutic approach for cholangiocarcinoma.

  6. Dicoumarol enhances gemcitabine-induced cytotoxicity in high NQO1-expressing cholangiocarcinoma cells

    Institute of Scientific and Technical Information of China (English)

    Benjaporn; Buranrat; Auemduan; Prawan; Upa; Kukongviriyapan; Sarinya; Kong-petch; Veerapol; Kukongviriyapan


    AIM: To investigate whether dicoumarol, a potent inhibitor of NAD(P)H quinone oxidoreductase-1 (NQO1), potentiates gemcitabine to induce cytotoxicity in chol-angiocarcinoma cells (CCA) and the role of reactive oxygen generation in sensitizing the cells. METHODS: Four human cell lines with different NQO1 activity were used; the human CCA cell lines, KKU-100, KKU-OCA17, KKU-M214, and Chang liver cells. NQO1 activity and mRNA expression were determined. The cells were pretreated with dicoumarol at relevant con...

  7. Molecular mechanisms of cholangiocarcinoma cell inhibition by medicinal plants (United States)

    Leelawat, Surang; Leelawat, Kawin


    Cholangiocarcinoma (CCA) is one of the most common causes of cancer-associated mortality in Thailand. Certain phytochemicals have been demonstrated to modulate apoptotic signaling pathways, which may be targeted for the prevention and treatment of cancer. Therefore, the aim of the present study was to investigate the effect of specific medicinal plants on the inhibition of CCA cell proliferation, and to identify the molecular mechanisms underlying this. A WST-1 cell proliferation assay was performed using an RMCCA1 cell line, and apoptotic signaling pathways were also investigated using a PathScan Stress and Apoptosis Signaling Antibody Array Kit. The cell proliferation assay indicated that extracts from the Phyllanthus emblica fruit pulp (PEf), Phyllanthus emblica seed (PEs), Terminalia chebula fruit pulp (TCf), Terminalia chebula seed (TCs), Areca catechu seed (ACs), Curcuma longa (CL) and Moringa oleifera seed (MOs) exerted anti-proliferative activity in RMCCA1 cells. In addition, the PathScan assay revealed that certain pro-apoptotic molecules, including caspase-3, poly (ADP-ribose) polymerase, checkpoint kinase 2 and tumor protein 53, exhibited increased activity in RMCCA1 cells treated with the aforementioned selected plant extracts, with the exception of PEf. The mitogen-activated protein kinase (MAPK) pathways (including ERK1/2 and p38 MAPK) expression level was significantly increased in RMCCA1 cells pre-treated with extracts of PEs, TCf, CL and MOs. The activation of protein kinase B (Akt) was significantly demonstrated in RMCCA1 cells pre-treated with extracts of TCf, ACs and MOs. In summary, the present study demonstrated that extracts of PEs, TCf, TCs, ACs, CL and MOs exhibited anti-proliferative effects in CCA cells by inducing pro-apoptotic signals and modulating signal transduction molecules. Further studies in vivo are required to demonstrate the potential applications of specific plant extracts for the treatment of human cancer.

  8. Establishment and characterization of a cholangiocarcinoma cell line (RMCCA-1) from a Thai patient

    Institute of Scientific and Technical Information of China (English)

    Panthip Rattanasinganchan; Kawin Leelawat; Sa-ard Treepongkaruna; Chintana Tocharoentanaphol; Somboon Subwongcharoen; Tuangporn Suthiphongchai; Rutaiwan Tohtong


    AIM: To establish and characterize a new cell line derived from peripheral cholangiocarcinoma of a Thai patient.METHODS: The peripheral cholangiocarcinoma specimen surgically obtained from the patient was aseptically processed by washing and mincing before culturing in Ham's F12 medium containing 10% fetal bovine serum. After 3 mo, when the cell line has become homogeneous and stabilized, several features were investigated, including growth characteristics,immunofluorescence staining for cytokeratins, expression of tumor markers, chromosomal analysis by G-banding and multicolour fluorescence in situ hybridization (mFISH), in vitro migration and invasion characteristics.RESULTS: The RMCCA-1 cell line has been established.These cells proliferated as a monolayer with a population doubling time of 48 h. Immunofluorescence staining showed positive staining for human cytokeratin 7 and 19 verifying the biliary epithelial origin. RMCCA-1secreted carbohydrate antigen 19-9 (CA19-9), but insignificant levels of carcinoembryonic antigen (CEA)and α-fetoprotein (AFP). Chromosome analysis identified aneuploidy karyotypes with a modal chromosome number of 59. RMCCA-1 exhibited a low level of in vitro invasiveness, but a high degree of motility. The cell line exhibited a significant number of chromosomal aberrations as shown by mFISH and G-banding methods.CONCLUSION: A new cell line derived from peripheral cholangiocarcinoma of a Thai patient has been established. This cell line shows a low level of in vitro invasiveness, but a high degree of motility. It will serve as a valuable tool for further studies on tumor biology,molecular pathogenesis, metastatic mechanism and response to therapeutic drugs of cholangiocarcinoma.

  9. MART-10 represses cholangiocarcinoma cell growth and high vitamin D receptor expression indicates better prognosis for cholangiocarcinoma (United States)

    Chiang, Kun-Chun; Yeh, Ta-Sen; Huang, Cheng-Cheng; Chang, Yu-Chan; Juang, Horng-Heng; Cheng, Chi-Tung; Pang, Jong-Hwei S.; Hsu, Jun-Te; Takano, Masashi; Chen, Tai C.; Kittaka, Atsushi; Hsiao, Michael; Yeh, Chun-Nan


    Cholangiocarcinoma (CCA) is a devastating disease due to no effective treatments available. Since the non-mineral functions of vitamin D emerges, 1α,25(OH)2D3, the active form of vitamin D, has been applied in anti-cancer researches. In this study, we demonstrated that both the 1α,25(OH)2D3 analog, MART-10, and 1α,25(OH)2D3 possessed anti-growth effect on human CCA cells with MART-10 much more potent than 1α,25(OH)2D3. The growth inhibition of both drugs were mediated by induction of G0/G1 cell cycle arrest through upregulation of p27 and downregulation of CDK4, CDK6, and cyclin D3. Human neutrophil gelatinase associated lipocalin (NGAL) was found to be involved in 1α,25(OH)2D3 and MART-10 meditated growth inhibition for CCA as knockdown of NGAL decreased Ki-67 expression in SNU308 cells and rendered SNU308 cells less responsive to 1α,25(OH)2D3 and MART-10 treatment. Vitamin D receptor (VDR) knockdown partly abolished MART-10-induced inhibition of NGAL and cell growth in SNU308 cells. The xenograft animal study demonstrated MART-10 could effectively repressed CCA growth in vivo without inducing obvious side effects. The IHC examination of human CCA specimen for VDR revealed that higher VDR expression was linked with better prognosis. Collectively, our results suggest that MART-10 could be a promising regimen for CCA treatment. PMID:28256614

  10. miR-29b, miR-205 and miR-221 enhance chemosensitivity to gemcitabine in HuH28 human cholangiocarcinoma cells.

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    Kinya Okamoto

    Full Text Available BACKGROUND AND AIMS: Cholangiocarcinoma (CCA is highly resistant to chemotherapy, including gemcitabine (Gem treatment. MicroRNAs (miRNAs are endogenous, non-coding, short RNAs that can regulate multiple genes expression. Some miRNAs play important roles in the chemosensitivity of tumors. Here, we examined the relationship between miRNA expression and the sensitivity of CCA cells to Gem. METHODS: Microarray analysis was used to determine the miRNA expression profiles of two CCA cell lines, HuH28 and HuCCT1. To determine the effect of candidate miRNAs on Gem sensitivity, expression of each candidate miRNA was modified via either transfection of a miRNA mimic or transfection of an anti-oligonucleotide. Ontology-based programs were used to identify potential target genes of candidate miRNAs that were confirmed to affect the Gem sensitivity of CCA cells. RESULTS: HuCCT1 cells were more sensitive to Gem than were HuH28 cells, and 18 miRNAs were differentially expressed whose ratios over ± 2log2 between HuH28 and HuCCT1. Among these 18 miRNAs, ectopic overexpression of each of three downregulated miRNAs in HuH28 (miR-29b, miR-205, miR-221 restored Gem sensitivity to HuH28. Suppression of one upregulated miRNA in HuH28, miR-125a-5p, inhibited HuH28 cell proliferation independently to Gem treatment. Selective siRNA-mediated downregulation of either of two software-predicted targets, PIK3R1 (target of miR-29b and miR-221 or MMP-2 (target of miR-29b, also conferred Gem sensitivity to HuH28. CONCLUSIONS: miRNA expression profiling was used to identify key miRNAs that regulate Gem sensitivity in CCA cells, and software that predicts miRNA targets was used to identify promising target genes for anti-tumor therapies.

  11. Matrine induces RIP3-dependent necroptosis in cholangiocarcinoma cells (United States)

    Xu, Beibei; Xu, Minying; Tian, Yuan; Yu, Qiang; Zhao, Yujie; Chen, Xiong; Mi, Panying; Cao, Hanwei; Zhang, Bing; Song, Gang; Zhan, Yan-yan; Hu, Tianhui


    The development of acquired resistance to pro-apoptotic antitumor agents is a major impediment to the cure of cholangiocarcinoma (CCA). Antitumor drugs inducing non-apoptotic cell death are considered as a new approach to overcome such drug resistance. Here, we reported for the first time that matrine-induced necroptosis in CCA cell lines, differing from its classical role to induce apoptosis in many other kinds of cancer cells. CCA cells under matrine treatment exhibited typical necrosis-like but not apoptotic morphologic change. These matrine-induced morphologic change and cell death in CCA cells were greatly attenuated by necroptosis inhibitor necrostatin-1, but not apoptosis inhibitor z-VAD-fmk. Unlike many cancer cells with negative receptor-interacting protein 3 (RIP3) expression, moderate expression of RIP3 in CCA cells was observed and was required for matrine to induce necroptosis, which was switched to apoptosis after knocking down endogenous RIP3. Moreover, matrine could increase RIP3 expression level, which may facilitate the necroptosis process. Translocation of mixed lineage kinase-domain like (MLKL) from cytoplasm to plasma membrane as a downstream event of RIP3, as well as the increased production of reactive oxygen species (ROS) by RIP3/MLKL, was critical for matrine to induce necroptosis. In clinical study, we found RIP3 was lower but still moderately expressed in most CCA tissue samples compared with adjacent normal tissues. Taken together, we identified matrine as a necroptosis inducer in CCA by enhancing RIP3 expression and the following RIP3/MLKL/ROS signaling pathway, which provided new individualized strategies based on RIP3 expression to overcome chemoresistance in CCA therapy. PMID:28179994

  12. YAP promotes proliferation, chemoresistance, and angiogenesis in human cholangiocarcinoma through TEAD transcription factors. (United States)

    Marti, Patricia; Stein, Claudia; Blumer, Tanja; Abraham, Yann; Dill, Michael T; Pikiolek, Monika; Orsini, Vanessa; Jurisic, Giorgia; Megel, Philippe; Makowska, Zuzanna; Agarinis, Claudia; Tornillo, Luigi; Bouwmeester, Tewis; Ruffner, Heinz; Bauer, Andreas; Parker, Christian N; Schmelzle, Tobias; Terracciano, Luigi M; Heim, Markus H; Tchorz, Jan S


    The Yes-associated protein (YAP)/Hippo pathway has been implicated in tissue development, regeneration, and tumorigenesis. However, its role in cholangiocarcinoma (CC) is not established. We show that YAP activation is a common feature in CC patient biopsies and human CC cell lines. Using microarray expression profiling of CC cells with overexpressed or down-regulated YAP, we show that YAP regulates genes involved in proliferation, apoptosis, and angiogenesis. YAP activity promotes CC growth in vitro and in vivo by functionally interacting with TEAD transcription factors (TEADs). YAP activity together with TEADs prevents apoptosis induced by cytotoxic drugs, whereas YAP knockdown sensitizes CC cells to drug-induced apoptosis. We further show that the proangiogenic microfibrillar-associated protein 5 (MFAP5) is a direct transcriptional target of YAP/TEAD in CC cells and that secreted MFAP5 promotes tube formation of human microvascular endothelial cells. High YAP activity in human CC xenografts and clinical samples correlates with increased MFAP5 expression and CD31(+) vasculature. These findings establish YAP as a key regulator of proliferation and antiapoptotic mechanisms in CC and provide first evidence that YAP promotes angiogenesis by regulating the expression of secreted proangiogenic proteins. © 2015 by the American Association for the Study of Liver Diseases.

  13. Inhibition of PI3K increases oxaliplatin sensitivity in cholangiocarcinoma cells

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    Udomchaiprasertkul Wandee


    Full Text Available Abstract Background Resistance of cholangiocarcinoma to chemotherapy is a major problem in cancer treatment. The mechanism of resistance is believed to involve phosphoinositide-3- kinase (PI3K/Akt activation. Although the platinum-containing compound oxaliplatin has been extensively used in the treatment of several solid tumors, recent data regarding its use to treat cholangiocarcinoma are ambiguous. Oxaliplatin resistance in this disease could potentially involve PI3K pathways. We, therefore, examined the effects of PI3K pathways in cholangiocarcinoma cells in modulating oxaliplatin resistance. Results After exposing the cholangiocarcinoma cell lines RMCCA1 and KKU100 to oxaliplatin, the levels of Akt and mTOR phosphorylation increased, as shown by western blot analysis. The WST-1 cell proliferation assay showed increased inhibition of cell growth under high concentrations of oxaliplatin. The combination of oxaliplatin with LY294002, an inhibitor of PI3K, resulted in a remarkable arrest of cell proliferation. Deactivation of mTOR by RAD001 was also synergistic with oxaliplatin, although to a lesser extent. The combination of oxaliplatin and a PI3K inhibitor also resulted in a significant induction of apoptosis, as demonstrated by the TUNEL assay. Conclusion Activation of PI3K might protect cholangiocarcinoma cells from oxaliplatininduced cytotoxicity. Although the inhibition of PI3K and the inhibition of mTOR both enhance oxaliplatin-induced cytotoxicity, PI3K inhibition has a greater effect. Targeting the PI3K pathway may be a useful approach to improve the chemotherapeutic sensitivity of cholangiocarcinoma.

  14. Establishment and characterization of an opisthorchiasis-associated cholangiocarcinoma cell line (KKU-100)

    Institute of Scientific and Technical Information of China (English)

    Banchob Sripa; Saman Leungwattanawanit; Takayuki Nitta; Chaisiri Wongkham; Vajarabhongsa Bhudhisawasdi; Anucha Puapairoj; Chongrak Sripa; Masanao Miwa


    AIM To establish and dharacterize a nev cholangiocarcinoma cell line from a patient living in the Opisthorchis viverrini (O. viverrini) endemic area of Northeast Thailand.METHODS: Fresh liver biopsy and bile specimens were obtained from a 65-year-old Thai woman with cholangiocarcinoma of the porta hepatis. After digestion, the cells were cultured in Ham's F12 media. The established cell line was then characterized for growth kinetics, cell morphology, imm unocytochemistry and cytogenetics. Tumorigenicity of the cell line was determined by heterotransplanting in nude mice. RESULTS: The primary tumor was a poorly differentiated tubular adenocarcinoma. Examination of the bile revealed malignant cells with O. viverrini eggs. The cholangiocarcinoma cell line KKU-100 was established 4 mo after the primary culture-population doubling time was 72 h. KKU-100 possesses compact and polygonal-shapedepithelial cells. Immunocytochemically, this cell line exhibited cytokeratin, EMA, CEA, and CA125, but not α-fetoprotein (AFP), CA19-9, desmin, c-met, or p53. Such protein expressions parallel those of the primary tumor. Cytogenetic analysis identified aneuploidy karyotypes with a modal chromosome number of 78 and marked chromosomal structural changes. Inoculation of KKU-100 cells into nude mice produced a transplantable, poorly differentiated aden-ocarcinoma, similar to the original tumor.CONCLUSION: KKJ-100 is the first egg-proven, Opisthorchis- associated cholangiocarcinoma cell line, which should prove useful for further investigations of the tumor biology of this cancer.

  15. Metallothionein overexpression and its prognostic relevance in intrahepatic cholangiocarcinoma and extrahepatic hilar cholangiocarcinoma (Klatskin tumors). (United States)

    Schmitz, Klaus Jürgen; Lang, Hauke; Kaiser, Gernot; Wohlschlaeger, Jeremias; Sotiropoulos, Georgios Charalambos; Baba, Hideo Andreas; Jasani, Bharat; Schmid, Kurt Werner


    Metallothionein is a group of small molecular weight cysteine-rich proteins with a broad variety of functions. Metallothionein has been shown to regulate apoptosis and proliferation. Overexpression of metallothionein frequently occurs in human tumors and is related to prognosis as well as therapy response. However, metallothionein expression and its clinical relevance in cholangiocarcinoma have not been investigated. The present study aimed to analyze metallothionein over-expression and its possible prognostic impact in intrahepatic cholangiocarcinoma and hilar extrahepatic cholangiocarcinoma (Klatskin tumors). We investigated the relationship of immunohistochemically demonstrated metallothionein expression with various clinicopathological parameters in a series of 56 intrahepatic and 56 extrahepatic cholangiocarcinoma. In noncancerous bile duct epithelia metallothionein was only occasionally weakly expressed; strong metallothionein overexpression (>50% metallothionein -positive tumor cells) was noted in 7 (12.5%) of 56 intrahepatic cholangiocarcinoma and 14 (25%) of 56 Klatskin tumors, which was associated with poor clinical outcome in univariate Kaplan-Meier testing in both intrahepatic cholangiocarcinoma (P = .002) and Klatskin tumors (P = .034). Moreover, strong metallothionein expression was identified as an independent prognostic parameter in multivariate Cox regression analysis in both intrahepatic cholangiocarcinoma (P = .005) and Klatskin tumors (P = .035). In contrast, cholangiocarcinoma with a papillary phenotype (8/112; 7.1%) exhibited a significant lack of strong metallothionein expression in all 8 of 8 cases. Strong metallothionein expression is identified as an independent poor prognostic parameter, and determination of the metallothionein expression may serve as an additional tool for the therapeutic management of patients with cholangiocarcinoma. In comparison, lack of metallothionein expression seems to be associated with cholangiocarcinoma with a

  16. Inlfuences of Resveratrol on the Proliferation of Cholangiocarcinoma Cells and Expression of p21 and p27 Proteins

    Institute of Scientific and Technical Information of China (English)

    Liu Yong; Qian Huaxiang; Zhuo Ma; Xu Jianzhong


    Objective:To explore the effects of resveratrol on the proliferation of cholangiocarcinoma cells and expression of p21 and p27 proteins. Methods:The influences of resveratrol on the proliferation of cholangiocarcinoma cells and expression of p21 and p27 proteins were detected by methyl thiazolyl tetrazolium (MTT) method and Western blotting method, respectively. Results:Resveratrol had a conspicuous inhibitory effect on the proliferation of cholangiocarcinoma QBC939 cells, and was concentration-and time-dependant (P Conclusion:Resveratrol can signiifcantly inhibit the proliferation of cholangiocarcinoma QBC939 cells, which may be related to regulation of p21 and p27 expression and negative regulation to cell cycles.

  17. Moderndiagnosticapproachesto cholangiocarcinoma

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    Larisa E Vasilieva; Stefanos I Papadhimitriou; Spyros P Dourakis


    BACKGROUND: Cholangiocarcinoma is a very aggressive tumor with poor survival. Therefore, early diagnosis and surgical resection are of paramount importance. Its diagnosis is dififcult because access to the tumor is not easy. Biopsy is possible only for intrahepatic cholangiocarcinoma, which accounts for 10%of cases. Routine brush cytology from endoscopic retrograde cholangiopancreatography (ERCP) has a high speciifcity of 100%but unfortunately a low sensitivity of 30%. In this review we brielfy describe new diagnostic techniques applicable to ERCP brush cytology specimens and targeting the genetic background of the disease, in particular lfuorescence in situ hybridization (FISH) and digital image analysis (DIA). DATE SOURCES: The PubMed database up to 2011 was used for the retrieval of relevant articles. The search terms FISH, lfuorescence in situ hybridization, DIA, digital image analysis and cholangiocarcinoma were used. Both original and review articles were used. RESULTS: FISH identiifes cells with chromosomal abnormalities, mainly numerical aberrations, using a mixture of lfuorescence-labeled probes. FISH offers a higher sensitivity than routine cytology, retaining a high level of speciifcity. The DIA criterion for malignancy is demonstration of aneuploidy. This technique increases the sensitivity to 40%, but the speciifcity remains low. Preliminary data from application to other tumors suggest that combination of FISH and DIA may be of further beneift. CONCLUSIONS: The new techniques offer a signiifcantly enhanced diagnostic efifcacy in the evaluation of ERCP brush specimens. Apart from contributing to a more timely diagnosis, their wider application to cholangiocarcinoma may also facilitate the genetic study of the disease and add to our understanding of oncogenesis at the molecular level, with the prospect of identifying targets for novel therapeutic interventions.

  18. Inflammatory cytokines suppress arylamine N-acetyltransferase 1 in cholangiocarcinoma cells

    Institute of Scientific and Technical Information of China (English)


    AIM: To evaluate the effect of inflammatory cytokines on arylamine N-acetyltransferase 1 (NAT1), which is a phase-Ⅱ enzyme involved in the biotransformation of aromatic and heterocyclic amines found in food, drugs and the environment.METHODS: Human cholangiocarcinoma KKU-100 cells were treated with a mixture of proinflammatory cytokines (interferon-y, interleukin-1β and tumor necrosis factor-α)for 48 h, and the effect on NAT1 activity was assessed by high performance liquid chromatography, while NAT1 expression was determined by reverse-transcription polymerase chain reaction. The oxidative stress on the cells was examined by the formation of nitric oxide,superoxide anion and glutathione (GSH) levels. The cells were also treated with S-nitroso-glutathione (GSNO), a nitric oxide donor, to see if the responses were similar to those obtained with the inflammatory cytokines.RESULTS: Cytokines suppressed NAT1 activity,reducing the Vmax without affecting the Km. Cytokines also had a significant impact on the induction of nitric oxide production and in reducing the redox ratios of glutathione (GSH) and GSH disulfide. Treatment with GSNO for 2-48 h reduced NAT1 activity without affecting the GSH ratio. Moreover, inflammatory cytokines and GSNO suppressed NAT1 mRNA expression.CONCLUSION: These findings indicate an association between inflammation and suppression of NAT1, which perhaps contributes to chemical-mediated toxicity and carcinogenesis.

  19. The E-cadherin repressor slug and progression of human extrahepatic hilar cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Wang Xin-sheng


    Full Text Available Abstract Objectives This study explored the expression and function of Slug in human extrahepatic hilar cholangiocarcinoma (EHC to identify its role in tumor progression. Methods The expression of Snail and Slug mRNA in 52 human tissue samples of EHC was investigated. The mRNA of Snail and Slug were quantified using reverse transcriptase-PCR, and correlations with E-cadherin expression and clinicopathological factors were investigated. We then investigated transfection of Slug cDNA in endogenous E-cadherin-positive human EHC FRH0201 cells, selectively induced the loss of E-cadherin protein expression, and then small interfering RNA (siRNA for inhibition of Slug expression in endogenous Slug-positive human EHC QBC939 cells, selectively induced the loss of Slug protein expression. A Boyden chamber transwell assay was used for invasion. Results Slug mRNA was overexpressed in 18 cases (34.6% of EHC compared with adjacent noncancerous tissue. E-Cadherin protein expression determined in the same 52 cases by immunohistochemistry was significantly down-regulated in those cases with Slug mRNA overexpression (P = 0.0001. The tumor and nontumor ratio of Slug mRNA was correlated with nodal metastasis(p = 0.0102, distant metastasis (p = 0.0001and Survival time(p = 0.0443. However, Snail mRNA correlated with neither E-cadherin expression nor tumor invasiveness. By inhibiting Slug expression by RNA interference, we found that reduced Slug levels upregulated E-cadherin and decreased invasion in QBC939 cell. When the QBC939 cells was infected with Slug cDNA,, significant E-cadherin was downregulated and increased invasion in QBC939 cell. Conclusions The results suggested that Slug expression plays an important role in both the regulation of E-cadherin expression and in the acquisition of invasive potential in human EHC. Slug is possibly a potential target for an antitumor therapy blocking the functions of invasion and metastasis in human EHCs.

  20. Integrin β6 serves as an immunohistochemical marker for lymph node metastasis and promotes cell invasiveness in cholangiocarcinoma (United States)

    Li, Zequn; Biswas, Siddhartha; Liang, Benjia; Zou, Xueqing; Shan, Liqun; Li, Yang; Fang, Ruliang; Niu, Jun


    Cholangiocarcinoma is a devastating malignancy that is notoriously difficult to diagnose and is associated with a high mortality. Despite extensive efforts to improve the diagnosis and treatment of this neoplasm, limited progress has been made. Integrin β6 is a subtype of integrin that is expressed exclusively on the surfaces of epithelial cells and is associated with a variety of tumors. In the present study, we investigated the expression and roles of integrin β6 in cholangiocarcinoma. β6 upregulation in cholangiocarcinoma was correlated with lymph node metastasis and distant metastasis. Moreover, integrin β6 was identified as a biomarker for the diagnosis of cholangiocarcinoma and an indicator of lymph node metastasis. Integrin β6 significantly promoted the proliferation, migration and invasion of cholangiocarcinoma cells. Furthermore, integrin β6 increased Rac1-GTPase, resulting in the upregulation of metalloproteinase-9 (MMP9) and F-actin polymerization. Taken together, our results indicate that integrin β6 promotes tumor invasiveness in a Rac1-dependent manner and is a potential biomarker for tumor metastasis. Integrin β6 may help to improve the diagnostic accuracy, and targeting β6 may be a novel strategy for the treatment of cholangiocarcinoma. PMID:27440504

  1. Expression of growth factor receptors and targeting of EGFR in cholangiocarcinoma cell lines

    Directory of Open Access Journals (Sweden)

    Kellermeier Silvia


    Full Text Available Abstract Background Cholangiocarcinoma (CC is a malignant neoplasm of the bile ducts or the gallbladder. Targeting of growth factor receptors showed therapeutic potential in palliative settings for many solid tumors. The aim of this study was to determine the expression of seven growth factor receptors in CC cell lines and to assess the effect of blocking the EGFR receptor in vitro. Methods Expression of EGFR (epithelial growth factor receptor, HGFR (hepatocyte growth factor receptor IGF1R (insulin-like growth factor 1 receptor, IGF2R (insulin-like growth factor 2 receptor and VEGFR1-3 (vascular endothelial growth factor receptor 1-3 were examined in four human CC cell lines (EGI-1, HuH28, OZ and TFK-1. The effect of the anti-EGFR-antibody cetuximab on cell growth and apoptosis was studied and cell lines were examined for KRAS mutations. Results EGFR, HGFR and IGFR1 were present in all four cell lines tested. IGFR2 expression was confirmed in EGI-1 and TFK-1. No growth-inhibitory effect was found in EGI-1 cells after incubation with cetuximab. Cetuximab dose-dependently inhibited growth in TFK-1. Increased apoptosis was only seen in TFK-1 cells at the highest cetuximab dose tested (1 mg/ml, with no dose-response-relationship at lower concentrations. In EGI-1 a heterozygous KRAS mutation was found in codon 12 (c.35G>A; p.G12D. HuH28, OZ and TFK-1 lacked KRAS mutation. Conclusion CC cell lines express a pattern of different growth receptors in vitro. Growth factor inhibitor treatment could be affected from the KRAS genotype in CC. The expression of EGFR itself does not allow prognoses on growth inhibition by cetuximab.

  2. Cyclophilin A enhances cell proliferation and tumor growth of liver fluke-associated cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Sawanyawisuth Kanlayanee


    Full Text Available Abstract Background Cyclophilin A (CypA expression is associated with malignant phenotypes in many cancers. However, the role and mechanisms of CypA in liver fluke-associated cholangiocarcinoma (CCA are not presently known. In this study, we investigated the expression of CypA in CCA tumor tissues and CCA cell lines as well as regulation mechanisms of CypA in tumor growth using CCA cell lines. Methods CypA expression was determined by real time RT-PCR, Western blot or immunohistochemistry. CypA silence or overexpression in CCA cells was achieved using gene delivery techniques. Cell proliferation was assessed using MTS assay or Ki-67 staining. The effect of silencing CypA on CCA tumor growth was determined in nude mice. The effect of CypA knockdown on ERK1/2 activation was assessed by Western blot. Results CypA was upregulated in 68% of CCA tumor tissues. Silencing CypA significantly suppressed cell proliferation in several CCA cell lines. Likewise, inhibition of CypA peptidyl-prolyl cis-trans isomerase (PPIase activity using cyclosporin A (CsA decreased cell proliferation. In contrast, overexpression of CypA resulted in 30% to 35% increases in proliferation of CCA cell lines. Interestingly, neither silence nor overexpression of CypA affected cell proliferation of a non-tumor human cholangiocyte cell line, MMNK1. Suppression of CypA expression attenuated ERK1/2 activity in CCA M139 cells by using both transient and stable knockdown methods. In the in vivo study, there was a 43% reduction in weight of tumors derived from CypA-silenced CCA cell lines compared with control vector CCA tumors in mice; these tumors with stable CypA silencing showed a reduced cell proliferation. Conclusions CypA is upregulated in majority of CCA patients' tissues and confers a significant growth advantage in CCA cells. Suppression of CypA expression decreases proliferation of CCA cell lines in vitro and reduces tumor growth in the nude mouse model. Inhibition of Cyp

  3. Omega-3 Polyunsaturated Fatty Acids Upregulate 15-PGDH Expression in Cholangiocarcinoma Cells by Inhibiting miR-26a/b Expression. (United States)

    Yao, Lu; Han, Chang; Song, Kyoungsub; Zhang, Jinqiang; Lim, Kyu; Wu, Tong


    Prostaglandin E2 (PGE2) is a proinflammatory lipid mediator that promotes cancer growth. The 15-hydroxyprostaglandin dehydrogenase (15-PGDH) catalyzes oxidation of the 15(S)-hydroxyl group of PGE2, leading to its inactivation. Therefore, 15-PGDH induction may offer a strategy to treat cancers that are driven by PGE2, such as human cholangiocarcinoma. Here, we report that omega-3 polyunsaturated fatty acids (ω-3 PUFA) upregulate 15-PGDH expression by inhibiting miR-26a and miR-26b, thereby contributing to ω-3 PUFA-induced inhibition of human cholangiocarcinoma cell growth. Treatment of human cholangiocarcinoma cells (CCLP1 and TFK-1) with ω-3 PUFA (DHA) or transfection of these cells with the Fat-1 gene (encoding Caenorhabditis elegans desaturase, which converts ω-6 PUFA to ω-3 PUFA) significantly increased 15-PGDH enzymes levels, but with little effect on the activity of the 15-PGDH gene promoter. Mechanistic investigations revealed that this increase in 15-PGDH levels in cells was mediated by a reduction in the expression of miR-26a and miR-26b, which target 15-PGDH mRNA and inhibit 15-PGDH translation. These findings were extended by the demonstration that overexpressing miR-26a or miR-26b decreased 15-PGDH protein levels, reversed ω-3 PUFA-induced accumulation of 15-PGDH protein, and prevented ω-3 PUFA-induced inhibition of cholangiocarcinoma cell growth. We further observed that ω-3 PUFA suppressed miR-26a and miR-26b by inhibiting c-myc, a transcription factor that regulates miR-26a/b. Accordingly, c-myc overexpression enhanced expression of miR-26a/b and ablated the ability of ω-3 PUFA to inhibit cell growth. Taken together, our results reveal a novel mechanism for ω-3 PUFA-induced expression of 15-PGDH in human cholangiocarcinoma and provide a preclinical rationale for the evaluation of ω-3 PUFA in treatment of this malignancy.

  4. Perihilar cholangiocarcinoma: Current therapy

    Institute of Scientific and Technical Information of China (English)

    Wei; Zhang; Lu-Nan; Yan


    Perihilar cholangiocarcinoma, which is a rare primary malignancy, originates from the epithelial cells of the bile duct. Usually invading the periductal tissues and the lymph nodes, perihilar cholangiocarcinoma is commonly diagnosed in the advanced stage of the disease and has a dismal prognosis. Currently, complete hepatectomy is the primary therapy for curing this disease. Perioperative assessment and available surgical procedures can be considered for achieving a negative margin resection, which is associated with long-term survival and better quality of life. For patients with unresectable cholangiocarcinoma, several palliative treatments have been demonstrated to produce a better outcome; and liver transplantation for selected patients with perihilar cholangiocarcinoma is promising and desirable. However, the role of palliative treatments and liver transplantation was controversial and requires more evidence and substantial validity from multiple institutions. In this article, we summarize the data from multiple institutions and discuss the resectability, mortality, morbidity and outcome with different approaches.

  5. Effect of mutated IκBα transfection on multidrug resistance in hilar cholangiocarcinoma cell lines

    Institute of Scientific and Technical Information of China (English)

    Ru-Fu Chen; Zhi-Hua Li; Xian-He Kong; Ji-Sheng Chen


    AIM: To explore the expression effect of mutated IκBαtransfection on multidrug resistance gene (MDR-1) in hilar cholangiocarcinoma cells by inhibiting the activity of nuclear transcription factor-κB (NF-κB).METHODS: We used the mutated IκBα plasmid to transfect QBC939HCVC+ cells and QBC939 cells, and electrophoretic gel mobility shift assay (EMSA) to detect the binding activity of NF-κB DNA and the effect of the transfrecting mutated IκBα plasmid on multidrug resistance gene (MDR-1) in hilar cholangiocarcinoma cells and its expression protein (P-GP).RFSULTS: Plasmid DNA was digested by restriction enzymes Xbal and Hand Ⅲ, and its product after electrophoresis showed two bands with a big difference in molecular weight,with a size of 4.9 kb and 1.55 kb respectively, which indicated that the carrier was successfully constructed and digested with enzymes. The radioactivity accumulation of QBC939HCVC+and QBC939 cells transfected with mutated IκBα plasmid was significantly lower than that of the control group not transfected with mutated IκBα plasmid. Double densimeter scanning showed that the relative signal density between the tansfection group and non-transfection group was significantly different, which proved that the mutated IκBα plasmid could inhibit the binding activity of NF-κB DNA in hilar cholangiocarcinoma cells. Compared to control group not transfected with m IκBα plasmid, the expression level of MDR-1mRNA in the QBC939 and QBC939HCVC+ cells transfected with mutated IκBα plasmid was lower. The expression intensity of P-GP protein in QBC939 and QBC939HCVC+ cells transfected with mutated IκBα was significantly lower than that of the control group not transfected with mutated IκBα plasmid.CONCLUSION: The mutated IκBα plasmid transfection can markedly reverse the multidrug resistance of hilar cholangiocarcinoma cells. Interruption of NF-κB activity may become a new target in gene therapy for hilar cholangiocarcinogenesic carcinoma.


    Institute of Scientific and Technical Information of China (English)

    Xiao-ran He; Xiao-peng Wu


    Objective To investigate and compare the biological characteristics and sensitivity to chemotherapy and radiother-apy of intrahepatic and extrahepatie cholangiocarcinoma cells in vitro.Methods The intrahepatic and extrahepatie eholangiocarcinoma cell lines were established, and cells with steady passage were chosen to study the biological characteristics including morphology, growth dynamics, chromosome, and levels of cancer antigen (CA)125, CA 19-9, alpha-fetoprotein (AFP), and carcino-embryonic antigen (CEA).M.eanwhile, MTT assay was used to determine the sensitivity of both kinds of cells to 6 chemotherapeutic drugs, inclu-ding cisplatin, paclitaxel, harringtonine, 5-fluorouracil, vincristine, and aelacimomycin, and the inhibitory rate of ceils under the irradiation of 10 Gy ray was also measured.Reanlts The intrahepatic cholangiocarcinoma cells were mostly fusiform in shape, and extrahepatic eholangiocar-cinoma cells were mostly round or polygon in shape. Their doubling time was 26. 3 hours and 23.1 hours, respectively.Their average number of chromosomes was 59 (range, 38-84) and 67 (range, 49-103 ), respectively. The chromo-some karyotypes of most intrahepatlc ebolangiocarcinoma cells were hyperdiploid and hypotriploid, while hypertriploid was predominant in extrahepatic cholangioearcinoma cells. The level of CA 125 in supernatant of extrahepatic cholangio-carcinoma cells increased obviously, while levels of other determined tumor markers in both kinds of cells were all with-in normal range. The intrahepatic cholangiocarcinoma cells were low sensitive to cisplatin and paclitaxel, but not sensi-tive to the other 4 chemotherapeutic drugs. The extrahepatic cholangiocarcinoma cells were high sensitive to eisplatin,but not sensitive to the other 5 drugs. Both kinds of cells had poor sensitivity to radiotherapy.Conelusions Intrahepatic and extrahepatie cbolangiocareinoma cells show differences in shape, doubling time,chromosome karyotype, tumor marker level, and

  7. Effect of SMYD3 over-expression on DNMT3B levels and proliferation ability in human cholangiocarcinoma cell line FRH0201%上调SMYD3对人胆管癌FRH0201细胞中DNMT3B表达及细胞增殖的影响

    Institute of Scientific and Technical Information of China (English)

    程帝; 李志花; 陈汝福; 郭宁; 廖巧芳; 郑礼平; 周泉波; 周嘉嘉


    目的:研究人胆管癌细胞株FRH0201中SET和MYND结构域含有蛋白3(SET and MYND domain-containing protein 3,SMYD3)过度表达对DNA甲基化转移酶3B(DNA methyltransferase 3B,DNMT3B)表达及细胞增殖能力的影响.方法:瞬时转染SMYD3真核表达质粒后,RT-PCR检测细胞中DNMT3B mRNA水平的变化;Western blotting检测细胞中DNMT3B蛋白水平的变化;CCK-8检测细胞增殖能力的改变;流式细胞术检测细胞周期的改变.结果:以未处理组为对照,胆管癌细胞FRH0201在转染pEGFP-C3-SMYD3质粒后,DNMT3B蛋白及mRNA表达均显著上升(P<0.01);细胞的增殖能力显著提高、细胞增殖速度加快(P<0.05);进入G2/M期的细胞明显增多(P<0.05).结论:过度表达SMYD3,可引起细胞中DNMT3B的表达上调并增强细胞增殖能力.%AIM:To explore ihe effect of SET and MYND domain - containing protein 3 ( SMYD3 ) over - expression on the expression of DNA methyltransferase 3B (DNMT3B) and the proliferation ability in human cholangiocarci-noma cell line FRH0201. METHODS; Transient transfection of SMYD3 eukaryotic expression plasmid pEGFP - C3 - SMYD3 into human cholangiocarcinoma cell line FRH0201 was performed. The expression of DNMT3B at mRNA and protein levels was detected by RT - PCR and Western blotting,respectively. Cell proliferation was examined by CCK -8 method and cell cycle situation was checked by flow cytometry. RESULTS; After transfected with SMYD3 eukaryotic expression plasmid pEGFP - C3 - SMYD3 , the over - expression of SMYD3 in FRH0201 cells was observed. Compared with the un-transfected cells, the expression of DNMT3B was significantly increased (P <0. 01) , the proliferation rate was obviously accelerated (P<0. 05) and the number of the cells in G2/M phase was significantly increased (P<0. 05) in FRH0201 cells transiently transfected with pEGFP - C3 - SMYD3 plasmid. CONCLUSION; The transient transfection of pEGFP -C3 - SMYD3 plasmid induces over - expression of DNMT3B and

  8. Tumour cell–derived extracellular vesicles interact with mesenchymal stem cells to modulate the microenvironment and enhance cholangiocarcinoma growth

    Directory of Open Access Journals (Sweden)

    Hiroaki Haga


    Full Text Available The contributions of mesenchymal stem cells (MSCs to tumour growth and stroma formation are poorly understood. Tumour cells can transfer genetic information and modulate cell signalling in other cells through the release of extracellular vesicles (EVs. We examined the contribution of EV-mediated inter-cellular signalling between bone marrow MSCs and tumour cells in human cholangiocarcinoma, highly desmoplastic cancers that are characterized by tumour cells closely intertwined within a dense fibrous stroma. Exposure of MSCs to tumour cell–derived EVs enhanced MSC migratory capability and expression of alpha-smooth muscle actin mRNA, in addition to mRNA expression and release of CXCL-1, CCL2 and IL-6. Conditioned media from MSCs exposed to tumour cell–derived EVs increased STAT-3 phosphorylation and proliferation in tumour cells. These effects were completely blocked by anti-IL-6R antibody. In conclusion, tumour cell–derived EVs can contribute to the generation of tumour stroma through fibroblastic differentiation of MSCs, and can also selectively modulate the cellular release of soluble factors such as IL-6 by MSCs that can, in turn, alter tumour cell proliferation. Thus, malignant cells can “educate” MSCs to induce local microenvironmental changes that enhance tumour cell growth.

  9. Fisetin Reduces Cell Viability Through Up-Regulation of Phosphorylation of ERK1/2 in Cholangiocarcinoma Cells. (United States)

    Kim, Nayoung; Lee, Sang Hyub; Son, Jun Hyuk; Lee, Jae Min; Kang, Min-Jung; Kim, Bo Hye; Lee, Jung-Su; Ryu, Ji Kon; Kim, Yong-Tae


    Cholangiocarcinoma (CCA) is a malignancy with poor prognosis and limited therapeutic options. Effective prevention and treatment of CCA require developing novel anticancer agents and improved therapeutic regimens. As natural products are concidered a rich source of potential anticancer agents, we investigated the anticancer effect of fisetin in combination with gemcitabine. Cytotoxic effect of fisetin and gemcitabine on a human CCA cell line SNU-308 was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and apoptosis assay using propidium iodine and annexin V. Molecular mechanisms of fisetin action in CCA were investigated by western blotting. Fisetin was found to inhibit survival of CCA cells, through strongly phosphorylating ERK. It also induced cellular apoptosis additively in combination with gemcitabine. Expression of cellular proliferative markers, such as phospho-p65 and myelocytomatosis (MYC), were reduced by fisetin. These results suggest fisetin in combination with gemcitabine as a candidate for use in improved anticancer regimens. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  10. Immune Responses of Dendritic Cells Loaded with Antigens from Apoptotic Cholangiocarcinoma Cells Caused by γ-Irradation

    Institute of Scientific and Technical Information of China (English)

    WUGang; HANBenli; PEIXuetao


    Objective:To investigate the induction cytotoxic T cells(CTLs) with antitumor activity and therapeutic efficacy after dendritic cells(DCs) acquired antigen from apoptotic cholangiocarcinoma cells caused by γ-irradiation. Methods:DCs from peripheral blood mononuclear cells (PBMC) that maintain the antigen capturing and processing capacity charateristic of immature cells have been established in vitro, using granulocyte macrophage colony stimulating factor (GM-CSF) and interleukin-4 (IL-4). Then, in cholangiocarcinoma cells apoptosis was induced by γ-irradiation. The experimental groups were as follows:(1)coculture of DCs and apoptotic cancer cells and T cells;(2)coculture of DCs and necrotic cancer cells and T cells;(3)coculture of DCs, cultured cancer cell and T cells. They are cocultured for 7 days.DCs and T cells were riched, isolated and their antitumor response was tested. Results:The cells had typical dendritic morphology, expressed high levels of CDla and B7, acquired antigen from apoptotic cells caused by γ-irradiation and induced an increased T cell stimulatory capacity in mixed lymphocyte reactions (MLR). Conclusion:DCs obtained from PBMCs using GM-CSF and IL-4 can efficiently present antigen derived from apoptotic cells caused by γ-irradiation and efficiently induce T cells.This strategy, therefore, may present an effective approach to transduce DCs with antigen.

  11. ZHX1 Promotes the Proliferation, Migration and Invasion of Cholangiocarcinoma Cells (United States)

    Kim, Ji-young; Liu, Liangwen; Kim, Yun-Hak; Jung, Jin-Sup; Oh, Sae-Ock


    Zinc-fingers and homeoboxes 1 (ZHX1) is a transcription repressor that has been associated with the progressions of hepatocellular carcinoma, gastric cancer, and breast cancer. However, the functional roles of ZHX1 in cholangiocarcinoma (CCA) have not been determined. We investigated the expression and roles of ZHX1 during the proliferation, migration, and invasion of CCA cells. In silico analysis and immunohistochemical studies showed amplification and overexpression of ZHX1 in CCA tissues. Furthermore, ZHX1 knockdown using specific siRNAs decreased CCA cell proliferation, migration, and invasion, whereas ZHX1 overexpression promoted all three characteristics. In addition, results suggested EGR1 might partially mediate the effect of ZHX1 on the proliferation of CCA cells. Taken together, these results show ZHX1 promotes CCA cell proliferation, migration, and invasion, and present ZHX1 as a potential target for the treatment of CCA. PMID:27835650

  12. Hepatitis C virus infection of cholangiocarcinoma cell lines

    NARCIS (Netherlands)

    Fletcher, Nicola F.; Humphreys, Elizabeth; Jennings, Elliott; Osburn, William; Lissauer, Samantha; Wilson, Garrick K.; van Ijzendoorn, Sven C. D.; Baumert, Thomas F.; Balfe, Peter; Afford, Simon; McKeating, Jane A.


    Hepatitis C virus (HCV) infects the liver and hepatocytes are the major cell type supporting viral replication. Hepatocytes and cholangiocytes derive from a common hepatic progenitor cell that proliferates during inflammatory conditions, raising the possibility that cholangiocytes may support HCV re

  13. The BH3-mimetic ABT-737 targets the apoptotic machinery in cholangiocarcinoma cell lines resulting in synergistic interactions with zoledronic acid


    ROMANI, ANTONELLO A.; Desenzani, Silvia; Morganti, Marina M.; Baroni, Maria Cristina; Borghetti, Angelo F.; Soliani, Paolo


    Abstract Purpose In TFK-1 and EGI-1 cholangiocarcinoma cell lines, zoledronic acid (ZOL) determines an S-phase block without apoptosis. Here, we investigated the occurrence of apoptosis stigmata when ZOL is associated to the BH3-mimetic ABT-737. Methods In EGI-1 and TFK-1 cholangiocarcinoma cell lines untreated or treated with ABT-737 alone or in combination with ZOL, the pro-survival protein?s pattern (BC...

  14. Development and characterization of a hydrogen peroxide-resistant cholangiocyte cell line: A novel model of oxidative stress-related cholangiocarcinoma genesis

    Energy Technology Data Exchange (ETDEWEB)

    Thanan, Raynoo [Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002 (Thailand); Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002 (Thailand); Techasen, Anchalee [Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002 (Thailand); Faculty of Associated Medical Science, Khon Kaen University, Khon Kaen 40002 (Thailand); Hou, Bo [Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507 (Japan); Jamnongkan, Wassana; Armartmuntree, Napat [Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002 (Thailand); Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002 (Thailand); Yongvanit, Puangrat, E-mail: [Department of Biochemistry, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002 (Thailand); Liver Fluke and Cholangiocarcinoma Research Center, Faculty of Medicine, Khon Kaen University, Khon Kaen 40002 (Thailand); Murata, Mariko, E-mail: [Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Tsu, Mie 514-8507 (Japan)


    Oxidative stress is a cause of inflammation–related diseases, including cancers. Cholangiocarcinoma is a liver cancer with bile duct epithelial cell phenotypes. Our previous studies in animal and human models indicated that oxidative stress is a major cause of cholangiocarcinoma development. Hydrogen peroxide (H{sub 2}O{sub 2}) can generate hydroxyl radicals, which damage lipids, proteins, and nucleic acids, leading to cell death. However, some cells can survive by adapting to oxidative stress conditions, and selective clonal expansion of these resistant cells would be involved in oxidative stress-related carcinogenesis. The present study aimed to establish H{sub 2}O{sub 2}-resistant cell line from an immortal cholangiocyte cell line (MMNK1) by chronic treatment with low-concentration H{sub 2}O{sub 2} (25 μM). After 72 days of induction, H{sub 2}O{sub 2}-resistant cell lines (ox-MMNK1-L) were obtained. The ox-MMNK1-L cell line showed H{sub 2}O{sub 2}-resistant properties, increasing the expression of the anti-oxidant genes catalase (CAT), superoxide dismutase-1 (SOD1), superoxide dismutase-2 (SOD2), and superoxide dismutase-3 (SOD3) and the enzyme activities of CAT and intracellular SODs. Furthermore, the resistant cells showed increased expression levels of an epigenetics-related gene, DNA methyltransferase-1 (DNMT1), when compared to the parental cells. Interestingly, the ox-MMNK1-L cell line had a significantly higher cell proliferation rate than the MMNK1 normal cell line. Moreover, ox-MMNK1-L cells showed pseudopodia formation and the loss of cell-to-cell adhesion (multi-layers) under additional oxidative stress (100 μM H{sub 2}O{sub 2}). These findings suggest that H{sub 2}O{sub 2}-resistant cells can be used as a model of oxidative stress-related cholangiocarcinoma genesis through molecular changes such as alteration of gene expression and epigenetic changes. - Highlights: • An H{sub 2}O{sub 2}-resistant ox-MMNK1-L cells was established from



    Wewer Albrechtsen; Da Zhang; A. Bashar Abdulkarim; Damjanov Ivan


    Sarcomatoid transformation is a rare but well documented change that may occur in intrahepatic cholangiocarcinomas. We report a tumor of this type to illustrate the difficulties in properly diagnosing cholangiocarcinoma when only the sarcomatous component is sampled by transcutaneous needle biopsy. The proper diagnosis of the tumor was suggested by finding the expression of cytokeratin 7 in the sarcomatous spindle cells. The surgically resected tumor comprised a spindle cell sarcomatous and a...

  16. Inhibitory Effects of Gallic Acid Isolated from Caesalpinia mimosoides Lamk on Cholangiocarcinoma Cell Lines and Foodborne Pathogenic Bacteria. (United States)

    Rattanata, Narintorn; Klaynongsruang, Sompong; Daduang, Sakda; Tavichakorntrakool, Ratree; Limpaiboon, Temduang; Lekphrom, Ratsami; Boonsiri, Patcharee; Daduang, Jureerut


    Gallic acid was isolated from Caesalpinia mimosoides Lamk and the structure s identified based on spectroscopic analysis and comparison with authentic compound. In this study we compared the ability of natural gallic acid (nGA) and commercial gallic acid (cGA) to inhibit the proliferation of cholangiocarcinoma cell lines (M213, M214) and foodborne pathogenic bacteria (Salmonella spp. and Plesiomonas shigelloides). Both nGA and cGA had the same inhibitory effects on cell proliferation by inducing apoptosis of cholangiocarcinoma cell lines. In addition, nGA inhibited growth of foodborne pathogenic bacteria in the same manner as cGA. Our results suggest that nGA from Caesalpinia mimosoides Lamk is a potential anticancer and antibacterial compound. However, in vivo studies are needed to elucidate the specific mechanisms involved.

  17. MiR-145 functions as a tumor suppressor targeting NUAK1 in human intrahepatic cholangiocarcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Xiong, Xinkui; Sun, Daoyi; Chai, Hao; Shan, Wengang [Liver Transplantation Center, First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu Province (China); Key Laboratory of Living Donor Liver Transplantation, Ministry of Public Health, Nanjing, Jiangsu Province (China); Yu, Yue [Key Laboratory of Living Donor Liver Transplantation, Ministry of Public Health, Nanjing, Jiangsu Province (China); Pu, Liyong [Liver Transplantation Center, First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu Province (China); Key Laboratory of Living Donor Liver Transplantation, Ministry of Public Health, Nanjing, Jiangsu Province (China); Cheng, Feng, E-mail: [Liver Transplantation Center, First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu Province (China); Key Laboratory of Living Donor Liver Transplantation, Ministry of Public Health, Nanjing, Jiangsu Province (China)


    The dysregulation of micro (mi)RNAs is associated with cancer development. The miRNA miR-145 is downregulated in intrahepatic cholangiocarcinoma (ICC); however, its precise role in tumor progression has not yet been elucidated. Novel (nua) kinase family (NUAK)1 functions as an oncogene in various cancers and is a putative target of miR-145 regulation. In this study, we investigated the regulation of NUAK1 by miR-145 in ICC. We found that miR-145 level was significantly decreased in ICC tissue and cell lines, which corresponded with an increase in NUAK1 expression. NUAK1 was found to be a direct target of miR-145 regulation. The overexpression of miR-145 in ICC cell lines inhibited proliferation, growth, and invasion by suppressing NUAK1 expression, which was associated with a decrease in Akt signaling and matrix metalloproteinase protein expression. Similar results were observed by inhibiting NUAK1 expression. These results demonstrate that miR-145 can prevent ICC progression by targeting NUAK1 and its downstream effectors, and can therefore be useful for clinical diagnosis and targeted therapy of ICC. - Highlights: • MiR-145 suppresses ICC proliferation and invasion abilities. • We demonstrated that miR-145 directly targets NUAK1 in ICC. • MiR-145 expression in ICC was associated with Akt signaling and MMPs expression.

  18. Hepatocarcinoma-Intestine-Pancreas/Pancreatic Associated Protein (HIP/PAP) Is Expressed and Secreted by Proliferating Ductules as well as by Hepatocarcinoma and Cholangiocarcinoma Cells (United States)

    Christa, Laurence; Simon, Marie-Thérèse; Brezault-Bonnet, Catherine; Bonte, Eric; Carnot, Françoise; Zylberberg, Hervé; Franco, Dominique; Capron, Frédérique; Roskams, Tania; Bréchot, Christian


    Hepatocarcinoma-intestine-pancreas/pancreatic associated protein (HIP/PAP) gene was identified because of its increased expression in 25% of human hepatocellular carcinoma. HIP/PAP protein, a C-type lectin, binds laminin, acts as an adhesion molecule for hepatocytes, and has also been described as an acute phase secretory protein during acute pancreatitis in humans and rats. We investigated HIP/PAP protein expression in patients with various liver diseases associated with ductular reaction. At the same time, we analyzed patients with hepatocellular carcinoma and cholangiocarcinoma, and tested HIP/PAP protein levels in sera to establish the pattern of secretion. Our data show that HIP/PAP expression was not restricted to hepatocellular carcinoma, but was also detected in cholangiocarcinoma cells as well as in reactive non-malignant bile ductules. In contrast, HIP/PAP protein expression was undetectable in normal mature hepatocytes, but some ductular cells localized at the interface of portal tracts with parenchyma were HIP/PAP immunoreactive in normal liver. Finally, we present evidence that HIP/PAP serum levels were increased in 21/28 (75%) patients with hepatocellular carcinoma, and in 25/51 (49%) patients with nonmalignant cirrhosis. Altogether, these results suggest that HIP/PAP protein may be implicated in hepatocytic and cholangiolar differentiation and proliferation. PMID:10550309

  19. Secreted cyclophilin A mediates G1/S phase transition of cholangiocarcinoma cells via CD147/ERK1/2 pathway. (United States)

    Obchoei, Sumalee; Sawanyawisuth, Kanlayanee; Wongkham, Chaisiri; Kasinrerk, Watchara; Yao, Qizhi; Chen, Changyi; Wongkham, Sopit


    Cyclophilin A (CypA) was shown to be upregulated in human cholangiocarcinoma (CCA) tissues. Suppression of intracellular CypA (inCypA) significantly reduces cell proliferation in vitro and tumor growth in nude mice. In the present study, the effect and potential mechanism of secreted CypA (sCypA) on cell proliferation of CCA cell lines were further investigated. CCA cells were treated with sCypA-containing conditioned media (CM) or with purified recombinant human CypA (rhCypA). Cell proliferation, cell cycle, ERK1/2, p38 MAPK, NF-κB, and STAT3 activities were examined by MTS assay, flow cytometry, and Western blot. sCypA was detected in CM from MMNK1 (an immortalized human cholangiocyte cell line) and six CCA cell lines. The sCypA levels corresponded to the inCypA levels indicating the intracellular origin of sCypA. Both sCypA-containing CM and rhCypA significantly increased proliferation of CCA cells. CD147 depletion by shRNA-knockdown or neutralizing with a CD147-monoclonal antibody significantly reduced sCypA-, and rhCypA-mediated cell proliferation. Upon rhCypA treatment, ERK1/2 was rapidly phosphorylated; whereas neutralizing CD147 inhibited ERK1/2 phosphorylation. Cell cycle analysis showed a significant increase in S phase and decrease in G1 population in rhCypA-treated cells. The expression levels of cyclin D1 and phosphorylated-retinoblastoma protein in the rhCypA-treated cells were increased compared with those in the non-treated control cells. p38 MAPK pathway was shown to be suppressed in siCypA-treated cells. In summary, CypA is secreted from CCA cells and enhances cell proliferation in an autocrine/paracrine manner, at least via direct binding with CD147, which may activate the ERK1/2 and p38 MAPK signaling pathways.

  20. The Prognostic Role of SOCS3 and A20 in Human Cholangiocarcinoma.

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    Yimin Wang

    Full Text Available As an antagonist of the JAK/STAT pathway, suppressor of cytokine signaling 3 (SOCS3 plays an integral role in shaping the inflammatory environment, tumorigenesis and disease progression in cholangiocarcinoma (CCA; however, its prognostic significance remains unclear. Although tumor necrosis factor α-induced protein 3 (TNFAIP3, also known as A20 can decrease SOCS3 expression and is involved in the regulation of tumorigenesis in certain malignancies, its role in CCA remains unknown. In this study, we investigated the expression of SOCS3 and A20 in human CCA tissues to assess the prognostic significance of these proteins. The expression of SOCS3 and A20 was initially detected by western blot in 22 cases of freshly frozen CCA tumors with corresponding peritumoral tissues and 22 control normal bile duct tissues. Then, these proteins were investigated in 86 CCA patients by immunohistochemistry (IHC and were evaluated for their association with clinicopathological parameters in human CCA. The results indicated that SOCS3 expression was significantly lower in CCA tumor tissues than in corresponding peritumoral biliary tissues and normal bile duct tissues. Conversely, A20 was overexpressed in CCA tissues. Thus, an inverse correlation between the expression of SOCS3 and A20 was discovered. Furthermore, patients with low SOCS3 expression or high A20 expression showed a dramatically lower overall survival rate. These proteins were both associated with CCA lymph node metastasis, postoperative recurrence and overall survival rate. However, only A20 showed a significant association with the tumor node metastasis (TNM stage, while SOCS3 showed a significant association with tumor differentiation. Multivariate Cox analysis revealed that SOCS3 and A20 were independent prognostic indicators for overall survival in CCA. Thus, our study demonstrated that SOCS3 and A20 represent novel prognostic factors for human CCA.

  1. Diagnostic approaches for cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)


    Cholangiocarcinomas arise from the epithelial cells of the bile ducts and are associated with poor prognosis. Despite new diagnostic approaches, the definite diagnosis of this malignancy continues to be challenging. Cholangiocarcinomas often grow longitudinally along the bile duct rather than in a radial direction. Thus, large tumor masses are frequently absent and imaging techniques, including ultrasound, CT, and MRI have only limited sensitivity. Tissue collection during endoscopic (ERCP) and/or percutaneous transhepatic (PTC) procedures are usually used to confirm a definitive diagnosis of cholangiocarcinoma. However, forceps biopsy and brush cytology provide positive results for malignancy in about only 50% of patients. Percutaneous and peroral cholangioscopy using fiber-optic techniques were therefore developed for direct visualization of the biliary tree, yielding additional information about endoscopic appearance and tumor extension, as well as a guided biopsy acquistion. Finally, endoscopic ultrasonography (EUS) complements endoscopic and percutaneous approaches and may provide a tissue diagnosis of tumors in the biliary region through fine- needle aspiration. In the future, new techniques allowing for early detection, including molecular markers, should be developed to improve the diagnostic sensitivity in this increasing tumor entity.

  2. Expression of c-kit receptor in human cholangiocarcinoma and in vivo treatment with imatinib mesilate in chimeric mice

    Institute of Scientific and Technical Information of China (English)

    Thomas Kamenz; Karel Caca; Thilo Blüthner; Andrea Tannapfel; Joachim M(o)ssner; Marcus Wiedmann


    AIM: To investigate the c-kit expression in biliary tract cancer cell lines and histological sections from patients with extrahepatic cholangiocarcinoma (CC) and to evaluate the efficacy of in vitro and in vitro treatment with imatinib mesilate.METHODS: The protein expression of c-kit in the human biliary tract cancer cell lines Mz-ChA-2 and EGI-1 and histological sections from 19 patients with extrahepatic CC was assessed by immunoblotting,immunocytochemistry, and immunohistochemistry. The anti-proliferative effect of imatinib mesilate on biliary tract cancer cell lines Mz-ChA-2 and EGI-1 was studied in vitro by automated cell counting. In addition, immunodeficient NMRI mice (TaconicTM) were subcutaneously injected with 5×106 cells of cell lines MzChA-2 and EGI-1. After having reached a tumour volume of 200 mm3, daily treatment was started intraperitoneally with imatinib mesilate at a dose of 50 mg/kg or normal saline (NS).Tumor volume was calculated with a Vernier caliper.After 14 d, mice were sacrificed with tumors excised and tumor mass determined.RESULTS: Immunoblotting revealed presence of c-kit in Mz-ChA-2 and absence in EGI-1 cells.Immunocytochemistry with c-kit antibodies displayed a cytoplasmatic and membraneous localization of receptor protein in Mz-ChA-2 cells and absence of c-kit in EGI-1 cells, c-kit was expressed in 7 of 19 (37%) extrahepatic humanCC tissue samples, 2 showed a moderate and 5 a rather weak immunostaining. Imatinib mesilate at a low concentration of 5 μmol/L caused a significant growth inhibition in the c-kit positive cell line Mz-ChA-2 (31%), but not in the c-kit negative cell line EGI-1 (0%) (P< 0.05). Imatinib mesilate at an intermediate concentration of 10 μmol/L inhibited cellular growth of both cell lines (51% vs 57%). Imatinib mesilate at a higher concentration of 20 μmol/L seemed to have a general toxic effect on both cell lines. The IC50 values were 9.7 μmol/L and 11 μmol/L, respectively. After 14 d of in vitro

  3. Sarcomatoid change in intrahepatic cholangiocarcinoma

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    Albrechtsen Wewer Nicolai


    Full Text Available Sarcomatoid transformation is a rare but well documented change that may occur in intrahepatic cholangiocarcinomas. We report a tumor of this type to illustrate the difficulties in properly diagnosing cholangiocarcinoma when only the sarcomatous component is sampled by transcutaneous needle biopsy. The proper diagnosis of the tumor was suggested by finding the expression of cytokeratin 7 in the sarcomatous spindle cells. The surgically resected tumor comprised a spindle cell sarcomatous and an adenocarcinomatous component accounting for approximately 30% and 70% of the total tumor mass respectively

  4. Investigation of thermo-sensitive amphiphilic micelles as drug carriers for chemotherapy in cholangiocarcinoma in vitro and in vivo. (United States)

    Wang, Xuefeng; Li, Songgang; Wan, Ziwei; Quan, Zhiwei; Tan, Qinggang


    Cholangiocarcinoma is an epithelial cancer of the bile ducts with poor prognosis and, in recent years, a rapidly increasing incidence. In this study, nano-sized thermo-sensitive micelles were investigated as drug carriers to improve chemotherapy in cholangiocarcinoma. Thermo-sensitive amphiphilic block copolymer, P-(N,N-isopropylacrylamide-co-N-hydroxymethylacrylamide)-b-caprolactone [P-(NIPAAm-co-NHMAAm)-b-PCL] with lower critical solution temperature (LCST) at about 38°C was synthesized. Doxorubicin (DOX)-loaded micelles were prepared by dialysis method. The micelles exhibited a sustained and temperature-dependent DOX release. Toxicity of the blank micelles for human cholangiocarcinoma (QBC939) cells was minimal both in vitro and in vivo. In contrast, the DOX-loaded micelles effectively inhibited proliferation and induced apoptosis of QBC939 cells in vitro (pthermo-sensitive amphiphilic micelles are a promising and effective drug carrier, and show potential for improving chemotherapy for cholangiocarcinoma.

  5. Genetics Home Reference: cholangiocarcinoma (United States)

    ... certain lifestyle factors, including smoking, alcohol use, and obesity, may also contribute to the risk of developing cholangiocarcinoma . Studies suggest that a combination of genetic, environmental, and lifestyle factors influence whether a person will develop cholangiocarcinoma . However, ...

  6. Establishment of an allo-transplantable hamster cholangiocarcinoma cell line and its application for in vivo screening of anti-cancer drugs. (United States)

    Puthdee, Nattapong; Vaeteewoottacharn, Kulthida; Seubwai, Wunchana; Wonkchalee, Orasa; Kaewkong, Worasak; Juasook, Amornrat; Pinlaor, Somchai; Pairojkul, Chawalit; Wongkham, Chaisiri; Okada, Seiji; Boonmars, Thidarut; Wongkham, Sopit


    Opisthorchis viverrini (O. viverrini) is a well-known causative agent of cholangiocarcinoma (CCA) in humans. CCA is very resistant to chemotherapy and is frequently fatal. To understand the pathogenesis of CCA in humans, a rodent model was developed. However, the development of CCA in rodents is time-consuming and the xenograft-transplantation model of human CCA in immunodeficient mice is costly. Therefore, the establishment of an in vivo screening model for O. viverrini-associated CCA treatment was of interest. We developed a hamster CCA cell line, Ham-1, derived from the CCA tissue of O. viverrini-infected and N-nitrosodimethylamine-treated Syrian golden hamsters. Ham-1 has been maintained in Dulbecco's Modified Essential Medium supplemented with 10% fetal bovine serum for more than 30 subcultures. These cells are mostly diploid (2n=44) with some being polyploid. Tumorigenic properties of Ham-1 were demonstrated by allograft transplantation in hamsters. The transplanted tissues were highly proliferative and exhibited a glandular-like structure retaining a bile duct marker, cytokeratin 19. The usefulness of this for in vivo model was demonstrated by berberine treatment, a traditional medicine that is active against various cancers. Growth inhibitory effects of berberine, mainly by an induction of G1 cell cycle arrest, were observed in vitro and in vivo. In summary, we developed the allo-transplantable hamster CCA cell line, which can be used for chemotherapeutic drug testing in vitro and in vivo.

  7. Mandibular metastasis of cholangiocarcinoma: A case report

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    You, Tae Min [Dept. of Advanced General Dentistry, Dankook University, Cheonan (Korea, Republic of); Kim, Kee Dong; Jeong, Ho Gui; Park, Won Se [Advanced General Dentistry, Dankook University, Cheonan (Korea, Republic of)


    Tumors metastasizing from distant regions to the oral and maxillofacial region are uncommon, comprising only 1%-2% of all malignancies. Cholangiocarcinoma is a malignancy that arises from cholangiocytes, which are epithelial cells that line the bile ducts. These cancers are difficult to diagnose and have a poor prognosis. In this paper, we report a rare case of mandibular metastasis of cholangiocarcinoma diagnosed at the primary site and discuss the radiographic findings observed in this case.

  8. Treatment of Leptomeningeal Carcinomatosis in a Patient With Metastatic Cholangiocarcinoma


    Jacobs, Ramon E.A.; McNeill, Katharine; Volpicelli, Frank M.; Warltier, Karin; Iturrate, Eduardo; Okamura, Charles; Adler, Nicole; Smith, Joshua; Sigmund, Alana; Mednick, Aron; Wertheimer, Benjamin; Hochman, Katherine


    A 49-year-old woman with cholangiocarcinoma metastatic to the lungs presented with new-onset unrelenting headaches. A lumbar puncture revealed malignant cells consistent with leptomeningeal metastasis from her cholangiocarcinoma. Magnetic resonance imaging (MRI) of the brain revealed leptomeningeal enhancement. An intrathecal (IT) catheter was placed and IT chemotherapy was initiated with methotrexate. Her case is notable for the rarity of cholangiocarcinoma spread to the leptomeninges, the u...

  9. Cholangiocarcinomas can originate from hepatocytes in mice. (United States)

    Fan, Biao; Malato, Yann; Calvisi, Diego F; Naqvi, Syed; Razumilava, Nataliya; Ribback, Silvia; Gores, Gregory J; Dombrowski, Frank; Evert, Matthias; Chen, Xin; Willenbring, Holger


    Intrahepatic cholangiocarcinomas (ICCs) are primary liver tumors with a poor prognosis. The development of effective therapies has been hampered by a limited understanding of the biology of ICCs. Although ICCs exhibit heterogeneity in location, histology, and marker expression, they are currently thought to derive invariably from the cells lining the bile ducts, biliary epithelial cells (BECs), or liver progenitor cells (LPCs). Despite lack of experimental evidence establishing BECs or LPCs as the origin of ICCs, other liver cell types have not been considered. Here we show that ICCs can originate from fully differentiated hepatocytes. Using a mouse model of hepatocyte fate tracing, we found that activated NOTCH and AKT signaling cooperate to convert normal hepatocytes into biliary cells that act as precursors of rapidly progressing, lethal ICCs. Our findings suggest a previously overlooked mechanism of human ICC formation that may be targetable for anti-ICC therapy.

  10. Genetic and epigenetic changes associated with cholangiocarcinoma: From DNA methylation to microRNAs

    Institute of Scientific and Technical Information of China (English)


    Cholangiocarcinomas are malignant epithelial liver tumors arising from the intra- and extra-hepatic bile ducts. Little is known about the molecular development of this disease, and very few effective treatment options are available. Thus, prognosis is poor. Genetic and epigenetic changes play an integral role in the neoplastic transformation of human cells to their malignant counterparts. This review summarizes some of the more prevalent genetic alterations (by microRNA expression)and epigenetic changes (hypermethylation of specific gene promoters) that are thought to contribute to the carcinogenic process in cholangiocarcinoma.

  11. Treatment of Leptomeningeal Carcinomatosis in a Patient With Metastatic Cholangiocarcinoma (United States)

    McNeill, Katharine; Volpicelli, Frank M.; Warltier, Karin; Iturrate, Eduardo; Okamura, Charles; Adler, Nicole; Smith, Joshua; Sigmund, Alana; Mednick, Aron; Wertheimer, Benjamin; Hochman, Katherine


    A 49-year-old woman with cholangiocarcinoma metastatic to the lungs presented with new-onset unrelenting headaches. A lumbar puncture revealed malignant cells consistent with leptomeningeal metastasis from her cholangiocarcinoma. Magnetic resonance imaging (MRI) of the brain revealed leptomeningeal enhancement. An intrathecal (IT) catheter was placed and IT chemotherapy was initiated with methotrexate. Her case is notable for the rarity of cholangiocarcinoma spread to the leptomeninges, the use of IT chemotherapy with cytologic and potentially symptomatic response, and a possible survival benefit in comparison to previously reported cases of leptomeningeal carcinomatosis secondary to cholangiocarcinoma. PMID:26157901

  12. Targeting the IL-6 dependent phenotype can identify novel therapies for cholangiocarcinoma.

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    Chiara Braconi

    Full Text Available BACKGROUND: The need for new therapies for cholangiocarcinoma is highlighted by their poor prognosis and refractoriness to chemotherapy. Increased production of Interleukin-6 promotes cholangiocarcinoma growth and contributes to chemoresistance by activating cell survival mechanisms. We sought to identify biologically active compounds capable of ameliorating the phenotypic effects of IL-6 expression and to explore their potential therapeutic use for cholangiocarcinoma. METHODOLOGY: A genomic signature associated with Interleukin-6 expression in Mz-ChA-1 human malignant cholangiocytes was derived. Computational bioinformatics analysis was performed to identify compounds that induced inverse gene changes to the signature. The effect of these compounds on cholangiocarcinoma growth was then experimentally verified in vitro and in vivo. Interactions with other therapeutic agents were evaluated using median effects analysis. PRINCIPAL FINDINGS: A group of structurally related compounds, nitrendipine, nifedipine and felodipine was identified. All three compounds were cytotoxic to Mz-ChA-1 cells with an IC50 for felodipine of 26 µM, nitrendipine, 44 µM and nifedipine, 15 µM. Similar results were observed in KMCH-1, CC-LP-1 and TFK-1 cholangiocarcinoma cell lines. At a fractional effect of 0.5, all three agents were synergistic with either camptothecin or gemcitabine in Mz-ChA-1 cells in vitro. Co-administration of felodipine and gemcitabine decreased the growth of Mz-ChA-1 cell xenografts in nude athymic mice. CONCLUSIONS: Computational bioinformatics analysis of phenotype-based genomic expression can be used to identify therapeutic agents. Using this drug discovery approach based on targeting a defined tumor associated phenotype, we identified compounds with the potential for therapeutic use in cholangiocarcinoma.

  13. Quinoline-based clioquinol and nitroxoline exhibit anticancer activity inducing FoxM1 inhibition in cholangiocarcinoma cells

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    Chan-on W


    Full Text Available Waraporn Chan-on,1 Nguyen Thi Bich Huyen,2 Napat Songtawee,3 Wilasinee Suwanjang,1 Supaluk Prachayasittikul,3 Virapong Prachayasittikul2 1Center for Research and Innovation, 2Department of Clinical Microbiology and Applied Technology, 3Center of Data Mining and Biomedical Informatics, Faculty of Medical Technology, Mahidol University, Bangkok, Thailand Purpose: Fork head box M1 (FoxM1 is an oncogenic transcription factor frequently elevated in numerous cancers, including cholangiocarcinoma (CCA. A growing body of evidence documents its diverse functions contributing to tumorigenesis and cancer progression. As such, discovery of agents that can target FoxM1 would be valuable for the treatment of CCA. The quinoline-based compounds, namely clioquinol (CQ and nitroxoline (NQ, represent a new class of anticancer drug. However, their efficacy and underlying mechanisms have not been elucidated in CCA. In this study, anticancer activities and inhibitory effects of CQ and NQ on FoxM1 signaling were explored using CCA cells.Methods: The effects of CQ and NQ on cell viability and proliferation were evaluated using the colorimetric 3-(4,5-dimethylthiazol-2yl-5-(3-carboxymethoxyphenyl-(4-sulfophenyl-2H-tetrazolium (MTS assay. Colony formation and cell migration affected by CQ and NQ were investigated using a clonogenic and a wound healing assay, respectively. To demonstrate the agents’ effects on FoxM1 signaling, expression levels of the target genes were quantitatively determined using real-time polymerase chain reaction.Results: CQ and NQ significantly inhibited cell survival of HuCCT1 and Huh28 in a dose- and a time-dependent fashion. Further investigations using the rapidly proliferating HuCCT1 cells revealed significant suppression of cell proliferation and colony formation induced by low doses of the compounds. Treatment of CQ and NQ repressed expression of cyclin D1 but enhanced expression of p21. Most importantly, upon CQ and NQ treatment

  14. Cholangiocarcinomas: New Insights from the Discovery of Stem Cell Niches in Peribiliary Glands of the Biliary Tree

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    Vincenzo Cardinale


    Full Text Available Peribiliary glands (PBGs are located in the large intrahepatic and extrahepatic bile ducts. Although they were described many years ago, their functions have been elucidated only in the last couple of years when our group demonstrated that PBGs are niches of multipotent stem/progenitor cells of endodermal origin. These cells express genes of multipotency and can be rapidly differentiated in vitro into hepatocytes, cholangiocytes, and endocrine pancreatic cells. PBGs share common features, in terms of stem/progenitor cell niches, with pancreatic duct glands and colon crypts, glandular structures representing in the adult life the endodermal remnants of fetal life. PBG stem/progenitor cells participate in the renewal of surface biliary epithelium and are active players in chronic pathologies of the biliary tree as well as in cholangiocarcinomas (CCA. Specifically, a large amount of recent evidence indicates that the pure mucin-CCA originates from PBGs; this could explain the similarities with pancreatic ductal adenocarcinoma and colorectal cancer, which also originate from transformed gland cells. In this paper, we summarized our recent findings concerning structure and functions of PBGs with the implications for liver pathophysiology and, specifically, for cancers of the biliary tree.

  15. Taurolithocholic acid promotes intrahepatic cholangiocarcinoma cell growth via muscarinic acetylcholine receptor and EGFR/ERK1/2 signaling pathway. (United States)

    Amonyingcharoen, Sumet; Suriyo, Tawit; Thiantanawat, Apinya; Watcharasit, Piyajit; Satayavivad, Jutamaad


    Cholangiocarcinoma (CCA) is a malignant cancer of the biliary tract and its occurrence is associated with chronic cholestasis which causes an elevation of bile acids in the liver and bile duct. The present study aimed to investigate the role and mechanistic effect of bile acids on the CCA cell growth. Intrahepatic CCA cell lines, RMCCA-1 and HuCCA-1, were treated with bile acids and their metabolites to determine the growth promoting effect. Cell viability, cell cycle analysis, EdU incorporation assays were conducted. Intracellular signaling proteins were detected by western immunoblotting. Among eleven forms of bile acids and their metabolites, only taurolithocholic acid (TLCA) concentration dependently (1-40 µM) increased the cell viability of RMCCA-1, but not HuCCA-1 cells. The cell cycle analysis showed induction of cells in the S phase and the EdU incorporation assay revealed induction of DNA synthesis in the TLCA-treated RMCCA-1 cells. Moreover, TLCA increased the phosphorylation of EGFR, ERK 1/2 and also increased the expression of cyclin D1 in RMCCA-1 cells. Furthermore, TLCA-induced RMCCA-1 cell growth could be inhibited by atropine, a non-selective muscarinic acetylcholine receptor (mAChR) antagonist, AG 1478, a specific EGFR inhibitor, or U 0126, a specific MEK 1/2 inhibitor. These results suggest that TLCA induces CCA cell growth via mAChR and EGFR/EKR1/2 signaling pathway. Moreover, the functional presence of cholinergic system plays a certain role in TLCA-induced CCA cell growth.

  16. Silencing of miR-370 in human cholangiocarcinoma by allelic loss and interleukin-6 induced maternal to paternal epigenotype switch.

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    Fangmei An

    Full Text Available Cholangiocarcinoma (CCA is a highly lethal malignant tumor arising from the biliary tract epithelium. Interleukin-6 (IL-6 is a major mediator of inflammation and contributor to carcinogenesis within the biliary tree. Previous studies suggested that enforced IL-6 contributes to cholangiocarcinogenesis through hypermethylation of several genes implicated in CCA. However, the precise mechanisms of IL-6 effects in CCA remain unclear. We now demonstrate that microRNA (miR-370 is underexpressed in a large cohort of human CCA vs. normal liver tissues. In addition, we show that IL-6 induces a time-dependent silencing of miR-370. In addition, demethylation of CCA cells results in upregulation of miR-370. Furthermore, we demonstrate that miR-370 is imprinted, and that the Intergenic Differentially Methylated Region (IG-DMR responsible for imprinting regulation of this genomic locus is hypermethylated in response to IL-6 treatment. In addition, the IG-DMR is hypermethylated in human CCA specimens compared to normal matched controls, in the same location as the IL-6 induced hypermethylation. Finally, miR-370 was found to regulate WNT10B in luciferase as well as western blotting experiments. Our data indicate that the paternal allele of miR-370 is normally silenced through genomic imprinting and that the overexpression of IL-6 in CCA effectively suppresses the expression of miR-370 from the maternal allele, lending support to the theory that miR-370 silencing in human CCA follows a classic two-hit mechanism.

  17. Hepatic progenitor cells in human liver tumor development

    Institute of Scientific and Technical Information of China (English)

    Louis Libbrecht


    In recent years, the results of several studies suggest that human liver tumors can be derived from hepatic progenitor cells rather than from mature cell types.The available data indeed strongly suggest that most combined hepatocellular-cholangiocarcinomas arise from hepatic progenitor cells that retained their potential to differentiate into the hepatocytic and biliary lineages.Hepatic progenitor cells could also be the basis for some hepatocellular carcinomas and hepatocellular adenomas, although it is very difficult to determine the origin of an individual hepatocellular carcinoma. There is currently not enough data to make statements regarding a hepatic progenitor cell origin of cholangiocarcinoma.The presence of hepatic progenitor cell markers and the presence and extent of the cholangiocellular component are factors that are related to the prognosis of hepatocellular carcinomas and combined hepatocellularcholangiocarcinomas, respectively.

  18. Fibroblast growth factor receptor 2 translocations in intrahepatic cholangiocarcinoma. (United States)

    Graham, Rondell P; Barr Fritcher, Emily G; Pestova, Ekaterina; Schulz, John; Sitailo, Leonid A; Vasmatzis, George; Murphy, Stephen J; McWilliams, Robert R; Hart, Steven N; Halling, Kevin C; Roberts, Lewis R; Gores, Gregory J; Couch, Fergus J; Zhang, Lizhi; Borad, Mitesh J; Kipp, Benjamin R


    Patients with cholangiocarcinoma often present with locally advanced or metastatic disease. There is a need for effective therapeutic strategies for advanced stage cholangiocarcinoma. Recently, FGFR2 translocations have been identified as a potential target for tyrosine kinase inhibitor therapies. This study evaluated 152 cholangiocarcinomas and 4 intraductal papillary biliary neoplasms of the bile duct for presence of FGFR2 translocations by fluorescence in situ hybridization and characterized the clinicopathologic features of cases with FGFR2 translocations. Thirteen (10 women, 3 men; 8%) of 156 biliary tumors harbored FGFR2 translocations, including 12 intrahepatic cholangiocarcinomas (12/96; 13%) and 1 intraductal papillary neoplasm of the bile duct. Histologically, cholangiocarcinomas with FGFR2 translocations displayed prominent intraductal growth (62%) or anastomosing tubular glands with desmoplasia (38%). Immunohistochemically, the tumors with FGFR2 translocations frequently showed weak and patchy expression of CK19 (77%). Markers of the stem cell phenotype in cholangiocarcinoma, HepPar1 and CK20, were negative in all cases. The median cancer-specific survival for patients whose tumors harbored FGFR2 translocations was 123 months compared to 37 months for cases without FGFR2 translocations (P = .039). This study also assessed 100 cholangiocarcinomas for ERBB2 amplification and ROS1 translocations. Of the cases tested, 3% and 1% were positive for ERBB2 amplification and ROS1 translocation, respectively. These results confirm that FGFR2, ERRB2, and ROS1 alterations are potential therapeutic targets for intrahepatic cholangiocarcinoma.

  19. Targeting the ∆133p53 isoform can restore chemosensitivity in 5-fluorouracil-resistant cholangiocarcinoma cells. (United States)

    Nutthasirikul, Nichapavee; Hahnvajanawong, Chariya; Techasen, Anchalee; Limpaiboon, Temduang; Leelayuwat, Chanvit; Chau-In, Siri; Jearanaikoon, Patcharee


    Lack of the normal p53 transactivation domain, ∆133p53 isoform exhibits anti-p53 function. Many studies report the correlation between ∆133p53 expression and poor survival in various cancers, including cholangiocarcinoma (CCA), which is a cancer of the bile ducts. CCA almost always results in short survival times. The relevance of ∆133p53 to drug resistance in CCA is not yet well understood. This study aimed to demonstrate the association between ∆133p53 and 5-fluorouracil (5-FU) resistance in CCA. ∆133p53 protein was highly expressed in CCA patients with poor outcome compared to favorable outcome but was not statistically significant. However, a significant correlation was found between normalized ∆133p53 levels and 5-FU resistance which was defined by an ex vivo histoculture drug response assay (P=0.019). Two stable 5-FU-resistant CCA cell lines, KKU-M139R (IC50 38.8 µM) and KKU-M214R (IC50 39.5 µM), were used as a model to evaluate the role of ∆133p53. Increased ∆133p53 was correlated with 5-FU in a dose-dependent manner. The transient knockdown of ∆133p53 expression can restore drug sensitivity in both resistant CCA cells with 11- to 45-fold reduction of IC50 compared to control. Upon ∆133p53 silencing, apoptotic signaling was enhanced by the upregulation of Bax and downregulation of Bcl-2. Additionally, p21 and p27 were upregulated, resulting in cell cycle arrest at G2. Inhibition of colony formation and prolong doubling time were also observed. Our findings demonstrated that chemosensitivity can be modulated via targeting of ∆133p53 suggesting the potential use of ∆133p53 as a candidate for targeting therapy in CCA.

  20. Pathology of intrahepatic cholangiocarcinoma (United States)

    Vijgen, Sandrine; Terris, Benoit


    Intrahepatic cholangiocarcinoma (iCC) is a primary carcinoma of the liver with increasing significance and major pathogenic, clinical and therapeutic challenges. Classically, it arises from malignant transformation of cholangiocytes bordering small portal bile duct (BD) to second-order segmental large BDs. It has three major macroscopic growth pattern [mass-forming (MF), periductal infiltrative (PI), and intraductal growth (IG)] and histologically is a desmoplastic stroma-rich adenocarcinoma with cholangiocyte differentiation. Recent data pointed out noteworthy degree of heterogeneity in regards of their epidemiology and risk factors, pathological and molecular features, pathogenesis, clinical behaviors and treatment. Notably, several histological variants are described and can coexist within the same tumor. Several different cells of origin have also been depicted in a fraction of iCCs, amongst which malignant transformation of ductules, of hepatic stem/progenitor cells, of periductal glands or through oncogenic reprogramming of adult hepatocytes. A degree of pathological overlap with hepatocellular carcinoma (HCC) may be observed in a portion of iCC. A series of precursor lesions are today characterized and emphasize the existence of a multistep carcinogenesis process. Overall, these new data have brought up in proposal of new histological or molecular classifications, which could soon replace current anatomic-based classification and could have major impact on establishment of prognosis and on development of novel target treatment approaches. PMID:28261592

  1. High expression of ErbB2 contributes to cholangiocarcinoma cell invasion and proliferation through AKT/p70S6K

    Institute of Scientific and Technical Information of China (English)

    Warapen; Treekitkarnmongkol; Tuangporn; Suthiphongchai


    AIM:To compare the impact of ErbB2 on cell invasion and proliferation in cholangiocarcinoma(CCA) cell lines.METHODS:Level of endogenous ErbB2 expression in three CCA cell lines,namely HuCCA-1,KKU-100 and KKU-M213,was determined by real-time reversetranscriptase polymerase chain reaction.Two ErbB2 inhibitory methods,a small molecule ErbB2 kinase inhibitor(AG825) and siRNA,were used to disrupt ErbB2 function in the cell lines.CCA cell invasion,motility and proliferation under ErbB2-disrupted conditions were d...

  2. Characterization of a novel rat cholangiocarcinoma cell culture model-CGCCA

    Institute of Scientific and Technical Information of China (English)

    Chun-Nan Yeh; Kun-Ju Lin; Tsung-Wen Chen; Ren-Ching Wu; Lee-Cheng Tsao; Ying-Tzu Chen; Wen-Hui Weng; Miin-Fu Chen


    AIM: To characterize a culture model of rat CCA cells, which were derived from a transplantable TTA-induced CCA and designated as Chang Gung CCA (CGCCA). METHODS: The CGCCA cells were cultured at in vitro passage 12 times on a culture dish in DMEM medium. To measure the doubling time, 103 cells were plated in a 96-well plate containing the growth medium. The cells were harvested 4 to 10 d after seeding, and a standard MTT assay was used to measure the growth. The phenotype of CACCA cell and xenograft was determined by immunohistochemical study. We also determine the chromosomal alterations of CGCCA, G-banding and spectral karyotyping studies were performed. The CGCCA cell line was transplanted into the nude mice for examining its tumorigenicity. 2-Deoxy-2-(18F)fluoro-Dglucose (FDG) autoradiography was also performed to evaluate the FDG uptake of the tumor xenograft. RESULTS: The doubling time for the CGCCA cell line was 32 h. After transplantation into nude mice, FDG autoradiography showed that the tumors formed at the cell transplantation site had a latency period of 4-6 wk with high FDG uptake excluding necrosis tissue. Moreover, immunohistochemical staining revealed prominent cytoplasmic expression of c-erb-B2, CK19, c-Met, COX-Ⅱ, EGFR, MUC4, and a negative expression of K-ras. All data confirmed the phenotypic features of the CGCCA cell line coincide with the xenograft mice tumors, indicating cells containing the tumorigenicity of CCA originated from CCA. In addition, karyotypic banding analysis showed that the diploid (2n) cell status combines with ring and giant rod marker chromosomes in these clones; either both types simultaneously appeared or only one type of marker chromosome in a pair appeared in a cell. The major materials contained in the marker chromosome were primarily identified from chromosome 4. CONCLUSION: The current CGCCA cell line may be used as a non-K-ras effect CCA model and to obtain information and reveal novel pathways for CCA. Further

  3. Characterization of a novel rat cholangiocarcinoma cell culture model-CGCCA. (United States)

    Yeh, Chun-Nan; Lin, Kun-Ju; Chen, Tsung-Wen; Wu, Ren-Ching; Tsao, Lee-Cheng; Chen, Ying-Tzu; Weng, Wen-Hui; Chen, Miin-Fu


    To characterize a culture model of rat CCA cells, which were derived from a transplantable TTA-induced CCA and designated as Chang Gung CCA (CGCCA). The CGCCA cells were cultured at in vitro passage 12 times on a culture dish in DMEM medium. To measure the doubling time, 10(3) cells were plated in a 96-well plate containing the growth medium. The cells were harvested 4 to 10 d after seeding, and a standard MTT assay was used to measure the growth. The phenotype of CACCA cell and xenograft was determined by immunohistochemical study. We also determine the chromosomal alterations of CGCCA, G-banding and spectral karyotyping studies were performed. The CGCCA cell line was transplanted into the nude mice for examining its tumorigenicity. 2-Deoxy-2-((18)F)fluoro-D-glucose (FDG) autoradiography was also performed to evaluate the FDG uptake of the tumor xenograft. The doubling time for the CGCCA cell line was 32 h. After transplantation into nude mice, FDG autoradiography showed that the tumors formed at the cell transplantation site had a latency period of 4-6 wk with high FDG uptake excluding necrosis tissue. Moreover, immunohistochemical staining revealed prominent cytoplasmic expression of c-erb-B2, CK19, c-Met, COX-II, EGFR, MUC4, and a negative expression of K-ras. All data confirmed the phenotypic features of the CGCCA cell line coincide with the xenograft mice tumors, indicating cells containing the tumorigenicity of CCA originated from CCA. In addition, karyotypic banding analysis showed that the diploid (2n) cell status combines with ring and giant rod marker chromosomes in these clones; either both types simultaneously appeared or only one type of marker chromosome in a pair appeared in a cell. The major materials contained in the marker chromosome were primarily identified from chromosome 4. The current CGCCA cell line may be used as a non-K-ras effect CCA model and to obtain information and reveal novel pathways for CCA. Further applications regarding tumor

  4. Endoscopic tissue diagnosis of cholangiocarcinoma.

    LENUS (Irish Health Repository)

    Harewood, Gavin C


    The extremely poor outcome in patients with cholangiocarcinoma, in large part, reflects the late presentation of these tumors and the challenging nature of establishing a tissue diagnosis. Establishing a diagnosis of cholangiocarcinoma requires obtaining evidence of malignancy from sampling of the epithelium of the biliary tract, which has proven to be challenging. Although endoscopic ultrasound-guided fine needle aspiration performs slightly better than endoscopic retrograde cholangiopancreatography in diagnosing cholangiocarcinoma, both endoscopic approaches demonstrate disappointing performance characteristics.

  5. Gold Nanoparticles Enhance the Anticancer Activity of Gallic Acid against Cholangiocarcinoma Cell Lines. (United States)

    Rattanata, Narintorn; Daduang, Sakda; Wongwattanakul, Molin; Leelayuwat, Chanvit; Limpaiboon, Temduang; Lekphrom, Ratsami; Sandee, Alisa; Boonsiri, Patcharee; Chio-Srichan, Sirinart; Daduang, Jureerut


    Gold nanoparticles (GNPs) were conjugated with gallic acid (GA) at various concentrations between 30 and 150 μM and characterized using transmission electron microscopy (TEM) and UV-Vis spectroscopy (UV-VIS). The anticancer activities of the gallic acid-stabilized gold nanoparticles against well-differentiated (M213) and moderately differentiated (M214) adenocarcinomas were then determined using a neutral red assay. The GA mechanism of action was evaluated using Fourier transform infrared (FTIR) microspectroscopy. Distinctive features of the FTIR spectra between the control and GA-treated cells were confirmed by principal component analysis (PCA). The surface plasmon resonance spectra of the GNPs had a maximum absorption at 520 nm, whereas GNPs-GA shifted the maximum absorption values. In an in vitro study, the complexed GNPs-GA had an increased ability to inhibit the proliferation of cancer cells that was statistically significant (Pacids, which may enhance the efficacy of this anticancer activity regarding apoptosis pathways.

  6. Interleukin-2 gene-encoded stromal cells inhibit the growth of metastatic cholangiocarcinomas

    Institute of Scientific and Technical Information of China (English)

    Myung-Hwan Kim; Sang Soo Lee; Sung Koo Lee; Seung-Gyu Lee; Chul-Won Suh; Gyung-Yub Gong; Jung-Sun Park; Young-Hoon Kim; Sang-Hee Kim


    AIM: To demonstrate bone marrow stromal cells (BMSCs) can be used as an attractive target for genetic modification in the treatment of malignant diseases.METHODS: Using a hamster model of biliary cancer,we investigated the therapeutic effects of interleukin-2 (IL-2) gene-modified BMSCs. Syrian golden hamsters were injected via the femoral vein with 5x10s cells of the KIGB-5 biliary cancer cell line (n=20). One week later,the hamsters were injected intraperitoneally with BMSCs containing Ad/hIL-2 and Ad/△E1, unmodified BMSCs, or RPMI only (control) and observed for 12 wk (n=5/each group).RESULTS: All hamsters treated with BMSCs containing Ad/hIL-2 survived with no evidence of the disease during this period. In contrast, hamsters in the other three groups showed disseminated metastases involving the lungs as early as 4 wk.CONCLUSION: Ad/IL-2 therapy is effective in the treatment of biliary cancer.

  7. Intrahepatic cholangiocarcinoma: current perspectives

    Directory of Open Access Journals (Sweden)

    Buettner S


    Full Text Available Stefan Buettner, Jeroen LA van Vugt, Jan NM IJzermans, Bas Groot Koerkamp Department of Surgery, Erasmus MC University Medical Center, Rotterdam, the Netherlands Abstract: Intrahepatic cholangiocarcinoma (ICC is the second most common malignancy arising from the liver. ICC makes up about 10% of all cholangiocarcinomas. It arises from the peripheral bile ducts within the liver parenchyma, proximal to the secondary biliary radicals. Histologically, the majority of ICCs are adenocarcinomas. Only a minority of patients (15% present with resectable disease, with a median survival of less than 3 years. Multidisciplinary management of ICC is complicated by large differences in disease course for individual patients both across and within tumor stages. Risk models and nomograms have been developed to more accurately predict survival of individual patients based on clinical parameters. Predictive risk factors are necessary to improve patient selection for systemic treatments. Molecular differences between tumors, such as in the epidermal growth factor receptor status, are promising, but their clinical applicability should be validated. For patients with locally advanced disease, several treatment strategies are being evaluated. Both hepatic arterial infusion chemotherapy with floxuridine and yttrium-90 embolization aim to downstage locally advanced ICC. Selected patients have resectable disease after downstaging, and other patients might benefit because of postponing widespread dissemination and biliary obstruction. Keywords: intrahepatic cholangiocarcinoma, diagnosis, treatment, developments 

  8. Sarcomatoid Change in Intrahepatic Cholangiocarcinoma

    DEFF Research Database (Denmark)

    Albrechtsen, Nicolai Jacob Wewer; Zhang, Da; Abdulkarim, Bashar;


    Sarcomatoid transformation is a rare but well documented change that may occur in intrahepatic cholangiocarcinomas.We report a tumor of this type to illustrate the difficulties in properly diagnosing cholangiocarcinoma when only the sarcomatous component is sampled by transcutaneous needle biopsy...

  9. Decrease of miR-622 expression promoted the proliferation, migration and invasion of cholangiocarcinoma cells by targeting regulation of c-Myc. (United States)

    Wu, Yi-Fei; Li, Zhuo-Ri; Cheng, Zhi-Qi; Yin, Xin-Min; Wu, Jin-Shu


    To explore the mechanism of miR-622 in regulating the proliferation, migration and invasion of cholangiocarcinoma (CCA) cells. Quantitative real-time PCR was conducted to measure the expression of miR-622 and c-Myc in CCA tissues and cell lines. Protein level of c-Myc was measured by Western blot. The effect of miR-622 on cell proliferation, migration and invasion was analyzed by MTT assay and Transwell chamber migration assay. Luciferase reporter assay was performed to measure the effect of miR-622 on c-Myc. miR-622 expression was downregulated in both CCA tissues and cell lines, while c-Myc expression was uregulated. Overexpression of miR-622 in CCA cells was statistically correlated with a decrease of cell proliferation, migration and invasion, while inhibition of miR-622 made an inverse result. We also proved c-Myc was identified as a target gene of miR-622 in CCA. Moreover, we found overexpression of c-Myc can strengthen the effects of miR-622 on the proliferation, migration and invasion of CCA cells. Decrease of miR-622 promotes the proliferation, migration and invasion of CCA cells by directly targeting c-Myc. Copyright © 2017. Published by Elsevier Masson SAS.

  10. 表观遗传学药物对肝内胆管癌的作用%Effects of epigenetic drugs in intrahepatic cholangiocarcinoma cells

    Institute of Scientific and Technical Information of China (English)

    周文杰; 张锦前; 贺轲; 段小鹏; 黄睿; 夏正林; 何景亮; 向国安


    Objective Screening of epigenetic drugs sensitive to intrahepatic cholangiocarcinoma and exploring the mechanism.Methods Screening a 96-epigenetic library in RBE cells through cell validity assay and comparing the inhibition effect of these sensitive drugs on multiple cell lines;Giemsa staining was used to observe the cell morphological change and flow cytometry to check the cell cycle distribution;Morphological observation,flow cytometry technique and Western blotting were combined to check the cell apoptosis.Results Twenty two drugs from the 96-epigenetic library had certain inhibition on RBE cells,including twelve histone deacetylase (HDAC) targeted species;HC-toxin' s inhibition effect on four intrahepatic cholangiocarcinoma cell lines were the most obvious among the HDAC inhibitors' and were superior to that of gemcitabine;cells at G0/G1 phases in HC toxin treated group was much more than that in control group (82.93% vs.48.51%) and HC toxin brought about diverse of morphological changes,apoptosis bodies and reduced mitosis included;flow cytometry indicated intensively apoptosis inducted by HC toxin and it was not associated with Caspase-3 depended pathways for the Caspase-3 up-regulated in the trial groups tested by Western blotting was not remarkable in statistics.Conclusion HDAC inhibitors,especially HC toxin,are of great potential in the treatment of intrahepatic cholangiocarcinoma.%目的 筛选肝内胆管癌敏感的表观遗传学药物并研究其作用机制.方法 采用细胞活力实验筛选对RBE细胞敏感的药物;在多种细胞株上比较其作用;选择最敏感的药物探讨其机制:通过Giemsa染色观察肝内胆管癌细胞经药物处理后的形态学变化,用流式细胞技术检测细胞周期分布,联合流式细胞技术、Western blot检测细胞的凋亡.结果 RBE细胞分别经96种表观遗传学药物处理48 h,27 μmol/L浓度下22种药物对RBE细胞有抑制作用(细胞活力<60%),组蛋白

  11. Resection of Perihilar Cholangiocarcinoma. (United States)

    Hartog, Hermien; Ijzermans, Jan N M; van Gulik, Thomas M; Groot Koerkamp, Bas


    Perihilar cholangiocarcinoma presents at the biliary and vascular junction of the hepatic hilum with a tendency to extend longitudinally into segmental bile ducts. Most patients show metastatic or unresectable disease at time of presentation or surgical exploration. In patients eligible for surgical resection, challenges are to achieve negative bile duct margins, adequate liver remnant function, and adequate portal and arterial inflow to the liver remnant. Surgical treatment is characterized by high rates of postoperative morbidity and mortality. This article reviews the various strategies and techniques, the role of staging laparoscopy, intraoperative frozen section, caudate lobectomy, and vascular reconstruction.

  12. Autocrine and Paracrine Mechanisms Promoting Chemoresistance in Cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Massimiliano Cadamuro


    Full Text Available Resistance to conventional chemotherapeutic agents, a typical feature of cholangiocarcinoma, prevents the efficacy of the therapeutic arsenal usually used to combat malignancy in humans. Mechanisms of chemoresistance by neoplastic cholangiocytes include evasion of drug-induced apoptosis mediated by autocrine and paracrine cues released in the tumor microenvironment. Here, recent evidence regarding molecular mechanisms of chemoresistance is reviewed, as well as associations between well-developed chemoresistance and activation of the cancer stem cell compartment. It is concluded that improved understanding of the complex interplay between apoptosis signaling and the promotion of cell survival represent potentially productive areas for active investigation, with the ultimate aim of encouraging future studies to unveil new, effective strategies able to overcome current limitations on treatment.

  13. [Radical surgery for hilar cholangiocarcinoma (Klatskin tumor)]. (United States)

    Ramos Rubio, Emilio


    In patients with hilar cholangiocarcinoma, long-term survival critically depends on complete tumor resection. Indeed, there are no long-term survivors with positive resection margins. Furthermore, hilar cholangiocarcinoma seems to have a low propensity for distant metastases and adjuvant therapy after surgery has not been shown to have clear clinical benefits. This evidence should be regarded as arguments for extended resections. The question remains of how to achieve an R0 resection. In the last few years greater use of major hepatectomy has increased resectability and has improved long-term results. Concomitant resection of the caudate lobe is recommended as this site is a prime area of local recurrence. Frozen sections should be routinely used to assess the remnant proximal and distal ductal stumps. However, if the proximal remnant is positive, additional ductal resection at the separating limits is not always feasible. Gross portal vein invasion has a negative impact on survival, but should not be a contraindication to resection. Hepatectomy with portal vein resection can offer long-term survival in some patients with advanced hilar cholangiocarcinoma. The incidence of nodal involvement in resected specimens has been reported to range from 30% to more than 50% and there is a correlation between primary tumor extension and nodal involvement. Lymphatic metastases from hilar cholangiocarcinoma appear to spread first to pericholedochal nodes in the hepatoduodenal ligament and then to spread widely toward the posteriorsuperior area around the pancreatic head, portal vein and common hepatic artery. Routine lymphadenectomy should include all these areas. The only factors precluding resection are involvement of celiac, superior mesenteric or para-aortic tumoral nodes. Survival is closely associated with the extent of nodal involvement. The no-touch technique including right trisegmentectomy combined with portal vein resection has been proposed as the surgical procedure

  14. Cholangiocarcinoma with respect to IgG4 Reaction

    Directory of Open Access Journals (Sweden)

    Kenichi Harada


    Full Text Available IgG4 reactions marked by infiltration of IgG4-positive plasma cells in affected organs occur in cancer patients and in patients with IgG4-related diseases. Extrahepatic cholangiocarcinomas including gall bladder cancer are often accompanied by significant IgG4 reactions; these reactions show a negative correlation with CD8-positive cytotoxic T cells, suggesting that the evasion of immune surveillance is associated with cytotoxic T cells. The regulatory cytokine IL-10 may induce IgG4-positive plasma cell differentiation or promote B cell switching to IgG4 in the presence of IL-4. Cholangiocarcinoma cells may function as nonprofessional antigen presenting cells that indirectly induce IgG4 reactions via the IL-10-producing cells and/or these may act as Foxp3-positive and IL-10-producing cells that directly induce IgG4 reactions. Moreover, IgG4-related disease is a high-risk factor for cancer development; IgG4-related sclerosing cholangitis (IgG4-SC cases associated with cholangiocarcinoma or its precursor lesion biliary intraepithelial neoplasia (BilIN have been reported. IgG4-positive cell infiltration is an important finding of IgG4-SC but is not a histological hallmark of IgG4-SC. For the diagnosis of IgG4-SC, its differentiation from cholangiocarcinoma remains important.

  15. Anticancer activity of streptochlorin, a novel antineoplastic agent, in cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Kwak TW


    Full Text Available Tae Won Kwak,1,* Hee Jae Shin,2,* Young-Il Jeong,1 Myoung-Eun Han,3 Sae-Ock Oh,3 Hyun-Jung Kim,4 Do Hyung Kim,5 Dae Hwan Kang1 1Biomedical Research Institute, Pusan National University Hospital, Busan, 2Marine Natural Products Chemistry Laboratory, Korea Institute of Ocean Science and Technology, Ansan, 3Department of Anatomy, School of Medicine, Pusan National University, Gyeongnam, 4Genewel Co Ltd. Gyeonggi-do, 5School of Medicine, Pusan National University, Yangsan, Gyeongnam, Republic of Korea *These authors contributed equally to this work Background: The aim of this study is to investigate the anticancer activity of streptochlorin, a novel antineoplastic agent, in cholangiocarcinoma. Methods: The anticancer activity of streptochlorin was evaluated in vitro in various cholangiocarcinoma cell lines for apoptosis, proliferation, invasiveness, and expression of various protein levels. A liver metastasis model was prepared by splenic injection of HuCC-T1 cholangiocarcinoma cells using a BALB/c nude mouse model to study the systemic antimetastatic efficacy of streptochlorin 5 mg/kg at 8 weeks. The antitumor efficacy of subcutaneously injected streptochlorin was also assessed using a solid tumor xenograft model of SNU478 cells for 22 days in the BALB/c nude mouse. Results: Streptochlorin inhibited growth and secretion of vascular endothelial growth factor by cholangiocarcinoma cells in a dose-dependent manner and induced apoptosis in vitro. In addition, streptochlorin effectively inhibited invasion and migration of cholangiocarcinoma cells. Secretion of vascular endothelial growth factor and activity of matrix metalloproteinase-9 in cholangiocarcinoma cells were also suppressed by treatment with streptochlorin. Streptochlorin effectively regulated metastasis of HuCC-T1 cells in a mouse model of liver metastasis. In a tumor xenograft study using SNU478 cells, streptochlorin significantly inhibited tumor growth without changes in body weight

  16. Notch1 is overexpressed in human intrahepatic cholangiocarcinoma and is associated with its proliferation, invasiveness and sensitivity to 5-fluorouracil in vitro. (United States)

    Wu, Wen-Rui; Zhang, Rui; Shi, Xiang-De; Zhu, Man-Sheng; Xu, Lei-Bo; Zeng, Hong; Liu, Chao


    The Notch signaling pathway has been reported to play crucial roles in inhibiting hepatocyte differentiation and allowing formation of intrahepatic bile ducts. However, little is known about its significance in intrahepatic cholangiocarcinoma (ICC). The aim of the present study was to investigate the effects of Notch1 expression in ICC tissues and cells. The expression of Notch1 was examined in paraffin-embedded sections of ICC (n=44) by immunohistochemistry. Notch1 was knocked down by RNA interference (RNAi) in cultured ICC cells (RBE and HCCC-9810). The proliferation, invasiveness and sensitivity to 5-fluorouracil (5-FU) were detected by Cell Counting Kit-8 (CCK-8), colony formation assays, Transwell assays and flow cytometry, respectively. The expression levels of several multidrug resistance (MDR)-related genes, MDR1-P-glycoprotein (ABCB‑1), breast cancer resistance protein (ABCG‑2) and the multidrug resistance protein isoform 1 (MRP‑1), were examined by quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting. Notch1 was overexpressed in cell membranes and cytoplasm of ICC compared with the adjacent liver tissue (35/44, 79.5%) and this was more common in cases with tumor size≥5 cm (p=0.021) and HBs-Ag positive (p=0.018). By silencing Notch1, the proliferation and invasiveness of ICC cells were inhibited and the inhibition rate of 5-FU was markedly increased. In addition, IC50 values of 5-FU in RBE cells were decreased from 148.74±0.72 to 5.37±0.28 µg/ml and the corresponding values for HCCC-9810 cells were 326.92±0.87 to 42.60±0.35 µg/ml, respectively. Furthermore, Notch1 silencing clearly increased the percentage of apoptotic cells treated by 5-FU compared with the control. Notch1 knockdown led to diminished expression levels of ABCB‑1 and MRP‑1. Therefore, Notch may play important roles in the development of ICC. Silencing Notch1 can inhibit the proliferation and invasiveness of ICC cells and increase their

  17. Current update on combined hepatocellular-cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Suresh Maximin


    Combined hepatocellular cholangiocarcinoma tends to present with an more aggressive behavior and a poorer prognosis than either hepatocellular carcinoma or cholangiocarcinoma. An accurate preoperative diagnosis and aggressive treatment planning can play crucial roles in appropriate patient management.

  18. [Perihilar cholangiocarcinoma (Klatskin tumor)]. (United States)

    Stavrou, G A; Donati, M; Faiss, S; Jenner, R M; Niehaus, K J; Oldhafer, K J


    Perihilar cholangiocarcinoma or Klatskin tumors are a rare entity arising from the extrahepatic bile duct bifurcation. Considering the close anatomical relationship of the bile duct bifurcation with the portal vein bifurcation and hepatic arteries, surgical treatment is demanding. With an incidence of only 2-4 cases/100,000 population/year patients should be referred to a specialized center. The tumors are usually poorly differentiated adenocarcinomas growing diffusely along the duct and also the perineural sheath. Only radical surgery offers a curative option and currently surgical strategy usually consists of en bloc resection of the bile duct, extended liver resection and portal vein resection. Proximal and lateral safety margin R0 resections are technically very demanding procedures because of the local anatomy.

  19. Cocktail treatment with EGFR-specific and CD133-specific chimeric antigen receptor-modified T cells in a patient with advanced cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Kai-chao Feng


    Full Text Available Abstract Background Cholangiocarcinoma (CCA is one of the most fatal malignant tumors with increasing incidence, mortality, and insensitivity to traditional chemo-radiotherapy and targeted therapy. Chimeric antigen receptor-modified T cell (CART immunotherapy represents a novel strategy for the management of many malignancies. However, the potential of CART therapy in treating advanced unresectable/metastatic CCA is uncharted so far. Case presentation In this case, a 52-year-old female who was diagnosed as advanced unresectable/metastatic CCA and resistant to the following chemotherapy and radiotherapy was treated with CART cocktail immunotherapy, which was composed of successive infusions of CART cells targeting epidermal growth factor receptor (EGFR and CD133, respectively. The patient finally achieved an 8.5-month partial response (PR from the CART-EGFR therapy and a 4.5-month-lasting PR from the CART133 treatment. The CART-EGFR cells induced acute infusion-related toxicities such as mild chills, fever, fatigue, vomiting and muscle soreness, and a 9-day duration of delayed lower fever, accompanied by escalation of IL-6 and C reactive protein (CRP, acute increase of glutamic-pyruvic transaminase and glutamic-oxalacetic transaminase, and grade 2 lichen striatus-like skin pathological changes. The CART133 cells induced an intermittent upper abdominal dull pain, chills, fever, and rapidly deteriorative grade 3 systemic subcutaneous hemorrhages and congestive rashes together with serum cytokine release, which needed emergent medical intervention including intravenous methylprednisolone. Conclusions This case suggests that CART cocktail immunotherapy may be feasible for the treatment of CCA as well as other solid malignancies; however, the toxicities, especially the epidermal/endothelial damages, require a further investigation. Trial registration NCT01869166 and NCT02541370 .

  20. Isocitrate dehydrogenase 1 and 2 mutations in cholangiocarcinoma. (United States)

    Kipp, Benjamin R; Voss, Jesse S; Kerr, Sarah E; Barr Fritcher, Emily G; Graham, Rondell P; Zhang, Lizhi; Highsmith, W Edward; Zhang, Jun; Roberts, Lewis R; Gores, Gregory J; Halling, Kevin C


    Somatic mutations in isocitrate dehydrogenase 1 and 2 genes are common in gliomas and help stratify patients with brain cancer into histologic and molecular subtypes. However, these mutations are considered rare in other solid tumors. The aims of this study were to determine the frequency of isocitrate dehydrogenase 1 and 2 mutations in cholangiocarcinoma and to assess histopathologic differences between specimens with and without an isocitrate dehydrogenase mutation. We sequenced 94 formalin-fixed, paraffin-embedded cholangiocarcinoma (67 intrahepatic and 27 extrahepatic) assessing for isocitrate dehydrogenase 1 (codon 132) and isocitrate dehydrogenase 2 (codons 140 and 172) mutations. Multiple histopathologic characteristics were also evaluated and compared with isocitrate dehydrogenase 1/2 mutation status. Of the 94 evaluated specimens, 21 (22%) had a mutation including 14 isocitrate dehydrogenase 1 and 7 isocitrate dehydrogenase 2 mutations. Isocitrate dehydrogenase mutations were more frequently observed in intrahepatic cholangiocarcinoma than in extrahepatic cholangiocarcinoma (28% versus 7%, respectively; P = .030). The 14 isocitrate dehydrogenase 1 mutations were R132C (n = 9), R132S (n = 2), R132G (n = 2), and R132L (n = 1). The 7 isocitrate dehydrogenase 2 mutations were R172K (n = 5), R172M (n = 1), and R172G (n = 1). Isocitrate dehydrogenase mutations were more frequently observed in tumors with clear cell change (P < .001) and poorly differentiated histology (P = .012). The results of this study show for the first time that isocitrate dehydrogenase 1 and 2 genes are mutated in cholangiocarcinoma. The results of this study are encouraging because it identifies a new potential target for genotype-directed therapeutic trials and may represent a potential biomarker for earlier detection of cholangiocarcinoma in a subset of cases.

  1. Circulating hTERT mRNA as a tumor marker in cholangiocarcinoma patients

    Institute of Scientific and Technical Information of China (English)

    Kawin Leelawat; Surang Leelawat; Thawee Ratanachu-Ek; Somboon Trubwongchareon; Jerasak Wannaprasert; Saad Tripongkaruna; Suchart Chantawibul; Panadda Tepaksorn


    AIM: To investigate human telomerase reverse transcriptase (hTERT) mRNA in the serum of cholangiocarcinoma patients.METHODS: The serum of thirty three cholangiocarcinoma patients, forty one benign biliary tract disease patients and ten healthy volunteers were collected and analyzed for the expression of hTERT mRNA by real-time reverse transcriptase-polymerase chain reaction (RT-PCR).We then examined the correlation between values of serum hTERT mRNA and the pathological staging of cholangiocarcinoma.RESULTS: hTERT mRNA was detected in 28 of 33(84.85%) of serum obtained from cholangiocarcinoma patients and 9 of 41 (21.9%) of serum obtained from benign biliary tract disease patients. hTERT mRNA was not detected in any serum obtained from healthy volunteers. on the other hand the common tumor marker, CA19-9 was detected in 20 of 33 (60.6%) of serum obtained from cholangiocarcinoma patients and 8 of 41 (19.5%) of serum obtained from benign biliary tract disease patients. However, no correlation was found between the present of serum hTERT mRNA and tumor staging.CONCLUSION: These results indicate that the detection of circulating hTERT mRNA was identified in almost all cholangiocarcinoma patients. It offers anovel tumor marker, which can be used as a complementary study for diagnosis of cholangiocarcinoma.

  2. Strong expression of CD133 is associated with increased cholangiocarcinoma progression

    Institute of Scientific and Technical Information of China (English)

    Kawin Leelawat; Taweesak Thongtawee; Siriluck Narong; Somboon Subwongcharoen; Sa-ad Treepongkaruna


    AIM: To determine the role of CD133 in cholangiocar-cinoma progression.METHODS: CD133 protein expression was evaluated by immunohistochemistry in 34 cholangiocarcinoma specimens. In addition, proliferation, chemoresistance and invasive properties of CD133-enriched (CD133+) and CD133-depleted (CD133-) RMCCA1 cholangiocarci-noma cells were studied and compared.RESULTS: Strong CD133 expression was observed in 67.6% (23/34) of the cholangiocarcinoma specimens. Strong expression of CD133 was significantly associat-ed with nodal metastasis (P = 0.009) and positive sur-gical margin status (P = 0.011). In the in vitro study, both the CD133+ and CD133- cells had similar prolifera-tion abilities and resistance to chemotherapeutic drugs. However, the CD133+ cells had a higher invasive ability compared with CD133- cells.CONCLUSION: CD133+ cells play an important role in the invasiveness of cholangiocarcinoma. Targeting of the CD133+ cells may be a useful approach to improve treatment against cholangiocarcinoma.

  3. ABT737 enhances cholangiocarcinoma sensitivity to cisplatin through regulation of mitochondrial dynamics

    Energy Technology Data Exchange (ETDEWEB)

    Fan, Zhongqi [Department of Hepatobiliary & Pancreas Surgery, The First Hospital, Jilin University, Changchun, Jilin 130021 (China); Yu, Huimei [Department of Pathophysiology, School of Basic Medical Sciences, Jilin University, Changchun, Jilin 130021 (China); Cui, Ni [Bethune Medical College, Jilin University, Changchun, Jilin 130021 (China); Kong, Xianggui; Liu, Xiaomin; Chang, Yulei [State Key Laboratory of Luminescence and Applications, Changchun Institute of Optics, Fine Mechanics and Physics, Chinese Academy of Sciences, Changchun, Jilin 130033 (China); Wu, Yao [Department of Pathophysiology, School of Basic Medical Sciences, Jilin University, Changchun, Jilin 130021 (China); Sun, Liankun, E-mail: [Department of Pathophysiology, School of Basic Medical Sciences, Jilin University, Changchun, Jilin 130021 (China); Wang, Guangyi, E-mail: [Department of Hepatobiliary & Pancreas Surgery, The First Hospital, Jilin University, Changchun, Jilin 130021 (China)


    Cholangiocarcinoma responses weakly to cisplatin. Mitochondrial dynamics participate in the response to various stresses, and mainly involve mitophagy and mitochondrial fusion and fission. Bcl-2 family proteins play critical roles in orchestrating mitochondrial dynamics, and are involved in the resistance to cisplatin. Here we reported that ABT737, combined with cisplatin, can promote cholangiocarcinoma cells to undergo apoptosis. We found that the combined treatment decreased the Mcl-1 pro-survival form and increased Bak. Cells undergoing cisplatin treatment showed hyperfused mitochondria, whereas fragmentation was dominant in the mitochondria of cells exposed to the combined treatment, with higher Fis1 levels, decreased Mfn2 and OPA1 levels, increased ratio of Drp1 60 kD to 80 kD form, and more Drp1 located on mitochondria. More p62 aggregates were observed in cells with fragmented mitochondria, and they gradually translocated to mitochondria. Mitophagy was induced by the combined treatment. Knockdown p62 decreased the Drp1 ratio, increased Tom20, and increased cell viability. Our data indicated that mitochondrial dynamics play an important role in the response of cholangiocarcinoma to cisplatin. ABT737 might enhance cholangiocarcinoma sensitivity to cisplatin through regulation of mitochondrial dynamics and the balance within Bcl-2 family proteins. Furthermore, p62 seems to be critical in the regulation of mitochondrial dynamics. - Highlights: • Cholangiocarcinoma may adapt to cisplatin through mitochondrial fusion. • ABT737 sensitizes cholangiocarcinoma to cisplatin by promoting fission and mitophagy. • p62 might participate in the regulation of mitochondrial fission and mitophagy.

  4. Cancer review: Cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Yezaz Ahmed Ghouri


    Full Text Available Cholangiocarcinoma (CCA is the most common biliary tract malignancy. CCA is classified as intrahepatic, perihilar or distal extrahepatic; the individual subtypes differ in their biologic behavior, clinical presentation, and management. Throughout the last decades, CCA incidence rates had significantly increased. In addition to known established risk factors, novel possible risk factors (i.e. obesity, hepatitis C virus have been identified that are of high importance in developed countries where CCA prevalence rates have been low. CCA tends to develop on the background of inflammation and cholestasis. In recent years, our understanding of the molecular mechanisms of cholangiocarcinogenesis has increased, thereby, providing the basis for molecularly targeted therapies. In its diagnostic evaluation, imaging techniques have improved, and the role of complementary techniques has been defined. There is a need for improved CCA biomarkers as currently used ones are suboptimal. Multiple staging systems have been developed, but none of these is optimal. The prognosis of CCA is considered dismal. However, treatment options have improved throughout the last two decades for carefully selected subgroups of CCA patients. Perihilar CCA can now be treated with orthotopic liver transplantation with neoadjuvant chemoradiation achieving 5-year survival rates of 68%. Classically considered chemotherapy-resistant, the ABC-02 trial has shown the therapeutic benefit of combination therapy with gemcitabine and cisplatin. The benefits of adjuvant treatments for resectable CCA, local ablative therapies and molecularly targeted therapies still need to be defined. In this article, we will provide the reader with an overview over CCA, and discuss the latest developments and controversies.

  5. Palliation:Hilar cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Mahesh; Kr; Goenka; Usha; Goenka


    Hilar cholangiocarcinomas are common tumors of the bile duct that are often unresectable at presentation. Palliation, therefore, remains the goal in the majority of these patients. Palliative treatment is particularly indicated in the presence of cholangitis and pruritus but is often also offered for high-grade jaundice and abdominal pain. Endoscopic drainage by placing stents at endoscopic retrograde cholangio-pancreatography(ERCP) is usually the preferred modality of palliation. However, for advanced disease, percutaneous stenting has been shown to be superior to endoscopic stenting. Endosonography-guided biliary drainage is emerging as an alternative technique, particularly when ERCP is not possible or fails. Metal stents are usually preferred over plastic stents, both for ERCP and for percutaneous bili-ary drainage. There is no consensus as to whether it is necessary to place multiple stents within advanced hi-lar blocks or whether unilateral stenting would suffice. However, recent data have suggested that, contrary to previous belief, it is useful to drain more than 50% of the liver volume for favorable long-term results. In the presence of cholangitis, it is beneficial to drain all of the obstructed biliary segments. Surgical bypass plays a limited role in palliation and is offered primarily as asegment Ⅲ bypass if, during a laparotomy for resec-tion, the tumor is found to be unresectable. Photody-namic therapy and, more recently, radiofrequency abla-tion have been used as adjuvant therapies to improve the results of biliary stenting. The exact technique to be used for palliation is guided by the extent of the bili-ary involvement(Bismuth class) and the availability of local expertise.

  6. Review to better understand the macroscopic subtypes and histogenesis of intrahepatic cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Yuichi; Sanada; Yujo; Kawashita; Satomi; Okada; Takashi; Azuma; Shigetoshi; Matsuo


    Intrahepatic cholangiocarcinoma is macroscopically classified into three subtypes, mass-forming-type, periductal infiltrating-type, and intraductal growth-type. Each subtype should be preoperatively differentiated to perform the valid surgical resection. Recent researches have revealed the clinical, radiologic, pathobiological characteristics of each subtype. We reviewed recently published studies covering various aspects of intrahepatic cholangiocarcinoma(ICC), focusing especially on the macroscopic subtypes and stem cell features to better understand the pathophysiology of ICC and to establish the valid therapeutic strategy.

  7. JAK-STAT pathway in carcinogenesis: Is it relevant to cholangiocarcinoma progression?

    Institute of Scientific and Technical Information of China (English)


    The features of JAK-STAT signaling in liver cells are discussed in the current review. The role of this signaling cascade in carcinogenesis is accentuated. The possible involvement of this pathway and alteration of its elements are compared for normal cholangiocytes,cholangiocarcinoma predisposition and development.Prolactin and interleukin-6 are described in detail as the best studied examples. In addition, the non-classical nuclear translocation of cytokine receptors is discussed in terms of its possible implication to cholangiocarcinoma development.

  8. LncRNAs H19 and HULC, activated by oxidative stress, promote cell migration and invasion in cholangiocarcinoma through a ceRNA manner

    Directory of Open Access Journals (Sweden)

    Wen-Tao Wang


    Full Text Available Abstract Background Long non-coding RNAs (lncRNAs are known to play important roles in different cell contexts, including cancers. However, little is known about lncRNAs in cholangiocarcinoma (CCA, a cholangiocyte malignancy with poor prognosis, associated with chronic inflammation and damage to the biliary epithelium. The aim of the study is to identify if any lncRNA might associate with inflammation or oxidative stress in CCA and regulate the disease progression. Methods In this study, RNA-seqs datasets were used to identify aberrantly expressed lncRNAs. Small interfering RNA and overexpressed plasmids were used to modulate the expression of lncRNAs, and luciferase target assay RNA immunoprecipitation (RIP was performed to explore the mechanism of miRNA-lncRNA sponging. Results We firstly analyzed five available RNA-seqs datasets to investigate aberrantly expressed lncRNAs which might associate with inflammation or oxidative stress. We identified that two lncRNAs, H19 and HULC, were differentially expressed among all the samples under the treatment of hypoxic or inflammatory factors, and they were shown to be stimulated by short-term oxidative stress responses to H2O2 and glucose oxidase in CCA cell lines. Further studies revealed that these two lncRNAs promoted cholangiocyte migration and invasion via the inflammation pathway. H19 and HULC functioned as competing endogenous RNAs (ceRNAs by sponging let-7a/let-7b and miR-372/miR-373, respectively, which activate pivotal inflammation cytokine IL-6 and chemokine receptor CXCR4. Conclusions Our study revealed that H19 and HULC, up-regulated by oxidative stress, regulate CCA cell migration and invasion by targeting IL-6 and CXCR4 via ceRNA patterns of sponging let-7a/let-7b and miR-372/miR-373, respectively. The results suggest that these lncRNAs might be the chief culprits of CCA pathogenesis and progression. The study provides new insight into the mechanism linking lncRNA function with CCA and

  9. Novel Silicone-Coated 125I Seeds for the Treatment of Extrahepatic Cholangiocarcinoma. (United States)

    Lin, Lizhou; Guo, Lili; Zhang, Weixing; Cai, Xiaobo; Chen, Dafan; Wan, Xinjian


    125I seeds coated with titanium are considered a safe and effective interstitial brachytherapy for tumors, while the cost of 125I seeds is a major problem for the patients implanting lots of seeds. The aim of this paper was to develop a novel silicone coating for 125I seeds with a lower cost. In order to show the radionuclide utilization ratio, the silicone was coated onto the seeds using the electro-spinning method and the radioactivity was evaluated, then the anti-tumor efficacy of silicone 125I seeds was compared with titanium 125I seeds. The seeds were divided into four groups: A (control), B (pure silicone), C (silicone 125I), D (titanium 125I) at 2 Gy or 4 Gy. Their anti-tumour activity and mechanism were assessed in vitro and in vivo using a human extrahepatic cholangiocarcinoma cell line FRH-0201 and tumor-bearing BALB/c nude mice. The silicone 125I seeds showed higher radioactivity; the rate of cell apoptosis in vitro and the histopathology in vivo demonstrated that the silicone 125I seeds shared similar anti-tumor efficacy with the titanium 125I seeds for the treatment of extrahepatic cholangiocarcinoma, while they have a much lower cost.

  10. Impact of classification of hilar cholangiocarcinomas (Klatskin tumors) on the incidence of intra- and extrahepatic cholangiocarcinoma in the United States. (United States)

    Welzel, Tania M; McGlynn, Katherine A; Hsing, Ann W; O'Brien, Thomas R; Pfeiffer, Ruth M


    Cholangiocarcinomas are topographically categorized as intrahepatic or extrahepatic by the International Classification of Diseases for Oncology (ICD-O). Although hilar cholangiocarcinomas (Klatskin tumors) are extrahepatic cholangiocarcinomas, the second edition of the ICD-O (ICD-O-2) assigned them a histology code 8162/3, Klatskin, which was cross-referenced to intrahepatic cholangiocarcinoma. Recent studies in the United States that included this code (8162/3, Klatskin) with intrahepatic cholangiocarcinoma reported an increasing incidence of intrahepatic cholangiocarcinoma and a decreasing incidence of extrahepatic cholangiocarcinoma. To investigate the impact of this misclassification on site-specific cholangiocarcinoma incidence rates, we calculated annual percent changes (APCs) with data from the Surveillance, Epidemiology, and End Results (SEER) program using a Poisson regression model that was age-adjusted to the year 2000 U.S. population. All statistical tests were two-sided. During 1992-2000, when SEER used ICD-O-2, 1710 intrahepatic cholangiocarcinomas, 1371 extrahepatic cholangiocarcinomas, and 269 hilar cholangiocarcinomas identified by code 8162/3, Klatskin were diagnosed. Ninety-one percent (246 of 269) of the hilar cholangiocarcinomas were incorrectly coded as intrahepatic cholangiocarcinomas, resulting in an overestimation of intrahepatic cholangiocarcinoma incidence by 13% and underestimation of extrahepatic cholangiocarcinomas incidence by 15%. However, even after the exclusion of tumors that were coded to the histology code 8162/3, Klatskin, age-adjusted annual intrahepatic cholangiocarcinoma incidence increased during this period (APC = 4%, 95% confidence interval = 2% to 6%, P<.001).

  11. Notch3 drives development and progression of cholangiocarcinoma. (United States)

    Guest, Rachel V; Boulter, Luke; Dwyer, Benjamin J; Kendall, Timothy J; Man, Tak-Yung; Minnis-Lyons, Sarah E; Lu, Wei-Yu; Robson, Andrew J; Gonzalez, Sofia Ferreira; Raven, Alexander; Wojtacha, Davina; Morton, Jennifer P; Komuta, Mina; Roskams, Tania; Wigmore, Stephen J; Sansom, Owen J; Forbes, Stuart J


    The prognosis of cholangiocarcinoma (CC) is dismal. Notch has been identified as a potential driver; forced exogenous overexpression of Notch1 in hepatocytes results in the formation of biliary tumors. In human disease, however, it is unknown which components of the endogenously signaling pathway are required for tumorigenesis, how these orchestrate cancer, and how they can be targeted for therapy. Here we characterize Notch in human-resected CC, a toxin-driven model in rats, and a transgenic mouse model in which p53 deletion is targeted to biliary epithelia and CC induced using the hepatocarcinogen thioacetamide. We find that across species, the atypical receptor NOTCH3 is differentially overexpressed; it is progressively up-regulated with disease development and promotes tumor cell survival via activation of PI3k-Akt. We use genetic KO studies to show that tumor growth significantly attenuates after Notch3 deletion and demonstrate signaling occurs via a noncanonical pathway independent of the mediator of classical Notch, Recombinant Signal Binding Protein for Immunoglobulin Kappa J Region (RBPJ). These data present an opportunity in this aggressive cancer to selectively target Notch, bypassing toxicities known to be RBPJ dependent.

  12. Reappraisal of the therapeutic role of celecoxib in cholangiocarcinoma.

    Directory of Open Access Journals (Sweden)

    Chun-Nan Yeh

    Full Text Available Cholangiocarcinoma (CCA, a lethal disease, affects many thousands worldwide yearly. Surgical resection provides the best chance for a cure; however, only one-third of CCA patients present with a resectable tumour at the time of diagnosis. Currently, no effective chemotherapy is available for advanced CCA. Cyclooxygenase-2 (COX-2 is a potential oncogene expressing in human CCA tissues and represents a candidate target for treatment; however, COX-2 inhibitors increase the risk of negative cardiovascular events as application for chemoprevention aim. Here, we re-evaluated the effectiveness and safety of celecoxib, one widely used COX-2 inhibitor, in treating CCA. We demonstrated that celecoxib exhibited an anti-proliferative effect on CGCCA cells via cell cycle arrest at G2 phase and apoptosis induction. Treatment for 5 weeks high dose celecoxib (160 mg/kg significantly repressed thioacetamide-induced CCA tumour growth in rats as monitored by animal positron emission tomography through apoptosis induction. No obviously observable side effects were noted during the therapeutic period. As retrospectively reviewing 78 intrahepatic mass-forming CCA patients, their survival was strongly and negatively associated with a positive resection margin and high COX-2 expression. Based on our result, we concluded that short-term high dose celecoxib may be a promising therapeutic regimen for CCA. Yet its clinical application still needs more studies to prove its safety.

  13. Long Noncoding RNA AFAP1-AS1 Promoted Tumor Growth and Invasion in Cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Xu Lu


    Full Text Available Background: Long non-coding RNAs (lncRNAs have been shown to play important roles in a wide range of pathophysiological processes, including cancer progression. Our previous study has shown that AFAP1-AS1 was upregulated and acted as an oncogene in hepatocellular carcinoma. However, the expression and biological functions of lncRNA AFAP1-AS1 in intrahepatic cholangiocarcinoma (CCA remains largely unknown. Methods: The expression level of AFAP1-AS1 was measured in 56 pairs of human cholangiocarcinoma tumor tissues and corresponding adjacent normal bile duct tissues. The correlation between AFAP1-AS1 and the clinicopathological features were evaluated by chi-square test. The effects of AFAP1-AS1 on CCA cells were determined by CCK-8 assay, clone formation assay, flow cytometry and transwell assay. Finally, to determine the effect of AFAP1-AS1 on tumor growth in vivo, AFAP1-AS1 knockdowned CCLP-1 cells were subcutaneously into nude mice to evaluate tumor growth. Results: In this study, we found that lncRNA AFAP1-AS1 was increased in CCA tissues and patients with high AFAP1-AS1 expression had a shorter overall survival. SiRNA-mediated AFAP1-AS1 knockdown significantly decreased cell proliferation of the CCA cells, with downregulation of C-myc and Cycling D1 in vitro. Furthermore, AFAP1-AS1 silencing inhibited cell migration partly due to decrease the expression of MMP-2 and MMP-9. In addition, CCLP-1 cells with AFAP1-AS1 knockdown were injected into nude mice to investigate the effect of AFAP1-AS1 on the tumorigenesis in vivo. Conclusions: Taken together, our findings suggested that AFAP1-AS1 might promote the CCA progression and provided a novel potential therapeutic target for CCA.

  14. Imbalanced adaptive responses associated with microsatellite instability in cholangiocarcinoma (United States)

    Loilome, Watcharin; Kadsanit, Sasithorn; Muisook, Kanha; Yongvanit, Puangrat; Namwat, Nisana; Techasen, Anchalee; Puapairoj, Anucha; Khuntikeo, Narong; Phonjit, Pichai


    The adaptive response of the genome protection mechanism occurs in cells when exposed to genotoxic stress due to the overproduction of free radicals via inflammation and infection. In such circumstances, cells attempt to maintain health via several genome protection mechanisms. However, evidence is increasing that this adaptive response may have deleterious effect; a reduction of antioxidant enzymes and/or imbalance in the DNA repair system generates microsatellite instability (MSI), which has procarcinogenic implications. Therefore, the present study hypothesized that MSI caused by imbalanced responses of antioxidant enzymes and/or DNA repair enzymes as a result of oxidative/nitrative stress arising from the inflammatory response is involved in liver fluke-associated cholangiocarcinogenesis. The present study investigated this hypothesis by identifying the expression patterns of antioxidant enzymes, including superoxide dismutase 2 (SOD2) and catalase (CAT), and DNA repair enzymes, including alkyladenine DNA glycosylase (AAG), apurinic endonuclease (APE) and DNA polymerase β (DNA pol β). In addition, the activities of the antioxidant enzymes, SOD2 and CAT, were examined in human cholangiocarcinoma (CCA) tissues using immunohistochemical staining. MSI was also analyzed in human CCA tissues. The resulting data demonstrated that the expression levels of the SOD2 and CAT enzymes decreased. The activities of SOD2 and CAT decreased significantly in the CCA tissues, compared with the hepatic tissue of cadaveric donors. In the DNA repairing enzymes, it was found that the expression levels of AAG and DNA pol β enzymes increased, whereas the expression of APE decreased. In addition, it was found that MSI-high was present in 69% of patients, whereas MSI-low was present in 31% of patients, with no patients classified as having microsatellite stability. In the patients, a MSI-high was correlated with poor prognosis, indicated by a shorter survival rate. These results

  15. Claudin-7-positive synchronous spontaneous intrahepatic cholangiocarcinoma, adenocarcinoma and adenomas of the gallbladder in a Bearded dragon (Pogona vitticeps). (United States)

    Jakab, Csaba; Rusvai, Miklós; Szabó, Zoltán; Gálfi, Péter; Marosán, Miklós; Kulka, Janina; Gál, János


    In this study, synchronous spontaneous, independent liver and gallbladder tumours were detected in a Bearded dragon (Pogona vitticeps). The multiple tumours consisted of intrahepatic cholangiocarcinoma as well as in situ adenocarcinoma and two adenomas of the gallbladder. The biliary epithelial cells and the cholangiocarcinoma showed membranous cross-immunoreactivity for claudin-7. The gallbladder epithelial cells, its adenoma and adenocarcinoma showed basolateral cross-reactivity for claudin-7. We think that the humanised anti-claudin-7 antibody is a good marker for the detection of different primary cholangiocellular and gallbladder tumours in Bearded dragons. The cholangiocytes, the cholangiocarcinoma, the endothelial cells of the liver and the epithelial cells and gallbladder tumours all showed claudin-5 cross-reactivity. The humanised anti-cytokeratin AE1-AE3 antibody showed cross-reactivity in the biliary epithelial cells, cholangiocarcinoma cells, epithelial cells and tumour cells of the gallbladder. It seems that this humanised antibody is a useful epithelial marker for the different neoplastic lesions of epithelial cells in reptiles. The humanised anti-α-smooth muscle actin (α-SMA) antibody showed intense cross-reactivity in the smooth muscle cells of the hepatic vessels and in the muscle layer of the gallbladder. The portal myofibroblasts, the endothelial cells of the sinusoids and the stromal cells of the cholangiocarcinoma and gallbladder tumours were positive for α-SMA. The antibovine anti-vimentin and humanised anti-Ki-67 antibodies did not show crossreactivity in the different samples from the Bearded dragon.

  16. Cholangiocarcinoma: Has There been Any Progress?

    Directory of Open Access Journals (Sweden)

    Judith Meza-Junco


    Full Text Available Cholangiocarcinoma is the second most common primary hepatic tumour after hepatocellular carcinoma. Primary sclerosing cholangitis is one of the most commonly recognized risk factors for cholangiocarcinoma; however, approximately 90% of patients have no identifiable risk factors. Extrahepatic type is its most common presentation. Cholangiocarcinoma is considered to be a devastating disease, with a poor survival rate and few therapeutic options. Although surgical resection has been considered the best treatment option for localized cholangiocarcinoma, local recurrences of this cancer are very common, and imply persistent micrometastatic disease in lymph nodes or at surgical margins, even after extended surgical resection. Consequently, the five-year survival rate after attempted curative resection is only 20% to 40%. Early studies of liver transplantation for cholangiocarcinoma did not show a survival benefit and, currently, this tumour is considered to be an absolute contraindication for liver transplantation in most transplant centres worldwide. Recently, neoadjuvant chemoradiation in combination with liver transplantation for highly selected patients with cholangiocarcinoma has shown impressive results, with five-year survival rates at approximately 76% to 82% – similar to other standard indications for liver transplantation, such as hepatocellular carcinoma or hepatitis C-induced cirrhosis. However, this success of liver transplantation applies to only a subset of patients and most of the data originated from a single centre. Wider application of this strategy, especially for patients with potentially resectable disease, will require validation by other centres.

  17. Pathological aspects of so called "hilar cholangiocarcinoma" (United States)

    Castellano-Megías, Víctor M; Ibarrola-de Andrés, Carolina; Colina-Ruizdelgado, Francisco


    Cholangiocarcinoma (CC) arising from the large intrahepatic bile ducts and extrahepatic hilar bile ducts share clinicopathological features and have been called hilar and perihilar CC as a group. However, “hilar and perihilar CC” are also used to refer exclusively to the intrahepatic hilar type CC or, more commonly, the extrahepatic hilar CC. Grossly, a major distinction can be made between papillary and non-papillary tumors. Histologically, most hilar CCs are well to moderately differentiated conventional type (biliary) carcinomas. Immunohistochemically, CK7, CK20, CEA and MUC1 are normally expressed, being MUC2 positive in less than 50% of cases. Two main premalignant lesions are known: biliary intraepithelial neoplasia (BilIN) and intraductal papillary neoplasm of the biliary tract (IPNB). IPNB includes the lesions previously named biliary papillomatosis and papillary carcinoma. A series of 29 resected hilar CC from our archives is reviewed. Most (82.8%) were conventional type adenocarcinomas, mostly well to moderately differentiated, although with a broad morphological spectrum; three cases exhibited a poorly differentiated cell component resembling signet ring cells. IPNB was observed in 5 (17.2%), four of them with an associated invasive carcinoma. A clear cell type carcinoma, an adenosquamous carcinoma and two gastric foveolar type carcinomas were observed. PMID:23919110

  18. Molecular profiling of intrahepatic cholangiocarcinoma

    DEFF Research Database (Denmark)

    Oliveira, Douglas V N P; Zhang, Shanshan; Chen, Xin


    INTRODUCTION: Intrahepatic cholangiocarcinoma (iCCA) is the second most frequent primary tumor of the liver and a highly lethal disease. Therapeutic options for advanced iCCA are limited and ineffective due to the largely incomplete understanding of the molecular pathogenesis of this deadly tumor....... Areas covered: The present review article outlines the main studies and resulting discoveries on the molecular profiling of iCCA, with a special emphasis on the different techniques used for this purpose, the diagnostic and prognostic markers identified, as well as the genes and pathways that could...... be potentially targeted with innovative therapies. Expert commentary: Molecular profiling has led to the identification of distinct iCCA subtypes, characterized by peculiar genetic alterations and transcriptomic features. Targeted therapies against some of the identified genes are ongoing and hold great promise...

  19. Current therapy of hilar cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Stephanie HiuYan Lau; WanYee Lau


    BACKGROUND: Hilar cholangiocarcinoma (HC) is an adeno-carcinoma of the extrahepatic biliary tree arising from the main left or right hepatic ducts or their confluence. This tumor is still considered to be difficult to treat or to cure. DATA SOURCES: We reviewed the medical literature on HC. Relevant and updated information on this tumor was analyzed in a concise and easy-to-read manner. The article is not intended to be a systematic review, but an extensive search was conducted on PubMed and MEDLINE using the keywords "hilar cholangiocarcinoma" and "Klatskin tumor" until July 2011. RESULTS: The selection and the timing of management options for patients with HC are determined by the degree of certainty of the diagnosis, the general condition of the patients, the underlying liver function and the stage of the disease. Current treatment of HC can be divided into curative and palliative treatment. For the curative treatment, local excision should only be used on small tumors which are confined to the bile duct wall and Bismuth I papillary carcinoma. Partial hepatectomy should be combined with caudate lobe resection and porta-hepatis lymph node dissection. The results of these major resections can be improved with portal vein embolization, and staging laparoscopy and laparoscopic ultrasound. The role of preoperative biliary drainage is controversial. Autotransplantation for HC gave disappointing results while the Mayo Protocol of chemoradiation for selecting patients with unresectable HC for orthotopic liver transplantation has been widely accepted. Palliative treatment included bypass surgery, endoscopic or percutaneous stenting, photodynamic therapy, intraluminal brachytherapy, and external radiation and systemic therapy. CONCLUSIONS: Adequate surgery with R0 resection should be the main goal of treatment. For patients with unresectable HC, treatment aims to improve the quality and quantity of their survival.

  20. Risk factors and classifications of hilar cholangiocarcinoma. (United States)

    Suarez-Munoz, Miguel Angel; Fernandez-Aguilar, Jose Luis; Sanchez-Perez, Belinda; Perez-Daga, Jose Antonio; Garcia-Albiach, Beatriz; Pulido-Roa, Ysabel; Marin-Camero, Naiara; Santoyo-Santoyo, Julio


    Cholangiocarcinoma is the second most common primary malignant tumor of the liver. Perihilar cholangiocarcinoma or Klatskin tumor represents more than 50% of all biliary tract cholangiocarcinomas. A wide range of risk factors have been identified among patients with Perihilar cholangiocarcinoma including advanced age, male gender, primary sclerosing cholangitis, choledochal cysts, cholelithiasis, cholecystitis, parasitic infection (Opisthorchis viverrini and Clonorchis sinensis), inflammatory bowel disease, alcoholic cirrhosis, nonalcoholic cirrhosis, chronic pancreatitis and metabolic syndrome. Various classifications have been used to describe the pathologic and radiologic appearance of cholangiocarcinoma. The three systems most commonly used to evaluate Perihilar cholangiocarcinoma are the Bismuth-Corlette (BC) system, the Memorial Sloan-Kettering Cancer Center and the TNM classification. The BC classification provides preoperative assessment of local spread. The Memorial Sloan-Kettering cancer center proposes a staging system according to three factors related to local tumor extent: the location and extent of bile duct involvement, the presence or absence of portal venous invasion, and the presence or absence of hepatic lobar atrophy. The TNM classification, besides the usual descriptors, tumor, node and metastases, provides additional information concerning the possibility for the residual tumor (R) and the histological grade (G). Recently, in 2011, a new consensus classification for the Perihilar cholangiocarcinoma had been published. The consensus was organised by the European Hepato-Pancreato-Biliary Association which identified the need for a new staging system for this type of tumors. The classification includes information concerning biliary or vascular (portal or arterial) involvement, lymph node status or metastases, but also other essential aspects related to the surgical risk, such as remnant hepatic volume or the possibility of underlying disease.

  1. Role of ErbB family receptor tyrosine kinases in intrahepatic cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)


    Aberrant expression and signaling of epidermal growth factor receptor (ErbB) family receptor tyrosine kinases, most notably that of ErbB2 and ErbB1, have been implicated in the molecular pathogenesis of intrahepatic cholangiocarcinoma. Constitutive overexpression of ErbB2 and/or ErbB1 in malignant cholangiocytes has raised interest in the possibility that agents which selectively target these receptors could potentially be effective in cholangiocarcinoma therapy. However, current experience with such ErbB-directed therapies have at best produced only modest responses in patients with biliary tract cancers. This review provides a comprehensive and critical analysis of both preclinical and clinical studies aimed at assessing the role of altered ErbB2 and/or ErbB1 expression, genetic modifications, and dysregulated signaling on cholangiocarcinoma development and progression. Specific limitations in experimental approaches that have been used to assess human cholangiocarcinoma specimens for ErbB2 and/or ErbB1 overexpression and gene amplification are discussed. In addition, current rodent models of intrahepatic cholangiocarcinogenesis associated with constitutive ErbB2 overexpression are reviewed. Select interactive relationships between ErbB2 or ErbB1 with other relevant molecular signaling pathways associated with intrahepatic cholangiocarcinoma development and progression are also detailed, including those linking ErbB receptors to bile acid, cyclooxygenase-2, interleukin-6/gp130, transmembrane mucins, hepatocyte growth factor/Met, and vascular endothelial growth factor signaling. Lastly, various factors that can limit therapeutic efficacy of ErbB-targeted agents against cholangiocarcinoma are considered.

  2. Common Molecular Subtypes Among Asian Hepatocellular Carcinoma and Cholangiocarcinoma

    DEFF Research Database (Denmark)

    Chaisaingmongkol, Jittiporn; Budhu, Anuradha; Dang, Hien


    Intrahepatic cholangiocarcinoma (ICC) and hepatocellular carcinoma (HCC) are clinically disparate primary liver cancers with etiological and biological heterogeneity. We identified common molecular subtypes linked to similar prognosis among 199 Thai ICC and HCC patients through systems integration...... of genomics, transcriptomics, and metabolomics. While ICC and HCC share recurrently mutated genes, including TP53, ARID1A, and ARID2, mitotic checkpoint anomalies distinguish the C1 subtype with key drivers PLK1 and ECT2, whereas the C2 subtype is linked to obesity, T cell infiltration, and bile acid...

  3. The anti-tumor activity of dendritic cell/cytotoxic T lymphocyte induced by cholangiocarcinoma-derived exosome and its ultrafiltered lysate%基于exosome裂解超滤液的树突状细胞/细胞毒性T淋巴细胞诱导及其抗胆管癌研究

    Institute of Scientific and Technical Information of China (English)

    陈炯煌; 丁国平; 陈文超; 曹利平


    Objective To observe the impact of cholangiocarcinoma-derived exosome and ultrafiltered exosome lysate on the anti-tumor activity of dendritic celL/cytotoxic T lymphocyte (DC/CTL) cells and discuss the mechanism involved.Methods Exosomes derived from RBE cells (human cholangiocarcinoma line) were collected by ultracentrifugation,ow-osmotic splitting followed by ultrafiltration was performed to remove exosomal microRNAs and purify the ultrafiltered exosome lysate.ImDCs (immature dendritic cells) were induced from peripheral blood and were co-cultured with CTL(cytotoxic lymphocyte),which were impulsed by exosome and ultrafiltered exosome lysate,respectively.The concentrations of tumor necrosis factor (TNF)-αt and perforin in culture medium supernatant were examined by enzyme linked immunosorbent assay (ELISA).Cell counting kit-8 (CCK-8) was used to evaluate the cytotoxic activity of DC/CTL to RBE cells.Results The concentrations of TNF-αand perforin in E-DC/CTL group were 138.61 ng/L and 2.41 μg/L respectively,lower than those of DC/CTL (194.08 ng/L and 3.39 μg/L) and EL-DC/CTL group (210.87 ng/L and 3.79 μg/L).The killing rate of E-DC/CTL was 33.35%,lower than that of DC/CTL (47.35%) and EL-DC/CTL (66.23%) significantly.The killing rate of EL-DC/CTL was significantly higher than that of DC/CTL (P < 0.01).Conclusion RBE cell-derived exosome inhibits the anti-tumor activity of DC/CTL by down-regulating TNF-α and perforin,exosomal miRNA may play important roles in the immune escape of cholangiocarcinoma.We built a new model based on ultrafiltered exosome lysate derived from cholangiocarcinoma to enhance the anti-tumor activity of DC/CTL.%目的 观察人胆管癌细胞来源的exosome及exosome裂解超滤液(UEL)对树突状细胞/细胞毒性T淋巴细胞诱导(DC/CTL)混合细胞抗肿瘤活性的影响及机制.方法 采用超速离心法提取人胆管癌细胞(RBE细胞)释放的exosome,低渗法裂解exosome,超

  4. Effect of histone deacetylase inhibitor on expression of HDAC1 in gallbladder carcinoma cell line and extrahepatic cholangiocarcinoma cell line in vivo and in vitro%组蛋白去乙酰化酶抑制剂对胆囊癌细胞系和肝外胆管癌细胞系HDAC1表达的影响

    Institute of Scientific and Technical Information of China (English)

    王欣; 黄凯; 徐立宁


    目的 观察组蛋白去乙酰化酶抑制剂-曲古抑菌素(TSA)在体外和体内对胆囊癌细胞和肝外胆管癌细胞HDAC1表达的影响.方法 用TSA作用于胆囊癌细胞系(Mz-ChA-1)和肝外胆管癌细胞系(QBC939、KMBC、OZ),然后用逆转录-聚合酶链反应(RT-PCR)检测HDAC1之mR-NA的表达变化,用Western blot检测其蛋白的表达变化.将这些细胞接种在裸鼠皮下建立胆囊癌和肝外胆管癌裸鼠种植瘤模型,用免疫组织化学方法 观察TSA在体内对裸鼠种植瘤组织中HDAC1蛋白表达的影响.结果 TSA可以减弱胆囊癌细胞系(Mz-ChA-1)和肝外胆管癌细胞系(QBC939、KMBC、OZ)HDAC1 mRNA和蛋白的表达;TSA作用前后的胆囊癌细胞系(Mz-ChA-1)和肝外胆管癌细胞系(KMBC)裸鼠成瘤组织中的各种蛋白的表达均无变化.结论 TSA在体外可以抑制胆囊癌和肝外胆管癌HDAC1的表达;TSA抑制HDAC1表达的作用可能受到体内环境的影响而消失.%Objective To study the effect of histone deacetylase inhibitor-trichostatin A (TSA) on the HDAC1 expression in gallbladder carcinoma and extrahepatic cholangioearcinoma cells in vivo and in vitro. Methods The cells from gallbladder carcinoma cell line ( Mz-ChA-1 ) and cholangiocarcinoma cell lines (QBC939, KMBC and OZ) were treated with TSA, and the expression of HDAC1 mRNA and protein was detected by RT-PCR assay and Western blot respectively. These cells were subcutaneously transplanted into nude mice to establish the transplanted cholangiocarcinoma and gallbladder carcinoma models. The effect of TSA on the expression of HDAC1 protein in transplanted cancer tissues in vivo was observed. Results TSA could down-regulate the expression of HDAC1 mRNA and protein in gallbladder carcinoma cells and cholangiocarcinoma cells. TSA could not down-or up-regulate the expression of HDAC1 protein in the transplanted biliary tract cancer models in nude mice. Conclusion TSA might down-regulate the expression of HDAC1 in

  5. Human mesenchymal stem cells

    DEFF Research Database (Denmark)

    Abdallah, Basem; Kassem, Moustapha


    Mesenchymal stem cells (MSC) are a group of clonogenic cells present among the bone marrow stroma and capable of multilineage differentiation into mesoderm-type cells such as osteoblasts, adipocytes and chondrocytes. Due to their ease of isolation and their differentiation potential, MSC are being...... introduced into clinical medicine in variety of applications and through different ways of administration. Here, we discuss approaches for isolation, characterization and directing differentiation of human mesenchymal stem cells (hMSC). An update of the current clinical use of the cells is also provided....

  6. Induction of apoptosis in cholangiocarcinoma by an andrographolide analogue is mediated through topoisomerase II alpha inhibition. (United States)

    Nateewattana, Jintapat; Dutta, Suman; Reabroi, Somrudee; Saeeng, Rungnapha; Kasemsook, Sakkasem; Chairoungdua, Arthit; Weerachayaphorn, Jittima; Wongkham, Sopit; Piyachaturawat, Pawinee


    Cholangiocarcinoma (CCA), the common primary malignant tumor of bile duct epithelial cells, is unresponsive to most chemotherapeutic drugs. Diagnosis with CCA has a poor prognosis, and therefore urgently requires effective therapeutic agents. In the present study we investigated anti-cancer effects of andrographolide analogue 3A.1 (19-tert-butyldiphenylsilyl-8, 17-epoxy andrographolide) and its mechanism in human CCA cell line KKU-M213 derived from a Thai CCA patient. By 24h after exposure, the analogue 3A.1 exhibited a potent cytotoxic effect on KKU-M213 cells with an inhibition concentration 50 (IC50) of approximately 8.0µM. Analogue 3A.1 suppressed DNA topoisomerase II α (Topo II α) protein expression, arrested the cell cycle at sub G0/G1 phase, induced cleavage of DNA repair protein poly (ADP-ribose) polymerases-1 (PARP-1), and enhanced expression of tumor suppressor protein p53 and pro-apoptotic protein Bax. In addition, analogue 3A.1 induced caspase 3 activity and inhibited cyclin D1, CDK6, and COX-2 protein expression. These results suggest that andrographolide analogue 3A.1, a novel topo II inhibitor, has significant potential to be developed as a new anticancer agent for the treatment of CCA.

  7. Antitumor effect of FGFR inhibitors on a novel cholangiocarcinoma patient derived xenograft mouse model endogenously expressing an FGFR2-CCDC6 fusion protein. (United States)

    Wang, Yu; Ding, Xiwei; Wang, Shaoqing; Moser, Catherine D; Shaleh, Hassan M; Mohamed, Essa A; Chaiteerakij, Roongruedee; Allotey, Loretta K; Chen, Gang; Miyabe, Katsuyuki; McNulty, Melissa S; Ndzengue, Albert; Barr Fritcher, Emily G; Knudson, Ryan A; Greipp, Patricia T; Clark, Karl J; Torbenson, Michael S; Kipp, Benjamin R; Zhou, Jie; Barrett, Michael T; Gustafson, Michael P; Alberts, Steven R; Borad, Mitesh J; Roberts, Lewis R


    Cholangiocarcinoma is a highly lethal cancer with limited therapeutic options. Recent genomic analysis of cholangiocarcinoma has revealed the presence of fibroblast growth factor receptor 2 (FGFR2) fusion proteins in up to 13% of intrahepatic cholangiocarcinoma (iCCA). FGFR fusions have been identified as a novel oncogenic and druggable target in a number of cancers. In this study, we established a novel cholangiocarcinoma patient derived xenograft (PDX) mouse model bearing an FGFR2-CCDC6 fusion protein from a metastatic lung nodule of an iCCA patient. Using this PDX model, we confirmed the ability of the FGFR inhibitors, ponatinib, dovitinib and BGJ398, to modulate FGFR signaling, inhibit cell proliferation and induce cell apoptosis in cholangiocarcinoma tumors harboring FGFR2 fusions. In addition, BGJ398 appeared to be superior in potency to ponatinib and dovitinib in this model. Our findings provide a strong rationale for the investigation of FGFR inhibitors, particularly BGJ398, as a therapeutic option for cholangiocarcinoma patients harboring FGFR2 fusions.

  8. Pharmacological activity of Kaempferia parviflora extract against human bile duct cancer cell lines. (United States)

    Leardkamolkarn, Vijittra; Tiamyuyen, Sunida; Sripanidkulchai, Bung-orn


    A crude ethanol extract of Kaemperia parviflora Wall. Ex Baker and a purified compound, 5,7,4-trimethoxyflavone (KP.8.10), were evaluated for pharmacological effects on human cholangiocarcinoma cell lines (HuCCA-1 and RMCCA-1). The cells were incubated with various concentrations of extract for various time periods and metabolic activity (MTT assay) was assessed for cell viability. The results showed a dose-dependent effect of both crude ethanol extract and the pure compound. CC50s for the crude extract on HuCCA-1 and RMCCA-1 cells were 46.1 microg/ml and 62.0 microg/ml, respectively. Values for the pure compound could not be determined because of solubility problems. Interestingly, K. parviflora ethanol extract and KP.8.10 at low concentrations (10-20 microg/ml and 2.5-5 microg/ml, respectively) markedly reduced rhHGF-induced invasion by HuCCA-1 and RMCCA-1 cells across matrix-coated transwell plates. Higher concentrations of K. parviflora ethanol extract (60 and 80 microg/ml) and KP.8.10 (20 microg /ml) dramatically changed the cellular morphology and caused death in both cell types. KP.8.10 further exhibited progressive action via caspase-3 mitochondrial enzyme activation, enhancing cellular toxicity in a time-dose dependent fashion. Therefore, 5,7,4-trimethoxyflavone appeared to be a bioactive component of K. parviflora extract capable of exerting anti-cancer action. The results suggested a benefit of this edible plant in prevention and treatment of cholangiocarcinoma.

  9. Autophagy in intra-hepatic cholangiocarcinoma


    Huang, Jing Li; Hezel, Aram F.


    Intra-hepatic cholangiocarcinoma (IHCC) is a primary cancer of the liver that shares histological features with the hepatic bile ducts from which it is thought to arise. The incidence of this disease is increasing, possibly related to increased inflammatory liver diseases such as viral hepatitis and steatohepatitis (commonly called “fatty liver”), induced by obesity, diabetes, and other metabolic derangements. Its prognosis is generally poor with early metastasis and presently there are limit...

  10. [Hilar cholangiocarcinoma (Klatskin tumor) - current treatment options]. (United States)

    Bělina, F


    Hilar cholangiocarcinoma is a rare tumor with a rather poor prognosis, and thus remains a challenge for diagnosis and treatment. The sole potentionally curative treatment is the complete resection of the tumor. A negative surgical margin is one of the most important factors in achieving prolonged survival. A preoperative evaluation of the tumor is important for the evaluation of resectability and the extent of surgery. Unfortunately, only a small number of patients with hilar cholangiocarcinoma is indicated for the radical procedure. Liver transplantation is not a standard method in the treatment of the Klatskin tumor and it is reserved only for carefully selected patients in a few transplant centres. The main aim of the palliative treatment is biliary drainage, reduction of the pain and pruritus and overall improvement of the quality of life. Adjuvant chemotherapy and radiotherapy are the important parts of the complex therapy, however, no definite regimen has been established to date. Further evidence is needed to define the role of liver transplantation and (neo)adjuvant new therapeutic modalities. Key words: hilar cholangiocarcinoma - radical surgery - palliative treatment - liver transplantation - (neo) adjuvant chemotherapy - radiotherapy.

  11. Effective enrichment of cholangiocarcinoma secretomes using the hollow fiber bioreactor culture system. (United States)

    Weeraphan, Churat; Diskul-Na-Ayudthaya, Penchatr; Chiablaem, Khajeelak; Khongmanee, Amnart; Chokchaichamnankit, Daranee; Subhasitanont, Pantipa; Svasti, Jisnuson; Srisomsap, Chantragan


    The Northeastern region of Thailand is well known to have high incidence of bile duct cancer known as cholangiocarcinoma. So there is a continued need to improve diagnosis and treatment, and discovery of biomarkers for early detection of bile duct cancer should greatly improve treatment outcome for these patients. The secretome, a collection of proteins secreted from cells, is a useful source for identifying circulating biomarkers in blood secreted from cancer cells. Here a Hollow Fiber Bioreactor culture system was used for enrichment of cholangiocarcinoma secretomes, since this culture system mimics the dense three-dimensional microenvironment of the tumor found in vivo. Two-dimensional fluorescence difference gel electrophoresis using a sensitive Fluor saturation dye staining, followed by LC/MS/MS, was used to compare protein expression in the secretomes of cells cultured in the Hollow Fiber system and cells cultured in the monolayer culture system. For the first time, the 2D-patterns of cholangiocarcinoma secretomes from the two culture systems could be compared. The Hollow Fiber system improved the quality and quantity of cholangiocarcinoma secreted proteins compared to conventional monolayer system, showing less interference by cytoplasmic proteins and yielding more secreted proteins. Overall, 75 spots were analyzed by LC/MS/MS and 106 secreted proteins were identified. Two novel secreted proteins (C19orf10 and cystatin B) were found only in the Hollow Fiber system and were absent from the traditional monolayer culture system. Among the highly expressed proteins, 22 secreted soluble proteins were enriched by 5 fold in Hollow Fiber system compared to monolayer culture system. The Hollow Fiber system is therefore useful for preparing a wide range of proteins from low-abundance cell secretomes. Copyright © 2012 Elsevier B.V. All rights reserved.

  12. The roles of Notch1 expression in the migration of intrahepatic cholangiocarcinoma


    Zhou, Qi; Wang, Yafeng; Peng, Baogang; Liang, Lijian; Li, Jiaping


    Background Notch signaling, a critical pathway for tissue development, contributes to tumorigenesis in many tissues; however, the roles of Notch signaling in Intrahepatic Cholangiocarcinoma (ICC) remains unclear. In this study, we evaluated the expression and effects of Notch1 on cell migration in ICC. Methods Multiple cellular and molecular approaches were performed including gene transfection, siRNA transfection, RT-PCR, Western blotting, Rac activation assays and immunofluorescence. Result...

  13. Hepatocarcinoma-Intestine-Pancreas/Pancreatic Associated Protein (HIP/PAP) Is Expressed and Secreted by Proliferating Ductules as well as by Hepatocarcinoma and Cholangiocarcinoma Cells


    Christa, Laurence; Simon, Marie-Thérèse; Brezault-Bonnet, Catherine; Bonte, Eric; Carnot, Françoise; Zylberberg, Hervé; Franco, Dominique; Capron, Frédérique; Roskams, Tania; Bréchot, Christian


    Hepatocarcinoma-intestine-pancreas/pancreatic associated protein (HIP/PAP) gene was identified because of its increased expression in 25% of human hepatocellular carcinoma. HIP/PAP protein, a C-type lectin, binds laminin, acts as an adhesion molecule for hepatocytes, and has also been described as an acute phase secretory protein during acute pancreatitis in humans and rats. We investigated HIP/PAP protein expression in patients with various liver diseases associated with ductular reaction. A...

  14. Fibroblast growth factor receptor 4 promotes progression and correlates to poor prognosis in cholangiocarcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Yun-Fei [Department of Hepatobiliary Surgery, Qilu Hospital of Shandong University (China); Yang, Xiao-Qing [Department of Pathology, Shandong University (China); Lu, Xiao-Fei [Department of Gastrointestinal Surgery, Jinan Central Hospital (China); Guo, Sen; Liu, Yi; Iqbal, Mohammad; Ning, Shang-Lei; Yang, Hui [Department of Hepatobiliary Surgery, Qilu Hospital of Shandong University (China); Suo, Ning [Department of Anatomy, Shandong University (China); Chen, Yu-Xin, E-mail: [Department of Hepatobiliary Surgery, Qilu Hospital of Shandong University (China)


    Highlights: • FGFR4 is significantly related with N stage in IHCC, with T stage and TNM stage in PHCC. • FGFR4 is an independent prognostic factor in IHCC and PHCC. • FGFR4 promotes proliferation, invasion and EMT in cholangiocarinoma cell lines. • Inhibitor AP24354 can decrease proliferation, invasion and induce apoptosis of CCA. - Abstract: Fibroblast growth factor receptor 4 (FGFR4) is related to poor prognosis of several cancers, but the correlation between FGFR4 expression and cholangiocarcinoma (CCA) has not been well elucidated. We investigated the expression of FGFR4 in 83 intrahepatic cholangiocarcinomas (IHCCs), 75 perihilar cholangiocarcinomas (PHCCs) and 41 distal cholangiocarcinomas (DCCs) by immunohistochemistry (IHC), and subsequently evaluated association of FGFR4 with clinicopathologic parameters and survival rate. The rate of FGFR4 higher expression was 61.4% (51/83) in IHCCs, 53.3% (40/75) in PHCCs and 56.1% (23/41) in DCCs. FGFR4 expression was significantly related to poor prognosis of IHCC (P = 0.002) and PHCC (P = 0.019) with univariate analysis, and also identified as an independent prognostic factor in IHCC (P = 0.045) and PHCC (P = 0.049) with multivariate analysis. Additionally, with functional assays in vitro, we found FGFR4 can induce proliferation, invasion and epithelial–mesenchymal transition (EMT) of CCA cell lines with FGF19 stimulation. Moreover, FGFR4 inhibitor AP24354 can suppress proliferation, invasion and induce apoptosis of CCA cells. In conclusion, FGFR4 expression can be identified as a significant independent prognostic biomarker of IHCC and PHCC. FGFR4 played a pivotal role in proliferation, invasion and EMT of CCA. FGFR4 inhibitor can suppress proliferation, invasion and induce apoptosis of CCA, indicating that FGFR4 may act as a potential therapeutic target.

  15. Polymeric photosensitizer-embedded self-expanding metal stent for repeatable endoscopic photodynamic therapy of cholangiocarcinoma. (United States)

    Bae, Byoung-chan; Yang, Su-Geun; Jeong, Seok; Lee, Don Haeng; Na, Kun; Kim, Joon Mee; Costamagna, Guido; Kozarek, Richard A; Isayama, Hiroyuki; Deviere, Jacques; Seo, Dong Wan; Nageshwar Reddy, D


    Photodynamic therapy (PDT) is a new therapeutic approach for the palliative treatment of malignant bile duct obstruction. In this study, we designed photosensitizer-embedded self-expanding nonvascular metal stent (PDT-stent) which allows repeatable photodynamic treatment of cholangiocarcinoma without systemic injection of photosensitizer. Polymeric photosensitizer (pullulan acetate-conjugated pheophorbide A; PPA) was incorporated in self-expanding nonvascular metal stent. Residence of PPA in the stent was estimated in buffer solution and subcutaneous implantation on mouse. Photodynamic activity of PDT-stent was evaluated through laserexposure on stent-layered tumor cell lines, HCT-116 tumor-xenograft mouse models and endoscopic intervention of PDT-stent on bile duct of mini pigs. Photo-fluorescence imaging of the PDT-stent demonstrated homogeneous embedding of polymeric Pheo-A (PPA) on stent membrane. PDT-stent sustained its photodynamic activities at least for 2 month. And which implies repeatable endoscopic PDT is possible after stent emplacement. The PDT-stent after light exposure successfully generated cytotoxic singlet oxygen in the surrounding tissues, inducing apoptotic degradation of tumor cells and regression of xenograft tumors on mouse models. Endoscopic biliary in-stent photodynamic treatments on minipigs also suggested the potential efficacy of PDT-stent on cholangiocarcinoma. In vivo and in vitro studies revealed our PDT-stent, allows repeatable endoscopic biliary PDT, has the potential for the combination therapy (stent plus PDT) of cholangiocarcinoma. Copyright © 2014 Elsevier Ltd. All rights reserved.

  16. Cell encoding recombinant human erythropoietin

    Energy Technology Data Exchange (ETDEWEB)

    Beck, A.K.; Withy, R.M.; Zabrecky, J.R.; Masiello, N.C.


    This patent describes a C127 cell transformed with a recombinant DNA vector. It comprises: a DNA sequence encoding human erythropoietin, the transformed cell being capable of producing N-linked and O-linked glycosylated human erythropoietin.

  17. Radiofrequency ablation for incidentally identified primary intrahepatic cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Witold Zgodzinski; N.Joseph Espat


    Cholangiocarcinoma is the second most common primary hepato-biliary malignancy. The majority of patients with primary hepatic tumors are not suitable candidates for resection, due to advanced stage of the disease at presentation, anatomic limitations and medical comorbidities. At present, radiofrequency ablation (RFA) may offer an alternative, feasible and safe therapy for selected patients with hepatic tumors, who are not otherwise candidates for hepatic resection. Herein, we present the case of successful RFA in a patient with a solitary, primary intrahepatic cholangiocarcinoma. The patient remained free of disease 24 mo after the procedure, and is still followed up. This is the first report of RFA application inthe treatment of primary intrahepatic cholangiocarcinoma.

  18. Trousseau's Syndrome Caused by Intrahepatic Cholangiocarcinoma: An Autopsy Case Report and Literature Review

    Directory of Open Access Journals (Sweden)

    Takashi Yuri


    Full Text Available An autopsy case report of Trousseau's syndrome caused by intrahepatic cholangiocarcinoma is presented, and seven previously reported cases are reviewed. A 73-year-old woman experiencing light-headedness and dementia of unknown cause for 6 months developed severe hypotonia. A hypointense lesion compatible with acute cerebral infarction was detected by magnetic resonance imaging. Abdominal computed tomography revealed an ill-defined large liver mass in the right lobe. The mass was not further investigated because of the patient's poor condition. She died of multiple organ failure, and an autopsy was conducted. Postmortem examination revealed intrahepatic cholangiocarcinoma, fibrous vegetations on the mitral valves and multiple thromboemboli in the cerebrum, spleen and rectum. Trousseau's syndrome is defined as an idiopathic thromboembolism in patients with undiagnosed or concomitantly diagnosed malignancy. This syndrome is encountered frequently in patients with mucin-producing carcinomas, while the incidence in patients with intrahepatic cholangiocarcinoma is uncommon. We found that tissue factor and mucin tumor marker (CA19-9, CA15-3 and CA-125 expression in cancer cells may be involved in the pathogenesis of thromboembolism. A patient with unexplained thromboembolism may have occult visceral malignancy; thus, mucin tumor markers may indicate the origin of a mucin-producing carcinoma, and postmortem examination may play an important role in revealing the hidden malignancy.

  19. Cholangiocarcinoma accompanied by desmoid-type fibromatosis

    Institute of Scientific and Technical Information of China (English)

    Nan Xu; Zhe-Yu Chen; Lu-NanYan; Jia-YingYang; Wen-TaoWang; Shu-Guang Jing


    BACKGROUND: Cholangiocarcinoma complicated by intra-abdominal desmoid-type fibromatosis (DTF) is uncommon. There are no reports on patients with this type of fibromatosis, in which the pre-operative treatment (including diagnosis), surgical approach, post-operative pathologic reports, and prognosis are discussed. METHOD: The clinicopathological features of a 49-year-old man were retrospectively analyzed. RESULTS: Cholangiocarcinoma located in the inferior segment of the bile duct was considered pre-operatively on the basis of clinical findings. At the time of pancreaticoduodenectomy, the mesojejunum was stiff without nodules or a mass at a distance of approximately 80 cm from the ligament of Treitz. Complete excision of the entire lesion of the intestinal mesenteric contracture and its subsidiary was performed. Post-operative pathologic findings confirmed an adenocarcinoma located at the extremity of the common bile duct and infiltrating the full thickness of the common bile duct as well as the deep muscular layer of the duodenum. The contracted jejunal mesentery was shown to have DTF. The patient was alive with no evidence of recurrence after a follow-up of 6 months. CONCLUSIONS: The patient had a rare hereditary disease with intra-abdominal DTF, which manifests the characteristics of an aggressive growth pattern and a high rate of local recurrence;conservative therapy is recommended. Complete excision of the fibromatous lesion during pancreaticoduodenectomy may maximally decrease the risk of local recurrence.

  20. Excretory/secretory products of the carcinogenic liver fluke are endocytosed by human cholangiocytes and drive cell proliferation and IL6 production. (United States)

    Chaiyadet, Sujittra; Smout, Michael; Johnson, Michael; Whitchurch, Cynthia; Turnbull, Lynne; Kaewkes, Sasithorn; Sotillo, Javier; Loukas, Alex; Sripa, Banchob


    Liver fluke infection caused by Opisthorchis viverrini remains a major public health problem in many parts of Asia including Thailand, Lao PDR, Vietnam and Cambodia, where there is a strikingly high incidence of cholangiocarcinoma (CCA - hepatic cancer of the bile duct epithelium). Among other factors, uptake of O. viverrini excretory/secretory products (OvES) by biliary epithelial cells has been postulated to be responsible for chronic inflammation and proliferation of cholangiocytes, but the mechanisms by which cells internalise O. viverrini excretory/secretory products are still unknown. Herein we incubated normal human cholangiocytes (H69), human cholangiocarcinoma cells (KKU-100, KKU-M156) and human colon cancer (Caco-2) cells with O. viverrini excretory/secretory products and analysed the effects of different endocytic inhibitors to address the mechanism of cellular uptake of ES proteins. Opisthorchis viverrini excretory/secretory products was internalised preferentially by liver cell lines, and most efficiently/rapidly by H69 cells. There was no evidence for trafficking of ES proteins to cholangiocyte organelles, and most of the fluorescence was detected in the cytoplasm. Pretreatment with clathrin inhibitors significantly reduced the uptake of O. viverrini excretory/secretory products, particularly by H69 cells. Opisthorchis viverrini excretory/secretory products induced proliferation of liver cells (H69 and CCA lines) but not intestinal (Caco-2) cells, and proliferation was blocked using inhibitors of the classical endocytic pathways (clathrin and caveolae). Opisthorchis viverrini excretory/secretory products drove IL6 secretion by H69 cells but not Caco-2 cells, and cytokine secretion was significantly reduced by endocytosis inhibitors. This the first known study to address the endocytosis of helminth ES proteins by host epithelial cells and sheds light on the pathways by which this parasite causes one of the most devastating forms of cancer in south

  1. Identification of Cancer Stem Cell Subpopulations of CD34+ PLC/PRF/5 That Result in Three Types of Human Liver Carcinomas (United States)

    Park, Su Cheol; Nguyen, Ngoc Tue; Eun, Jong Ryeol; Zhang, Yanling; Jung, Yong Jin; Tschudy-Seney, Benjamin; Trotsyuk, Artem; Lam, Alexander; Ramsamooj, Rajendra; Zhang, Yanghong; Theise, Neil D.; Zern, Mark A.


    CD34+ stem cells play an important role during liver development and regeneration. Thus, we hypothesized that some human liver carcinomas (HLCs) might be derived from transformed CD34+ stem cells. Here, we determined that a population of CD34+ cells isolated from PLC/PRF/5 hepatoma cells (PLC) appears to function as liver cancer stem cells (LCSCs) by forming HLCs in immunodeficient mice with as few as 100 cells. Moreover, the CD34+ PLC subpopulation cells had an advantage over CD34− PLCs at initiating tumors. Three types of HLCs were generated from CD34+ PLC: hepatocellular carcinomas (HCCs); cholangiocarcinomas (CC); and combined hepatocellular cholangiocarcinomas (CHCs). Tumors formed in mice transplanted with 12 subpopulations and 6 progeny subpopulations of CD34+ PLC cells. Interestingly, progenies with certain surface antigens (CD133, CD44, CD90, or EPCAM) predominantly yielded HCCs. CD34+ PLCs that also expressed OV6 and their progeny OV6+ cells primarily produced CHC and CC. This represents the first experiment to demonstrate that the OV6+ antigen is associated with human CHC and CC. CD34+ PLCs that also expressed CD31 and their progeny CD31+ cells formed CHCs. Gene expression patterns and tumor cell populations from all xenografts exhibited diverse patterns, indicating that tumor-initiating cells (TICs) with distinct antigenic profiles contribute to cancer cell heterogeneity. Therefore, we identified CD34+ PLC cells functioning as LCSCs generating three types of HLCs. Eighteen subpopulations from one origin had the capacity independently to initiate tumors, thus functioning as TICs. This finding has broad implications for better understanding of the multistep model of tumor initiation and progression. Our finding also indicates that CD34+ PLCs that also express OV6 or CD31 result in types of HLCs. This is the first report that PLC/PRF/5 subpopulations expressing CD34 in combination with particular antigens defines categories of HLCs, implicating a

  2. Identification of cancer stem cell subpopulations of CD34(+) PLC/PRF/5 that result in three types of human liver carcinomas. (United States)

    Park, Su Cheol; Nguyen, Ngoc Tue; Eun, Jong Ryeol; Zhang, Yanling; Jung, Yong Jin; Tschudy-Seney, Benjamin; Trotsyuk, Artem; Lam, Alexander; Ramsamooj, Rajendra; Zhang, Yanghong; Theise, Neil D; Zern, Mark A; Duan, Yuyou


    CD34(+) stem cells play an important role during liver development and regeneration. Thus, we hypothesized that some human liver carcinomas (HLCs) might be derived from transformed CD34(+) stem cells. Here, we determined that a population of CD34(+) cells isolated from PLC/PRF/5 hepatoma cells (PLC) appears to function as liver cancer stem cells (LCSCs) by forming HLCs in immunodeficient mice with as few as 100 cells. Moreover, the CD34(+) PLC subpopulation cells had an advantage over CD34(-) PLCs at initiating tumors. Three types of HLCs were generated from CD34(+) PLC: hepatocellular carcinomas (HCCs); cholangiocarcinomas (CC); and combined hepatocellular cholangiocarcinomas (CHCs). Tumors formed in mice transplanted with 12 subpopulations and 6 progeny subpopulations of CD34(+) PLC cells. Interestingly, progenies with certain surface antigens (CD133, CD44, CD90, or EPCAM) predominantly yielded HCCs. CD34(+) PLCs that also expressed OV6 and their progeny OV6(+) cells primarily produced CHC and CC. This represents the first experiment to demonstrate that the OV6(+) antigen is associated with human CHC and CC. CD34(+) PLCs that also expressed CD31 and their progeny CD31(+) cells formed CHCs. Gene expression patterns and tumor cell populations from all xenografts exhibited diverse patterns, indicating that tumor-initiating cells (TICs) with distinct antigenic profiles contribute to cancer cell heterogeneity. Therefore, we identified CD34(+) PLC cells functioning as LCSCs generating three types of HLCs. Eighteen subpopulations from one origin had the capacity independently to initiate tumors, thus functioning as TICs. This finding has broad implications for better understanding of the multistep model of tumor initiation and progression. Our finding also indicates that CD34(+) PLCs that also express OV6 or CD31 result in types of HLCs. This is the first report that PLC/PRF/5 subpopulations expressing CD34 in combination with particular antigens defines categories of

  3. CRM-1 knockdown inhibits extrahepatic cholangiocarcinoma tumor growth by blocking the nuclear export of p27Kip1. (United States)

    Luo, Jian; Chen, Yongjun; Li, Qiang; Wang, Bing; Zhou, Yanqiong; Lan, Hongzhen


    Cholangiocarcinoma is a deadly disease which responds poorly to surgery and conventional chemotherapy or radiotherapy. Early diagnosis is difficult due to the anatomical and biological characteristics of cholangiocarcinoma. Cyclin-dependent kinase inhibitor 1B (p27Kip1) is a cyclin‑dependent kinase inhibitor and in the present study, we found that p27Kip1 expression was suppressed in the nucleus and increased in the cytoplasm in 53 samples of cholangiocarcinoma from patients with highly malignant tumors (poorly-differentiated and tumor-node-metastsis (TNM) stage III-IV) compared with that in samples from 10 patients with chronic cholangitis. The expression of phosphorylated (p-)p27Kip1 (Ser10), one of the phosphorylated forms of p27Kip1, was increased in the patient samples with increasing malignancy and clinical stage. Coincidentally, chromosome region maintenance 1 (CRM-1; also referred to as exportin 1 or Xpo1), a critical protein responsible for protein translocation from the nucleus to the cytoplasm, was also overexpressed in the tumor samples which were poorly differentiated and of a higher clinical stage. Through specific short hairpin RNA (shRNA)-mediated knockdown of CRM-1 in the cholangiocarcinoma cell line QBC939, we identified an elevation of cytoplasmic p27Kip1 and a decrease of nuclear p27Kip1. Furthermore, the viability and colony formation ability of QBC939 cells was largely reduced with G1 arrest. Consistent with the findings of the in vitro experiments, in a xenograft mouse model, the tumors formed in the CRM-1 knockdown group were markedly smaller and weighed less than those in the control group in vivo. Taken together, these findings demonstrated that the interplay between CRM-1 and p27Kip1 may provide potentially potent biomarkers and functional targets for the development of future cholangiocarcinoma treatments.

  4. 人胆管癌CD24+ CD44+ EpCAMhigh细胞亚群的分选及其肿瘤干细胞样特性的鉴定%Sorting of CD24+ CD44+ EpCAMhigh subset cells in human human cholangiocardnoma and the identificantion of their cancer stem cell-like properties

    Institute of Scientific and Technical Information of China (English)

    朱峰; 王敏; 秦仁义; 申铭; 王欣; 田锐; 江建新; 石程剑


    Objective To sort CD24+ CD44+ EpCAMhigh subset cells in human cholangiocarcinoma and identify their cancer stem cell-like properties. Methods The expression and rate of CD24, CD44 and EpCAM in 6 cases of human cholangiocacinomas were assayed by flow cytometry. The fresh specimens from two cases of cholangiocarcinoma were obtained and implanted subcutaneously into NOD/SCID mice for the establishment of xenografts model. CD24+ CD44+ EpCAMhigh subset cells were sorted from xenografts by flow cytometry and their tumorigenic potential, self-renewal ability and differentiation ability were assessed. Results The expression rate of CD24+ CD44+ EpCAMhigh cells ranged from 0. 58% to 2.43% (mean= 0. 94% ) in 6 cholangiocarcinoma specimens and 2 xenografts. CD24+ CD44+ EpCAMhigh subset cells sorted from 2 xenografts were found to be highly tumorigenic in NOD/SCID mice. CD24+ CD44+ EpCAMhigh cells consistently formed tumors with 1000 cells in 3/8 mice. In contrast, CD24+ CD44+ EpCAMhigh tumor cells were less tumorigenic and formed tumors with 50 000 cells in 1/8 mice. CD24+ CD44+ EpCAMhigh cells were passaged in NOD/SCID mice and formed tumors that recapitulated the histological features and heterogeneity of the original patient tumor. Conclusion CD24+ CD44+ EpCAMhigh subset cells were discriminated in human cholangiocarcinoma, and they had highly tumorigenic, self-renewal ability and differentiation ability. It was first confirmed that CD24+ CD44+ EpCAMhigh cells may be human cholangiocarcinoma cancer stem cells.%目的 检测及分选人胆管癌中的CD24+ CD44+ EpCAMhigh细胞亚群,探讨其是否具有肿瘤干细胞样生物学特性.方法 流式细胞术检测6例人胆管癌中CD24、CD44、EpCAM的表达率;取2例人胆管癌新鲜标本种植到NOD/SCID鼠皮下,建立荷人胆管癌小鼠模型.流式细胞术分选CD24+ CD44+ EpCAMhigh亚群细胞,NOD/SCID鼠移植瘤试验鉴定其成瘤和分化能力.结果 6例人胆管癌组织标本和2例移植瘤中,CD24

  5. Targeting hexokinase II as a possible therapy for cholangiocarcinoma. (United States)

    Thamrongwaranggoon, Ubonrat; Seubwai, Wunchana; Phoomak, Chatchai; Sangkhamanon, Sakkarn; Cha'on, Ubon; Boonmars, Thidarat; Wongkham, Sopit


    Overexpression of hexokinase 2 (HKII) has been demonstrated in various cancers. A number of in vitro and in vivo studies in several cancers show the significance of HKII in many cellular processes including proliferation, metastasis and apoptosis. However, the role of HKII in Opisthorchis viverrini (Ov) associated cholangiocarcinoma (CCA) is still unknown. In the present study, the expression and roles of HKII were determined in Ov associated CCA. The expression of HKII was investigated in 82 patients with histologically proven CCAs by immunohistochemistry. HKII was distinctively expressed in CCA tissues. It was rarely expressed in normal bile duct epithelium, but was expressed in hyperplastic/dysplastic and in 82% of CCA bile ducts. The observation was confirmed in the Ov associated hamster model. Suppression of HKII expression using siRNA significantly decreased cell proliferation, migration and invasion of CCA cell lines. Similar results were obtained using lonidamine (LND), an inhibitor of HK. LND significantly inhibited growth of 4 CCA cell lines tested in dose and time dependent fashion. Comparison the cytotoxic effects of LND and siRNA-HKII suggests the off target of LND above 100 μM. In addition, LND in non-cytotoxic doses could suppress migration and invasion of CCA cells. These results indicate the association of HKII in cholangiocarcinogenesis and progression and suggest the possibility of HKII as a therapeutic target for CCA.

  6. γ-射线诱导癌细胞凋亡致敏树突状细胞后的免疫应答%Immune Responses of Dendritic Cells Loaded with Antigens from Apoptotic Cholangiocarcinoma Cells Caused by γ-Irradation

    Institute of Scientific and Technical Information of China (English)

    吴刚; 韩本立; 裴雪涛


    Objective To investigate the induction cytotoxic T cells (CTLs) with antitumor activity and therapeutic efficacy after dendritic cells(DCs) acquired antigen from apoptotic cholangiocarcinoma cells caused by γ-irradiation.Methods DCs from peripheral blood mononuclear cells (PBMC) that maintain the antigen capturing and processing capacity charateristicof immature cells have been established in vitro, using granulocyte macrophage colony stimulating factor (GM-CSF) and interleukin-4 (IL-4). Then, in cholangiocarcinoma cells apoptosis was induced by γ-irradiation. The experimental groups were as follows: (1) coculture ofDCs and apoptotic cancer cells and T cells; (2) coculture of DCs and necrotic cancer cells and T cells; (3) coculture of DCs, culturedcancer cell and T cells. They are cocultured for 7 days. DCs and T cells were riched, isolated and their antitumor response was tested.Results The cells had typical dendritic morphology, expressed high levels of CD1a and B7, acquired antigen from apoptotic cells causedby γ-irradiation and induced an increased T cell stimulatory capacity in mixed lymphocyte reactions (MLR).Conclusion DCs obtained from PBMCs using GM-CSF and IL-4 can efficiently present antigen derived from apoptotic cells caused by γ-irradiation and efficiently induce T cells. This strategy, therefore, may present an effective approach to transduce DCs with antigen.%目的观察树突状细胞(DCs)从γ-射线诱导的凋亡胆管癌细胞获取抗原后,抗肿瘤免疫应答及对胆管癌细胞的特异性免疫杀伤效果.方法用粒-巨噬细胞集落刺激因子(GM-CSF)加白介素-4(IL-4)从人外周血分化、诱导DCs,γ-射线在体外诱导培养的人胆管癌细胞凋亡,将DCs、T淋巴细胞和凋亡胆管癌细胞共培养,同时设计不同类型肿瘤细胞(坏死胆管癌细胞及培养胆管癌细胞)作对照,7d后,分离、富集DCs、T淋巴细胞进行免疫应答及肿瘤细胞杀伤试验.结果与凋亡胆管癌细胞共

  7. Trefoil factors: Tumor progression markers and mitogens via EGFR/MAPK activation in cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Kanuengnuch Kosriwong; Trevelyan R Menheniott; Andrew S Giraud; Patcharee Jearanaikoon; Banchob Sripa; Temduang Limpaiboon


    AIM: To investigate trefoil factor (TFF ) gene copy number, mRNA and protein expression as potential biomarkers in cholangiocarcinoma (CCA). METHODS: TFF mRNA levels, gene copy number and protein expression were determined respectively by quantitative reverse transcription polymerase chain reaction (PCR), quantitative PCR and immunohistochemistry in bile duct epithelium biopsies collected from individuals with CCA, precancerous bile duct dysplasia and from disease-free controls. The functional impact of recombinant human (rh)TFF2 peptide treatment on proliferation and epidermal growth factor receptor (EGFR)/mitogenactivated protein kinase (MAPK) signaling was assessed in the CCA cell line, KMBC, by viable cell counting and immunoblotting, respectively. RESULTS: TFF1 , TFF2 and TFF3 mRNA expression was significantly increased in CCA tissue compared to disease-free controls, and was unrelated to gene copy number. TFF1 immunoreactivity was strongly increased in both dysplasia and CCA, whereas TFF2 immunoreactivity was increased only in CCA compared to diseasefree controls. By contrast, TFF3 immunoreactivity was moderately decreased in dysplasia and further decreased in CCA. Kaplan-Meier analysis found no association of TFF mRNA, protein and copy number with age, gender, histological subtype, and patient survival time. Treatment of KMBC cells with rhTFF2 stimulated proliferation, triggered phosphorylation of EGFR and downstream extracellular signal related kinase (ERK), whereas co-incubation with the EGFR tyrosine kinase inhibitor, PD153035, blocked rhTFF2-dependent proliferation and EGFR/ERK responses. CONCLUSION: TFF mRNA/protein expression is indicative of CCA tumor progression, but not predictive for histological sub-type or survival time. TFF2 is mitogenic in CCA via EGFR/MAPK activation.

  8. Current Status on Cholangiocarcinoma and Gallbladder Cancer (United States)

    Ebata, Tomoki; Ercolani, Giorgio; Alvaro, Domenico; Ribero, Dario; Di Tommaso, Luca; Valle, Juan W.


    Background Cholangiocarcinomas (CC) as well as gallbladder cancers are relatively rare and intractable diseases. Clinical, pathological, and epidemiological studies on these tumors have been under investigation. The current status and/or topics on biliary tract cancers have been reported in the East West Association of Liver Tumor (EWALT), held in Milano, Italy in 2015. Summary All the authors, herein, specifcally reported the current status and leading-edge findings on biliary tract cancers as the following sequence: epidemiology of CC, surgical therapy for intrahepatic CC, surgical therapy for perihilar CC, surgical therapy for gallblad der cancer, chemotherapy for biliary tract cancers, and new histological features in CC. Key Message The present review article will update the knowledge on biliary tract cancers, en hancing the quality of daily clinical practice. However, many features about these cancers remain unknown; further studies are required to establish disease-specific optimal treatment strategies. PMID:27995089

  9. Combined hepatocellular-cholangiocarcinoma in a Yellow-headed Amazon (Amazona oratrix). (United States)

    Tennakoon, Anusha Hemamali; Izawa, Takeshi; Fujita, Daisuke; Denda, Yuki; Seto, Eiko; Sasai, Hiroshi; Kuwamura, Mitsuru; Yamate, Jyoji


    A 9-year-old male Yellow-headed Amazon (Amazona oratrix) with a history of anorexia and vomiting died of a liver tumor. The tumor consisted of neoplastic cells with hepatocellular and cholangiocellular differentiations and their intermingled areas. Neoplastic hepatocytes showed islands or trabecular growth with vacuolated eosinophilic cytoplasm. Cells showing biliary differentiation formed ducts or tubules lined by cytokeratin AE1/AE3-positive epithelia, accompanied by desmoplasia consisting of myofibroblasts reacting to α-smooth muscle actin and desmin. The tumor was diagnosed as a combined hepatocellular-cholangiocarcinoma, which is very rare in the avian.

  10. Genome engineering in human cells. (United States)

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


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

  11. In-vivo monitoring of development of cholangiocarcinoma induced with C. sinensis and N-nitrosodimethylamine in Syrian golen hamsters using ultrasonography and magnetic resonance imaging: a preliminary study

    Energy Technology Data Exchange (ETDEWEB)

    Woo, Hyunsik [SMG-SNU Boramae Medical Center, Department of Radiology, Seoul (Korea, Republic of); Han, Joon Koo; Kim, Jung Hoon [Seoul National University Hospital, Department of Radiology, Seoul (Korea, Republic of); Hong, Sung-Tae [Seoul National University, Department of Parasitology, College of Medicine, Seoul (Korea, Republic of); Uddin, M.H. [Seoul National University, Adult Stem Cell Research Center, Laboratory of Stem Cell and Tumor Biology, Department of Veterinary Public Health, College of Veterinary Medicine, Seoul (Korea, Republic of); Jang, Ja-June [Seoul National University, Department of Pathology, College of Medicine, Seoul (Korea, Republic of)


    The purpose of this study is to evaluate high-resolution ultrasound and magnetic resonance imaging (MRI) in monitoring of cholangiocarcinoma in the hamsters with C. sinensis infection and N-nitrosodimethylamine (NDMA). Twenty-four male Syrian golden hamsters of were divided into four groups composed of five hamsters as control, five hamsters receiving 30 metacercariae of C. sinensis per each hamster, five hamsters receiving NDMA in drinking water, and nine hamsters receiving both metacercariae and NDMA. Ultrasound was performed every other week from baseline to the 12th week of infection. MRI and histopathologic examination was done from the 4th week to 12th week. Cholangiocarcinomas appeared as early as the 6th week of infection. There were 12 cholangiocarcinomas, nine and ten of which were demonstrated by ultrasound and MRI, respectively. Ultrasound and MRI findings of cholangiocarcinomas in the hamsters were similar to those of the mass-forming intrahepatic cholangiocarcinomas in humans. Ultrasound and MRI also showed other findings of disease progression such as periductal increased echogenicity or signal intensity, ductal dilatation, complicated cysts, and sludges in the gallbladder. High-resolution ultrasound and MRI can monitor and detect the occurrence of cholangiocarcinoma in the hamsters non-invasively. (orig.)

  12. Different carcinogenic process in cholangiocarcinoma cases epidemically developing among workers of a printing company in Japan. (United States)

    Sato, Yasunori; Kubo, Shoji; Takemura, Shigekazu; Sugawara, Yasuhiko; Tanaka, Shogo; Fujikawa, Masahiro; Arimoto, Akira; Harada, Kenichi; Sasaki, Motoko; Nakanuma, Yasuni


    Recently, cholangiocarcinoma has epidemically developed among young adult workers of a printing company in Japan. Exposure to organic solvents including 1,2-dichloropropane and/or dichloromethane is supposed to be associated with the carcinoma development. The metabolism of dichloromethane proceeds through a Theta-class glutathione S-transferase (GST) T1-1-catalyzed pathway, where its reactive intermediates have been implicated in genotoxicity and carcinogenicity. This study examined features of the carcinogenic process of the cholangiocarcinoma developed in the printing company. Surgically resected specimens of the cholangiocarcinoma cases were analyzed, where all cases were associated with precursor lesions such as biliary intraepithelial neoplasia (BilIN) and/or intraductal papillary neoplasm of the bile duct (IPNB). Immunohistochemical analysis confirmed constitutional expression of GST T1-1 in normal hepatobiliary tract. Immunostaining of γ-H2AX, a marker of DNA double strand break, showed that its expression was significantly increased in foci of BilIN, IPNB and invasive carcinoma as well as in non-neoplastic biliary epithelial cells of the printing company cases when compared to that of control groups. In the printing company cases, immunohistochemical expression of p53 was observed in non-neoplastic biliary epithelial cells and BilIN-1. Mutations of KRAS and GNAS were detected in foci of BilIN in one out of 3 cases of the printing company. These results revealed different carcinogenic process of the printing company cases, suggesting that the exposed organic solvents might act as a carcinogen for biliary epithelial cells by causing DNA damage, thereby contributing to the carcinoma development.

  13. Combination of Praziquantel and Aspirin Minimizes Liver Pathology of Hamster Opisthorchis viverrini Infection Associated Cholangiocarcinoma. (United States)

    Sudsarn, Pakkayanee; Boonmars, Thidarut; Ruangjirachuporn, Wipaporn; Namwat, Nisana; Loilome, Watcharin; Sriraj, Pranee; Aukkanimart, Ratchadawan; Nadchanan, Wonkchalee; Jiraporn, Songsri


    Opisthorchiasis is one of the major risk factors for cholangiocarcinoma (CCA) in northeastern Thailand. An effective drug for killing this parasite is praziquantel. Recently, several reports have shown that with frequent use, praziquantel may itself be a CCA risk and can cause liver cell damage from an immunopathological response after parasite death. Aspirin has many properties including anti-inflammation and anti-cancer. Therefore, we use of aspirin (As) and praziquantel (Pz) to improve hepatobiliary system function in hamsters infected with Opisthorchis viverrini (OV) and or administered N-nitrosodimethylamine (ND). Livers of OVNDAsPz, appeared healthy macroscopically, suggesting slow progression of cholangiocarcinoma evident by extent of fibrosis and bile duct cell proliferation was less than OVND although aggregations of inflammatory cells remained. Proliferating cell nuclear antigen (PCNA), cytokeratin 19 (CK19), and cancer antigen (CA19-9) staining were strongly positive in OVND, but were only slight in OVNDAs. Moreover, OVNDAsPz, appeared a few inflammatory infiltrations, bile duct proliferation, fibrosis and CCA area than the OVNDAs group. Thirty seven point five percent of hamster in this group could not develop CCA. These findings suggest that using aspirin combination with praziquantel treatment can improve the hepatobiliary system after O. viverrini infection and reduce the risk of CCA.

  14. Imaging and interventions in hilar cholangiocarcinoma: A review

    Institute of Scientific and Technical Information of China (English)

    Kumble; Seetharama; Madhusudhan; Shivanand; Gamanagatti; Arun; Kumar; Gupta


    Hilar cholangiocarcinoma is a common malignant tumor of the biliary tree. It has poor prognosis with very low 5-year survival rates. Various imaging modalities are available for detection and staging of the hilar cholangiocarcinoma. Although ultrasonography is the initial investigation of choice, imaging with contrast enhanced computed tomography scan or magnetic resonance imaging is needed prior to management. Surgery is curative wherever possible. Radiological interventions play a role in operable patients in theform of biliary drainage and/or portal vein embolization. In inoperable cases, palliative interventions include biliary drainage, biliary stenting and intra-biliary palliative treatment techniques. Complete knowledge of application of various imaging modalities available and about the possible radiological interventions is important for a radiologist to play a critical role in appropriate management of such patients.We review the various imaging techniques and appearances of hilar cholangiocarcinoma and the possible radiological interventions.

  15. The role of liver transplantation for hilar cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Durgatosh Pandey; Kang-Hoe Lee; Kai-Chah Tan


    BACKGROUND:Hilar cholangiocarcinoma is a devastating disease. Surgery is the only potentially curative modality. However, the results of surgical resection for hilar cholangiocarcinomas are disappointing. The introduction of liver transplantation for this condition has brought new hope for the management of this disease. The aim of this review is to discuss the role of liver transplantation in this disease. DATA SOURCES:A MEDLINE search was conducted for the articles on liver transplantation for hilar cholangiocarcinoma. Their results have been compiled and compared with the existing literature on resection for this disease. RESULTS:The earlier series on liver transplantation for hilar cholangiocarcinoma were not encouraging because of poor patient selection. The Mayo Clinic protocol of neoadjuvant chemoradiation followed by liver transplantation has shown remarkable success (survival at 1-, 3-, and 5-year post-transplantation being 92%, 82%, and 82%, respectively). With better patient selection and integration of neoadjuvant chemoradiation, the long-term survival is superior to that of the patients who undergo resection, as shown by the published literature on resection. The limitations of organ availability can be overcome by the living donor liver transplantation programme. This review article discusses the rationale, pros and cons of liver transplantation vis-à-vis resection for hilar cholangiocarcinoma. CONCLUSIONS:Liver transplantation, especially living donor liver transplantation, is a new and exciting alternative to resection for hilar cholangiocarcinoma. Integration of neoadjuvant chemoradiation has the potential to further improve the curative potential of liver transplantation. The strategy of combining neoadjuvant chemoradiation and liver transplantation brings new hope for the treatment of this dififcult disease.

  16. Diffusion inside living human cells

    DEFF Research Database (Denmark)

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


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

  17. Intrahepatic cholangiocarcinoma: current management and emerging therapies. (United States)

    Rahnemai-Azar, Amir A; Weisbrod, Allison B; Dillhoff, Mary; Schmidt, Carl; Pawlik, Timothy M


    Intrahepatic cholangiocarcinoma (iCCA) is a malignancy with an increasing incidence and a high-case fatality. While surgery offers the best hope at long-term survival, only one-third of tumors are amenable to surgical resection at the time of the diagnosis. Unfortunately, conventional chemotherapy offers limited survival benefit in the management of unresectable or metastatic disease. Recent advances in understanding the molecular pathogenesis of iCCA and the use of next-generation sequencing techniques have provided a chance to identify 'target-able' molecular aberrations. These novel molecular therapies offer the promise to personalize therapy for patients with iCCA and, in turn, improve the outcomes of patients. Area covered: We herein review the current management options for iCCA with a focus on defining both established and emerging therapies. Expert commentary: Surgical resection remains as an only hope for cure in iCCA patients. However, frequently the diagnosis is delayed till advanced stages when surgery cannot be offered; signifying the urge for specific diagnostic tumor biomarkers and targeted therapies. New advances in genomic profiling have contributed to a better understanding of the landscape of molecular alterations in iCCA and offer hope for the development of novel diagnostic biomarkers and targeted therapies.

  18. Characterization of CD133~+ parenchymal cells in the liver: Histology and culture

    Institute of Scientific and Technical Information of China (English)

    Seiichi Yoshikawa; Yoh Zen; Takahiko Fujii; Yasunori Sato; Tetsuo Ohta; Yutaka Aoyagi; Yasuni Nakanuma


    AIM: To reveal the characteristics of CD133~+ cells in the liver.METHODS: This study examined the histological characteristics of CD133~+ cells in non-neoplastic and neoplastic liver tissues by immunostaining, and also analyzed the biological characteristics of CD133~+ cells derived from human hepatocellular carcinoma (HCC) or cholangiocarcinoma cell lines.RESULTS: Immunostaining revealed constant expression of CD133 in non-neoplastic and neoplastic biliary epithelium, and these cells had the immunophenotype CD133~+/CK19+/HepPar-1~-. A small number of CD133~+/CK19~-/HepPar-1~+ cells were also identified in HCC and combined hepatocellular and cholangiocarcinoma. In addition, small ductal structures, resembling the canal of Hering, partly surrounded by hepatocytes were positive for CD133. CD133 expression was observed in three HCC (HuH7, PLC5 and HepG2) and two cholangiocarcinoma cell and HepG2) and two cholangiocarcinoma cell lines (HuCCT1 and CCKS1). Fluorescence-activated cell sorting (FACS) revealed that CD133~+ and CD133~-cells derived from HuH7 and HuCCT1 cells similarly produced CD133~+ and CD133~-cells during subculture. To examine the relationship between CD133~+ cells and the side population (SP) phenotype, FACS was performed using Hoechst 33342 and a monoclonal antibody against CD133. The ratios of CD133~+/CD133~-cells were almost identical in the SP and non-SP in HuH7. In addition, four different cellular populations (SP/CD133~+, SP/CD133~-, non-SP/CD133~+, and non-SP/CD133~-) could similarly produce CD133~+ and CD133~-cells during subculture. CONCLUSION: This study revealed that CD133 could be a biliary and progenitor cell marker in vivo. However, CD133 alone is not sufficient to detect tumor-initiating cells in cell lines.

  19. Sensitivity-increasing effect of recommbinant analgesic-antitumor peptide on cholangiocarcinoma cells to chemotherapy%重组东亚钳蝎镇痛抗肿瘤肽对胆管癌细胞化疗的增敏作用

    Institute of Scientific and Technical Information of China (English)

    葛贤秀; 曹鹏; 缪林; 李全鹏; 汪婷婷; 于泓; 季国忠


    目的 探讨重组东亚钳蝎镇痛抗肿瘤肽(rAGAP)对胆管癌细胞化疗的增敏作用及其机制.方法 体外培养人胆管癌细胞株HuCC-T1、QBC939和胆管上皮细胞HIBepic,单独用药:不同浓度的rAGAP、顺铂(DDP)和5-氟尿嘧啶(5-FU)处理三株细胞,四甲基偶氮唑盐比色法(MTT法)观察细胞的增殖抑制率及半抑制浓度(IC50).联合用药:选用rAGAP(5μg/ml)、DDP(4μg/ml)、5-FU(50μg/ml)处理三株细胞,分为对照(A)组、rAGAP(B)组、DDP(C)组、rAGAP与DDP联用(D)组、5-FU(E)组、rAGAP与5-FU联用(F)组.MTT法检测细胞存活率;荧光定量PCR方法检测细胞多药耐药基因1(MDR-1)及多药耐药相关蛋白1(MRP-1)的mRNA表达.结果 单独用药结果显示rAGAP抑制细胞增殖呈时间剂量依赖性,且对癌细胞具有一定的选择性;rAGAP组的IC50值明显小于DDP组和5-FU组(P<0.01).联合用药结果显示,胆管癌细胞株HuCC-T1和QBC939中,D组较B、C组以及F组较B、E组细胞存活减少;而且MDR-1和MRP-1表达下调(P<0.01).而在胆管上皮细胞HIBepic细胞系里,联合用药并未表现明显的增加化疗药效果的作用.结论 rAGAP可选择性抑制胆管癌细胞,与化疗药DDP和5 FU联用可增强化疗敏感性.%Objective To investigate the sensitivity-increasing effect of recommbinant analgesicantitumor peptide (rAGAP)on cholangiocarcinoma cells to chemotherapy and its underlying mechanism.Methods The cholangiocarcinoma cell lines HuCC-T1,QBC939 and bile duct epithelial cells HIBepic were cultured in vitro,and then exposed to rAGAP,cisplatin and 5-FU,respectively.The cells inhibition rate and 50% inhibiting concentration(IC5) were detected.The cells were divided into 6 groups of A(control),B(treated with rAGAP 5 μg/ml),C(treated with DDP 4μg/ml),D(treated with 5-FU 50μg/ml plus DDP),E(treated with 5-FU 50μg/ml) and F(treated with rAGAP plus 5FU).The cell survival rates were detected using MTT method and the mRNA expressions of

  20. Human stromal (mesenchymal) stem cells

    DEFF Research Database (Denmark)

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


    Human stromal (mesenchymal) stem cells (hMSC) represent a group of non-hematopoietic stem cells present in the bone marrow stroma and the stroma of other organs including subcutaneous adipose tissue, placenta, and muscles. They exhibit the characteristics of somatic stem cells of self-renewal and......Human stromal (mesenchymal) stem cells (hMSC) represent a group of non-hematopoietic stem cells present in the bone marrow stroma and the stroma of other organs including subcutaneous adipose tissue, placenta, and muscles. They exhibit the characteristics of somatic stem cells of self...... of clinical applications, e.g., non-healing bone fractures and defects and also non-skeletal degenerative diseases like heart failure. Currently, the numbers of clinical trials that employ MSC are increasing. However, several biological and biotechnological challenges need to be overcome to benefit from...

  1. Cholangiocarcinoma: A compact review of the literature

    Institute of Scientific and Technical Information of China (English)

    Yucel Ustundag; Yusuf Bayraktar


    Cholangiocarcinoma (CC) is a devastating cancer aris-ing from biliary epithelia. Unfortunately, the incidence of this disease is increasing in Western countries. These tumors progress insidiously, and liver failure, biliary sepsis, malnutrition and cancer cachexia are general modes of death associated with this disease. To dale, no established therapy for advanced dis-ease has been established or validated. However, our knowledge in tumor biology is increasing dramatically and new drugs are under investigation for treatment of this notorious tumor. In clinical practice, there are better diagnostic tools in use to facilitate an earlier diagnosis of CC, at least in those patients with known risk factors. CC is resectable for cure in only a small percentage of patients. Preoperative staging for vas-cular and biliary extension of CC is very important in this tumor. Laparoscopy and recently endosonography seem to protect against unnecessary laparotomies in these patients. During the last 15 years, aggressive surgical approaches, including combined liver resec-tions and vascular reconstructive surgical expertise, have improved survival in patients with CC. Surgery is contraindicated in CC cases having primary sclerosing cholangitis (PSC). Although CC was previously consid-ered a contraindication to liver transplantation, new cautious protocols, including neo-adjuvant chemora-diation therapies and staging procedures before the transplantation, have made it possible to achieve long-term survival after liver transplantation in this disease. New ablative therapies with photodynamic therapy, intraductal high-intensity ultrasonography and chemo- therapy-impregnated plastic biliary endoprosthesis are important steps in the palliative management of extra-hepatic CCs. Radiofrequency and chemo-embolization methods are also applicable for intra-hepatic CCs as palliative modes of treatment. We need more prospec-tive randomized controlled trials to evaluate the role of the new

  2. Combining biological agents and chemotherapy in the treatment of cholangiocarcinoma

    DEFF Research Database (Denmark)

    Jensen, Lars Henrik; Jakobsen, Anders


    is not always possible. Chemotherapy is effective and the combination of cisplatin and gemcitabine is considered a standard treatment of inoperable cholangiocarcinoma. Biological targeted treatment to date has minor effect when given as monotherapy, but some of the drugs hold promise as an adjunct...

  3. Human embryonic stem cells handbook

    Directory of Open Access Journals (Sweden)

    Carlo Alberto Redi


    Full Text Available After the Nobel prize in physiology or medicine was awarded jointly to Sir John Gurdon and Shinya Yamanaka for the discovery that mature cells can be reprogrammed to become pluripotent it became imperative to write down the review for a book entirely devoted to human embryonic stem cells (hES, those cells that are a urgent need for researchers, those cells that rekindle the ethical debates and finally, last but not least, those cells whose study paved the way to obtain induced pluripotent stem cells by the OSKC’s Yamanaka method (the OSKC acronim refers, for those not familiar with the topic, to the four stemness genes used to transfect somatic fibroblasts: Oct4, Sox2, Klf4 and c-Myc....

  4. Long-term survival after intraluminal brachytherapy for inoperable hilar cholangiocarcinoma: A case report

    Institute of Scientific and Technical Information of China (English)

    Siu-Yin Chan; Ronnie T. Poon; Kelvin K. Ng; Chi-Leung Liu; Raymond T. Chan; Sheung-Tat Fan


    Surgical resection with a tumor-free margin is the onlycurative treatment for hilar cholangiocarcinoma (Klatskin tumor). However, over half of the patients present late with unresectable tumors. Radiotherapy using external beamirradiation or intraluminal brachytherapy (ILBT) has been used to treat unresectable hilar cholangiocarcinoma with satisfactory outcome. We reported a patient with unresectable hilar cholangiocarcinoma surviving more than 6 years after combined external beam irradiation and ILBT.

  5. Review of endoscopic techniques in the diagnosis and management of cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Katherine Nguyen; James T Sing Jr


    Cholangiocarcinoma is a rare malignancy of the biliary tract. Key factors in determining therapeutic options include knowledge of tumor extent, anatomy and obtaining tissue diagnosis. Endoscopically, there are three modalities available to make the diagnosis of cholangiocarcinoma. These include endoscopic retrograde cholangiopancreatography, endoscopic ultrasound with fine needle aspiration and cholangioscopy. Management of cholangiocarcinoma endoscopically is typically confined to stent placement for palliative purposes or as a bridge to surgery. In this article, we will review the endoscopic techniques available for the diagnosis and management of cholangiocarcinoma.

  6. [Control of Opisthorchis viverrini infection for cholangiocarcinoma prevention]. (United States)

    Buisson, Y


    The International Agency for Research on Cancer (IARC) has classified two liver flukes as carcinogenic to humans (Group 1): Opisthorchis viverrini in 1994 and Clonorchis sinensis in 2009. This review is focused on O. viverrini, the most studied of these two trematodes, which infects nearly 10 million people in Southeast Asia. The life cycle involves two intermediate hosts living in fresh water: a snail of the genus Bithynia and a ciprinid fish. The definitive hosts (human, cat, dog) become infected by ingesting raw fish containing metacercariae, the infective stage of the parasite. Adult flukes attach to the epithelium of the bile ducts where they feed for as long as 10 to 30 years, resulting in chronic inflammation, epithelial hyperplasia, periductal fibrosis and formation of granuloma. For a long asymptomatic, the distomatosis is revealed by a chronic cholangitis when the parasite load becomes high. Complications can occur with time: gallstones, cholangitis, liver abscess, pancreatitis and, after a few decades, cholangiocarcinoma (CCA). The epidemiological correlation between the prevalence of O. viverrini infection and the incidence of CCA has been demonstrated in the northeast of Thailand. Specifically, the Khon Kaen province has the highest incidence rate in the world. The CCA can develop asymptomatically for a long time, especially in intrahepatic locations. It is often discovered at a late stage, unresectable. Its prognosis is dreadful with a survival rate less than 5% at 5 years. The phenomenon of carcinogenesis induced by O. viverrini is multifactorial. It has been specially studied using experimental infection on the Syrian golden hamster. Three intricated mechanisms are involved: (i) the direct damage caused by adult worms on the bile duct epithelium, (ii) the immunopathologic processes related to chronic inflammation (oxidative stress) and (iii) the mitogenic and anti-apoptotic effects of the proteins secreted by the parasite. Exogenous cofactors are

  7. Effects of microRNA-21 on invasion and migration of cholangiocarcinoma cells through regulating epithelial-mesenchymal transition%微小RNA-21通过调控上皮-间充质转化对胆管癌细胞侵袭与转移的影响

    Institute of Scientific and Technical Information of China (English)

    刘振; 黄强; 刘臣海; 林先盛; 谢放; 朱成林; 邵峰


    目的 观察微小RNA-21(miR-21)通过调控上皮-间充质(EMT)转化进程对胆管癌细胞的侵袭转移的影响.方法 构建合成无关序列、miR-21 mimics和miR-21 inhibitor并转染到胆管癌QBC939及RBE细胞中,实验设自然生长组(Cell组)、转染无关序列组(Cell-NC组)、转染miR-21mimics组(Cell-21M组)及转染miR-21 inhibitor组(Cell-21I组).通过实时定量逆转录聚合酶链反应(RT-qPCR)检测miR-21在各组细胞中的表达;采用划痕实验、Transwell侵袭及迁移实验检测各组细胞侵袭力;RT-qPCR、Western blot检测EMT相关分子E-钙黏蛋白(E-cadherin)、N-钙黏蛋白(N-cadherin)及波形蛋白(Vimentin)的mRNA及蛋白表达.结果 在胆管癌QBC939和RBE细胞中,Cell组与Cell-NC组miR-21表达差异无统计学意义(P>0.05),Cell-21M组较Cell组表达增高,而Cell-21I组表达降低(P<0.05).Cell-21M组细胞侵袭迁移能力增加,Cell-21I组细胞侵袭迁移能力减弱(P<0.05).同时,Cell组与Cell-NC组E-cadherin、N-cadherin及Vimentin mRNA及蛋白表达差异无统计学意义(P>0.05),Cell-21M组较Cell组E-cadherin mRNA及蛋白表达降低,N-cadherin及Vimentin mRNA及蛋白表达增高,Cell-21I组E-cadherin mRNA及蛋白表达增高,N-cadherin及Vimentin mRNA及蛋白表达均降低(P<0.05).结论 miR-21能够诱导胆管癌细胞发生EMT转化,促进胆管癌细胞的侵袭与转移.%Objective To discuss wheather microRNA-21 (miR-21) promotes invasion and rnigration in cholangiocarcinoma via inducing epithelial-mesenchymal transition (EMT).Methods Scamble sequence,miR-21 mimics and inhibitor were constructed and transfected into QBC939 and RBE cells.Cell group treated with nothing,Cell-NC group treated with scamble sequence,Cell-21M group treated with miR-21 mimics and Cell-21I group treated with miR-21 inhibitor were respectively designed in this study.The expression of miR-21 in each group was determined by real-time reverse transcription polymerase chain reaction (RT

  8. Human fetal mesenchymal stem cells. (United States)

    O'Donoghue, Keelin; Chan, Jerry


    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

  9. Clinical and biological significance of precursor lesions ofintrahepatic cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)


    Cholangiocarcinoma (CC) is primarily a malignant tumorof older adults most prevalent in Southeast Asia, whereliver fluke infestation is high. However the etiology inwestern countries is unknown. Although the incidence ofextrahepatic cholangiocarcinoma has remained constant,incidence of intrahepatic CC (ICC) which differs inmorphology, pathogenesis, risk factors, treatment andprognosis is increasing. While this increase is associatedwith hepatitis C virus infection, chronic nonalcoholicliver disease, obesity, and smoking, the pathogenesisof ICC and molecular alterations underlying the carcinogenesisare not completely elucidated. Benignbiliary lesions such as biliary intraepithelial neoplasia,intraductal papillary neoplasm of the bile duct, vonMeyenburg complex or bile duct hamartoma, and bileduct adenoma have been associated with ICC. For eachof these entities, evidence suggests or supports a roleas premalignant lesions. This article summarized theimportant biological significance of the precursor lesionsof ICC and the molecular mechanisms that may beinvolved in intrahepatic cholangiocarcinogenesis.

  10. A novel carbohydrate antigen expression during development of Opisthorchis viverrini- associated cholangiocarcinoma in golden hamster: a potential marker for early diagnosis. (United States)

    Sawanyawisuth, Kanlayanee; Silsirivanit, Atit; Kunlabut, Kunlathida; Tantapotinan, Nattawat; Vaeteewoottacharn, Kulthida; Wongkham, Sopit


    Poor prognosis of cholangiocarcinoma (CCA) is primarily due to delayed diagnosis because of the lack of appropriate tumor marker(s) to detect cancer development at an early stage. We have recently established a S121 monoclonal antibody (mAb) which recognizes an unidentified glycan epitope on MUC5AC, designated as CCA-associated carbohydrate antigen (CCA-CA). This antigen is expressed in human CCA cells but not in normal biliary epithelia. Detection of CCA-CA effectively distinguished CCA patients' sera from normal control sera with high specificity and sensitivity. In the present study, we examined a time profile of the expression of CCA-CA by immunohistochemical methods in the liver tissues of Opisthorchis viverrini (Ov)-associated CCA in a hamster model. Hamsters were divided into four groups; non-treated, Ov infected, NDMA (N-nitrosodimethamine) treated and Ov+NDMA treated groups, and animals from each group were euthanized at 1, 3 and 6 months post-treatment. CCA-CA was not detected in normal biliary cells of non-treated hamsters throughout the course of experiment. CCA-CA became detectable in the cytoplasm and apical surface of biliary cells of the NDMA and Ov+NDMA groups at early stage (1 month) of tumor development and increased with tumor progression. In contrast, CCA-CA was detected as nuclear staining at the 1 month post Ov infection and declined thereafter. These results suggest the possibility of CCA-CA as an early marker for CCA.

  11. Preoperative evaluation with T-staging system for hilar cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)


    AIM: To investigate the clinical value of T-staging system in the preoperative assessment of hilar cholangiocarcinoma.METHODS: From March 1993 to January 2006, 85 patients who had cholangiocarcinoma diagnosed by operative tissue-biopsy were placed into one of three stages based on the new T-staging system, and it was evaluated the resectability and survival correlated with T-staging.RESULTS: The likelihood of resection and achieving tumor-free margin decreased progressively with increasing T stage (P < 0.05). The cumulative 1-year survival rates of T1, T2 and T3 patients were 71.8%,50.8% and 12.9% respectively, and the cumulative 3-year survival rate was 34.4%, 18.2% and 0%respectively; the survival of different stage patients differed markedly (P < 0.001). Median survival in the hepatic resection group was greater than in the group that did not undergo hepatic resection (28 mo vs 18 mo;P < 0.05). The overall accuracy for combined MRCP and color Doppler Ultrasonagraphy detecting disease was higher than that of combined using CT and color Doppler Ultrasonagraphy (91.4% vs 68%; P < 0.05 ). And it was also higher in detecting port vein involvement (90% vs54.5%; P < 0.05).CONCLUSION: The proposed staging system for hilar cholangiocarcinoma can accurately predict resectability,the likelihood of metastatic disease, and survival.A concomitant partial hepatectomy would help to attain curative resection and the possibility of longterm survival. MRCP/MRA coupled with color Doppler Ultrasonagraphy was necessary for preoperative evaluation of hilar cholangiocarcinoma.

  12. Orthotopic liver transplantation with hepatopancreatoduodenectomy for hilar cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    HE Xiao-shun; ZHANG Shao; ZHU Xiao-feng; WANG Dong-ping; MA Yi; WANG Guo-dong; JU Wei-qiang; WU Lin-wei; HUANG Jie-fu


    @@ Although the surgical treatment for hilar cholangiocarcinoma represents the only potentially curative option, local excision and major hepatectomy1 have failed to produce a favorable survival figure over the last decade. This is partly due to local tumor recurrences within the resection margin, perineural sheaths infiltration,and the regional lymph nodes metastasis after the surgery.Recently, an extended bile duct resection combined with total hepatectomy, pancreatoduodenectomy, and

  13. Differentiated human stem cells resemble fetal, not adult, β cells. (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


    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.

  14. Sonography-guided percutaneous microwave ablation of intrahepatic primary cholangiocarcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Yu Mingan [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China); Liang Ping, E-mail: [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China); Yu Xiaoling; Cheng Zhigang; Han Zhiyu; Liu Fangyi; Yu Jie [Department of Interventional Ultrasound, Chinese PLA General Hospital, 28 Fuxing Road, Beijing, 100853 (China)


    Objective: To evaluate the efficacy and safety of sonography-guided percutaneous microwave ablation of intrahepatic primary cholangiocarcinoma. Materials and methods: From May 2006 to March 2010, 15 patients (11 men, 4 women; mean age, 57.4 years) with 24 histologically proven intrahepatic primary cholangiocarcinoma lesions (mean tumor size, 3.2 {+-} 1.9 cm; range, 1.3-9.9 cm) were treated with microwave ablation. Results: Thirty-eight sessions were performed for 24 nodules in 15 patients. The follow-up period was 4-31 months (mean, 12.8 {+-} 8.0 months). The ablation success rate, the technique effectiveness rate, and the local tumor progression rate were 91.7% (22/24), 87.5% (21/24), and 25% (6/24) respectively according to the results of follow-up. The cumulative overall 6, 12, 24 month survival rates were 78.8%, 60.0%, and 60.0%, respectively. Major complication occurred including liver abscess in two patients (13.3%) and needle seeding in one patient (6.7%). Both complications were cured satisfied with antibiotic treatment combined to catheter drainage for abscess and resection for needle seeding. The minor complications and side effects were experienced by most patients which subsided with supportive treatment. Conclusion: Microwave ablation can be used as a safe and effective technique to treat intrahepatic primary cholangiocarcinoma.

  15. Survival analysis of cholangiocarcinoma: A 10-year experience in Malaysia

    Institute of Scientific and Technical Information of China (English)

    Ahmad Ramzi Yusoff; Mohd Muzammil Abdul Razak; Yoong Boon Koon; R Vijeyasingam; Siti Zuraidah Mahmud


    AIM: To investigate the clinical features and survival of patients treated for cholangiocarcinoma in our institution and to analyze the factors affecting their survival. METHODS: This retrospective cohort study assessed patients diagnosed with cholangiocarcinoma between January 1997 and December 2007 at the University Malaya Medical Centre in Malaysia. The clinical data and associated outcomes were collected using a structured proforma. RESULTS: Of the 69 patients diagnosed with cholangiocarcinoma, 38 (55%) were male; mean patient age was 61 years. Twelve patients(17%) had intrahepatic, 38 (55%) had perihilar and 19 (28%) had distal tumors. Only 12 patients underwent curative surgery, including seven R0 resections. Only one patient died within 30 d after surgery. The overall median survival was 4 mo, whereas the median survival of R0 resected patients was 16 mo. The overall 1-, 2- and 3-year cumulative survival rates were 67%, 17% and 17%, respectively. Survival rates were significantly associated with curative resection (P= 0.002), intrahepatic tumor (P = 0.003), negative margin status (P = 0.013), early tumor stage (P = 0.016), higher tumor differentiation (P = 0.032) and absence of jaundice (P = 0.038). Multivariate analysis showed that tumor location was a significant independent predictor of patient survival. CONCLUSION: Curative, margin-negative resection of early stage, well-differentiated intrahepatic tumors is associated with improved patient survival.

  16. Liver transplantation for cholangiocarcinoma:Current status and new insights

    Institute of Scientific and Technical Information of China (English)

    Gonzalo; Sapisochín; Elena; Fernández; de; Sevilla; Juan; Echeverri; Ramón; Charco


    Cholangiocarcinoma is a malignant tumor of the biliary system that can be classified into intrahepatic(i CCA),perihiliar(ph CCA) and distal. Initial experiences with orthotopic liver transplantation(OLT) for patientswith i CCA and ph CCA had very poor results and this treatment strategy was abandoned. In the last decade,thanks to a strict selection process and a neoadjuvant chemoradiation protocol,the results of OLT for patients with non-resectable phC CA have been shown to be excellent and this strategy has been extended worldwide in selected transplant centers. Intrahepatic cholangiocarcinoma is a growing disease in most countries and can be diagnosed both in cirrhotic and in non-cirrhotic livers. Even though OLT is contraindicated in most centers,recent investigations analyzing patients that were transplanted with a misdiagnosis of HCC and were found to have an iC CA have shown encouraging results. There is some information suggesting that patients with early stages of the disease could benefit from OLT. In this review we analyze the current stateof-the-art of OLT for cholangiocarcinoma as well as the new insights and future perspectives.

  17. A combination of liver fluke infection and traditional northeastern Thai foods associated with cholangiocarcinoma development. (United States)

    Sriraj, Pranee; Boonmars, Thidarut; Aukkanimart, Ratchadawan; Songsri, Jiraporn; Sripan, Panupan; Ratanasuwan, Panaratana; Boonjaraspinyo, Sirintip; Wongchalee, Nadchanan; Laummaunwai, Porntip


    Opisthorchis viverrini infection is one of the risk factors for cholangiocarcinoma (CCA) in northeast Thailand, a region with one of the highest reported incidence rates of CCA. The traditional practice of eating raw fish, repeated exposure to liver flukes, and consumption of nitrosamine-contaminated food are major risk factors for CCA. So far, there have been no reports about which northeastern traditional dishes may be involved in CCA development. The present study, thus, investigated the effects of traditional foods. It focused specifically on the consumption of fermented foods in combination with O. viverrini infection in hamsters. Syrian hamsters were divided into six groups: (i) normal hamsters, (ii) O. viverrini infection only and (iii)-(vi) O. viverrini infection plus fermented foods (pla som-fish fermented for 1 day), som wua-fermented beef, som phag-fermented vegetables, and pla ra-fish fermented for 6 months. Syrian hamster livers were used for analysis of histopathological changes through hematoxylin and eosin; Sirius Red; and immunohistostaining for cytokeratin-19, proliferating cell nuclear antigen, and CA19-9. Hamster sera were used for liver and kidney function tests. Results of all O. viverrini-infected groups and fermented food groups showed that histopathological changes consisted primarily of aggregations of inflammatory cells surrounding the hepatic bile duct, especially at the hilar region. However, there was a difference in virulence. Interestingly, aggregations of inflammatory cells, new bile duct formation, and fibrosis were observed in subcapsular hepatic tissue, which correlated to positive immunohistochemical staining and increased liver function test. The present study suggests that fermented food consumption can exacerbate cholangitis and cholangiofibrosis, which are risk factors for cholangiocarcinoma-associated opisthorchiasis.

  18. 混合型肝癌伴干细胞特征(细胆管癌型)26例临床病理学观察%Combined hepatocellular-cholangiocarcinoma (cholangiolocellular type) with stem-cell features:a clinicopathologic analysis of 26 cases

    Institute of Scientific and Technical Information of China (English)

    徐菁; 张翠明; 乔爱秀; 郗彦凤


    Objective To study the clinicopathologic features of combined hepatocellular-cholangiocarcinoma ( cholangiolocellular type, CLC type ) with stem cell features and its relationship to hepatic progenitor cells ( HPCs ).Methods Clinical and histologic features of 26 cases of combined hepatocellular-cholangiocarcinoma ( CLC type) were reviewed.Histochemistry was performed to confirm the type of mucin and immunohistochemical study was carried out for hepatocytic markers ( Hep Par-1 and AFP) and biliary/HPCs markers ( CK7, CK9, EMA, EpCAM, NCAM, CKIT).Results The age of patients ranged from 51 to 82 years ( mean 64 years ).All 26 cases contained CLC and hepatocellular carcinoma components.CLC area was composed of mixtures of small monotonous glands with abundant fibrous stroma and lymphocytic infiltrate.Tumor cells were cuboidal, smaller in size than normal hepatocytes, with basophilic cytoplasm and round nuclei.All cases, especially at the tumor boundary, showed HCC-like trabecular areas characterized by mildly atypical tumor cells with abundant eosinophilic cytoplasm and little stroma.Out of 26 cases, 21 showed definite glandular formation with mucin production, representing intrahepatic cholangiocarcinoma areas.The three distinct areas showed transitional zones merging with each other.The surrounding liver tissue showed cirrhosis and chronic hepatitis with varying degrees of fibrosis and periportal ductular reaction.Immunohistochemistry showed that biliary/HPC markers ( CK7, CK9, EMA, EpCAM, NCAM and CKIT) were strongly positive in CLC area in almost all cases, similar to the staining pattern of ductular reaction.In HCC-like areas, CK7 and CK19 were positive in all cases and the expression rates of EMA, EpCAM, NCAM, CKIT, AFP, Hep Par-1 were 80.8%(21/26), 88.5%(23/26), 84.6%(22/26), 88.5%(23/26), 46.2%(12/26) and 53.8%(14/26) respectively, similar to the staining pattern of intermediate hepatocytes.In ICC areas, CK7, CK9, EMA and EpCAM were positive in all cases

  19. Adjuvant therapy for intrahepatic cholangiocarcinoma: the debate continues. (United States)

    Zhu, Andrew X; Knox, Jennifer J


    Presentation of the Case A 37-year-old woman presented at 35 weeks of gestation with her third child with failure to adequately gain weight and was noted by her obstetrician to have delay in the growth of her baby. Ultrasound of the abdomen incidentally revealed the presence of a liver lesion. After additional evaluation, she ultimately delivered her daughter at 36 weeks uneventfully. She subsequently underwent additional evaluation. Liver magnetic resonance imaging (MRI) revealed a 5-cm solitary solid mass in segment 4A of the liver, concerning for malignancy. Serum α-fetoprotein, carcinoembryonic antigen, cancer antigen (CA)19-9, CA15-3, and CA125 were all normal. Liver biopsy was positive for adenocarcinoma. The tumor cells demonstrated a phenotype suggesting a possible breast primary, although the immunohistochemistry did not support that diagnosis and the tumor was negative for mammaglobin, gross cystic disease fluid protein (GCDFP)-15, estrogen receptor (ER), and progesterone receptor (PR) (Table 1). The tumor was also CDX2 and cardiotrophin-1 negative, but cytokeratin (CK) 19 positive. Her endoscopic retrograde cholangiopancreatography, upper endoscopy, colonoscopy, breast mammogram, and breast MRI were completely normal. A positron emission tomography-computed tomography scan showed a fluorodeoxyglucose-avid 5.8-cm × 6.0-cm hypoattenuating lesion with peripheral enhancement involving segment 4 and segment 8 at the dome. In addition, central necrosis within the lesion was noted. The left main portal vein was mildly attenuated by the mass. She eventually underwent a left hepatectomy en bloc with caudate resection, portal lymphadenectomy, cholecystectomy, and omental pedicle flap. On exploration of the abdomen, no additional disease was noted. The final pathology revealed a 9.4-cm moderately to poorly differentiated adenocarcinoma of the intrahepatic bile ducts. Venous invasion was present. Perineural invasion was absent. The margins were negative. Thirteen

  20. Prognostic Significance of Neutrophil to Lymphocyte Ratio in Oncologic Outcomes of Cholangiocarcinoma: A Meta-analysis. (United States)

    Tan, De-Wen; Fu, Yan; Su, Qi; Guan, Ming-Jun; Kong, Po; Wang, Sheng-Qiang; Wang, He-Ling


    Increasing evidence indicates that the neutrophil to lymphocyte ratio (NLR) is a useful biomarker of long-term outcomes in patients with cholangiocarcinoma. However, the prognostic role of NLR in patients with cholangiocarcinoma remains unclear. Thus, the current meta-analysis was undertaken to clarify the correlation between NLR and overall survival (OS) in cholangiocarcinoma, and a comprehensive literature research was conducted to understand the association of NLR and prognosis of cholangiocarcinoma. The hazard ratio (HR) with 95% confidence interval (CI) was used to assess OS. The synthesized HR of 1.449 (95% CI: 1.296-1.619, P analysis suggested that elevated preoperative NLR is associated with poorer rates of survival in cholangiocarcinoma patients.

  1. Distant skeletal muscle metastasis from intrahepatic cholangiocarcinoma presenting as Budd-Chiari syndrome

    Institute of Scientific and Technical Information of China (English)

    Oh Sung Kwon; Young Sook Park; Jong Eun Joo; Dae Won Jun; Sang Heum Kim; Mee Yeon Chung; Nam In Kim; Moon Hee Song; Han Hyo Lee; Seung Hwan Kim; Yoon Ju Jo


    Intrahepatic cholangiocarcinoma is a malignant neoplasm arising from the biliary epithelium, which frequently invades adjacent organs or metastasizes to other visceral organs such as the lungs, bones, adrenals, and brain. However, distant skeletal muscle metastasis of cholangiocarcinoma has never been described before to the best of our knowledge and, furthermore, Budd-Chiari syndrome secondary to intrahepatic cholangiocarcinoma is also extremely rare. Here we present the first case overall of distant muscle metastasis from intrahepatic cholangiocarcinoma presenting as Budd-Chiari syndrome. A 44-year-old man admitted to the hospital with complaints of abdominal distension, edema of both legs, back pain and anorexia of 30 o" duration. Computed tomography and ultrasonography-guided percutaneous muscle biopsy established intrahepatic cholangiocarcinoma with disseminated thrombosis from inferior vena cava to bilateral iliac and femoral veins, and multiple skeletal muscle metastases in bilateral buttock and erector spinal muscle.

  2. The potency of human testicular stem cells

    NARCIS (Netherlands)

    Chikhovskaya, J.V.


    In this thesis, we evaluate the stem cell state of cells present in primary human testicular cell cultures as well as their origin and relation to germ or somatic lineages within testicular tissue. We conclude that human testis-derived embryonic stem cell-like (htES-like) colonies arising in primary

  3. Opposing actions of endocannabinoids on cholangiocarcinoma growth is via the differential activation of Notch signaling

    Energy Technology Data Exchange (ETDEWEB)

    Frampton, Gabriel; Coufal, Monique [Department of Internal Medicine, Texas A and M Health Science Center College of Medicine, Temple, TX (United States); Li, Huang [Department of Internal Medicine, Texas A and M Health Science Center College of Medicine, Temple, TX (United States); Department of Hepatobiliary Surgery, First Affiliated Hospital, Sun Yat-Sen University, Guangzhou (China); Ramirez, Jonathan [Digestive Disease Research Center, Scott and White Hospital, Temple, TX (United States); DeMorrow, Sharon, E-mail: [Department of Internal Medicine, Texas A and M Health Science Center College of Medicine, Temple, TX (United States); Digestive Disease Research Center, Scott and White Hospital, Temple, TX (United States)


    The endocannabinoids anandamide (AEA) and 2-arachidonylglycerol (2-AG) have opposing effects on cholangiocarcinoma growth. Implicated in cancer, Notch signaling requires the {gamma}-secretase complex for activation. The aims of this study were to determine if the opposing effects of endocannabinoids depend on the differential activation of the Notch receptors and to demonstrate that the differential activation of these receptors are due to presenilin 1 containing- and presenilin 2 containing-{gamma}-secretase complexes. Mz-ChA-1 cells were treated with AEA or 2-AG. Notch receptor expression, activation, and nuclear translocation were determined. Specific roles for Notch 1 and 2 on cannabinoid-induced effects were determined by transient transfection of Notch 1 or 2 shRNA vectors before stimulation with AEA or 2-AG. Expression of presenilin 1 and 2 was determined after AEA or 2-AG treatment, and the involvement of presenilin 1 and 2 in the cannabinoid-induced effects was demonstrated in cell lines with low presenilin 1 or 2 expression. Antiproliferative effects of AEA required increased Notch 1 mRNA, activation, and nuclear translocation, whereas the growth-promoting effects induced by 2-AG required increased Notch 2 mRNA expression, activation, and nuclear translocation. AEA increased presenilin 1 expression and recruitment into the {gamma}-secretase complex, whereas 2-AG increased expression and recruitment of presenilin 2. The development of novel therapeutic strategies aimed at modulating the endocannabinoid system or mimicking the mode of action of AEA on Notch signaling pathways would prove beneficial for cholangiocarcinoma management.

  4. Stem cell differentiation and human liver disease

    Institute of Scientific and Technical Information of China (English)

    Wen-Li Zhou; Claire N Medine; Liang Zhu; David C Hay


    Human stem cells are scalable cell populations capable of cellular differentiation.This makes them a very attractive in vitro cellular resource and in theory provides unlimited amounts of primary cells.Such an approach has the potential to improve our understanding of human biology and treating disease.In the future it may be possible to deploy novel stem cell-based approaches to treat human liver diseases.In recent years,efficient hepatic differentiation from human stem cells has been achieved by several research groups including our own.In this review we provide an overview of the field and discuss the future potential and limitations of stem cell technology.

  5. The human airway epithelial basal cell transcriptome.

    Directory of Open Access Journals (Sweden)

    Neil R Hackett

    Full Text Available BACKGROUND: The human airway epithelium consists of 4 major cell types: ciliated, secretory, columnar and basal cells. During natural turnover and in response to injury, the airway basal cells function as stem/progenitor cells for the other airway cell types. The objective of this study is to better understand human airway epithelial basal cell biology by defining the gene expression signature of this cell population. METHODOLOGY/PRINCIPAL FINDINGS: Bronchial brushing was used to obtain airway epithelium from healthy nonsmokers. Microarrays were used to assess the transcriptome of basal cells purified from the airway epithelium in comparison to the transcriptome of the differentiated airway epithelium. This analysis identified the "human airway basal cell signature" as 1,161 unique genes with >5-fold higher expression level in basal cells compared to differentiated epithelium. The basal cell signature was suppressed when the basal cells differentiated into a ciliated airway epithelium in vitro. The basal cell signature displayed overlap with genes expressed in basal-like cells from other human tissues and with that of murine airway basal cells. Consistent with self-modulation as well as signaling to other airway cell types, the human airway basal cell signature was characterized by genes encoding extracellular matrix components, growth factors and growth factor receptors, including genes related to the EGF and VEGF pathways. Interestingly, while the basal cell signature overlaps that of basal-like cells of other organs, the human airway basal cell signature has features not previously associated with this cell type, including a unique pattern of genes encoding extracellular matrix components, G protein-coupled receptors, neuroactive ligands and receptors, and ion channels. CONCLUSION/SIGNIFICANCE: The human airway epithelial basal cell signature identified in the present study provides novel insights into the molecular phenotype and biology of

  6. Clinicopathological and prognostic significance of Yes-associated protein expression in hepatocellular carcinoma and hepatic cholangiocarcinoma. (United States)

    Wu, Hao; Liu, Yan; Jiang, Xiao-Wei; Li, Wen-Fang; Guo, Gang; Gong, Jian-Ping; Ding, Xiong


    Hepatocellular carcinoma (HCC) and hepatic cholangiocarcinoma (CC) are the most aggressive malignancies with a poor prognosis in humans, and hepatic cholangiocarcinoma (CC) exhibits greater malignant behaviour. Yes-associated protein (YAP) is an important downstream target of the Hippo signalling pathway. As an oncogene, it plays a vital role in the occurrence and development of tumours. Our study focuses on the clinical significance of YAP protein expression in HCC and CC. Furthermore, we sought to explore the different survival rates between HCC and CC. A total of 137 patients with HCC and 122 with CC after resection were evaluated by immunohistochemistry for the expression of YAP. Our results showed that positive expression rates of YAP were more frequently noted in CC 67.2 % (82/122) than in HCC 56.9 % (78/137) (P = 0.024). High YAP expression in HCC and CC was significantly associated with tumour size (P < 0.001 and P = 0.019, respectively), liver cirrhosis (P = 0.002 and P = 0.009, respectively), vascular invasion (P = 0.047 and P = 0.018, respectively), multiplicity (P = 0.019 and P = 0.015, respectively), and intrahepatic metastasis (P = 0.015 and P = 0.047, respectively). Importantly, recurrence-free survival and disease-specific survival rates were lower in CC with high YAP expression than in HCC with high YAP expression (P < 0.001 and P < 0.001, respectively). Overall, high YAP expression was more frequently found in CC than in HCC, and YAP overexpression was associated with poor survival rates in patients with HCC and CC. Targeting YAP treatment requires further prospective investigations in larger patient populations.

  7. Isolated implant metastasis in chest wall due to seeding of transpleurally placed PTBD catheter tract in a case of hilar cholangiocarcinoma. (United States)

    Talukder, Shibojit; Behera, Arunanshu; Tandup, Cherring; Mitra, Suvradeep


    Percutaneous transhepatic biliary drainage (PTBD) catheter site metastasis in cases of cholangiocarcinoma is reported sporadically. But it is unusual to see left-sided tumour metastasising to the right PTBD catheter site. Metastasis, in general, has a poor prognosis, but recurrence along the catheter tract in the absence of other systemic diseases can be a different scenario altogether. To date, there is no consensus on the management of this form of metastasis. But carefully selected patients can benefit from aggressive surgical resection. We report a case of a young patient with isolated chest wall metastasis 1 year after resection of left-sided hilar cholangiocarcinoma. The metastasis was resected and, on pathological analysis, was confirmed to be due to implantation of malignant cells along the tract of the PTBD catheter placed via a transpleural route. © BMJ Publishing Group Ltd (unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  8. Endocannabinoids and Human Sperm Cells

    Directory of Open Access Journals (Sweden)

    Giovanna Zolese


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

  9. Human regulatory B cells control the TFH cell response. (United States)

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


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

  10. Impaired degradation followed by enhanced recycling of epidermal growth factor receptor caused by hypo-phosphorylation of tyrosine 1045 in RBE cells

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    Gui Anping


    Full Text Available Abstract Background Since cholangiocarcinoma has a poor prognosis, several epidermal growth factor receptor (EGFR-targeted therapies with antibody or small molecule inhibitor treatment have been proposed. However, their effect remains limited. The present study sought to understand the molecular genetic characteristics of cholangiocarcinoma related to EGFR, with emphasis on its degradation and recycling. Methods We evaluated EGFR expression and colocalization by immunoblotting and immunofluorescence, cell surface EGFR expression by fluorescence-activated cell sorting (FACS, and EGFR ubiquitination and protein binding by immunoprecipitation in the human cholangiocarcinoma RBE and immortalized cholangiocyte MMNK-1 cell lines. Monensin treatment and Rab11a depletion by siRNA were adopted for inhibition of EGFR recycling. Results Upon stimulation with EGF, ligand-induced EGFR degradation was impaired and the expression of phospho-tyrosine 1068 and phospho-p44/42 MAPK was sustained in RBE cells as compared with MMNK-1 cells. In RBE cells, the process of EGFR sorting for lysosomal degradation was blocked at the early endosome stage, and non-degradated EGFR was recycled to the cell surface. A disrupted association between EGFR and the E3 ubiquitin ligase c-Cbl, as well as hypo-phosphorylation of EGFR at tyrosine 1045 (Tyr1045, were also observed in RBE cells. Conclusion In RBE cells, up-regulation of EGFR Tyr1045 phosphorylation is a potentially useful molecular alteration in EGFR-targeted therapy. The combination of molecular-targeted therapy determined by the characteristics of individual EGFR phosphorylation events and EGFR recycling inhibition show promise in future treatments of cholangiocarcinoma.

  11. Multigene mutational profiling of cholangiocarcinomas identifies actionable molecular subgroups (United States)

    Mafficini, Andrea; Wood, Laura D.; Corbo, Vincenzo; Melisi, Davide; Malleo, Giuseppe; Vicentini, Caterina; Malpeli, Giorgio; Antonello, Davide; Sperandio, Nicola; Capelli, Paola; Tomezzoli, Anna; Iacono, Calogero; Lawlor, Rita T.; Bassi, Claudio; Hruban, Ralph H.; Guglielmi, Alfredo; Tortora, Giampaolo; de Braud, Filippo; Scarpa, Aldo


    One-hundred-fifty-three biliary cancers, including 70 intrahepatic cholangiocarcinomas (ICC), 57 extrahepatic cholangiocarcinomas (ECC) and 26 gallbladder carcinomas (GBC) were assessed for mutations in 56 genes using multigene next-generation sequencing. Expression of EGFR and mTOR pathway genes was investigated by immunohistochemistry. At least one mutated gene was observed in 118/153 (77%) cancers. The genes most frequently involved were KRAS (28%), TP53 (18%), ARID1A (12%), IDH1/2 (9%), PBRM1 (9%), BAP1 (7%), and PIK3CA (7%). IDH1/2 (p=0.0005) and BAP1 (p=0.0097) mutations were characteristic of ICC, while KRAS (p=0.0019) and TP53 (p=0.0019) were more frequent in ECC and GBC. Multivariate analysis identified tumour stage and TP53 mutations as independent predictors of survival. Alterations in chromatin remodeling genes (ARID1A, BAP1, PBRM1, SMARCB1) were seen in 31% of cases. Potentially actionable mutations were seen in 104/153 (68%) cancers: i) KRAS/NRAS/BRAF mutations were found in 34% of cancers; ii) mTOR pathway activation was documented by immunohistochemistry in 51% of cases and by mutations in mTOR pathway genes in 19% of cancers; iii) TGF-ß/Smad signaling was altered in 10.5% cancers; iv) mutations in tyrosine kinase receptors were found in 9% cases. Our study identified molecular subgroups of cholangiocarcinomas that can be explored for specific drug targeting in clinical trials. PMID:24867389

  12. Impact of Microenvironment and Stem-Like Plasticity in Cholangiocarcinoma

    DEFF Research Database (Denmark)

    Raggi, Chiara; Invernizzi, Pietro; Andersen, Jesper Bøje


    Clinical complexity, anatomic diversity and molecular heterogeneity of cholangiocarcinoma (CCA) represent a major challenge in the assessment of effective targeted therapies. Molecular and cellular mechanisms underlying diversity of CCA growth patterns remain a key issue and a clinical concern...... or tumor microenvironment (TME) likely promotes initiation and progression of this malignancy contributing to its heterogeneity. This review will emphasize the dynamic interplay between stem-like intrinsic and TME-extrinsic pathways, which may represent novel options for multi-targeted therapies in CCA....

  13. Etiological and clinicopathologic characteristics of intrahepatic cholangiocarcinoma in young patients

    Institute of Scientific and Technical Information of China (English)


    AIM:To investigate the prevalence,risk factors,and clinicopathologic characteristics of intrahepatic cholangiocarcinoma(ICC)in young patients.METHODS:A retrospective analysis was performed in ICC patients referred to the Eastern Hepatobiliary Surgery Hospital in Shanghai,China.Among 317 consecutively enrolled patients,40 patients were aged ≤40 years(12.61%).We compared the risk factors and clinicopathologic characteristics of these patients(groupⅠ:n=40)with those aged>40 years(group Ⅱ:n=277).RESULTS:Group I...

  14. Opisthorchiasis and cholangiocarcinoma in Southeast Asia: an unresolved problem

    Directory of Open Access Journals (Sweden)

    Hughes T


    Full Text Available Thomas Hughes,1,* Thomas O’Connor,1,* Anchalee Techasen,2,3 Nisana Namwat,2,3 Watcharin Loilome,2,3 Ross H Andrews,2–4 Narong Khuntikeo,3,5 Puangrat Yongvanit,3,6 Paiboon Sithithaworn,3,7 Simon D Taylor-Robinson1 1Division of Digestive Health, Department of Surgery and Cancer, Imperial College London, London, UK; 2Department of Biochemistry, Faculty of Medicine, Liver Fluke and Cholangiocarcinoma Centre, 3Cholangiocarcinoma Screening and Care Program (CASCAP, Khon Kaen University, Khon Kaen, Thailand; 4Faculty of Medicine, St Mary’s Campus, Imperial College, London, UK; 5Department of Surgery, 6Department of Biochemistry, 7Department of Parasitology, Faculty of Medicine, Liver Fluke and Cholangiocarcinoma Centre, Khon Kaen University, Khon Kaen, Thailand *These authors contributed equally to this work Abstract: The prevalence of cholangiocarcinoma (CCA in Southeast Asia is much higher than other areas of the world. Eating raw, fermented, or undercooked cyprinid fish, infected with the liver fluke, Opisthorchis viverrini sensu lato (sl, results in chronic biliary inflammation, periductal fibrosis, and increased cancer risk. There may be associated glomerulonephritis. The process of infection is difficult to disrupt because eating practices have proven extremely difficult to change, and the life cycle of the fluke cannot be broken due to high prevalence in canine and feline reservoir hosts. Fecal analysis and enzyme-linked immunosorbent assay tests can be used to diagnose opisthorchiasis. Diagnosis of CCA is complex, partly due to the lack of definitive imaging characteristics but also due to the difficulty of obtaining samples for cytology or histology. This cancer has proven to be resistant to common chemotherapy treatments and so the two avenues of treatment available are surgical resection and liver transplantation, both requiring early detection of the tumor for the best chances of success. Late presentation of symptoms reduces the

  15. Satellite cells in human skeletal muscle plasticity. (United States)

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


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

  16. Hypermutation and unique mutational signatures of occupational cholangiocarcinoma in printing workers exposed to haloalkanes (United States)

    Mimaki, Sachiyo; Totsuka, Yukari; Suzuki, Yutaka; Nakai, Chikako; Goto, Masanori; Kojima, Motohiro; Arakawa, Hirofumi; Takemura, Shigekazu; Tanaka, Shogo; Marubashi, Shigeru; Kinoshita, Masahiko; Matsuda, Tomonari; Shibata, Tatsuhiro; Nakagama, Hitoshi; Ochiai, Atsushi; Kubo, Shoji; Nakamori, Shoji; Esumi, Hiroyasu; Tsuchihara, Katsuya


    Cholangiocarcinoma is a relatively rare cancer, but its incidence is increasing worldwide. Although several risk factors have been suggested, the etiology and pathogenesis of the majority of cholangiocarcinomas remain unclear. Recently, a high incidence of early-onset cholangiocarcinoma was reported among the workers of a printing company in Osaka, Japan. These workers underwent high exposure to organic solvents, mainly haloalkanes such as 1,2-dichloropropane (1,2-DCP) and/or dichloromethane. We performed whole-exome analysis on four cases of cholangiocarcinoma among the printing workers. An average of 44.8 somatic mutations was detected per Mb in the genome of the printing workers’ cholangiocarcinoma tissues, approximately 30-fold higher than that found in control common cholangiocarcinoma tissues. Furthermore, C:G-to-T:A transitions with substantial strand bias as well as unique trinucleotide mutational changes of GpCpY to GpTpY and NpCpY to NpTpY or NpApY were predominant in all of the printing workers’ cholangiocarcinoma genomes. These results were consistent with the epidemiological observation that they had been exposed to high concentrations of chemical compounds. Whole-genome analysis of Salmonella typhimurium strain TA100 exposed to 1,2-DCP revealed a partial recapitulation of the mutational signature in the printing workers’ cholangiocarcinoma. Although our results provide mutational signatures unique to occupational cholangiocarcinoma, the underlying mechanisms of the disease should be further investigated by using appropriate model systems and by comparison with genomic data from other cancers. PMID:27267998


    Institute of Scientific and Technical Information of China (English)

    陈力真; 王树蕙; 张云; 王世真


    HuPBL-SCID mice were used to explore how they would response to human ttmoor cells of 801/MLC.Living 801/MLC cells appeared to be fetal to the the mice due to the production of human TNF. The huP-BL-SCID rniee did not generate any noticeable amotmt of specific human immunoglobttlin either by single immunization with living 801/MLC cells or by repeated immunization with irradiated 801/MLC cells. Our preliminary experiments with huPBL-SCID mice showed that such chimeras would he a very useful models for tumor immunological researches.

  18. Search for naive human pluripotent stem cells

    Institute of Scientific and Technical Information of China (English)

    Simone Aparecida Siqueira Fonseca; Roberta Montero Costas; Lygia Veiga Pereira


    Normal mouse pluripotent stem cells were originallyderived from the inner cell mass (ICM) of blastocystsand shown to be the in vitro equivalent of those preimplantationembryonic cells, and thus were calledembryonic stem cells (ESCs). More than a decade later,pluripotent cells were isolated from the ICM of humanblastocysts. Despite being called human ESCs, thesecells differ significantly from mouse ESCs, includingdifferent morphology and mechanisms of control ofpluripotency, suggesting distinct embryonic originsof ESCs from the two species. Subsequently, mousepluripotent stem cells were established from the ICMderivedepiblast of post-implantation embryos. Thesemouse epiblast stem cells (EpiSCs) are morphologicaland epigenetically more similar to human ESCs. Thisraised the question of whether cells from the humanICM are in a more advanced differentiation stage thantheir murine counterpart, or whether the availableculture conditions were not adequate to maintain thosehuman cells in their in vivo state, leading to a transitioninto EpiSC-like cells in vitro . More recently, novel cultureconditions allowed the conversion of human ESCs intomouse ESC-like cells called naive (or ground state)human ESCs, and the derivation of naive human ESCsfrom blastocysts. Here we will review the characteristicsof each type of pluripotent stem cells, how (andwhether) these relate to different stages of embryonicdevelopment, and discuss the potential implications ofnaive human ESCs in research and therapy.

  19. Cholangiocarcinoma with Lymphoepithelioma-like Component not Associated with Epstein-Barr Virus

    Directory of Open Access Journals (Sweden)

    Wen-Han Chang


    Full Text Available Lymphoepithelioma-like cholangiocarcinoma (LELCC is a rare variant of intrahepatic cholangiocarcinoma. We herein have reported an unusual case of LELCC in a 71-year-old Taiwanese women with cirrhotic liver disease and chronic hepatitis C. The patient's liver tumor was unexpectedly discovered during a regular abdominal ultrasound exam without obvious clinical symptoms and signs; she thereafter received surgical resection. Histologically, the liver tumor showed lymphoepithelial-like appearance and features of cholangiocarcinoma without association with Epstein-Barr virus (EBV. We maintained regular follow-up with the patient for 3 years, who at that time was alive without tumor recurrence.

  20. Endothelial cells derived from human embryonic stem cells (United States)

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


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

  1. Induced pluripotent stem cell lines derived from human somatic cells. (United States)

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


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

  2. Hilar Cholangiocarcinoma Diagnosed and Treated Early, in Prejaundice Phase

    Directory of Open Access Journals (Sweden)

    Denes M.


    Full Text Available Hilar cholangiocarcinoma, Klatskin tumor or proximal bile duct cancer, is a tumor growing in the right hepatic duct, left hepatic duct or at their confluence. It is a relatively rare but devastating disease. The tight stricture of the biliary ducts and the development of obstructive jaundice are the main characteristics of the disease. In the early phase, symptoms are nonspecific and jaundice is not present, leading to delayed diagnosis and denying the possibility of curative treatment. We present the case of a 74 years old woman who was referred to us with ambiguous symptomatology and without jaundice. The ultrasound and CT scan showed dilation of the left biliary tree, without increase of the cholestatic enzymes. Magnetic resonance cholangiography depicted a tumor in the left hepatic duct (3X3 cm. with enlargement of the bile ducts above. The surgical treatment consisted of left hepatectomy and hilar lymph nodes dissection. The pathology findings showed a cholangiocarcinoma with a few hilar nodes involvement. Our approach was potentially curative. Unfortunately these situations are seldom because in the majority of cases the patients have obstructive jaundice at presentation and the tumors are unresectable. We consider that a magnetic resonance cholangiography made when we suspect a bile duct tumor, leads us to an early diagnosis and gives us the possibility of a potential curative surgical treatment.

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

    Energy Technology Data Exchange (ETDEWEB)

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


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

  4. Transcriptomic and histopathological analysis of cholangiolocellular differentiation trait in intrahepatic cholangiocarcinoma

    DEFF Research Database (Denmark)

    Rhee, Hyungjin; Ko, Jung Eun; Chung, Taek


    BACKGROUND & AIMS: Intrahepatic cholangiocarcinoma (iCCA) is a heterogeneous entity with diverse etiologies, morphologies, and clinical outcomes. Recently, histopathological distinction of cholangiolocellular differentiation (CD) of iCCA has been suggested. However, its genome-wide molecular feat...

  5. Colon Mass as a Secondary Metastasis from Cholangiocarcinoma: A Diagnostic and Therapeutic Dilemma. (United States)

    Niazi, Azfar; Saif, Muhammad W


    Cholangiocarcinoma (bile ducts cancer) is a rare and aggressive form of cancer. It metastasizes frequently to liver, peritoneum, and lungs. Colon metastasis is extremely uncommon. We report here a 70-year-old male who was diagnosed with cholangiocarcinoma for which he underwent a Whipple procedure. Fifteen months later, a CT scan revealed mural thickening in the colon; this was supplemented with a PET scan, which confirmed this mass. Histological diagnosis of metastatic cholangiocarcinoma to the colon was made and the patient was treated with chemotherapy. Although rare, cholangiocarcinoma metastasis can be found in the colon. A high index of suspicion is required to diagnose and treat early. More cases need to be reported to find out further about the prognosis of the disease.

  6. Differential effects of FXR or TGR5 activation in cholangiocarcinoma progression

    DEFF Research Database (Denmark)

    Erice, O; Labiano, I; Arbelaiz, A


    BACKGROUND AND AIMS: Cholangiocarcinoma (CCA) is an aggressive tumor type affecting cholangiocytes. CCAs frequently arise under certain cholestatic liver conditions. Intrahepatic accumulation of bile acids may facilitate cocarcinogenic effects by triggering an inflammatory response and cholangioc...

  7. Human Neural Cell-Based Biosensor (United States)


    including incubation with factors such as SHH ) and proceed to Human Neural Progenitor Cells Dopaminergic Differentiation β-III Tubulin/TH...exposure in human embryonic stem cells. J Recept Signal Transduct Res. 2011 Jun;31(3):206-13. Gerwe BA, Angel PM, West FD, Hasneen K, Young A

  8. Radiofrequency ablation of recurrent cholangiocarcinoma after orthotopic liver transplantation - a case report

    Institute of Scientific and Technical Information of China (English)

    Rakesh Rai; Derek Manas; John Rose


    AIM: To report the use of radiofrequency ablation in the treatment of recurrenct cholangiocarcinoma in the transplanted liver.METHODS: A lady who underwent orthotopic liver transplantation (OLT) for intrahepatic cholangiocarcinoma recurrence of tumour 13 mo after tralsplantation inspite of adjuvant chemotherapy. Her recurrent tumour was treated with radiofrequency ablation.RESULTS: She survived for 18 mo following the recurrence of her tumour.CONCLUSION: Radiofrequency ablation can be used safely in the transplanted liver to treat recurrent tumour.

  9. Capsaicin Enhances the Drug Sensitivity of Cholangiocarcinoma through the Inhibition of Chemotherapeutic-Induced Autophagy. (United States)

    Hong, Zai-Fa; Zhao, Wen-Xiu; Yin, Zhen-Yu; Xie, Cheng-Rong; Xu, Ya-Ping; Chi, Xiao-Qin; Zhang, Sheng; Wang, Xiao-Min


    Cholangiocarcinoma (CCA), a devastating cancer with a poor prognosis, is resistant to the currently available chemotherapeutic agents. Capsaicin, the major pungent ingredient found in hot red chili peppers of the genus Capsicum, suppresses the growth of several malignant cell lines. Our aims were to investigate the role and mechanism of capsaicin with respect to the sensitivity of CCA cells to chemotherapeutic agents. The effect of capsaicin on CCA tumor sensitivity to 5-fluorouracil (5-FU) was assessed in vitro in CCA cells and in vivo in a xenograft model. The drug sensitivity of QBC939 to 5-FU was significantly enhanced by capsaicin compared with either agent alone. In addition, the combination of capsaicin with 5-FU was synergistic, with a combination index (CI) capsaicin. Moreover, the decrease in AKT and S6 phosphorylation induced by 5-FU was effectively reversed by capsaicin, indicating that capsaicin inhibits 5-FU-induced autophagy by activating the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway in CCA cells. Taken together, these results demonstrate that capsaicin may be a useful adjunct therapy to improve chemosensitivity in CCA. This effect likely occurs via PI3K/AKT/mTOR pathway activation, suggesting a promising strategy for the development of combination drugs for CCA.

  10. Cortical network from human embryonic stem cells



    Abstract The connection of embryonic stem cell technology and developmental biology provides valuable tools to decipher the mechanisms underlying human brain development and diseases, especially among neuronal populations, that are not readily available in primary cultures. It is obviously the case of neurons forming the human cerebral cortex. In the images that are presented, the neurons were generated in vitro from human embryonic stem cells via forebrain-like progenitors. Maintained in cul...

  11. Cholangiocarcinoma presenting as hemobilia and recurrent iron-deficiency anemia: a case report

    Directory of Open Access Journals (Sweden)

    Hariraj Radhakrishnan


    Full Text Available Abstract Introduction Iron-deficiency anemia is a relatively common presenting feature of several gastrointestinal malignancies. However, cholangiocarcinoma has rarely been reported as an underlying cause. The association of cholangiocarcinoma with the rare clinical finding of hemobilia is also highly unusual. To our knowledge, this is the first case report of cholangiocarcinoma presenting with acute hemobilia and chronic iron-deficiency anemia. Case presentation We report the case of a Caucasian, 84-year-old woman presenting with recurrent, severe iron-deficiency anemia who was eventually diagnosed with intra-hepatic cholangiocarcinoma, following an acute episode of hemobilia. A right hepatectomy was subsequently performed with curative intent, and our patient has now fully recovered. Conclusion This is a rare example of hemobilia and chronic iron-deficiency anemia in association with cholangiocarcinoma. We suggest that a diagnosis of cholangiocarcinoma should be considered in patients who present with iron-deficiency anemia of unknown cause, particularly in the presence of abnormal liver function.

  12. Clinicopathological significance of altered Notch signaling in extrahepatic cholangiocarcinoma and gallbladder carcinoma

    Institute of Scientific and Technical Information of China (English)

    Hyun Ah Yoon; Myung Hwan Noh; Byung Geun Kim; Ji Sun Han; Jin Seok Jang; Seok Ryeol Choi; Jin Sook Jeong; Jin Ho Chun


    AIM: To investigate the role and clinicopathological significance of aberrant expression of Notch receptors and Delta-like ligand-4 (DLL4) in extrahepatic cholangiocarcinoma and gallbladder carcinoma.METHODS: One hundred and ten patients had surgically resected extrahepatic cholangiocarcinoma (CC) and gallbladder carcinoma specimens examined by immunohistochemistry of available paraffin blocks. Immunohistochemistry was performed using anti-Notch receptors 1-4 and anti-DLL4 antibodies. We scored the immunopositivity of Notch receptors and DLL4 expression by percentage of positive tumor cells with cytoplasmic expression and intensity of immunostaining. Coexistent nuclear localization was evaluated. Clinicopathological parameters and survival data were compared with the expression of Notch receptors 1-4 and DLL4.RESULTS: Notch receptor proteins showed in the cytoplasm with or without nuclear expression in cancer cells, as well as showing weak cytoplasmic expression in non-neoplastic cells. By semiquantitative evaluation, positive immunostaining of Notch receptor 1 was detected in 96 cases (87.3%), Notch receptor 2 in 97 (88.2%), Notch receptor 3 in 97 (88.2%), Notch receptor 4 in 103 (93.6), and DLL4 in 84 (76.4%). In addition, coexistent nuclear localization was noted [Notch receptor 1; 18 cases (18.8%), Notch receptor 2; 40 (41.2%), Notch receptor 3; 32 (33.0%), Notch receptor 4; 99 (96.1%), DLL4; 48 (57.1%)]. Notch receptor 1 expression was correlated with advanced tumor, node, metastasis (TNM) stage (P = 0.043), Notch receptor 3 with advanced T stage (P = 0.017), tendency to express in cases with nodal metastasis (P = 0.065) and advanced TNM stage (P = 0.052). DLL4 expression tended to be related to less histological differentiation (P = 0.095). Coexistent nuclear localization of Notch receptor 3 was related to no nodal metastasis (P = 0.027) and Notch receptor 4 with less histological differentiation (P = 0.036), while DLL4 tended to be related inversely with T

  13. Human hematopoietic cell culture, transduction, and analyses

    DEFF Research Database (Denmark)

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


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

  14. Human embryonic stem cells derived by somatic cell nuclear transfer. (United States)

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


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

  15. Kaempferol inhibits the growth and metastasis of cholangiocarcinoma in vitro and in vivo. (United States)

    Qin, Youyou; Cui, Wu; Yang, Xuewei; Tong, Baifeng


    Kaempferol is a flavonoid that has been reported to exhibit antitumor activity in various malignant tumors. However, the role of kaempferol on cholangiocarcinoma (CCA) is largely unknown. In this article, we found that kaempferol inhibited proliferation, reduced colony formation ability, and induced apoptosis in HCCC9810 and QBC939 cells in vitro. Results from transwell assay and wound-healing assay demonstrated that kaempferol significantly suppressed the migration and invasion abilities of HCCC9810 and QBC939 cells in vitro. Kaempferol was found to decrease the expression of Bcl-2 and increase the expressions of Bax, Fas, cleaved-caspase 3, cleaved-caspase 8, cleaved-caspase 9, and cleaved-PARP. In addition, kaempferol also downregulated the levels of phosphorylated AKT, TIMP2, and MMP2. In vivo, it was found that the volume of subcutaneous xenograft (0.15 cm(3)) in the kaempferol-treated group was smaller than that (0.6 cm(3)) in the control group. Kaempferol also suppressed the number and volume of metastasis foci in the lung metastasis model, with no marked effects on body weight of mice. Immunohistochemistry assay showed that the number of Ki-67-positive cells was lower in the kaempferol-treated group than that in the control group. We further confirmed that the changes of apoptosis- and invasion-related proteins after kaempferol treatment in vivo were similar to the results in vitro. These data suggest that kaempferol may be a promising candidate agent for the treatment of CCA.

  16. Human Neuroepithelial Cells Express NMDA Receptors

    Directory of Open Access Journals (Sweden)

    Cappell B


    Full Text Available Abstract L-glutamate, an excitatory neurotransmitter, binds to both ionotropic and metabotropic glutamate receptors. In certain parts of the brain the BBB contains two normally impermeable barriers: 1 cerebral endothelial barrier and 2 cerebral epithelial barrier. Human cerebral endothelial cells express NMDA receptors; however, to date, human cerebral epithelial cells (neuroepithelial cells have not been shown to express NMDA receptor message or protein. In this study, human hypothalamic sections were examined for NMDA receptors (NMDAR expression via immunohistochemistry and murine neuroepithelial cell line (V1 were examined for NMDAR via RT-PCR and Western analysis. We found that human cerebral epithelium express protein and cultured mouse neuroepithelial cells express both mRNA and protein for the NMDA receptor. These findings may have important consequences for neuroepithelial responses during excitotoxicity and in disease.

  17. Cholangiocarcinoma in Magnetic Resonance Cholangiopancreatography and Fascioliasis in Endoscopic Ultrasonography

    Directory of Open Access Journals (Sweden)

    Amir Houshang Mohammad Alizadeh


    Full Text Available Fascioliasis is a worldwide zoonotic infection with Fasciola hepatica and Fasciola gigantica. The zoonoses are particularly endemic in sheep-raising countries and are also endemic in Iran. Typical symptoms that may be associated with fascioliasis can be divided by phases of the disease, including the acute or liver phase, the chronic or biliary phase, and ectopic or pharyngeal fascioliasis. Cholestatic symptoms may be absent, and in some cases diagnosis and treatment may be preceded by a long period of abdominal pain, eosinophilia and vague gastrointestinal symptoms. We report a case with epigastric and upper quadrant abdominal pain for the last 4 years, with imaging suggesting cholangiocarcinoma. Considering a new concept of endoscopic ultrasonography, at last F. hepatica was extracted with endoscopic retrograde cholangiography.

  18. Cholangiocarcinoma: Correlation between Molecular Profiling and Imaging Phenotypes.

    Directory of Open Access Journals (Sweden)

    Eran Sadot

    Full Text Available To investigate associations between imaging features of cholangiocarcinoma by visual assessment and texture analysis, which quantifies heterogeneity in tumor enhancement patterns, with molecular profiles based on hypoxia markers.The institutional review board approved this HIPAA-compliant retrospective study of CT images of intrahepatic cholangiocarcinoma, obtained before surgery. Immunostaining for hypoxia markers (EGFR, VEGF, CD24, P53, MDM2, MRP-1, HIF-1α, CA-IX, and GLUT1 was performed on pre-treatment liver biopsies. Quantitative imaging phenotypes were determined by texture analysis with gray level co-occurrence matrixes. The correlations between quantitative imaging phenotypes, qualitative imaging features (measured by radiographic inspection alone, and expression levels of the hypoxia markers from the 25 tumors were assessed.Twenty-five patients were included with a median age of 62 years (range: 54-84. The median tumor size was 10.2 cm (range: 4-14, 10 (40% were single tumors, and 90% were moderately differentiated. Positive immunostaining was recorded for VEGF in 67% of the cases, EGFR in 75%, and CD24 in 55%. On multiple linear regression analysis, quantitative imaging phenotypes correlated significantly with EGFR and VEGF expression levels (R2 = 0.4, p<0.05 and R2 = 0.2, p<0.05, respectively, while a trend was demonstrated with CD24 expression (R2 = 0.33, p = 0.1. Three qualitative imaging features correlated with VEGF and CD24 expression (P<0.05, however, none of the qualitative features correlated with the quantitative imaging phenotypes.Quantitative imaging phenotypes, as defined by texture analysis, correlated with expression of specific markers of hypoxia, regardless of conventional imaging features.

  19. Tumor markers as a diagnostic key for hilar Cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Juntermanns B


    Full Text Available Abstract Objective Hilar cholangiocarcinoma is the fourth most common gastrointestinal malignancy. CA19-9 and CEA are helpful devices in the management of gastrointestinal malignancies and belong to clinical routine in surgical oncology. But the validity of these parameters in terms of tumor extension and prognosis of bile duct malignancies still remains unclear. Methods From 1998 to 2008, we obtained preoperative CA19-9 and CEA serum levels in 136 patients with hilar cholangiocarcinoma. We correlated tumor stage, resectability rate and survival with preoperative CA 19-9 and CEA serum levels. Results CA19-9 (UICC I: 253 ± 561 U/ml; UICC II: 742 ± 1572 U/ml; UICC III: 906 ± 1708 U/ml; UICC IV: 1707 ± 3053 U/ml and CEA levels (UICC I: 2.9 ± 3.8 U/ml; UICC II: 4.6 ± 6.5 U/ml; UICC III: 18.1 ± 29.6 U/ml; UICC IV: 22.7 ± 53.9 U/ml increase significantly with rising tumor stage. Patients with pre operative serum levels of CA19-9 (> 1000 U/ml and CEA (> 14.4 ng/ml showed a significant poorer resectability rate and survival than patients with lower CA19-9 and CEA serum levels respectively. Conclusion CA19-9 and CEA serum levels are associated with the tumor stage. If preoperatively obtained CA19-9 and CEA serum levels are highly elevated patients have an even worse survival and the frequency of irresectability is significantly higher.

  20. The miRNAome of Opisthorchis viverrini induced intrahepatic cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Jin Peng


    Full Text Available Intrahepatic cholangiocarcinoma (ICC is an aggressive cancer, arising in the biliary ducts that extend into the liver. The highest incidence of ICC occurs in Southeast Asia, particularly in the Mekong River Basin countries of Thailand, Laos, Cambodia, and Vietnam, where it is strongly associated with chronic infection by the food-borne liver fluke Opisthorchis viverrini (OV, one of only three eukaryote pathogens considered Group one carcinogens. Intrahepatic cholangiocarcinoma is usually diagnosed at an advanced stage, with a poor prognosis and survival often less than 24 months. Hence, biomarkers that enable the early detection of ICC would be desirable and have a potentially important impact on the public health in the resource-poor regions where this cancer is most prevalent. As microRNAs (miRNAs remain well preserved after formalin fixation, there is much interest in developing them as biomarkers that can be investigated using tumor biopsy samples preserved in formalin fixed paraffin embedded (FFPE tumor blocks. Recently, we reported the first comprehensive profiling of tissue-based miRNA expression using FFPE from the three most common subtypes of OV-induced ICC tumors: moderately differentiated ICC, papillary ICC, and well-differentiated ICC. We observed that each subtype of OV-induced ICC exhibited a distinct miRNA profile, which suggested the involvement of specific sets of miRNAs in the progression of this cancer. In addition, non-tumor tissue adjacent to ICC tumor tissue on the same FFPE block shared a similar miRNA dysregulation profile with the tumor tissue than with normal (non-tumor liver tissue (individuals without ICC or OV infection. Herein, we provide a detailed description of the microarray analysis procedures used to derive these findings.

  1. Cell Cycle Progression of Human Cells Cultured in Rotating Bioreactor (United States)

    Parks, Kelsey


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

  2. Cell Cycle Progression of Human Cells Cultured in Rotating Bioreactor (United States)

    Parks, Kelsey


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

  3. Derivation of naive human embryonic stem cells. (United States)

    Ware, Carol B; Nelson, Angelique M; Mecham, Brigham; Hesson, Jennifer; Zhou, Wenyu; Jonlin, Erica C; Jimenez-Caliani, Antonio J; Deng, Xinxian; Cavanaugh, Christopher; Cook, Savannah; Tesar, Paul J; Okada, Jeffrey; Margaretha, Lilyana; Sperber, Henrik; Choi, Michael; Blau, C Anthony; Treuting, Piper M; Hawkins, R David; Cirulli, Vincenzo; Ruohola-Baker, Hannele


    The naïve pluripotent state has been shown in mice to lead to broad and more robust developmental potential relative to primed mouse epiblast cells. The human naïve ES cell state has eluded derivation without the use of transgenes, and forced expression of OCT4, KLF4, and KLF2 allows maintenance of human cells in a naïve state [Hanna J, et al. (2010) Proc Natl Acad Sci USA 107(20):9222-9227]. We describe two routes to generate nontransgenic naïve human ES cells (hESCs). The first is by reverse toggling of preexisting primed hESC lines by preculture in the histone deacetylase inhibitors butyrate and suberoylanilide hydroxamic acid, followed by culture in MEK/ERK and GSK3 inhibitors (2i) with FGF2. The second route is by direct derivation from a human embryo in 2i with FGF2. We show that human naïve cells meet mouse criteria for the naïve state by growth characteristics, antibody labeling profile, gene expression, X-inactivation profile, mitochondrial morphology, microRNA profile and development in the context of teratomas. hESCs can exist in a naïve state without the need for transgenes. Direct derivation is an elusive, but attainable, process, leading to cells at the earliest stage of in vitro pluripotency described for humans. Reverse toggling of primed cells to naïve is efficient and reproducible.

  4. Regulatory T Cells in Human Ovarian Cancer

    Directory of Open Access Journals (Sweden)

    Dong-Jun Peng


    Full Text Available Multiple layers of suppressive components including regulatory T (TReg cells, suppressive antigen-presenting cells, and inhibitory cytokines form suppressive networks in the ovarian cancer microenvironment. It has been demonstrated that as a major suppressive element, TReg cells infiltrate tumor, interact with several types of immune cells, and mediate immune suppression through different molecular and cellular mechanisms. In this paper, we focus on human ovarian cancer and will discuss the nature of TReg cells including their subsets, trafficking, expansion, and function. We will briefly review the development of manipulation of TReg cells in preclinical and clinical settings.

  5. 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


    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.

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

    Institute of Scientific and Technical Information of China (English)


    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.

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

    NARCIS (Netherlands)

    Dormeyer, W.; van Hoof, D.; Braam, S.R.; Heck, A.J.R.; Mummery, C.L.; Krijgsveld, J.


    Human embryonic stem cells (hESCs) are of immense interest in regenerative medicine as they can self-renew indefinitely and can give rise to any adult cell type. Human embryonal carcinoma cells (hECCs) are the malignant counterparts of hESCs found in testis tumors. hESCs that have acquired chromosom

  8. Untangling the Complexity of Liver Fluke Infection and Cholangiocarcinoma in NE Thailand Through Transdisciplinary Learning. (United States)

    Ziegler, A D; Echaubard, P; Lee, Y T; Chuah, C J; Wilcox, B A; Grundy-Warr, C; Sithithaworn, P; Petney, T N; Laithevewat, L; Ong, X; Andrews, R H; Ismail, T; Sripa, B; Khuntikeo, N; Poonpon, K; Tungtang, P; Tuamsuk, K


    This study demonstrates how a transdisciplinary learning approach provided new insights for explaining persistent Opisthorchis viverrini infection in northern Thailand, as well as elucidating problems of focusing solely on the parasite as a means of addressing high prevalence of cholangiocarcinoma. Researchers from diverse backgrounds collaborated to design an investigative homestay program for 72 Singaporean and Thai university students in five northeast Thai villages. The students explored how liver fluke infection and potential cholangiocarcinoma development are influenced by local landscape dynamics, aquatic ecology, livelihoods, food culture and health education. Qualitative fieldwork was guided daily by the researchers in a collaborative, co-learning process that led to viewing this health issue as a complex system, influenced by interlinked multidimensional factors. Our transdisciplinary experience has led us to believe that an incomplete understanding of these linkages may reduce the efficacy of interventions. Further, viewing liver fluke infection and cholangiocarcinoma as the same issue is inadvisable. Although O. viverrini infection is an established risk factor for the development of cholangiocarcinoma, multiple factors are known to influence the likelihood of acquiring either. Understanding the importance of the current livelihood transition, landscape modification and the resulting mismatch between local cultures and new socio-ecological settings on cholangiocarcinoma initiation and liver fluke transmission is of critical importance as it may help readjust our view of the respective role of O. viverrini and other socioeconomic risk factors in cholangiocarcinoma etiology and refine intervention strategies. As demonstrated in this study, transdisciplinary approaches have the potential to yield more nuanced perspectives to complex diseases than research that focuses on specific aspects of their epidemiology. They may therefore be valuable when designing

  9. Derivation of Human Skin Fibroblast Lines for Feeder Cells of Human Embryonic Stem Cells. (United States)

    Unger, Christian; Felldin, Ulrika; Rodin, Sergey; Nordenskjöld, Agneta; Dilber, Sirac; Hovatta, Outi


    After the first derivations of human embryonic stem cell (hESC) lines on fetal mouse feeder cell layers, the idea of using human cells instead of mouse cells as feeder cells soon arose. Mouse cells bear a risk of microbial contamination, and nonhuman immunogenic proteins are absorbed from the feeders to hESCs. Human skin fibroblasts can be effectively used as feeder cells for hESCs. The same primary cell line, which can be safely used for up to 15 passages after stock preparations, can be expanded and used for large numbers of hESC derivations and cultures. These cells are relatively easy to handle and maintain. No animal facilities or animal work is needed. Here, we describe the derivation, culture, and cryopreservation procedures for research-grade human skin fibroblast lines. We also describe how to make feeder layers for hESCs using these fibroblasts.

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

    Energy Technology Data Exchange (ETDEWEB)

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


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

  11. Factors Associated with Diffusely Increased Splenic F-18 FDG Uptake in Patients with Cholangiocarcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Keunyoung; Kim, Seongjang; Kim, Injoo; Kim, Dong Uk; Kim, Heeyoung; Kim, Sojung; Ahn, Sang Hyun [Pusan National Univ. Hospital, Busan (Korea, Republic of)


    Although diffuse splenic {sup 18}F-fluorodeoxyglucose (F-18 FDG) uptake exceeding hepatic activity, is considered abnormal, its clinical significance is rarely discussed in the literature. The aim of this study was to determine the contributing factors causing diffusely increased splenic FDG uptake in patients with cholangiocarcinoma. From January 2010 to March 2013, 140 patients (84 men, 56 women) were enrolled in this study. All patients had been diagnosed with cholangiocarcinoma and underwent F-18 FDG positron emission tomography/computed tomography (PET/CT) for the pretreatment staging work up. Clinical records were reviewed retrospectively. Various hematological parameters, C-reactive protein (CRP) level, CEA, CA19-9, pancreatic enzymes and liver function tests were conducted within 2 days after the F-18 FDG PET/CT study. Diffuse splenic uptake was observed in 23 patients (16.4%). Of those, 19 patients (82.6%) underwent endoscopic retrograde cholangiopancreastography (ERCP) 7 days before F-18 FDG PET/CT. The CRP level (p <0.001) and white blood cell count (p =0.023) were significantly higher in the group of patients with diffuse splenic FDG uptake. The hemoglobin (p <0.001) and the hematocrit (p <0.001) were significantly lower in patients with diffuse splenic FDG uptake. Pancreatic enzymes, liver function test results, and tumor markers were not significantly different between the patients who did or did not have diffusely increased splenic FDG uptake. The significant factors for diffuse splenic F-18 FDG uptake exceeding hepatic F-18 FDG uptake on multivariate analysis included: performing ERCP before F-18 FDG PET-CT (odds ratio [OR], 77.510; 95% CI, 7.624-132.105), and the presence of leukocytosis (OR, 12.436; 95% CI, 2.438-63.445) or anemia (OR, 1.211; 95% CI, 1.051-1.871). In conclusion, our study demonstrated that concurrent inflammation could be associated with diffusely increased splenic FDG uptake. We suggest that performing ERCP before F-18 FDG PET

  12. Rapid and high-resolution imaging of human liver specimens by full-field optical coherence tomography (United States)

    Zhu, Yue; Gao, Wanrong; Zhou, Yuan; Guo, Yingcheng; Guo, Feng; He, Yong


    We report rapid and high-resolution tomographic en face imaging of human liver specimens by full-field optical coherence tomography (FF-OCT). First, the arrangement of the FF-OCT system was described and the performance of the system was measured. The measured axial and lateral resolutions of the system are 0.8 and 0.9 μm, respectively. The system has a sensitivity of ˜60 dB and can achieve an imaging rate of 7 fps and a penetration depth of ˜80 μm. The histological structures of normal liver can be seen clearly in the en face tomographic images, including central veins, cords of hepatocytes separated by sinusoidal spaces, and portal area (portal vein, the hepatic arteriole, and the bile duct). A wide variety of histological subtypes of hepatocellular carcinoma was observed in en face tomographic images, revealing notable cancerous features, including the nuclear atypia (enlarged convoluted nuclei), the polygonal tumor cells with obvious resemblance to hepatocytes with enlarged nuclei. In addition, thicker fibrous bands, which make the cytoplasmic plump vesicular nuclei indistinct, were also seen in the images. Finally, comparison between the portal vein in a normal specimen versus that seen in the rare type of cholangiocarcinoma was made. The results show that the cholangiocarcinoma presents with a blurred pattern of portal vein in the lateral direction and an aggregated distribution in the axial direction; the surrounding sinusoidal spaces and nuclei of cholangiocarcinoma are absent. The findings in this work may be used as additional signs of liver cancer or cholangiocarcinoma, demonstrating capacity of FF-OCT device for early cancer diagnosis and many other tumor-related studies in biopsy.

  13. Stem cells in the human breast

    DEFF Research Database (Denmark)

    Petersen, Ole William; Polyak, Kornelia


    The origins of the epithelial cells participating in the development, tissue homeostasis, and cancer of the human breast are poorly understood. However, emerging evidence suggests a role for adult tissue-specific stem cells in these processes. In a hierarchical manner, these generate the two main...

  14. Human hair genealogies and stem cell latency

    Directory of Open Access Journals (Sweden)

    Tavaré Simon


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

  15. Detection of salivary antibodies to crude antigens of Opisthorchis viverrini in opisthorchiasis and cholangiocarcinoma patients. (United States)

    Chaiyarit, Ponlatham; Sithithaworn, Paiboon; Thuwajit, Chanitra; Yongvanit, Puangrat


    Opisthorchis viverrini (O. viverrini; known as human liver fluke) is a major health problem in the northeastern region of Thailand. Infection with O. viverrini is the cause of hepatobiliary disease and cholangiocarcinoma (CCA). Previous studies demonstrated specific antibodies to crude O. viverrini antigens in serum from O. viverrini-infected patients. However, no studies have measured specific antibodies to O. viverrini antigens in saliva from patients with opisthorchiasis and CCA. The objective of the study was to detect specific antibodies to crude O. viverrini antigens in saliva from patients with opisthorchiasis and CCA, and to evaluate their use for diagnosis of O. viverrini infection. Saliva samples from 23 control subjects, 30 opisthorchiasis patients, and 38 CCA patients were collected. ELISA was established for detection of salivary IgA and IgG to crude O. viverrini antigens. ANOVA was used to compare salivary IgA and IgG levels among groups. Salivary IgA to crude O. viverrini antigens in CCA patients was significantly higher than controls (p = 0.007). Salivary IgG in CCA patients was significantly higher than opisthorchiasis patients and controls (p = 0.010 and p viverrini infection than salivary IgA. In conclusion, specific antibodies to crude O. viverrini antigens were detected in saliva of patients with opisthorchiasis and CCA. Salivary antibodies reflect serum immune response to O. viverrini infection, and salivary IgG tends to be a good candidate for diagnosis of O. viverrini infection.

  16. GNAS1 T393C Polymorphism Is Associated with Clinical Course in Patients with Intrahepatic Cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Klaus J. Schmitz


    Full Text Available BACKGROUND/AIMS: The GNAS1 locus encodes the Gas protein, which stimulates the formation of cycloadenosinemonophosphate (cAMP. The cAMP pathway mediates pleiotropic effects, including the regulation of apoptosis and proliferation. We have recently shown that TT genotypes of the single-nucleotide polymorphism T393C in the gene GNAS1 predict the clinical outcome of patients with various carcinomas. METHODS: Eighty-seven patients with intrahepatic cholangiocarcinoma (ICC were retrospectively genotyped to elucidate a potential association between T393C genotypes and clinical outcome. RESULTS: ICCs of patients with homozygous TT genotypes revealed a higher proliferation rate and a lower apoptotic rate. Homozygous TT patients were at highest risk for cancer-related deaths (hazard ratio = 2.74; 95% confidence interval = 1.03-7.28 compared with C-allele carriers. Kaplan-Meier curves for disease-specific overall and local recurrence-free survival in a subgroup with Ro-resected ICC showed a significant association of T393 homozygosity with outcome, which was confirmed in multivariate Cox regression analysis. CONCLUSIONS: GNAS1 T393C is a novel independent host factor for disease progression in patients with ICC. Our finding that TT homozygosity (and not CC homozygosity was associated with unfavorable clinical outcome points to the complex and differing functional effects induced by GNAS1 T393C polymorphism in various human carcinomas.

  17. Human neutrophils facilitate tumor cell transendothelial migration.

    LENUS (Irish Health Repository)

    Wu, Q D


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

  18. Clonorchis sinensis ferritin heavy chain triggers free radicals and mediates inflammation signaling in human hepatic stellate cells. (United States)

    Mao, Qiang; Xie, Zhizhi; Wang, Xiaoyun; Chen, Wenjun; Ren, Mengyu; Shang, Mei; Lei, Huali; Tian, Yanli; Li, Shan; Liang, Pei; Chen, Tingjin; Liang, Chi; Xu, Jin; Li, Xuerong; Huang, Yan; Yu, Xinbing


    Clonorchiasis, caused by direct and continuous contact with Clonorchis sinensis, is associated with hepatobiliary damage, inflammation, periductal fibrosis, and the development of cholangiocarcinoma. Hepatic stellate cells respond to liver injury through production of proinflammatory mediators which drive fibrogenesis; however, their endogenous sources and pathophysiological roles in host cells were not determined. C. sinensis ferritin heavy chain (CsFHC) was previously confirmed as a component of excretory/secretory products and exhibited a number of extrahepatic immunomodulatory properties in various diseases. In this study, we investigated the expression pattern and biological role of CsFHC in C. sinensis. CsFHC was expressed throughout life stages of C. sinensis. More importantly, we found that treatment of human hepatic stellate cell line LX-2 with CsFHC triggered the production of free radicals via time-dependent activation of NADPH oxidase, xanthine oxidase, and inducible nitric oxide synthase. The increase in free radicals substantially promoted the degradation of cytosolic IκBα and nuclear translocation of NF-κB subunits (p65 and p50). CsFHC-induced NF-κB activation was markedly attenuated by preincubation with specific inhibitors of corresponding free radical-producing enzyme or the antioxidant. In addition, CsFHC induced an increased expression level of proinflammatory cytokines, IL-1β and IL-6, in NF-κB-dependent manner. Our results indicate that CsFHC-triggered free radical-mediated NF-κB signaling is an important factor in the chronic inflammation caused by C. sinensis infection.

  19. Intrahepatic and hilar mass-forming cholangiocarcinoma: Qualitative and quantitative evaluation with diffusion-weighted MR imaging

    Energy Technology Data Exchange (ETDEWEB)

    Fattach, Hassan El, E-mail: [Department of Abdominal Imaging, Hôpital Lariboisière, Assistance Publique-Hôpitaux de Paris, 2 rue Ambroise Paré, 75010 Paris (France); Dohan, Anthony, E-mail: [Department of Abdominal Imaging, Hôpital Lariboisière, Assistance Publique-Hôpitaux de Paris, 2 rue Ambroise Paré, 75010 Paris (France); Université Paris-Diderot, Sorbonne Paris Cité, 10 Avenue de Verdun, 75010 Paris (France); UMR INSERM 965-Paris 7 “Angiogenèse et recherche translationnelle”, 2 rue Amboise Paré, 75010 Paris (France); Guerrache, Youcef, E-mail: [Department of Abdominal Imaging, Hôpital Lariboisière, Assistance Publique-Hôpitaux de Paris, 2 rue Ambroise Paré, 75010 Paris (France); Dautry, Raphael, E-mail: [Department of Abdominal Imaging, Hôpital Lariboisière, Assistance Publique-Hôpitaux de Paris, 2 rue Ambroise Paré, 75010 Paris (France); Université Paris-Diderot, Sorbonne Paris Cité, 10 Avenue de Verdun, 75010 Paris (France); and others


    Highlights: • DW-MR imaging helps depicts all intrahepatic or hilar mass-forming cholangiocarcinomas. • DW-MRI provides best conspicuity of intrahepatic or hilar mass-forming cholangiocarcinomas than the other MRI sequences (P < 0.001). • The use of normalized ADC using the liver as reference organ results in the most restricted distribution of ADC values of intrahepatic or hilar mass-forming cholangiocarcinomas (variation coefficient = 16.6%). - Abstract: Objective: To qualitatively and quantitatively analyze the presentation of intrahepatic and hilar mass-forming cholangiocarcinoma with diffusion-weighted magnetic resonance imaging (DW-MRI). Materials and methods: Twenty-eight patients with histopathologically proven mass-forming cholangiocarcinoma (hilar, n = 17; intrahepatic, n = 11) underwent hepatic DW-MRI at 1.5-T using free-breathing acquisition and three b-values (0,400,800 s/mm{sup 2}). Cholangiocarcinomas were evaluated qualitatively using visual analysis of DW-MR images and quantitatively with conventional ADC and normalized ADC measurements using liver and spleen as reference organs. Results: All cholangiocarcinomas (28/28; 100%) were visible on DW-MR images. DW-MRI yielded best conspicuity of cholangiocarcinomas than the other MRI sequences (P < 0.001). Seven cholangiocarcinomas (7/11; 64%) showed hypointense central area on DW-MR images. Conventional ADC value of cholangiocarcinomas (1.042 × 10{sup −3} mm{sup 2}/s ± 0.221 × 10{sup −3} mm{sup 2}/s; range: 0.616 × 10{sup −3} mm{sup 2}/s to 2.050 × 10{sup −3} mm{sup 2}/s) was significantly lower than that of apparently normal hepatic parenchyma (1.362 × 10{sup −3} mm{sup 2}/s ± 0.187 × 10{sup −3} mm{sup 2}/s) (P < 0.0001), although substantial overlap was found. No significant differences in ADC and normalized ADC values were found between intrahepatic and hilar cholangiocarcinomas. The use of normalized ADC using the liver as reference organ resulted in the most restricted

  20. Macrophage stimulating protein variation enhances the risk of sporadic extrahepatic cholangiocarcinoma. (United States)

    Krawczyk, Marcin; Höblinger, Aksana; Mihalache, Florentina; Grünhage, Frank; Acalovschi, Monica; Lammert, Frank; Zimmer, Vincent


    Primary sclerosing cholangitis confers risk of cholangiocarcinoma. Here, we assessed the primary sclerosing cholangitis-associated variant rs3197999 in the MST1 gene, coding for RON receptor tyrosine kinase ligand macrophage stimulating protein, in a large European cholangiocarcinoma cohort. 223 cholangiocarcinoma patients including three primary sclerosing cholangitis individuals and 355 cancer- and primary sclerosing cholangitis-free controls were genotyped for MST1 rs3197999. The cancer group departed from Hardy-Weinberg equilibrium (p = 0.022) and exhibited a trend for rs3197999 [A] overrepresentation (31% vs. 26%: p = 0.10). Homozygous rs3197999 [AA] carrier status significantly increased overall (OR = 1.97; p = 0.023) and primary sclerosing cholangitis-unrelated biliary tract cancer risk (OR = 1.84; p = 0.044), relative to homozygous common allele carriers. The association was most pronounced in patients with extrahepatic tumours. This finding was robust to multivariate analysis (p < 0.05), validating the [AA] genotype as an independent cholangiocarcinoma risk factor. These results suggest that the [AA] genotype of the common MST1 variant rs3197999 enhances genetic risk of sporadic extrahepatic cholangiocarcinoma irrespective of primary sclerosing cholangitis status, presumably by modulating inflammatory responses and/or altered MSP/RON signalling. Copyright © 2013 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.

  1. Cholangiocarcinoma in Italy: A national survey on clinical characteristics, diagnostic modalities and treatment. Results from the "Cholangiocarcinoma" committee of the Italian Association for the Study of Liver disease. (United States)

    Alvaro, Domenico; Bragazzi, Maria Consiglia; Benedetti, Antonio; Fabris, Luca; Fava, Giammarco; Invernizzi, Pietro; Marzioni, Marco; Nuzzo, Gennaro; Strazzabosco, Mario; Stroffolini, Tommaso


    Very few studies assessed cholangiocarcinoma clinical characteristics. To evaluate the clinical characteristics of intra-hepatic (IH) and extra-hepatic (EH)-CCA. We performed a national survey based on a questionnaire. 218 cholangiocarcinomas were observed (47% EH-CCA, 53% IH-CCA) with an age at the diagnosis higher for EH-CCA. Coexistence of cirrhosis or viral cirrhosis was more frequent in IH-CCA than EH-CCA. An incidental asymptomatic presentation occurred in 28% of IH-CCA vs 4% EH-CCA whilst, 74% EH-CCA vs 28% IH-CCA presented with jaundice. 91% of IH-CCA presented as a single intra-hepatic mass, whilst 50% of EH-CCA was peri-hilar. In the diagnostic work-up, 70% of all cholangiocarcinoma cases received at least 3 different imaging procedures. Tissue-proven diagnosis was obtained in 80% cholangiocarcinoma. Open surgery with curative intent was performed in 45% of IH-CCA and 29% EH-CCA. 18% IH-CCA vs 4% EH-CCA did not received treatment. In Italy IH-CCA is managed as frequently as EH-CCA. In comparison to EH-CCA, IH-CCA occurs at younger age and is more frequently associated with cirrhosis and with an incidental asymptomatic presentation. In contrast, most EH-CCAs are jaundiced at the diagnosis. Cholangiocarcinoma diagnostic management is cost- and time-consuming with curative surgical treatment applicable more frequently in IH-CCA. Copyright © 2010 Editrice Gastroenterologica Italiana S.r.l. Published by Elsevier Ltd. All rights reserved.


    Institute of Scientific and Technical Information of China (English)


    Objective To definite the interactions between the human gastric carcinoma cell and the human vascular endothelial cell during the establishment and maintenance of the tumor vascular system and the tumor hematogenous metastasis.Methods We prepared the conditioned mediums of each cell so as to study the effect of the conditioned medium on itself or others by MTT colorimetry. The comprehensive effect of interactions between two cells was determined by stratified transfilter co-culture or direct contact co-culture.Results The conditioned medium of human gastric carcinoma cell can stimulate the proliferation of the human vascular endothelial cell, but the CM of HVEC can inhibit the growth of HGCC. Both kinds of cells can inhibit the growth of itself. The ultimate comprehensive effect of the interactions between two kinds of cells was increase of total cell numbers.Conclusion There exist the complicated interactions between the human gastric carcinoma cell and the human vascular endothelial cell during the tumor angiogenesis and the tumor hematogenous metastasis. The ultimate comprehensive effect of the interactions is increase of total cells numbers and tumor volume.

  3. Pulmonary Metastasis of Combined Hepatocellular and Cholangiocarcinoma: A Unique Radiographic Presentation with Air-space Consolidation Masquerading as Pneumonia and Primary Pulmonary Adenocarcinoma. (United States)

    Ishii, Takashi; Goto, Yasushi; Matsuzaki, Hirotaka; Ohishi, Nobuya; Sakamoto, Yoshihiro; Saito, Ruri; Matsusaka, Keisuke; Shibahara, Junji; Nagase, Takahide


    Lung metastasis showing radiographic findings of air-space consolidation is considered to be rare. This report describes the case of a man with progressive left lower lobe air-space consolidation with a history of hepatocellular carcinoma. The pulmonary lesion was initially suspected to be infection and later clinically diagnosed as primary adenocarcinoma of the lung. Although the patient was treated with systemic chemotherapy, the disease progressed very rapidly. A postmortem examination revealed that the alveolar spaces were filled with neoplastic cells subsequently proven to be metastases of combined hepatocellular and cholangiocarcinoma.

  4. Fibronectin production by human mammary cells

    Energy Technology Data Exchange (ETDEWEB)

    Stampfer, M.R. (Univ. of California, Berkeley); Vlodavsky, I.; Smith, H.S.; Ford, R.; Becker, F.F.; Riggs, J.


    Human mammary cells were examined for the presence of the high-molecular-weight surface glycoprotein fibronectin. Early passage mammary epithelial cell and fibroblast cultures from both carcinomas and normal tissues were tested for the presence of cell-associated fibronectin by immunofluorescence microscopy and for the synthesis and secretion of fibronectin by specific immunoprecipitation of metabolically labeled protein. In vivo frozen sections of primary carcinomas and normal tissues were tested for the localization of fibronectin by immunofluorescence microscopy. In contrast to the extensive fibrillar networks of fibronectin found in the fibroblast cultures, the epithelial cell cultures from both tissue sources displayed a pattern of cell-associated fibronectin characterizd by powdery, punctate staining. However, the cultured epithelial cells, as well as the fibroblasts, secreted large quantities of fibronectin into the medium. Putative myoepithelial cells also displayed extensive fibrillar networks of fibronectin. The difference in cell-associated fibronectin distribution between the epithelial cells and the fibroblasts and putative myoepithelial cells provided a simple means of quantitating stromal and myoepithelial cell contamination of the mammary epithelial cells in culture. In vivo, normal tissues showed fibronectin primarily localized in the basement membrane surrounding the epithelial cells and in the stroma. Most primary carcinomas displayed powdery, punctate staining on the epithelial cells in addition to the fibronectin present in the surrounding stroma.

  5. Laser printing of skin cells and human stem cells. (United States)

    Koch, Lothar; Kuhn, Stefanie; Sorg, Heiko; Gruene, Martin; Schlie, Sabrina; Gaebel, Ralf; Polchow, Bianca; Reimers, Kerstin; Stoelting, Stephanie; Ma, Nan; Vogt, Peter M; Steinhoff, Gustav; Chichkov, Boris


    Laser printing based on laser-induced forward transfer (LIFT) is a new biofabrication technique for the arrangement of biological materials or living cells in well-defined patterns. In the current study, skin cell lines (fibroblasts/keratinocytes) and human mesenchymal stem cells (hMSC) were chosen for laser printing experiments due to their high potential in regeneration of human skin and new application possibilities of stem cell therapy. To evaluate the influence of LIFT on the cells, their survival rate, their proliferation and apoptotic activity, and the DNA damages and modifications of their cell surface markers were assessed and statistically evaluated over several days. The cells survived the transfer procedure with a rate of 98%  +/- 1% standard error of the mean (skin cells) and 90%  +/- 10% (hMSC), respectively. All used cell types maintain their ability to proliferate after LIFT. Further, skin cells and hMSC did not show an increase of apoptosis or DNA fragmentation. In addition, the hMSC keep their phenotype as proven by fluorescence activated cell sorting (FACS) analysis. This study demonstrates LIFT as a suitable technique for unharmed computer-controlled positioning of different cell types and a promising tool for future applications in the ex vivo generation of tissue replacements.

  6. Human spleen and red blood cells (United States)

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


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

  7. Rotating cell culture systems for human cell culture: human trophoblast cells as a model. (United States)

    Zwezdaryk, Kevin J; Warner, Jessica A; Machado, Heather L; Morris, Cindy A; Höner zu Bentrup, Kerstin


    The field of human trophoblast research aids in understanding the complex environment established during placentation. Due to the nature of these studies, human in vivo experimentation is impossible. A combination of primary cultures, explant cultures and trophoblast cell lines support our understanding of invasion of the uterine wall and remodeling of uterine spiral arteries by extravillous trophoblast cells (EVTs), which is required for successful establishment of pregnancy. Despite the wealth of knowledge gleaned from such models, it is accepted that in vitro cell culture models using EVT-like cell lines display altered cellular properties when compared to their in vivo counterparts. Cells cultured in the rotating cell culture system (RCCS) display morphological, phenotypic, and functional properties of EVT-like cell lines that more closely mimic differentiating in utero EVTs, with increased expression of genes mediating invasion (e.g. matrix metalloproteinases (MMPs)) and trophoblast differentiation. The Saint Georges Hospital Placental cell Line-4 (SGHPL-4) (kindly donated by Dr. Guy Whitley and Dr. Judith Cartwright) is an EVT-like cell line that was used for testing in the RCCS. The design of the RCCS culture vessel is based on the principle that organs and tissues function in a three-dimensional (3-D) environment. Due to the dynamic culture conditions in the vessel, including conditions of physiologically relevant shear, cells grown in three dimensions form aggregates based on natural cellular affinities and differentiate into organotypic tissue-like assemblies. The maintenance of a fluid orbit provides a low-shear, low-turbulence environment similar to conditions found in vivo. Sedimentation of the cultured cells is countered by adjusting the rotation speed of the RCCS to ensure a constant free-fall of cells. Gas exchange occurs through a permeable hydrophobic membrane located on the back of the bioreactor. Like their parental tissue in vivo, RCCS

  8. Autophagy in human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Thien Tra

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

  9. Human embryonic stem cells for neuronal repair. (United States)

    Ben-Hur, Tamir


    Human embryonic stem cells may serve as a potentially endeless source of transplantable cells to treat various neurologic disorders. Accumulating data have shown the therapeutic value of various neural precursor cell types in experimental models of neurologic diseases. Tailoring cell therapy for specific disorders requires the generation of cells that are committed to specific neural lineages. To this end, protocols were recently developed for the derivation of dopaminergic neurons, spinal motor neurons and oligodendrocytes from hESC. These protocols recapitulate normal development in culture conditions. However, a novel concept emerging from these studies is that the beneficial effect of transplanted stem cells is not only via cell replacement in damaged host tissue, but also by trophic and protective effects, as well as by an immunomodulatory effect that down-regulates detrimental brain inflammation.

  10. Novel agents inhibit human leukemic cells

    Institute of Scientific and Technical Information of China (English)

    Wei-ping YU; Juan LI


    Ouabain (OUA) and pyrithione zinc (PZ) have been proved as the potential drugs for treating acute myeloid leukemia (AML).Selected from a screening among 1040 Food and Drug Administration-approved pharmacological agents,both drugs showability to induce apoptosis of the culturing AML cells,exhibiting the poisoning effect on the cells.Studies also reveal the efficiency of the drugs in inhibiting the growth of human AML cells injected into the mice lacking of immunity and killing primary AML cells from the peripheral blood of AML patients[1].

  11. Capsaicin Enhances the Drug Sensitivity of Cholangiocarcinoma through the Inhibition of Chemotherapeutic-Induced Autophagy.

    Directory of Open Access Journals (Sweden)

    Zai-Fa Hong

    Full Text Available Cholangiocarcinoma (CCA, a devastating cancer with a poor prognosis, is resistant to the currently available chemotherapeutic agents. Capsaicin, the major pungent ingredient found in hot red chili peppers of the genus Capsicum, suppresses the growth of several malignant cell lines. Our aims were to investigate the role and mechanism of capsaicin with respect to the sensitivity of CCA cells to chemotherapeutic agents. The effect of capsaicin on CCA tumor sensitivity to 5-fluorouracil (5-FU was assessed in vitro in CCA cells and in vivo in a xenograft model. The drug sensitivity of QBC939 to 5-FU was significantly enhanced by capsaicin compared with either agent alone. In addition, the combination of capsaicin with 5-FU was synergistic, with a combination index (CI < 1, and the combined treatment also suppressed tumor growth in the CCA xenograft to a greater extent than 5-FU alone. Further investigation revealed that the autophagy induced by 5-FU was inhibited by capsaicin. Moreover, the decrease in AKT and S6 phosphorylation induced by 5-FU was effectively reversed by capsaicin, indicating that capsaicin inhibits 5-FU-induced autophagy by activating the phosphoinositide 3-kinase (PI3K/protein kinase B (AKT/mammalian target of rapamycin (mTOR pathway in CCA cells. Taken together, these results demonstrate that capsaicin may be a useful adjunct therapy to improve chemosensitivity in CCA. This effect likely occurs via PI3K/AKT/mTOR pathway activation, suggesting a promising strategy for the development of combination drugs for CCA.

  12. Differential membrane proteomics using 18O-labeling to identify biomarkers for cholangiocarcinoma

    DEFF Research Database (Denmark)

    Kristiansen, Troels Zakarias; Harsha, H C; Grønborg, Mads


    Quantitative proteomic methodologies allow profiling of hundreds to thousands of proteins in a high-throughput fashion. This approach is increasingly applied to cancer biomarker discovery to identify proteins that are differentially regulated in cancers. Fractionation of protein samples based...... on enrichment of cellular subproteomes prior to mass spectrometric analysis can provide increased coverage of certain classes of molecules. We used a membrane protein enrichment strategy coupled with 18O labeling based quantitative proteomics to identify proteins that are highly expressed in cholangiocarcinomas....... In addition to identifying several proteins previously known to be overexpressed in cholangiocarcinoma, we discovered a number of molecules that were previously not associated with cholangiocarcinoma. Using immunoblotting and immunohistochemical labeling of tissue microarrays, we validated Golgi membrane...

  13. 3 CFR - Guidelines for Human Stem Cell Research (United States)


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

  14. Myristoylation profiling in human cells and zebrafish

    Directory of Open Access Journals (Sweden)

    Malgorzata Broncel


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

  15. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins (United States)

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


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

  16. Establishment of Human Neural Progenitor Cells from Human Induced Pluripotent Stem Cells with Diverse Tissue Origins. (United States)

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


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

  17. Efficient derivation and genetic modifications of human pluripotent stem cells on engineered human feeder cell lines. (United States)

    Zou, Chunlin; Chou, Bin-Kuan; Dowey, Sarah N; Tsang, Kitman; Huang, Xiaosong; Liu, Cyndi F; Smith, Cory; Yen, Jonathan; Mali, Prashant; Zhang, Yu Alex; Cheng, Linzhao; Ye, Zhaohui


    Derivation of pluripotent stem cells (iPSCs) induced from somatic cell types and the subsequent genetic modifications of disease-specific or patient-specific iPSCs are crucial steps in their applications for disease modeling as well as future cell and gene therapies. Conventional procedures of these processes require co-culture with primary mouse embryonic fibroblasts (MEFs) to support self-renewal and clonal growth of human iPSCs as well as embryonic stem cells (ESCs). However, the variability of MEF quality affects the efficiencies of all these steps. Furthermore, animal sourced feeders may hinder the clinical applications of human stem cells. In order to overcome these hurdles, we established immortalized human feeder cell lines by stably expressing human telomerase reverse transcriptase, Wnt3a, and drug resistance genes in adult mesenchymal stem cells. Here, we show that these immortalized human feeders support efficient derivation of virus-free, integration-free human iPSCs and long-term expansion of human iPSCs and ESCs. Moreover, the drug-resistance feature of these feeders also supports nonviral gene transfer and expression at a high efficiency, mediated by piggyBac DNA transposition. Importantly, these human feeders exhibit superior ability over MEFs in supporting homologous recombination-mediated gene targeting in human iPSCs, allowing us to efficiently target a transgene into the AAVS1 safe harbor locus in recently derived integration-free iPSCs. Our results have great implications in disease modeling and translational applications of human iPSCs, as these engineered human cell lines provide a more efficient tool for genetic modifications and a safer alternative for supporting self-renewal of human iPSCs and ESCs.

  18. [Human pluripotent stem cell and neural differentiation]. (United States)

    Wataya, Takafumi; Muguruma, Keiko; Sasai, Yoshiki


    Recovery of lost brain function is an important issue in medical studies because neurons of the central nervous system (CNS) have poor potential for regeneration. Since few CNS diseases can be treated completely by medicines, regenerative therapy by using stem cells should be studied as a new type of therapeutic intervention. The efficacy of cell replacement therapy in Parkinson's disease has been well investigated. Several studies on fetal tissue transplantation have revealed that quantity and purity of transplanted cells are necessary for recovery of symptoms. SFEB (Serum-free floating culture of embryoid body-like aggregates) method is capable of inducing multi-potential CNS progenitors that can be steered to differentiate into region-specific tissues. On the basis of the existing knowledge of embryology, we have succeeded in the generating of various types of neurons such as telencephalic, cerebeller (Purkinje and granule cells), retinal (photoreceptor cells) and hypothalamic neurons. Application of this culture method to human ES (hES) cells is necessary for clinical purpose: however, poor survival of hES cells in SFEB culture might limit the possibility of using these cells for future medical applications. We found that a selective Rho-associated kinase (ROCK) inhibitor, Y-27632, markedly diminished the dissociation-induced apoptosis of hES cells and enabled the cells to form aggregates in SFEB culture. For both mouse and human ES cells, SFEB culture is a favorable method that can generate large amounts of region-specific neurons. However, stem cell-based therapy continues to face several obstacles. It is important that researchers in the basic sciences and clinical medicine should discuss these problems together to overcome both scientific and ethical issues related to stem cells.

  19. Clinicopathological and prognostic significance of epithelial mesenchymal transition-related protein expression in intrahepatic cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Yao X


    Full Text Available Xing Yao,1,* Xiang Wang,1,* Zishu Wang,2,* Licheng Dai,1 Guolei Zhang,1 Qiang Yan,1 Weimin Zhou11Huzhou Central Hospital, Zhejiang Huzhou, 2Department of Medical Oncology, First Affiliated Hospital, Bengbu Medical College, Anhui, People’s Republic of China *These authors contributed equally to this workBackground: The aim of this study was to examine the patterns of expression of epithelial-mesenchymal transition (EMT-related proteins in intrahepatic cholangiocarcinoma. The clinicopathological and prognostic value of these markers was also evaluated.Methods: We detected the expression status of three EMT-related proteins, ie, E-cadherin, vimentin, and N-cadherin, by immunohistochemistry in consecutive intrahepatic cholangiocarcinoma specimens from 96 patients.Results: The frequency of loss of the epithelial marker E-cadherin, and acquisition of mesenchymal markers, vimentin and N-cadherin, in intrahepatic cholangiocarcinoma was 43.8%, 37.5% and 57.3%, respectively. Altered expression of EMT markers was associated with aggressive tumor behavior, including lymph node metastasis, undifferentiated-type histology, advanced tumor stage, venous invasion, and shorter overall survival. Moreover, loss of E-cadherin was retained as an independent prognostic factor for patients with intrahepatic cholangiocarcinoma in multivariate analysis.Conclusion: Our results suggest that the EMT process is associated with tumor progression and a poor outcome in patients with intrahepatic cholangiocarcinoma, and inhibition of EMT might offer novel promising molecular targets for the treatment of affected patients.Keywords: intrahepatic cholangiocarcinoma, epithelial-mesenchymal transition, expression, prognosis, immunohistochemistry

  20. T-cell response in human leishmaniasis

    DEFF Research Database (Denmark)

    Kharazmi, A; Kemp, K; Ismail, A


    In the present communication we provide evidence for the existence of a Th1/Th2 dichotomy in the T-cell response to Leishmania antigens in human leishmaniasis. Our data suggest that the pattern of IL-4 and IFN-gamma response is polarised in these patients. Lymphocytes from individuals recovered......+. Furthermore, IL-10 plays an important role in the development of post kala azar dermal leishmaniasis (PKDL) from VL. The balance between the parasitic-specific T-cell response plays an important regulatory role in determining the outcome of Leishmania infections in humans....

  1. Characterization of human pluripotent stem cells. (United States)

    Gokhale, Paul J; Andrews, Peter W


    Human pluripotent stem cells (PSCs), whether embryonic stem cells or induced PSCs, offer enormous opportunities for regenerative medicine and other biomedical applications once we have developed the ability to harness their capacity for extensive differentiation. Central to this is our ability to identify and characterize such PSCs, but this is fraught with potential difficulties that arise from a tension between functional definitions of pluripotency and the more convenient use of 'markers', a problem exacerbated by ethical issues, our lack of knowledge of early human embryonic development, and differences from the mouse paradigm.


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

  3. Human pluripotent stem cells in contemporary medicine

    Directory of Open Access Journals (Sweden)

    S. A. Rodin


    Full Text Available Human pluripotent stem cells (hPSCs are capable of indefinite proliferation and can be differentiated into any cell type of the human body. Therefore, they are a promising source of cells for treatment of numerous degenerative diseases and injuries. Pluripotent stem cells are also associated with a number of ethical, safety and technological issues. In this review, we describe various types of hPSCs, safety issues that concern all or some types of hPSCs and methods of clinical-grade hPSC line development. Also, we discuss current and past clinical trials involving hPSCs, their outcomes and future perspectives of hPSC-based therapy. 


    Directory of Open Access Journals (Sweden)

    Daniel V. Kostov


    Full Text Available Purpose: To compare the opportunities of the intraoperative ultrasound examination of the liver with the preoperative imaging methods – Computer tomography (CT, Ultrasound examination (US in detecting hepatic lesions in patients with tumor of Klatskin (hilar cholangiocarcinoma.Patients and methods: This is a prospective study of 7 patients, which had undergone open operative intervention for tumor of Klatskin. The affecting of the liver by the tumor process was examined preoperatively with Ultrasound examination and Computer tomography with contrast amplification, and by inspection and palpation and intra operative ultrasound during the operative intervention.Results: The maximum numbers of malignant lesions are found by IOUS - totally 21. Preoperative ultrasound found 5 lesions or 23,80% of the maximum number and preoperative CT – 9 or 33,33%. The information from IOUS changed the operative plan in 2/7 of the patients (28,57%. A biliary stent was placed under the control of IOUS in one patient with a heavy hyperbilirubinemia – 791,6 µmol/l.Conclusions: IOUS is significantly helpful in surgery of tumor of Klatskin. Besides the precise localization and staging of the tumor the information of IOUS changes the surgical strategy in some patients.

  5. An inguinal hernia sac tumor of extrahepatic cholangiocarcinoma origin

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    Yamazaki Hidehiro


    Full Text Available Abstract Background Metastatic hernia sac tumor from biliary malignancy is extremely rare with only one such case previously reported. We herein report an additional case of extrahepatic cholangiocarcinoma presenting as a hernia sac tumor. Case presentation A 78-year-old man presented with an irreducible right inguinal hernia associated with a firm tumor, 2.0 cm in diameter. A computed tomography scan demonstrated a soft tissue density mass with heterogeneous enhancement within the right inguinal canal. The patient underwent a hernia repair and the hernia sac tumor was resected. Histological examination of the tumor revealed a metastatic adenocarcinoma suggesting the tumor was of pancreato-biliary origin. Further investigation using imaging studies disclosed a primary tumor in the upper bile duct. The patient died of the disease nine months after the resection. Conclusion Hernia sac tumors should be considered when an irreducible, growing mass appears within an inguinal hernia. Computed tomography may be useful for the early detection of hernia sac tumors from undiagnosed intra-abdominal malignancies.

  6. Outcome of curative resection for perihilar cholangiocarcinoma in Northeast Thailand

    Institute of Scientific and Technical Information of China (English)

    Attapol; Titapun; Ake; Pugkhem; Vor; Luvira; Tharatip; Srisuk; Ongart; Somintara; O-tur; Saeseow; Anan; Sripanuskul; Anongporn; Nimboriboonporn; Bandit; Thinkhamrop; Narong; Khuntikeo


    AIM: To examine survival outcomes of perihilar cholangiocarcinoma(PCCA) resection including mortality, morbidity and prognostic factors. METHODS: Multivariate analyses were carried out based on the survival data of all patients with histologically confirmed PCCA who underwent curative resection at Srinagarind Hospital from January 2006 to December 2011. RESULTS: There were 29(19%) cases of intrahepatic CCA that involved hilar and 124(81%) with hilar bileduct cancer. R0 resection was carried out on 66(43.1%) patients of whom 50(32.7%) also had lymph node metastasis. The other patients underwent R1 resection. The overall 5-year survival rate was 20.6%(95%CI: 13.8-28.4) and median survival time was 19.9 mo. Postoperative mortality was 2%, and 30% of patients had complications. Patients without lymph node metastasis were 60% less likely to die than those with metastasis. Achieving R0 led to a 58% reduction in the chance of mortality as compared to R1. CONCLUSION: To achieve a better survival outcome, focus should center on performing radical surgery and detection of patients with early stage cancer.

  7. Cell pattern in adult human corneal endothelium.

    Directory of Open Access Journals (Sweden)

    Carlos H Wörner

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

  8. Merkel cell distribution in the human eyelid

    Directory of Open Access Journals (Sweden)

    C.A. May


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

  9. Natural killer cells in human autoimmune disorders (United States)


    Natural killer (NK) cells are innate lymphocytes that play a critical role in early host defense against viruses. Through their cytolytic capacity and generation of cytokines and chemokines, NK cells modulate the activity of other components of the innate and adaptive immune systems and have been implicated in the initiation or maintenance of autoimmune responses. This review focuses on recent research elucidating a potential immunoregulatory role for NK cells in T-cell and B-cell-mediated autoimmune disorders in humans, with a particular focus on multiple sclerosis, rheumatoid arthritis, and systemic lupus erythematous. A better understanding of the contributions of NK cells to the development of autoimmunity may lead to novel therapeutic targets in these diseases. PMID:23856014

  10. Human Colon Cancer Cells Cultivated in Space (United States)


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

  11. Repetitive response to gemcitabine that led to curative resection in cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Seong Hun Kim; In Hee Kim; Sang Wook Kim; Seung-Ok Lee


    This study reports a case of unresectable intrahepatic mass-forming cholangiocarcinoma which showed a dramatic response to gemcitabine that led to curative resection and a long-term survival of more than five years. Six and five cycles of gemcitabine monotherapy were administered separately over a three-year period and a radical excision was performed at 4.5 years after diagnosis. This case indicates the role of gemcitabine as a neoadjuvant chemotherapeutic agent for cholangiocarcinoma and guarantees a randomized controlled prospective study.

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

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    Haifeng Lan


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

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

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


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

  14. Genetic Manipulation of Human Embryonic Stem Cells. (United States)

    Eiges, Rachel


    One of the great advantages of embryonic stem (ES) cells over other cell types is their accessibility to genetic manipulation. They can easily undergo genetic modifications while remaining pluripotent, and can be selectively propagated, allowing the clonal expansion of genetically altered cells in culture. Since the first isolation of ES cells in mice, many effective techniques have been developed for gene delivery and manipulation of ES cells. These include transfection, electroporation, and infection protocols, as well as different approaches for inserting, deleting, or changing the expression of genes. These methods proved to be extremely useful in mouse ES cells, for monitoring and directing differentiation, discovering unknown genes, and studying their function, and are now being extensively implemented in human ES cells (HESCs). This chapter describes the different approaches and methodologies that have been applied for the genetic manipulation of HESCs and their applications. Detailed protocols for generating clones of genetically modified HESCs by transfection, electroporation, and infection will be described, with special emphasis on the important technical details that are required for this purpose. All protocols are equally effective in human-induced pluripotent stem (iPS) cells.

  15. 内磁场在磁靶向治疗胆道恶性肿瘤中作用的实验研究%Experimental study on magnetic drug targeting in treating cholangiocarcinoma based on internal magnetic fields

    Institute of Scientific and Technical Information of China (English)

    郑建伟; 王剑明; 唐涛


    Objective: To evaluate the effect of magnetic nanoparticle containing 5-fluorouracil (5-FU) targeting in treating cholangiocarcinoma based on internal magnetic fields built inside the tumor.Methods: 32 nude mice of BABL/C bearing ectopic tumor were built by subcutaneouly injecting cholangiocarcinoma cell line QBC 939.Three weeks after tumor inoculation,the animal models were divided equally into four groups at random including: (a) group A,consisting of internal magnetic field built by magnetic biliary stent wires inserted into tumor tissue and receiving magnetic nanoparticles containing 5-FU administered via tail vein injection at 250 mg/kg for consecutive five days; (b) group B,receiving placebo (sodium chloride); (c) group C,receiving pure magnetic biliary stent wires without the applying of magnetic nanoparticles; (d) group D,consisting of external magnetic fields and the same treatment of magnetic nanoparticles containing 5-FU as group A.The tumor volumes were measured every 3 days,totally six times from treatment started.Tumor tissues were observed by transmission electron microscope when the nude mice were killed after the observation period.Results: The experimental group (group A) showed significantly therapeutic efficacy.Moreover,apoptosis of tumor cells could be easily detected in this group.Conclusion: Magnetic particles containing5-FU combined with internal magnetic field can effectively treat cholangiocarcinoma,and its therapeutic efficacy is better than that of the traditional method based on external magnetic fields.

  16. Enriched retinal ganglion cells derived from human embryonic stem cells (United States)

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


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

  17. Human ES cells: starting culture from frozen cells. (United States)

    Trish, Erin; Dimos, John; Eggan, Kevin


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

  18. Sodium Valproate Induces Cell Senescence in Human Hepatocarcinoma Cells

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    Hong-Mei An


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

  19. Human embryonic stem cell lines model experimental human cytomegalovirus latency. (United States)

    Penkert, Rhiannon R; Kalejta, Robert F


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

  20. Mutation inactivation of Nijmegen breakage syndrome gene (NBS1 in hepatocellular carcinoma and intrahepatic cholangiocarcinoma.

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    Yan Wang

    Full Text Available Nijmegen breakage syndrome (NBS with NBS1 germ-line mutation is a human autosomal recessive disease characterized by genomic instability and enhanced cancer predisposition. The NBS1 gene codes for a protein, Nbs1(p95/Nibrin, involved in the processing/repair of DNA double-strand breaks. Hepatocellular carcinoma (HCC is a complex and heterogeneous tumor with several genomic alterations. Recent studies have shown that heterozygous NBS1 mice exhibited a higher incidence of HCC than did wild-type mice. The objective of the present study is to assess whether NBS1 mutations play a role in the pathogenesis of human primary liver cancer, including HBV-associated HCC and intrahepatic cholangiocarcinoma (ICC. Eight missense NBS1 mutations were identified in six of 64 (9.4% HCCs and two of 18 (11.1% ICCs, whereas only one synonymous mutation was found in 89 control cases of cirrhosis and chronic hepatitis B. Analysis of the functional consequences of the identified NBS1 mutations in Mre11-binding domain showed loss of nuclear localization of Nbs1 partner Mre11, one of the hallmarks for Nbs1 deficiency, in one HCC and two ICCs with NBS1 mutations. Moreover, seven of the eight tumors with NBS1 mutations had at least one genetic alteration in the TP53 pathway, including TP53 mutation, MDM2 amplification, p14ARF homozygous deletion and promoter methylation, implying a synergistic effect of Nbs1 disruption and p53 inactivation. Our findings provide novel insight on the molecular pathogenesis of primary liver cancer characterized by mutation inactivation of NBS1, a DNA repair associated gene.

  1. [Immune system evolution. (From cells to humans)]. (United States)

    Belek, A S


    The great variety of cells and molecules observed in the mammalian immune system can be explained by stepwise acquisition of them during phylogeny. Self/nonself discrimination and cell-mediated immunity have been present since the early stages of evolution. Although some inducible antimicrobial molecules have been demonstrated in invertebrates, immunoglobulins appear in vertebrates. T and B cell diversity, development of the lymphoid organs, MHC molecules, complement and cytokines are the characteristics that appear through the evolution of vertebrates. Further knowledge that will be obtained from phylogenetic studies will improve our understanding of the immune system of human.

  2. Centre for human development, stem cells & regeneration. (United States)

    Oreffo, Richard O C


    The Centre for Human Development, Stem Cells and Regeneration (CHDSCR) was founded in 2004 as a cross-disciplinary research and translational program within the Faculty of Medicine at the University of Southampton. The Centre undertakes fundamental research into early development and stem cells together with applied translational research for patient benefit. The Centre has vibrant and thriving multidisciplinary research programs that harness the translational strength of the Faculty together with an innovative Stem Cell PhD program, outstanding clinical infrastructure and enterprise to deliver on this vision.

  3. Biobanking human embryonic stem cell lines

    DEFF Research Database (Denmark)

    Holm, Søren


    Stem cell banks curating and distributing human embryonic stem cells have been established in a number of countries and by a number of private institutions. This paper identifies and critically discusses a number of arguments that are used to justify the importance of such banks in policy...... are curiously absent from the particular stem cell banking policy discourse. This to some extent artificially isolates this discourse from the broader discussions about the flows of reproductive materials and tissues in modern society, and such isolation may lead to the interests of important actors being...

  4. A granulin-like growth factor secreted by the carcinogenic liver fluke, Opisthorchis viverrini, promotes proliferation of host cells.

    Directory of Open Access Journals (Sweden)

    Michael J Smout


    Full Text Available The human liver fluke, Opisthorchis viverrini, infects millions of people throughout south-east Asia and is a major cause of cholangiocarcinoma, or cancer of the bile ducts. The mechanisms by which chronic infection with O. viverrini results in cholangiocarcinogenesis are multi-factorial, but one such mechanism is the secretion of parasite proteins with mitogenic properties into the bile ducts, driving cell proliferation and creating a tumorigenic environment. Using a proteomic approach, we identified a homologue of human granulin, a potent growth factor involved in cell proliferation and wound healing, in the excretory/secretory (ES products of the parasite. O. viverrini granulin, termed Ov-GRN-1, was expressed in most parasite tissues, particularly the gut and tegument. Furthermore, Ov-GRN-1 was detected in situ on the surface of biliary epithelial cells of hamsters experimentally infected with O. viverrini. Recombinant Ov-GRN-1 was expressed in E. coli and refolded from inclusion bodies. Refolded protein stimulated proliferation of murine fibroblasts at nanomolar concentrations, and proliferation was inhibited by the MAPK kinase inhibitor, U0126. Antibodies raised to recombinant Ov-GRN-1 inhibited the ability of O. viverrini ES products to induce proliferation of murine fibroblasts and a human cholangiocarcinoma cell line in vitro, indicating that Ov-GRN-1 is the major growth factor present in O. viverrini ES products. This is the first report of a secreted growth factor from a parasitic worm that induces proliferation of host cells, and supports a role for this fluke protein in establishment of a tumorigenic environment that may ultimately manifest as cholangiocarcinoma.

  5. A granulin-like growth factor secreted by the carcinogenic liver fluke, Opisthorchis viverrini, promotes proliferation of host cells.

    Directory of Open Access Journals (Sweden)

    Michael J Smout


    Full Text Available The human liver fluke, Opisthorchis viverrini, infects millions of people throughout south-east Asia and is a major cause of cholangiocarcinoma, or cancer of the bile ducts. The mechanisms by which chronic infection with O. viverrini results in cholangiocarcinogenesis are multi-factorial, but one such mechanism is the secretion of parasite proteins with mitogenic properties into the bile ducts, driving cell proliferation and creating a tumorigenic environment. Using a proteomic approach, we identified a homologue of human granulin, a potent growth factor involved in cell proliferation and wound healing, in the excretory/secretory (ES products of the parasite. O. viverrini granulin, termed Ov-GRN-1, was expressed in most parasite tissues, particularly the gut and tegument. Furthermore, Ov-GRN-1 was detected in situ on the surface of biliary epithelial cells of hamsters experimentally infected with O. viverrini. Recombinant Ov-GRN-1 was expressed in E. coli and refolded from inclusion bodies. Refolded protein stimulated proliferation of murine fibroblasts at nanomolar concentrations, and proliferation was inhibited by the MAPK kinase inhibitor, U0126. Antibodies raised to recombinant Ov-GRN-1 inhibited the ability of O. viverrini ES products to induce proliferation of murine fibroblasts and a human cholangiocarcinoma cell line in vitro, indicating that Ov-GRN-1 is the major growth factor present in O. viverrini ES products. This is the first report of a secreted growth factor from a parasitic worm that induces proliferation of host cells, and supports a role for this fluke protein in establishment of a tumorigenic environment that may ultimately manifest as cholangiocarcinoma.

  6. Advances in human B cell phenotypic profiling

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    Denise A Kaminski


    Full Text Available To advance our understanding and treatment of disease, research immunologists have been called-upon to place more centralized emphasis on impactful human studies. Such endeavors will inevitably require large-scale study execution and data management regulation (Big Biology, necessitating standardized and reliable metrics of immune status and function. A well-known example setting this large-scale effort in-motion is identifying correlations between eventual disease outcome and T lymphocyte phenotype in large HIV-patient cohorts using multiparameter flow cytometry. However, infection, immunodeficiency, and autoimmunity are also characterized by correlative and functional contributions of B lymphocytes, which to-date have received much less attention in the human Big Biology enterprise. Here, we review progress in human B cell phenotyping, analysis, and bioinformatics tools that constitute valuable resources for the B cell research community to effectively join in this effort.

  7. Human plasma cells express granzyme B. (United States)

    Xu, Wei; Narayanan, Priya; Kang, Ning; Clayton, Sandra; Ohne, Yoichiro; Shi, Peiqing; Herve, Marie-Cecile; Balderas, Robert; Picard, Capucine; Casanova, Jean-Laurent; Gorvel, Jean-Pierre; Oh, Sangkon; Pascual, Virginia; Banchereau, Jacques


    While studying the plasma cell (PC) compartment in human tonsils, we identified that immunoglobulin kappa or lambda chain-expressing PCs are the main cells expressing granzyme B (GrzB). In vitro studies revealed that activated B cells differentiated into GrzB-expressing PCs when co-cultured with macrophages and follicular helper T cells. This effect could be reproduced on combined stimulation of IL-15 (produced by macrophages) and IL-21 (produced by T follicular helper cells) in a STAT3-dependent manner. Whereas IL-21 triggers the transcription of mRNA of GrzB, IL-15 synergizes the translation of GrzB proteins. The precise role of GrzB in PC biology remains to be understood and studies in mice will not help as their PCs do not express GrzB.

  8. Human CD56bright NK Cells

    DEFF Research Database (Denmark)

    Michel, Tatiana; Poli, Aurélie; Cuapio, Angelica


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

  9. Human embryonic stem cells: preclinical perspectives

    Directory of Open Access Journals (Sweden)

    Sarda Kanchan


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

  10. Stem cell factor and c-Kit in human primordial germ cells and fetal ovaries

    DEFF Research Database (Denmark)

    Høyer, Poul Erik; Byskov, Anne Grete; Møllgård, Kjeld


    Prenatal ovary (human), Primordial germ cells, Folliculogenesis, c-Kit, Stem cell factor, immunohistochemistry......Prenatal ovary (human), Primordial germ cells, Folliculogenesis, c-Kit, Stem cell factor, immunohistochemistry...

  11. The Expression and Significance of DNMT3b Gene in Extrahepatic Cholangiocarcinoma Tissues%DNA甲基转移酶3b基因在肝外胆管癌组织中的表达

    Institute of Scientific and Technical Information of China (English)

    左石; 邹声泉; 孙诚谊


    Objective: To study the expression of DNA methyltransferase 3b (DNMT3b) in human extrahepatic cholangiocarcinoma tissues, analyze the relationship between its expression and clinico-pathologic features of the disease, and to explore the roles of DNMT3b during tumorigenesis and development of the disease. Methods: The expression of DNMT3b protein was detected in 24 extrahepatic cholangiocarcinoma specimens and 20 chronic cholangeitis tissue specimens by immunohistochemistry method. The results were compared between groups, and analyzed in association with clinicopatholigic features of extrahepatic cholangiocarcinoma. Results; (1) The positive cell rates of DNMT3b expression were (46. 56 ±12. 12)% and (3. 34 ±2. 73)% in extrahepatic cholangiocarcinoma specimens and in chronic cholangeitis tissues respectively. The difference between the two groups ( P = 0. 000) was significant; (2) The expression of DNMT3b protein was correlated with differentiation grade of the carcinoma. The protein levels of DNMT3b increased progressively along with the adenocarcinoma differentiation from high to moderate, and to poor. The abnormal expression of DNMT3b protein did not correlate with gender, age, tumor sizes, tumor location, metastasis of lymph node or clinical TNM stage. Conclusions: The expression of DNMT3b protein in extrahepatic cholangiocarcinoma is signifi-cantly higher than that in chronic cholangeitis tissues. DNMT3b protein level is correlated with differentiation grade of tumor, but not correlated with gender, age, tumor size, tumor location, metastasis of lymph node, or clinical TNM stage. This result suggests that over-expression of DNMT3b may relate to the pathogenesis of biliary tract carcinoma and might be an early molecular event during genesis of biliary tract carcinoma.%目的:研究DNA甲基转移酶3b( DNMT3b)在人肝外胆管癌组织中的表达情况,并分析DNMT3b的表达与胆管癌临床病理因素之间的关系,探讨其在胆管癌发生发

  12. Molecular and cytogenetic abnormality on cholangiocarcinoma%肝门胆管癌细胞分子遗传学异常的研究

    Institute of Scientific and Technical Information of China (English)

    高戈; 邹声泉


    Objective To study alterations of molecular cytogenetics of cholangiocarcinoma on chromosome and gene,study the possible involvement of chromosome abnormalities and the genetic susceptibility in pathogenesis of cholangiocarcinoma,investigate chromosome instability and expression of fragile sites of cholangiocarcinoma and relationship each other,and find marker chromosome of cholangiocarcinoma.Methods Chromosomal analysis and fluorescence in situ hybridization (FISH) were used with 9th chromosome libraries to investigate the chromosomal aberration,the chromosome instability and expressiom of fragile sites in patients with cholangiocarcinoma.The genetic alteration and expression of p16 gene were analyzed by polymerase chain reaction single strand comformation polymorphism (PCR-SSCP) and immunohistochemistry.Results Chromosomal aberrations were found in 10 case of patients and 1 case of cell line of cholangiocarcinoma.The chromosome model number was 62.3 and predominantly hyperdiploid,with a chromosome number aberration rate of 83.67%,and a structure aberration rate of 62.8%.9p deletion appeared more frequently(50.8%).p16 gene mutation rate was 58.3% (21/36).The expression of p16 gene was not significantly related with sex and age of the patients,and had a relation with Bismuth type,the depth of invasion and lymph node metastasis.The chromosome gaps (8.75 ± 3.30),chromosome breakpoints (7.63 ± 2.76),abnormal cell rate (13.35 ± 4.73) and expression rate of fragile sites of the patients were markedly higher in patients with cholgnagiocarcinoma than those of normal persons (3.17 ±1.82,2.04±1.76,3.65 ±1.97) (P<0.01).Conclusion Chromosomal aberrations and gene mutation may play an important role in the pathogenesis of cholangiocarcinoma.The alteration of p16 gene and abnormal expression of p16 protein are significantly correlated with the biological behaviors and clinical staging of cholangiocarcinorma and may hence be helpful to prognosis.The increase of

  13. Differences in the microrheology of human embryonic stem cells and human induced pluripotent stem cells. (United States)

    Daniels, Brian R; Hale, Christopher M; Khatau, Shyam B; Kusuma, Sravanti; Dobrowsky, Terrence M; Gerecht, Sharon; Wirtz, Denis


    Embryonic and adult fibroblasts can be returned to pluripotency by the expression of reprogramming genes. Multiple lines of evidence suggest that these human induced pluripotent stem (hiPS) cells and human embryonic stem (hES) cells are behaviorally, karyotypically, and morphologically similar. Here we sought to determine whether the physical properties of hiPS cells, including their micromechanical properties, are different from those of hES cells. To this end, we use the method of particle tracking microrheology to compare the viscoelastic properties of the cytoplasm of hES cells, hiPS cells, and the terminally differentiated parental human fibroblasts from which our hiPS cells are derived. Our results indicate that although the cytoplasm of parental fibroblasts is both viscous and elastic, the cytoplasm of hiPS cells does not exhibit any measurable elasticity and is purely viscous over a wide range of timescales. The viscous phenotype of hiPS cells is recapitulated in parental cells with disassembled actin filament network. The cytoplasm of hES cells is predominantly viscous but contains subcellular regions that are also elastic. This study supports the hypothesis that intracellular elasticity correlates with the degree of cellular differentiation and reveals significant differences in the mechanical properties of hiPS cells and hES cells. Because mechanical stimuli have been shown to mediate the precise fate of differentiating stem cells, our results support the concept that stem cell "softness" is a key feature of force-mediated differentiation of stem cells and suggest there may be subtle functional differences between force-mediated differentiation of hiPS cells and hES cells.

  14. Exome sequencing identifies frequent inactivating mutations in BAP1, ARID1A and PBRM1 in intrahepatic cholangiocarcinomas (United States)

    Selaru, Florin M; Streppel, Mirte M; Lucas, Donald J; Niknafs, Noushin; Guthrie, Violeta Beleva; Maitra, Anirban; Argani, Pedram; Offerhaus, G Johan A; Roa, Juan Carlos; Roberts, Lewis R; Gores, Gregory J; Popescu, Irinel; Alexandrescu, Sorin T; Dima, Simona; Fassan, Matteo; Simbolo, Michele; Mafficini, Andrea; Capelli, Paola; Lawlor, Rita T; Ruzzenente, Andrea; Guglielmi, Alfredo; Tortora, Giampaolo; de Braud, Filippo; Scarpa, Aldo; Jarnagin, William; Klimstra, David; Karchin, Rachel; Velculescu, Victor E; Hruban, Ralph H; Vogelstein, Bert; Kinzler, Kenneth W; Papadopoulos, Nickolas; Wood, Laura D


    Through exomic sequencing of 32 intrahepatic cholangiocarcinomas, we discovered frequent inactivating mutations in multiple chromatin-remodeling genes (including BAP1, ARID1A and PBRM1), and mutation in one of these genes occurred in almost half of the carcinomas sequenced. We also identified frequent mutations at previously reported hotspots in the IDH1 and IDH2 genes encoding metabolic enzymes in intrahepatic cholangiocarcinomas. In contrast, TP53 was the most frequently altered gene in a series of nine gallbladder carcinomas. These discoveries highlight the key role of dysregulated chromatin remodeling in intrahepatic cholangiocarcinomas. PMID:24185509

  15. Generation of mature hematopoietic cells from human pluripotent stem cells. (United States)

    Togarrati, Padma Priya; Suknuntha, Kran


    A number of malignant and non-malignant hematological disorders are associated with the abnormal production of mature blood cells or primitive hematopoietic precursors. Their capacity for continuous self-renewal without loss of pluripotency and the ability to differentiate into adult cell types from all three primitive germ layers make human embryonic stem cells and induced pluripotent stem cells (hiPSCs) attractive complementary cell sources for large-scale production of transfusable mature blood cell components in cell replacement therapies. The generation of patient-specific hematopoietic stem/precursor cells from iPSCs by the regulated manipulation of various factors involved in reprograming to ensure complete pluripotency, and developing innovative differentiation strategies for generating unlimited supply of clinically safe, transplantable, HLA-matched cells from hiPSCs to outnumber the inadequate source of hematopoietic stem cells obtained from cord blood, bone marrow and peripheral blood, would have a major impact on the field of regenerative and personalized medicine leading to translation of these results from bench to bedside.

  16. Human colostral cells. I. Separation and characterization. (United States)

    Crago, S S; Prince, S J; Pretlow, T G; McGhee, J R; Mestecky, J


    Analyses of the cells present in human colostrum obtained from fifty-four healthy donors during the first four days of lactation revealed that there were 3.3 x 10(6) (range 1.1 x 10(5)--1.2 x 10(7)) cells per ml of colostrum. Based on histochemical examinations, it was found that this population consisted of 30--47% macrophages, 40--60% polymorphonuclear leucocytes, 5.2--8.9% lymphocytes, and 1.3--2.8% colostral corpuscles; epithelial cells were rarely encountered. The identity of various cell types was confirmed by Wright's stain and by a series of histochemical techniques which disclosed the presence of non-specific esterase, peroxidase, and lipids. For further characterization, the different types of cells were separated by various methods, such as Ficoll-Hypaque density centrifugation, isokinetic centrifugation on a linear Ficoll gradient, adherence to glass or plastic, and phagocytosis of carbonyl iron. Immunohistochemical staining with FITC- and/or TRITC-labelled reagents to IgA, IgM, IgG, K- and lambda-chains, secretory component, lactoferrin, and alpha-lactalbumin were applied to unseparated as well as separated colostral cells. Polymorphonuclear leucocytes (staining for peroxidase) as well as macrophages and colostral corpuscles (staining for non-specific esterase) exhibited numerous intracellular vesicles that contained lipids as well as various combinations of milk proteins. Lymphoid cells did not stain with any of these reagents and plasma cells were not detected among the colostral cells. Individual phagocytic cells contained immunoglobulins of the IgA and IgM classes, both K and lambda light chains, secretory component, lactoferrin, and alpha-lactalbumin. The coincidental appearance of these proteins in single, phagocytic cells but not in lymphoid cells indicate that the cells acquired these proteins by ingestion from the environment. Markers commonly used for the identification of B lymphocytes (surface immunoglobulins) and T lymphocytes (receptors

  17. 21 CFR 864.2280 - Cultured animal and human cells. (United States)


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

  18. Peritumoral SPARC expression and patient outcome with resectable intrahepatic cholangiocarcinoma

    Directory of Open Access Journals (Sweden)

    Cheng CT


    Full Text Available Chi-Tung Cheng,1,* Yin-Yi Chu,2,* Chun-Nan Yeh,1 Shih-Chiang Huang,3 Ming Huang Chen,4 Shang-Yu Wang,1 Chun-Yi Tsai,1 Kun-Chun Chiang,5 Yen-Yang Chen,6,7 Ming-Chun Ma,6,7 Chien-Ting Liu,6,7 Tsung-Wen Chen,1 Ta-Sen Yeh11Department of Surgery, 2Department of Gastroenterology and Hepatology, 3Department of Pathology, Chang Gung Memorial Hospital, Chang Gung University, Taoyuan, Taiwan; 4Division of Hematology and Oncology, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan; 5Department of Surgery, Chang Gung Memorial Hospital, Chang Gung University, Keelung, Taiwan; 6Division of Hematology and Oncology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan; 7Chang Gung University, Taoyuan, Taiwan*These authors contributed equally to this workBackground and objectives: Cholangiocarcinoma (CCA affects thousands worldwide with increasing incidence. SPARC (secreted protein acidic and rich in cysteine plays an important role in cellular matrix interactions, wound repair, and cellular migration, and has been reported to prevent malignancy from growth. SPARC undergoes epigenetic silencing in pancreatic malignancy, but is frequently expressed by stromal fibroblasts adjacent to infiltrating pancreatic adenocarcinomas. CCA is also a desmoplastic tumor, similar to pancreatic adenocarcinoma. SPARC’s clinical influence on clinicopathological characteristics of mass-forming (MF-CCA still remains unclear. In this study, we evaluate the expression of SPARC in tumor and stromal tissue to clarity its relation with prognosis.Methods: Seventy-eight MF-CCA patients who underwent hepatectomy with curative intent were enrolled for an immunohistochemical study of SPARC. The expression of immunostaining of SPARC was characterized for both tumor and stromal tissues. We conducted survival analysis with 16 clinicopathological variables. The overall survival (OS was analyzed by Kaplan–Meier analysis and Cox

  19. DNA repair responses in human skin cells

    Energy Technology Data Exchange (ETDEWEB)

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


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

  20. Prognostic significance of snail expression in hilar cholangiocarcinoma

    Energy Technology Data Exchange (ETDEWEB)

    Kong, Dalu [Department of Hepatobiliary Surgery, Tianjin Medical University Cancer Institute and Hospital, Hexi District, Tianjin (China); Liang, Jun [Department of Oncology, Affiliated Hospital of Medical College, Qingdao University, Qingdao, Shandong Province (China); Li, Rong [Department of Hepatobiliary Surgery, Tianjin Medical University Cancer Institute and Hospital, Hexi District, Tianjin (China); Liu, Shihai [Department of Laboratory Center, Affiliated Hospital of Medical College, Qingdao University, Qingdao, Shandong Province (China); Wang, Jigang [Department of Oncology, Affiliated Hospital of Medical College, Qingdao University, Qingdao, Shandong Province (China); Zhang, Kejun; Chen, Dong [Department of General Surgery, Affiliated Hospital of Medical College, Qingdao University, Qingdao, Shandong Province (China)


    Many patients with hilar cholangiocarcinoma (HC) have a poor prognosis. Snail, a transcription factor and E-cadherin repressor, is a novel prognostic factor in many cancers. The aim of this study was to evaluate the relationship between snail and E-cadherin protein expression and the prognostic significance of snail expression in HC. We examined the protein expression of snail and E-cadherin in HC tissues from 47 patients (22 males and 25 females, mean age 61.2 years) using immunohistochemistry and RT-PCR. Proliferation rate was also evaluated in the same cases by the MIB1 index. High, low and negative snail protein expression was recorded in 18 (38%), 17 (36%), and 12 (26%) cases, respectively, and 40.4% (19/47) cases showed reduced E-cadherin protein expression in HC samples. No significant correlation was found between snail and E-cadherin protein expression levels (P = 0.056). No significant correlation was found between snail protein expression levels and gender, age, tumor grade, vascular or perineural invasion, nodal metastasis and invasion, or proliferative index. Cancer samples with positive snail protein expression were associated with poor survival compared with the negative expresser groups. Kaplan-Meier curves comparing different snail protein expression levels to survival showed highly significant separation (P < 0.0001, log-rank test). With multivariate analysis, only snail protein expression among all parameters was found to influence survival (P = 0.0003). We suggest that snail expression levels can predict poor survival regardless of pathological features and tumor proliferation. Immunohistochemical detection of snail protein expression levels in routine sections may provide the first biological prognostic marker.

  1. Low dose mTHPC photodynamic therapy for cholangiocarcinoma (United States)

    Stepp, Herbert; Kniebühler, Gesa; Pongratz, Thomas; Betz, Christian S.; Göke, Burkhard; Sroka, Ronald; Schirra, Jörg


    Objective: Demonstration of whether a low dose of mTHPC (temoporfin , Foscan) is sufficient to induce an efficient clinical response in palliative PDT of non-resectable cholangiocarcinoma (CC), while showing a low side effect profile as compared to the standard Photofrin PDT. Materials and Methods: 13 patients (14 treatment sessions) with non-resectable CC were treated with stenting and PDT (3 mg Foscan per treatment, 0.032-0.063 mg/kg body weight, 652 nm, 50 J/cm). Fluorescence measurements were performed with a single bare fiber for 5/13 patients prior to PDT at the tumor site to determine the fluorescence contrast. For another 7/13 patients, long-term fluorescence-kinetics were measured on the oral mucosa to determine the time of maximal relative fluorescence intensity. Results: Foscan fluorescence could clearly be identified spectroscopically as early as 20 hours after administration. It was not significantly different between lesion and normal tissue within the bile duct. Fluorescence kinetics assessed at the oral mucosa were highest at 72-96 hours after administration. The DLI was therefore extended from 20 hours to approx. 70 hours for the last 5 patients treated. The treatment effect was promising with a median survival of 11 months for the higher grade tumors (Bismuth types III and IV). Local side effects occurred in one patient (pancreatitis), systemic side effects were much reduced compared to prior experience with Photofrin. Conclusion: Combined stenting and photodynamic therapy (PDT) performed with a low dose of Foscan results in comparable survival times relative to standard Photofrin PDT, while lowering the risk of side effects significantly.

  2. Hepatitis viruses and risk of cholangiocarcinoma in northeast Thailand. (United States)

    Srivatanakul, Petcharin; Honjo, Satoshi; Kittiwatanachot, Pacharin; Jedpiyawongse, Adisorn; Khuhaprema, Thiravud; Miwa, Masanao


    Liver cancer is the most common cancer in males in Thailand and the third in females. A high incidence of cholangiocarcinoma (CCA) is estimated in the northeast of Thailand. Chronic infection with Opisthorchis viverrini (OV) is the major risk factor for development of CCA. It has been demonstrated that HCV infection is a risk factor for CCA in non - endemic area of OV infection. We examined the association of HBV and HCV and risk of CCA in the northeast Thailand. All cases of CCA were recruited between 1999 and 2001 from Nakhon Phanom provincial hospital and all community hospitals in the province. One control per case was selected, matched by sex, age (∓5 years) and residence. 106 case-control pairs were obtained. Anti-OV, HBsAg, and Anti HCV were determined by ELISA. Among 103 age-sex-place of residence matched case-control pairs, there were 7, 0, 0, 96 pairs for anti-HCV (+) case vs. (-) control, (+) case vs. (+) control, (-) case vs. (+) control and (-) case vs. (-) control combinations (OR=7/0). Among 106 matched pairs, there were 9, 2, 4, 91 pairs for the similar four combinations of HBsAg (OR=2.25 (95%CI: 0.63-10.0). If the subject had anti-HCV and/or HBsAg, the OR for CCA was 4.00 (95%CI: 1.29-16.4). Even after adjustment for anti-OV, risk for HBsAg and/or anti-HCV positive was still marginally increased with an OR of 4.69 although not reaching statistical significance (95%CI: 0.98-22.5). Hepatitis B and C virus infection may also play role in the development of CCA in northeast Thailand.

  3. Genetic and environmental determinants of risk for cholangiocarcinoma in Thailand

    Institute of Scientific and Technical Information of China (English)

    Masanao; Miwa; Satoshi; Honjo; Gyokukou; You; Masakazu; Tanaka; Kazuhiko; Uchida; Petcharin; Srivatanakul; Thiravud; Khuhaprema; Watcharin; Loilome; Anchalee; Techasen; Chaisiri; Wongkham; Temduang; Limpaiboon; Puangrat; Yongvanit; Sopit; Wongkham


    Cholangiocarcinoma(CCA) is a difficult cancer to diagnose in the early stage and to treat by curative resec-tion. The incidence of CCA in the northeast of Thailand is the highest in the world. To make progress in detecting a high risk group and in the prevention and detection of CCA, we have been analyzing the risk factors for CCA. Although liver fluke infection is known to be a risk factor, there are patients who are not infected with the liver fluke and not all people infected with the liver fluke will suffer from the disease. Therefore, it is of the utmost importance to analyze the risk factors and the mechanism to prevent the disease and also to detect the disease in its early stage to save patients’ lives. Through collaboration among Thai and Japanese researchers, we analyzed the genetic and environmental determinants of risks for CCA. Also, we have been trying to develop methods to detect the disease in a non-invasive way. Without repeating findings reported in various reviews on CCA, we will first discuss the environmental and genetic determinants of the risks for CCA. Second, we will discuss the properties of CCA, including the etiological agents and the mechanism of cholangiocarcinogenesis, and finally, we will discuss future approaches to prevent and cure CCA from the standpoint of evidence-based medicine. We will discuss these points by including the data from our laboratories. We would like to emphasize the importance of the genetic data, especially whole genome approaches, to understand the properties of CCA, to find a high risk population for CCA and to develop effective preventative methods to stop the carcinogenic steps toward CCA in the near future. In addition, it is of the upmost importance to develop a non-invasive, specific and sensitive method to detect CCA in its early stage for the application of modern medical approaches to help patients with CCA.

  4. Human T Cell Memory: A Dynamic View

    Directory of Open Access Journals (Sweden)

    Derek C. Macallan


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

  5. Biological impact of human embryonic stem cells. (United States)

    Martín, Miguel; Menéndez, Pablo


    Research on human embryonic stem cells (hESCs) and induced pluripotent (iPS) stem cells is currently a field of great potential in biomedicine. These cells represent a highly valuable tool for developmental biology studies, disease models, and drug screening and toxicity. The ultimate goal of hESCs and iPS cell research is the treatment of diseases or disorders for which there is currently no treatment or existing therapies are only partially effective. Despite the disproportionate short-term hopes generated, which are putting too much pressure on scientists, the international scientific community is making rapid progress in understanding hESCs and iPS cells. Nonetheless, great efforts have to be made to provide an answer to still quite basic questions concerning their biology. Moreover, translation to clinical applications in cell replacement therapy requires prior solution to ethical barriers. The recent development of iPS cells has provided a strong alternative to overcome ethical issues concerning hESCs. However, an in-depth characterization of their genetic and epigenetic features, as well as their differentiation potential still remains to be undertaken. This chapter will describe, precisely, what the critical issues are, where scientific and ethical barriers stand, and how we are to overcome them. Only then, we shall finally discover whether hESCs and iPS cells will allow building reproducible disease models, and whether they really are a safe tool, with great potential for regenerative medicine.

  6. Bile proteomic profiles differentiate cholangiocarcinoma from primary sclerosing cholangitis and choledocholithiasis

    NARCIS (Netherlands)

    Lankisch, Tim O; Metzger, Jochen; Negm, Ahmed A; Vosskuhl, Katja; Schiffer, Eric; Siwy, Justyna; Weismüller, Tobias J; Schneider, Andrea S; Thedieck, Kathrin; Baumeister, Ralf; Zürbig, Petra; Weissinger, Eva M; Manns, Michael P; Mischak, Harald; Wedemeyer, Jochen


    UNLABELLED: Early detection of malignant biliary tract diseases, especially cholangiocarcinoma (CC) in patients with primary sclerosing cholangitis (PSC), is very difficult and often comes too late to give the patient a therapeutic benefit. We hypothesize that bile proteomic analysis distinguishes C

  7. Oral Floor and Gingival Metastasis of Cholangiocarcinoma: A Case Report and Review of the Literature

    Directory of Open Access Journals (Sweden)

    Yukihiro Nakanishi


    Full Text Available The oral cavity is very unusual site of metastases even though wide spread metastatic disease may be present. The most common primary sites that metastasize to the oral cavity are lung, kidney, breast, and hepatocellular carcinoma. We present a rare case of a 77-year-old Caucasian female with metastasis from a cholangiocarcinoma to the oral floor contiguous with lingual gingival mucosa. The patient presented with left sided rib pain. A CT scan of the chest, abdomen, and pelvis showed multiple pulmonary nodules and a single dominant mass in the right lobe of the liver. This tumor was 6.5 cm with multiple satellite lesions surrounding it. The liver biopsy was diagnostic of a moderately to poorly differentiated adenocarcinoma, consistent with a primary cholangiocarcinoma. After undergoing one cycle of gemcitabine chemotherapy, the patient noticed an extremely rapidly growing mass involving her right lower gingiva and the entire right floor of her mouth. The biopsy of that mass also showed a moderately to poorly differentiated adenocarcinoma. The gingival tumor had a similar cytomorphology and immunophenotype as her cholangiocarcinoma. Therefore, an unusual site for metastatic cholangiocarcinoma was confirmed.

  8. Intraoperative frozen section analysis of the proximal bile ducts in hilar cholangiocarcinoma is of limited value

    NARCIS (Netherlands)

    Mantel, Hendrik T J; Westerkamp, Andrie C; Sieders, Egbert; Peeters, Paul M J G; de Jong, Koert P; Boer, de Marieke; de Kleine, Ruben H; Gouw, Annette S H; Porte, Robert J


    Frozen section analysis (FS) during cancer surgery is widely used to assess resection margins. However, in hilar cholangiocarcinoma (HCCA), FS may be less reliable because of the specific growth characteristics of the tumor. The aim of this study was to determine the accuracy and consequences of int

  9. Behavioral Modification Regarding Liver Fluke and Cholangiocarcinoma with a Health Belief Model Using Integrated Learning. (United States)

    Phatisena, Panida; Eaksanti, Tawatchai; Wichantuk, Pitsanee; Tritipsombut, Jaruwan; Kaewpitoon, Soraya J; Rujirakul, Ratana; Wakkhuwattapong, Parichart; Tongtawee, Taweesak; Matrakool, Likit; Panpimanmas, Sukij; Norkaew, Jun; Kujapun, Jirawoot; Chavengkun, Wasugree; Kompor, Porntip; Pothipim, Mali; Ponphimai, Sukanya; Padchasuwan, Natnapa; Kaewpitoon, Natthawut


    This study aimed to modify behavior regarding liver fluke and cholangiocarcinoma prevention in Chumphuang district, Nakhon Ratchasima province, Thailand through integrated learning. A total of 180 participants were included through purposive selection of high-risk scores on verbal screening. Participants attended the health education program which applied the health belief model included family based, knowledge station based, academic merit based and community based learning. Data were collected using a questionnaire composed of 4 parts: 1) personal information, 2) knowledge, 3) perceived susceptibility, severity, benefits, and barriers, 4) practice regarding liver fluke and cholangiocarcinoma prevention. The result revealed that the majority were female (79.9%), age ≥60 years old (33.2%), primary school educational level (76.1%), and agricultural occupation (70.1%). The mean scores of knowledge, perception, and practice to liver fluke and cholangiocarcinoma prevention, before participated the integrative learning were low, moderate, and low, respectively. Meanwhile, the mean score of knowledge, perceived susceptibility, severity, benefits, and barriers, and practice regarding liver fluke and cholangiocarcinoma prevention, were higher with statistical significance after participation in the integrated learning. This finding indicates that health education programs may successfully modify health behavior in the rural communities. Therefore they may useful for further work behavior modification in other epidemic areas.

  10. Catheter tract implantation metastases associated with percutaneous biliary drainage for extrahepatic cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Jun Sakata; Yoshio Shirai; Toshifumi Wakai; Tatsuya Nomura; Eiko Sakata; Katsuyoshi Hatakeyama


    AIM: To estimate the incidence of catheter tract implantation metastasis among patients undergoing percutaneous transhepatic biliary drainage (PTBD) for extrahepatic cholangiocarcinoma, and to provide data regarding the management of this unusual complication of PTBD by reviewing cases reported in the literature.METHODS: A retrospective analysis of 67 consecutive patients who underwent PTBD before the resection of extrahepatic cholangiocarcinoma was conducted. The median follow-up period after PTBD was 106 mo. The English language literature (PubMed, National Library of Medicine, Bethesda, MD, USA), from January 1966through December 2004, was reviewed.RESULTS: Catheter tract implantation metastasis developed in three patients. The cumulative incidence of implantation metastasis reached a plateau (6%)at 20 mo after PTBD. All of the three patients with implantation metastasis died of tumor progression at 3, 9, and 20 mo after the detection of this complication. Among the 10 reported patients with catheter tract implantation metastasis from extrahepatic cholangiocarcinoma (including our three patients), two survived for more than 5 years after the excision of isolated catheter tract metastases.CONCLUSION: Catheter tract implantation metastasis is not a rare complication following PTBD for extrahepatic cholangiocarcinoma. Although the prognosis for patients with this complication is generally poor, the excision of the catheter tract may enable survival in selected patients with isolated metastases along the catheter tract.

  11. Survival and an overview of decision-making in patients with cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Håvard Mjørud Forsmo; Arild Horn; Asgaut Viste; Dag Hoem; Kjell Øvrebø


    BACKGROUND: Cholangiocarcinoma is rare, accounting for approximately 3%of all gastrointestinal cancers. This study aimed to identify the survival rate among surgically treated and palliated patients, and secondly to identify parameters that could predict a curative resection. METHODS: A total of 121 patients, 55 men and 66 women, median age 70 years (range 31-91), who had been treated for cholangiocarcinoma in the period of 1990-2005 were evaluated retrospectively. RESULTS: Curative resection was performed in 40 patients (33%), whereas 81 received palliative treatment (67%). 16%(19 of 121) of the patients had an explorative laparotomy without tumour resection. Age above 65 years (OR 3.4;95%CI 1.4-8.4;P=0.008), weight loss (OR 8.5;95%CI 1.5-46;P=0.01) or tumour location (The resection rate of hilar cholangiocarcinoma was lower than that of intrapancreatic cancer.) (OR 2.7; 95% CI 1.7-4.5; P=0.001) predicted palliative treatment. The adjusted 5-year survival rate of patients who received tumour resection and palliative treatment was 30%and 1.2%, respectively (P CONCLUSIONS: In few patients with a resectable cholangiocarcinoma, an explorative laparotomy is often necessary to evaluate resectability. However, long-term survival is signiifcantly better in patients who received radical surgical resection.

  12. Reversal of Jaundice in Two Patients with Inoperable Cholangiocarcinoma Treated with Cisplatin and Gemcitabine Combination

    Directory of Open Access Journals (Sweden)

    Maarten Criel


    Full Text Available Two patients are presented with severe jaundice, due to inoperable cholangiocarcinoma. The chemotherapeutic approach in patients with severe jaundice is discussed. Many schedules of chemotherapy were developed in this tumor type with normal serum bilirubin. We report here the first successful use of cisplatin and gemcitabine combination chemotherapy in these patients. Tolerability was good and liver function tests gradually improved.

  13. Cholangiocarcinoma and malignant bile duct obstruction: Areview of last decades advances in therapeutic endoscopy

    Institute of Scientific and Technical Information of China (English)


    In the last decades many advances have been achievedin endoscopy, in the diagnosis and therapy ofcholangiocarcinoma,however blood test, magneticresonance imaging, computed tomography scan mayfail to detect neoplastic disease at early stage, thus thediagnosis of cholangiocarcinoma is achieved usuallyat unresectable stage. In the last decades the roleof endoscopy has moved from a diagnostic role toan invaluable therapeutic tool for patients affectedby malignant bile duct obstruction. One of the majorissues for cholangiocarcinoma is bile ducts occlusion,leading to jaundice, cholangitis and hepatic failure.Currently, endoscopy has a key role in the work upof cholangiocarcinoma, both in patients amenable tosurgical intervention as well as in those unfit for surgeryor not amenable to immediate surgical curative resectionowing to locally advanced or advanced disease, withpalliative intention. Endoscopy allows successful biliarydrainage and stenting in more than 90% of patientswith malignant bile duct obstruction, and allows rapidreduction of jaundice decreasing the risk of biliary sepsis.When biliary drainage and stenting cannot be achievedwith endoscopy alone, endoscopic ultrasound-guidedbiliary drainage represents an effective alternativemethod affording successful biliary drainage in morethan 80% of cases. The purpose of this review is tofocus on the currently available endoscopic managementoptions in patients with cholangiocarcinoma.

  14. SOX17 Regulates Cholangiocyte Differentiation and Acts as a Tumor Suppressor in Cholangiocarcinoma

    DEFF Research Database (Denmark)

    Merino-Azpitarte, M; Lozano, E; Perugorria, M J


    BACKGROUND & AIMS: Cholangiocarcinoma (CCA) is a biliary malignancy linked to genetic and epigenetic abnormalities, such as hypermethylation of SOX17 promoter. Here, the role of SOX17 in cholangiocyte differentiation and cholangiocarcinogenesis was studied. METHODS: SOX17 expression/function was ...

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


    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.

  16. Cell phoney: human cloning after Quintavalle. (United States)

    Morgan, Derek; Ford, Mary


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

  17. Angiopoietin-2 and biliary diseases: elevated serum, but not bile levels are associated with cholangiocarcinoma.

    Directory of Open Access Journals (Sweden)

    Torsten Voigtländer

    Full Text Available BACKGROUND: The diagnosis of cholangiocarcinoma (CC is challenging especially in patients with primary sclerosing cholangitis (PSC and often delayed due to the lack of reliable markers. Angiopoietin-2 (Angpt-2 has been employed as a biomarker of angiogenesis and might be involved in tumor neoangiogenesis. AIM: To evaluate the diagnostic potential of Angpt-2 as a biomarker to detect patients with CC. METHODS: Bile and serum Angpt-2 levels were measured in patients with CC (n=45, PSC (n=74, CC complicating PSC (CC/PSC (n=11 and patients with bile duct stones (n=37 in a cross sectional study. Diagnostic accuracy of Angpt-2 was compared to carbohydrate antigen 19-9 (CA19-9. Fluorescent immunohistochemistry from human CC liver tissue samples was performed to localize the origin of Angpt-2. RESULTS: Serum Angpt-2 concentration was significantly elevated in patients with CC compared to control patients (p<0.05. Diagnostic accuracy of Angpt-2 as determined by receiver operating characteristic (ROC analysis resulted in a higher area under the curve (AUC value compared to CA19-9 (AUC: 0.85 versus 0.77; 95% confidence interval (CI: 0.74-0.93 versus 0.65-0.87, respectively. Angpt-2 was also detectable in bile, but was not associated with the presence of CC. Immunohistochemistry revealed a strong induction of Angpt-2 expression in the tumor vasculature. CONCLUSIONS: Circulating Angpt-2 in serum might be a promising protein candidate locally derived from the tumor vasculature in patients with CC. Measurement of Angpt-2 in serum may be useful for diagnosis and further clinical management of patients with CC.

  18. Microbiota studies in the bile duct strongly suggest a role for Helicobacter pylori in extrahepatic cholangiocarcinoma. (United States)

    Avilés-Jiménez, F; Guitron, A; Segura-López, F; Méndez-Tenorio, A; Iwai, S; Hernández-Guerrero, A; Torres, J


    Biliary tract cancer or extrahepatic cholangiocarcinoma (ECCA) represents the sixth commonest cause of cancer in the gastrointestinal tract in western countries. We aimed to characterize the microbiota and its predicted associated functions in the biliary tract of ECCA and benign biliary pathology (BBP). Samples were taken from 100 patients with ECCA and 100 patients with BBP by endoscopic cholangio-pancreatography for DNA extraction. Ten patients with ECCA and ten with BBP were selected for microbiota studies using the V4-16S rRNA gene and sequenced in Illumina platform. Microbiota analyses included sample-to-sample distance metrics, ordination/clustering and prediction of functions. Presence of Nesterenkonia sp. and Helicobacter pylori cagA and vacA genes were tested in the 100 ECCA and 100 BBP samples. Phylum Proteobacteria dominated all samples (60.4% average). Ordination multicomponent analyses showed significant microbiota separation between ECCA and BBP (p 0.010). Analyses of 4002 operational taxonomic units with presence variation in at least one category probed a separation of ECCA from BBP. Among these, Nesterenkonia decreased, whereas Methylophilaceae, Fusobacterium, Prevotella, Actinomyces, Novosphingobium and H. pylori increased in ECCA. Predicted associated functions showed increased abundance of H. pylori virulence genes in ECCA. cagA and vacA genes were confirmed by PCR in ECCA and BBP samples. This is the first microbiota report in ECCA and BBP to show significant changes in microbial composition. Bacterial species unusual for human flora were found: Methylophilaceae and Nesterenkonia are reported in hypersaline soils, and Mesorhizobium is a nitrogen-fixing bacterium. Enrichment of virulence genes confirms previous studies suggesting that H. pylori might be associated with ECCA.

  19. Lymphoid Cell-Glioma Cell Interaction Enhances Cell Coat Production by Human Gliomas: Novel Suppressor Mechanism (United States)

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


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

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

    Directory of Open Access Journals (Sweden)

    Leonardo D'Aiuto

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

  1. Human somatic cell nuclear transfer and cloning. (United States)


    This document presents arguments that conclude that it is unethical to use somatic cell nuclear transfer (SCNT) for infertility treatment due to concerns about safety; the unknown impact of SCNT on children, families, and society; and the availability of other ethically acceptable means of assisted reproduction. This document replaces the ASRM Ethics Committee report titled, "Human somatic cell nuclear transfer (cloning)," last published in Fertil Steril 2000;74:873-6. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  2. Dopamine receptor repertoire of human granulosa cells

    Directory of Open Access Journals (Sweden)

    Kunz Lars


    Full Text Available Abstract Background High levels of dopamine (DA were described in human ovary and recently evidence for DA receptors in granulosa and luteal cells has been provided, as well. However, neither the full repertoire of ovarian receptors for DA, nor their specific role, is established. Human granulosa cells (GCs derived from women undergoing in vitro fertilization (IVF are an adequate model for endocrine cells of the follicle and the corpus luteum and were therefore employed in an attempt to decipher their DA receptor repertoire and functionality. Methods Cells were obtained from patients undergoing IVF and examined using cDNA-array, RT-PCR, Western blotting and immunocytochemistry. In addition, calcium measurements (with FLUO-4 were employed. Expression of two DA receptors was also examined by in-situ hybridization in rat ovary. Effects of DA on cell viability and cell volume were studied by using an ATP assay and an electronic cell counter system. Results We found members of the two DA receptor families (D1- and D2 -like associated with different signaling pathways in human GCs, namely D1 (as expected and D5 (both are Gs coupled and linked to cAMP increase and D2, D4 (Gi/Gq coupled and linked to IP3/DAG. D3 was not found. The presence of the trophic hormone hCG (10 IU/ml in the culture medium for several days did not alter mRNA (semiquantitative RT-PCR or protein levels (immunocytochemistry/Western blotting of D1,2,4,5 DA receptors. Expression of prototype receptors for the two families, D1 and D2, was furthermore shown in rat granulosa and luteal cells by in situ hybridization. Among the DA receptors found in human GCs, D2 expression was marked both at mRNA and protein levels and it was therefore further studied. Results of additional RT-PCR and Western blots showed two splice variants (D2L, D2S. Irrespective of these variants, D2 proved to be functional, as DA raised intracellular calcium levels. This calcium mobilizing effect of DA was observed

  3. TALEN-Induced Translocations in Human Cells. (United States)

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


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

  4. The core regulatory network in human cells. (United States)

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


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

  5. Cell Culture Assay for Human Noroviruses [response

    Energy Technology Data Exchange (ETDEWEB)

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


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

  6. A massive hepatic tumor demonstrating hepatocellular, cholangiocarcinoma and neuroendocrine lineages: A case report and review of the literature

    Directory of Open Access Journals (Sweden)

    Rachel E. Beard


    Conclusion: This is one of the only reports of a hepatic tumor arising from hepatocellular carcinoma, cholangiocarcinoma and neuroendocrine lineages. Increased awareness of this tumor type may optimize improve future management.

  7. Preliminary study on human fibroblasts as feeder layer for human embryonic stem cells culture in vitro

    Institute of Scientific and Technical Information of China (English)


    To avoid the direct contact with mouse cells and possible heterogeneous pathogen in future application, we need to replace mouse embryonic fibroblastswith human fibroblasts as the feeder layer to maintain human embryonic stem cells growth in the undifferentiated state. We successfully use human fibroblasts derived from aborted fetus and adult prepuce as feeder layer to maintain human embryonic stem cells growth. During the passage and growth on this feeder layer, the human embryonic stem cells can keep their undifferentiated state.

  8. Primitive cardiac cells from human embryonic stem cells. (United States)

    Hudson, James; Titmarsh, Drew; Hidalgo, Alejandro; Wolvetang, Ernst; Cooper-White, Justin


    Pluripotent stem cell-derived cardiomyocytes are currently being investigated for in vitro human heart models and as potential therapeutics for heart failure. In this study, we have developed a differentiation protocol that minimizes the need for specific human embryonic stem cell (hESC) line optimization. We first reduced the heterogeneity that exists within the starting population of bulk cultured hESCs by using cells adapted to single-cell passaging in a 2-dimensional (2D) culture format. Compared with bulk cultures, single-cell cultures comprised larger fractions of TG30(hi)/OCT4(hi) cells, corresponding to an increased expression of pluripotency markers OCT4 and NANOG, and reduced expression of early lineage-specific markers. A 2D temporal differentiation protocol was then developed, aimed at reducing the inherent heterogeneity and variability of embryoid body-based protocols, with induction of primitive streak cells using bone morphogenetic protein 4 and activin A, followed by cardiogenesis via inhibition of Wnt signaling using the small molecules IWP-4 or IWR-1. IWP-4 treatment resulted in a large percentage of cells expressing low amounts of cardiac myosin heavy chain and expression of early cardiac progenitor markers ISL1 and NKX2-5, thus indicating the production of large numbers of immature cardiomyocytes (~65,000/cm(2) or ~1.5 per input hESC). This protocol was shown to be effective in HES3, H9, and, to a lesser, extent, MEL1 hESC lines. In addition, we observed that IWR-1 induced predominantly atrial myosin light chain (MLC2a) expression, whereas IWP-4 induced expression of both atrial (MLC2a) and ventricular (MLC2v) forms. The intrinsic flexibility and scalability of this 2D protocol mean that the output population of primitive cardiomyocytes will be particularly accessible and useful for the investigation of molecular mechanisms driving terminal cardiomyocyte differentiation, and potentially for the future treatment of heart failure.

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



    Relatively little is known for the differentiation and maturation process of human B cells to plasma cells. This is particularly important in reconstitution work involving transfer of autoantibodies. To address this issue, we transplanted human peripheral blood mononuclear cells (PBMC) directly into the spleen of irradiated NOD/SCID mice depleted of natural killer cell activity. Within 6 weeks, naïve B cells differentiated into memory B cells and, importantly, the numbers of human CD138+ plas...

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

    Directory of Open Access Journals (Sweden)

    Hayato Fukusumi


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

  11. Human microglial cells synthesize albumin in brain.

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    Sung-Min Ahn

    Full Text Available Albumin, an abundant plasma protein with multifunctional properties, is mainly synthesized in the liver. Albumin has been implicated in Alzheimer's disease (AD since it can bind to and transport amyloid beta (Abeta, the causative agent of AD; albumin is also a potent inhibitor of Abeta polymerization. Despite evidence of non-hepatic transcription of albumin in many tissues including kidney and pancreas, non-hepatic synthesis of albumin at the protein level has been rarely confirmed. In a pilot phase study of Human Brain Proteome Project, we found evidence that microglial cells in brain may synthesize albumin. Here we report, for the first time, the de novo synthesis of albumin in human microglial cells in brain. Furthermore, we demonstrate that the synthesis and secretion of albumin from microglial cells is enhanced upon microglial activation by Abeta(1-42- or lipopolysaccharide (LPS-treatment. These data indicate that microglial cells may play a beneficial role in AD by secreting albumin that not only inhibits Abeta polymerization but also increases its clearance.

  12. Neocortical glial cell numbers in human brains

    DEFF Research Database (Denmark)

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


    and neurons and counting were done in each of the four lobes. The study showed that the different subpopulations of glial cells behave differently as a function of age; the number of oligodendrocytes showed a significant 27% decrease over adult life and a strong correlation to the total number of neurons...... while the total astrocyte number is constant through life; finally males have a 28% higher number of neocortical glial cells and a 19% higher neocortical neuron number than females. The overall total number of neocortical neurons and glial cells was 49.3 billion in females and 65.2 billion in males......, a difference of 24% with a high biological variance. These numbers can serve as reference values in quantitative studies of the human neocortex. (C) 2007 Elsevier Inc. All rights reserved Udgivelsesdato: 2008/11...

  13. Characterizing motility dynamics in human RPE cells (United States)

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


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

  14. Human cell culture in a space bioreactor (United States)

    Morrison, Dennis R.


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

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

    Directory of Open Access Journals (Sweden)

    Kentaro Kikuchi


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

  16. Human somatic cell nuclear transfer is alive and well. (United States)

    Cibelli, Jose B


    In this issue, Chung et al. (2014) generate human embryonic stem cells by fusing an adult somatic cell to a previously enucleated human oocyte, in agreement with recent reports by the Mitalipov and Egli groups. We can now safely say that human somatic cell nuclear transfer is alive and well.

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

    DEFF Research Database (Denmark)

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


    There is emerging evidence of the association between human papillomavirus and a subset of head and neck cancers. However, the role of human papillomavirus as a causal factor is still debated. This review addresses the association between human papillomavirus and oropharyngeal squamous cell...... of well-defined premalignant lesions. However, a causal relationship between human papillomavirus infection and oropharyngeal squamous cell carcinoma seems evident....

  18. Cell cycle regulation in human embryonic stem cells: links to adaptation to cell culture. (United States)

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


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

  19. Mutation profiling in cholangiocarcinoma: prognostic and therapeutic implications.

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    Chaitanya R Churi

    Full Text Available Cholangiocarcinoma (CCA is clinically heterogeneous; intra and extrahepatic CCA have diverse clinical presentations. Next generation sequencing (NGS technology may identify the genetic differences between these entities and identify molecular subgroups for targeted therapeutics.We describe successful NGS-based testing of 75 CCA patients along with the prognostic and therapeutic implications of findings. Mutation profiling was performed using either a NGS panel of hotspot regions in 46 cancer-related genes using a 318-chip on Ion PGM Sequencer or b Illumina HiSeq 2000 sequencing platform for 3,769 exons of 236 cancer-related genes plus 47 introns from 19 genes to an average depth of 1000X. Clinical data was abstracted and correlated with clinical outcome. Patients with targetable mutations were referred to appropriate clinical trials.There were significant differences between intrahepatic (n = 55 and extrahepatic CCA (n = 20 in regard to the nature and frequency of the genetic aberrations (GAs. IDH1 and DNA repair gene alterations occurred more frequently in intrahepatic CCA, while ERBB2 GAs occurred in the extrahepatic group. Commonly occurring GAs in intrahepatic CCA were TP53 (35%, KRAS (24%, ARID1A (20%, IDH1 (18%, MCL1 (16% and PBRM1 (11%. Most frequent GAs in extrahepatic CCA (n = 20 were TP53 (45%, KRAS (40%, ERBB2 (25%, SMAD4 (25%, FBXW7 (15% and CDKN2A (15%. In intrahepatic CCA, KRAS, TP53 or MAPK/mTOR GAs were significantly associated with a worse prognosis while FGFR GAs correlated with a relatively indolent disease course. IDH1 GAs did not have any prognostic significance. GAs in the chromatin modulating genes, BAP1 and PBRM1 were associated with bone metastases and worse survival in extrahepatic CCA. Radiologic responses and clinical benefit was noted with EGFR, FGFR, C-met, B-RAF and MEK inhibitors.There are significant genetic differences between intra and extrahepatic CCA. NGS can potentially identify disease subsets with distinct

  20. Cell adhesion molecules P-cadherin and CD24 are markers for carcinoma and dysplasia in the biliary tract. (United States)

    Riener, Marc-Oliver; Vogetseder, Alexander; Pestalozzi, Bernhard C; Clavien, Pierre-Alain; Probst-Hensch, Nicole; Kristiansen, Glen; Jochum, Wolfram


    P-cadherin (CDH3) and CD24 are cell adhesion molecules that control morphogenic processes, cell motility, and invasive growth of tumor cells. The aim of our study was to investigate P-cadherin and CD24 expression in carcinomas and dysplastic lesions of the biliary tract and to evaluate the potential diagnostic usefulness of these cell adhesion molecules. Using immunohistochemistry on tissue microarrays, we analyzed P-cadherin, CD24, and p53 expression in 117 carcinomas of the biliary tract (19 intrahepatic cholangiocarcinomas, 59 extrahepatic cholangiocarcinomas, and 39 gallbladder carcinomas) and correlated our findings with clinicopathologic parameters. We found P-cadherin positivity in 37% of intrahepatic cholangiocarcinomas, 73% of extrahepatic cholangiocarcinomas, and 64% of gallbladder carcinomas, respectively. CD24 reactivity was observed in 21% of intrahepatic cholangiocarcinomas, 58% of extrahepatic cholangiocarcinomas, and 42% of gallbladder carcinomas. Nuclear p53 expression was found in 37% of intrahepatic cholangiocarcinomas, 46% of extrahepatic cholangiocarcinomas, and 45% of gallbladder carcinomas. We also studied P-cadherin, CD24, and p53 expression in normal (n = 30), inflamed (n = 22), and dysplastic (n = 21) biliary epithelium of extrahepatic bile ducts. Dysplastic biliary epithelium was positive for P-cadherin in 91%, for CD24 in 71%, and for p53 in 24% of lesions, respectively. In contrast, normal and inflamed epithelia were negative for all 3 proteins. We conclude that P-cadherin and CD24 are expressed in carcinomas of the biliary tract with high frequency and at an early stage of carcinogenesis. Therefore, they may be useful markers for early detection and as targets for therapy of cholangiocarcinoma.

  1. Glycomics of human embryonic stem cells and human induced pluripotent stem cells. (United States)

    Furukawa, Jun-Ichi; Okada, Kazue; Shinohara, Yasuro


    Most cells are coated by a dense glycocalyx composed of glycoconjugates such as glycosphingolipids, glycoproteins, and proteoglycans. The overall glycomic profile is believed to be crucial for the diverse roles of glycans, which are mediated by specific interactions that regulate cell-cell adhesion, the immune response, microbial pathogenesis, and other cellular events. Many cell surface markers were discovered and identified as glycoconjugates such as stage-specific embryonic antigen, Tra-1-60/81 and various other cell surface molecules (e.g., cluster of differentiation). Recent progress in the development of analytical methodologies and strategies has begun to clarify the cellular glycomics of various cells including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs). The glycomic profiles of these cells are highly cell type-specific and reflect cellular alterations, such as development, differentiation and cancerous change. In this mini review, we briefly summarize the glycosylation spectra specific to hESCs and hiPSCs, which cover glycans of all major glycoconjugates (i.e., glycosphingolipids, N- and O-glycans of glycoproteins, and glycosaminoglycans) and free oligosaccharides.

  2. Isolation and characterization of human spermatogonial stem cells

    Directory of Open Access Journals (Sweden)

    Liu Shixue


    Full Text Available Abstract Background To isolate and characterization of human spermatogonial stem cells from stem spermatogonium. Methods The disassociation of spermatogonial stem cells (SSCs were performed using enzymatic digestion of type I collagenase and trypsin. The SSCs were isolated by using Percoll density gradient centrifugation, followed by differential surface-attachment method. Octamer-4(OCT4-positive SSC cells were further identified using immunofluorescence staining and flow cytometry technques. The purity of the human SSCs was also determined, and a co-culture system for SSCs and Sertoli cells was established. Results The cell viability was 91.07% for the suspension of human spermatogonial stem cells dissociated using a two-step enzymatic digestion process. The cells isolated from Percoll density gradient coupled with differential surface-attachement purification were OCT4 positive, indicating the cells were human spermatogonial stem cells. The purity of isolated human spermatogonial stem cells was 86.7% as assessed by flow cytometry. The isolated SSCs were shown to form stable human spermatogonial stem cell colonies on the feeder layer of the Sertoli cells. Conclusions The two-step enzyme digestion (by type I collagenase and trypsin process is an economical, simple and reproducible technique for isolating human spermatogonial stem cells. With little contamination and less cell damage, this method facilitates isolated human spermatogonial stem cells to form a stable cell colony on the supporting cell layer.

  3. Cell entry by human pathogenic arenaviruses. (United States)

    Rojek, Jillian M; Kunz, Stefan


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

  4. Markers of T Cell Senescence in Humans

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    Weili Xu


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

  5. Human embryonic stem cells and microenvironment

    Directory of Open Access Journals (Sweden)

    Banu İskender


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

  6. Arecoline is cytotoxic for human endothelial cells. (United States)

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


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

  7. DNA Damage in Inflammation-Related Carcinogenesis and Cancer Stem Cells

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    Shiho Ohnishi


    Full Text Available Infection and chronic inflammation have been recognized as important factors for carcinogenesis. Under inflammatory conditions, reactive oxygen species (ROS and reactive nitrogen species (RNS are generated from inflammatory and epithelial cells and result in oxidative and nitrative DNA damage, such as 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG and 8-nitroguanine. The DNA damage can cause mutations and has been implicated in the initiation and/or promotion of inflammation-mediated carcinogenesis. It has been estimated that various infectious agents are carcinogenic to humans (IARC group 1, including parasites (Schistosoma haematobium (SH and Opisthorchis viverrini (OV, viruses (hepatitis C virus (HCV, human papillomavirus (HPV, and Epstein-Barr virus (EBV, and bacterium Helicobacter pylori (HP. SH, OV, HCV, HPV, EBV, and HP are important risk factors for bladder cancer, cholangiocarcinoma, hepatocellular carcinoma, cervical cancer, nasopharyngeal carcinoma, and gastric cancer, respectively. We demonstrated that 8-nitroguanine was strongly formed via inducible nitric oxide synthase (iNOS expression at these cancer sites of patients. Moreover, 8-nitroguanine was formed in Oct3/4-positive stem cells in SH-associated bladder cancer tissues and in Oct3/4- and CD133-positive stem cells in OV-associated cholangiocarcinoma tissues. Therefore, it is considered that oxidative and nitrative DNA damage in stem cells may play a key role in inflammation-related carcinogenesis.

  8. The complex role of NOTCH receptors and their ligands in the development of hepatoblastoma, cholangiocarcinoma and hepatocellular carcinoma. (United States)

    Gil-García, Borja; Baladrón, Victoriano


    The NOTCH signalling pathway is one of the key molecular pathways of embryonic development and adult tissues homeostasis in mammals. Mammals have four NOTCH receptors and various ligands that modulate their activity. Many cell disorders, whose genesis involves the NOTCH signalling pathway, have been discovered, including cancer. The mechanisms by which these receptors and their ligands affect liver cell transformation are not yet well understood, and they seem to behave as both oncogenes and tumour-suppressor proteins. In this review, we discuss the published data regarding the role of these proteins in the development of hepatoblastoma, cholangiocarcinoma and hepatocellular carcinoma malignancies. The alteration of the NOTCH signalling pathway may be one of the main drivers of hepatic neoplastic growth. However, this signalling pathway might also modulate the development of specific liver tumour features. The complexity of the function of NOTCH receptors and their ligands may be due to their interactions with many other cell signalling pathways. Furthermore, the different levels of expression and activation of these receptors could be a reason for their distinct and sometimes contradictory effects.

  9. Identification of a candidate stem cell in human gallbladder

    Directory of Open Access Journals (Sweden)

    Rohan Manohar


    In conclusion, we have isolated a distinct clonogenic population of epithelial cells from primary human fetal gallbladder with stem cell characteristics and found it to be unique compared to IHBD cells.

  10. Cell cycle regulation by glucosamine in human pulmonary epithelial cells. (United States)

    Chuang, Kun-Han; Lu, Chih-Shen; Kou, Yu Ru; Wu, Yuh-Lin


    Airway epithelial cells play an important role against intruding pathogens. Glucosamine, a commonly used supplemental compound, has recently begun to be regarded as a potential anti-inflammatory molecule. This study aimed to uncover how glucosamine impacts on cellular proliferation in human alveolar epithelial cells (A549) and bronchial epithelial cells (HBECs). With trypan blue-exclusion assay, we observed that glucosamine (10, 20, 50 mM) caused a decrease in cell number at 24 and 48 h; with a flow cytometric analysis, we also noted an enhanced cell accumulation within the G(0)/G(1) phase at 24 h and induction of late apoptosis at 24 and 48 h by glucosamine (10, 20, 50 mM) in A549 cells and HBECs. Examination of phosphorylation in retinoblastoma (Rb) protein, we found an inhibitory effect by glucosamine at 20 and 50 mM. Glucosamine at 50 mM was demonstrated to elevate both the mRNA and protein expression of p53 and heme oxygenase-1 (HO-1), but also caused a reduction in p21 protein expression. In addition, glucosamine attenuated p21 protein stability via the proteasomal proteolytic pathway, as well as inducing p21 nuclear accumulation. Altogether, our results suggest that a high dose of glucosamine may inhibit cell proliferation through apoptosis and disturb cell cycle progression with a halt at G(0)/G(1) phase, and that this occurs, at least in part, by a reduction in Rb phosphorylation together with modulation of p21, p53 and HO-1 expression, and nuclear p21 accumulation.

  11. Human embryonic stem cells and patent protection

    Directory of Open Access Journals (Sweden)

    Radovanović Sanja M.


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

  12. Generation of human induced pluripotent stem cells from dermal fibroblasts



    The generation of patient-specific pluripotent stem cells has the potential to accelerate the implementation of stem cells for clinical treatment of degenerative diseases. Technologies including somatic cell nuclear transfer and cell fusion might generate such cells but are hindered by issues that might prevent them from being used clinically. Here, we describe methods to use dermal fibroblasts easily obtained from an individual human to generate human induced pluripotent stem (iPS) cells by ...

  13. Restriction of human adenovirus replication in Chinese hamster cell lines and their hybrids with human cells. (United States)

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


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

  14. Phenotypic changes of human cells in human-rat liver during partial hepatectomy-induced regeneration

    Institute of Scientific and Technical Information of China (English)

    Yan Sun; Dong Xiao; Hong-An Li; Jin-Fang Jiang; Qing Li; Ruo-Shuang Zhang; Xi-Gu Chen


    AIM: To examine the human hepatic parenchymal and stromal components in rat liver and the phenotypic changes of human cells in liver of human-rat chimera (HRC) generated by in utero transplantation of human cells during partial hepatectomy (PHx)-induced liver regeneration. METHODS: Human hepatic parenchymal and stromal components and phenotypic changes of human cells during liver regeneration were examined by flow cytometry, in situ hybridization and immunohistochemistry. RESULTS: ISH analysis demonstrated human Alupositive cells in hepatic parenchyma and stroma of recipient liver. Functional human hepatocytes generated in this model potentially constituted human hepatic functional units with the presence of donor-derived human endothelial and biliary duct cells in host liver. Alpha fetoprotein (AFP)+, CD34+ and CD45+ cells were observed in the chimeric liver on day 10 after PHxinduced liver regeneration and then disappeared in PHx group, but not in non-PHx group, suggesting that dynamic phenotypic changes of human cells expressing AFP, CD34 and CD45 cells may occur during the chimeric liver regeneration. Additionally, immunostaining for human proliferating cell nuclear antigen (PCNA) showed that the number of PCNA-positive cells in the chimeric liver of PHx group was markedly increased, as compared to that of control group, indicating that donor-derived human cells are actively proliferated during PHx-induced regeneration of HRC liver.

  15. HIF induces human embryonic stem cell markers in cancer cells. (United States)

    Mathieu, Julie; Zhang, Zhan; Zhou, Wenyu; Wang, Amy J; Heddleston, John M; Pinna, Claudia M A; Hubaud, Alexis; Stadler, Bradford; Choi, Michael; Bar, Merav; Tewari, Muneesh; Liu, Alvin; Vessella, Robert; Rostomily, Robert; Born, Donald; Horwitz, Marshall; Ware, Carol; Blau, C Anthony; Cleary, Michele A; Rich, Jeremy N; Ruohola-Baker, Hannele


    Low oxygen levels have been shown to promote self-renewal in many stem cells. In tumors, hypoxia is associated with aggressive disease course and poor clinical outcomes. Furthermore, many aggressive tumors have been shown to display gene expression signatures characteristic of human embryonic stem cells (hESC). We now tested whether hypoxia might be responsible for the hESC signature observed in aggressive tumors. We show that hypoxia, through hypoxia-inducible factor (HIF), can induce an hESC-like transcriptional program, including the induced pluripotent stem cell (iPSC) inducers, OCT4, NANOG, SOX2, KLF4, cMYC, and microRNA-302 in 11 cancer cell lines (from prostate, brain, kidney, cervix, lung, colon, liver, and breast tumors). Furthermore, nondegradable forms of HIFα, combined with the traditional iPSC inducers, are highly efficient in generating A549 iPSC-like colonies that have high tumorigenic capacity. To test potential correlation between iPSC inducers and HIF expression in primary tumors, we analyzed primary prostate tumors and found a significant correlation between NANOG-, OCT4-, and HIF1α-positive regions. Furthermore, NANOG and OCT4 expressions positively correlated with increased prostate tumor Gleason score. In primary glioma-derived CD133 negative cells, hypoxia was able to induce neurospheres and hESC markers. Together, these findings suggest that HIF targets may act as key inducers of a dynamic state of stemness in pathologic conditions.

  16. Derivation of human embryonic stem cells in defined conditions. (United States)

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


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

  17. Introduction: characterization and functions of human T regulatory cells. (United States)

    Romagnani, Sergio


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

  18. Human pancreatic cell autotransplantation following total pancreatectomy. (United States)

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


    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

  19. Sourcing human embryos for embryonic stem cell lines: Problems & perspectives


    Mehta, Rajvi H.


    The ability to successfully derive human embryonic stem cells (hESC) lines from human embryos following in vitro fertilization (IVF) opened up a plethora of potential applications of this technique. These cell lines could have been successfully used to increase our understanding of human developmental biology, transplantation medicine and the emerging science of regenerative medicine. The main source for human embryos has been ′discarded′ or ′spare′ fresh or frozen human embryos following IVF...

  20. Efficient Induction and Isolation of Human Primordial Germ Cell-Like Cells from Competent Human Pluripotent Stem Cells. (United States)

    Irie, Naoko; Surani, M Azim


    We recently reported a robust and defined culture system for the specification of human primordial germ cell-like cells (hPGCLCs) from human pluripotent stem cells (hPSCs), both embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) in vitro (Irie et al. Cell 160: 253-268, 2015). Similar attempts previously produced hPGCLCs from hPSCs at a very low efficiency, and the resulting cells were not fully characterized. A key step, which facilitated efficient hPGCLC specification from hPSCs, was the induction of a "competent" state for PGC fate via the medium containing a cocktail of four inhibitors. The competency of hPSCs can be maintained indefinitely and interchangeably with the conventional/low-competent hPSCs. Specification of hPGCLC occurs following sequential expression of key germ cell fate regulators, notably SOX17 and BLIMP1, as well as initiation of epigenetic resetting over 5 days. The hPGCLCs can be isolated using specific cell surface markers without the need for generating germ cell-specific reporter hPSC lines. This powerful method for the induction and isolation of hPGCLCs can be applied to both hESCs and iPSCs, which can be used for advances in human germ line biology.

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

    Institute of Scientific and Technical Information of China (English)

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


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

  2. Hepatopancreatoduodenectomy for anastomotic recurrence from residual cholangiocarcinoma: report of a case. (United States)

    Natsume, Seiji; Ebata, Tomoki; Yokoyama, Yukihiro; Igami, Tsuyoshi; Sugawara, Gen; Takahashi, Yu; Nagino, Masato


    Resection of cholangiocarcinoma often results in a positive ductal margin, from carcinoma in situ (CIS) near the main tumor; however, the biological behavior of the residual CIS after surgical resection remains equivocal. We report a case of late local recurrence of CIS, defined as long-term tumor progression from CIS residue at the ductal stump. The patient, a 73-year-old man, had undergone bile duct resection for distal cholangiocarcinoma, leaving positive ductal margins with CIS. A biliary stricture was found 10 years later at the site of anastomosis, and right hepatectomy with pancreatoduodenectomy was performed. Based on histological analogy and the evidence of remnant CIS, a final diagnosis of late local recurrence from the CIS foci was made. This uncommon mode of recurrence should be considered in patients with early-stage disease with expected favorable survival because salvage surgery is feasible for selected patients.

  3. Endopancreatic Bile Duct Cholangiocarcinoma in a Patient with Peutz-Jeghers Syndrome

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    Alexandros K. Charalabopoulos


    Full Text Available Peutz-Jeghers syndrome is a rare autosomal dominant inherited disease characterized by a special type of hamartomatous gastrointestinal polyps combined with mucocutaneous melanin pigmentations. Patients with the syndrome have a high risk of developing neoplasia, with colon, small bowel, and stomach being the most common gastrointestinal sites. Herein, we present the occurrence of a rare tumor in patients with Peutz-Jeghers syndrome; a cholangiocarcinoma of the endopancreatic bile duct. A minireview is also presented. It can be concluded that cholangiocarcinoma remains a possible diagnosis in PJS patients, as in others that present with biliary obstruction. PJS patients may be at higher risk than others in view of their propensity for malignancy.

  4. Hemobilia as the initial manifestation of cholangiocarcinoma in a hemophilia B patient

    Institute of Scientific and Technical Information of China (English)


    Hemobilia is a rare manifestation of hemophilia and is usually iatrogenic following liver biopsy. There are only few reports of spontaneous hemobilia in hemophilia patients. Cholangiocarcinoma is a well- established cause of hemobilia. We describe a case of a 70-year-old male, with known haemophilia B and a past history of papillotomy, who presented with classical symptoms of hemobilia. The initial diagnostic work-up failed to demonstrate a potential cause of bleeding other than the coagulopathy. Three months later, he was readmitted to our hospital with a second episode of hemobilia. During the second work-up, a cholangiocarcinoma was diagnosed both by imaging studies and by a significant elevation of cancer antigen 19-9. Although hemobilia could be attributed to hemophilia, especially in a patient with previous papillotomy, an underlying malignancy of the biliary tree should be suspected.

  5. Integrative Genomic Analysis of Cholangiocarcinoma Identifies Distinct IDH-Mutant Molecular Profiles

    Directory of Open Access Journals (Sweden)

    Farshad Farshidfar


    Full Text Available Cholangiocarcinoma (CCA is an aggressive malignancy of the bile ducts, with poor prognosis and limited treatment options. Here, we describe the integrated analysis of somatic mutations, RNA expression, copy number, and DNA methylation by The Cancer Genome Atlas of a set of predominantly intrahepatic CCA cases and propose a molecular classification scheme. We identified an IDH mutant-enriched subtype with distinct molecular features including low expression of chromatin modifiers, elevated expression of mitochondrial genes, and increased mitochondrial DNA copy number. Leveraging the multi-platform data, we observed that ARID1A exhibited DNA hypermethylation and decreased expression in the IDH mutant subtype. More broadly, we found that IDH mutations are associated with an expanded histological spectrum of liver tumors with molecular features that stratify with CCA. Our studies reveal insights into the molecular pathogenesis and heterogeneity of cholangiocarcinoma and provide classification information of potential therapeutic significance.

  6. Radiosensitivity of Human Melanoma Cell Lines

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


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

  7. Generation of human induced pluripotent stem cells from dermal fibroblasts. (United States)

    Lowry, W E; Richter, L; Yachechko, R; Pyle, A D; Tchieu, J; Sridharan, R; Clark, A T; Plath, K


    The generation of patient-specific pluripotent stem cells has the potential to accelerate the implementation of stem cells for clinical treatment of degenerative diseases. Technologies including somatic cell nuclear transfer and cell fusion might generate such cells but are hindered by issues that might prevent them from being used clinically. Here, we describe methods to use dermal fibroblasts easily obtained from an individual human to generate human induced pluripotent stem (iPS) cells by ectopic expression of the defined transcription factors KLF4, OCT4, SOX2, and C-MYC. The resultant cell lines are morphologically indistinguishable from human embryonic stem cells (HESC) generated from the inner cell mass of a human preimplantation embryo. Consistent with these observations, human iPS cells share a nearly identical gene-expression profile with two established HESC lines. Importantly, DNA fingerprinting indicates that the human iPS cells were derived from the donor material and are not a result of contamination. Karyotypic analyses demonstrate that reprogramming of human cells by defined factors does not induce, or require, chromosomal abnormalities. Finally, we provide evidence that human iPS cells can be induced to differentiate along lineages representative of the three embryonic germ layers indicating the pluripotency of these cells. Our findings are an important step toward manipulating somatic human cells to generate an unlimited supply of patient-specific pluripotent stem cells. In the future, the use of defined factors to change cell fate may be the key to routine nuclear reprogramming of human somatic cells.

  8. Role of diffusion-weighted magnetic resonance imaging in the diagnosis of extrahepatic cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)


    AIM: To determine the clinical value of diffusion-weight- ed imaging (DWI) for the diagnosis of extrahepatic cholangiocarcinoma (EHCC) by comparing the diagnostic sensitivity of DWI and magnetic resonance cholan-giopancreatography (MRCP). METHODS: Magnetic resonance imaging examination was performed in 56 patients with suspected EHCC. T1- weighted imaging, T2-weighted imaging, MRCP and DWI sequence, DWI using single-shot spin-echo echoplanar imaging sequence with different b values (100, 300, 500, 800 and 1...

  9. [Classification and staging systems for hilar cholangio-carcinoma (Klatskin tumors): clinical application and practical relevance]. (United States)

    Gavrilovici, V; Grecu, F; Seripcariu, V; Dragomir, Cr


    Hilar cholangiocarcinomas or Klatskin tumors have been classified in 1975 by French surgeons Henri Bismuth and Marvin B. Corlette and this remains largely used in clinical practice. The authors present the TNM classification and the changes introduced by the sixth and seventh edition of Union for International Cancer Control regarding the tumors of the proximal bile duct and describe Blumgart classification for tumors of this site. The usefulness of these systems is assessed considering the last six years experience of the service.

  10. Prospective study of MMP7 serum levels in the diagnosis of cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Kawin; Leelawat; Siriluck; Narong; Jerasak; Wannaprasert; Thawee; Ratanashu-ek


    AIM: To determine whether the serum level of matrix metalloproteinase-7 (MMP7) has the potential to diagnosis cholangiocarcinoma from benign biliary tract diseases. METHODS: This study was performed according to the PRoBE (a prospective-specimen-collection, retrospectiveblinded-evaluation) design. A total of 187 patients with obstructive jaundice were consecutively enrolled. After the diagnostic status of these patients was ascertained, their levels of serum MMP7 were assayed and compared with serum carbohy...

  11. Heterozygosity for the alpha1-antitrypsin Z allele may confer genetic risk of cholangiocarcinoma. (United States)

    Mihalache, F; Höblinger, A; Grünhage, F; Krawczyk, M; Gärtner, B C; Acalovschi, M; Sauerbruch, T; Lammert, F; Zimmer, V


    Alpha1-antitrypsin (α1AT) deficiency caused by Z allele homozygosity represents a well-established risk factor for hepatocellular carcinoma. Previous studies have also implicated α1AT Z heterozygosity in cholangiocarcinogenesis. To assess the 'common' Z and S alleles as well as the promoter variant rs8004738 for association with cholangiocarcinoma. We genotyped 182 Caucasian patients and 350 controls for rs28929474 (Z), rs17580 (S) and the variant rs8004738. Exploratory analyses were performed in relation to gender and cholangiocarcinoma localisation. rs28929474 was significantly enriched in the cholangiocarcinoma group (4.1 vs. 1.7%; OR 2.46, 95% CI 1.14-5.32; Bonferroni corrected p(c) = 0.036), reinforced by Armitage trend testing (OR 2.53; p(c) = 0.032). The rs8004738 (promoter) minor allele tended to be overrepresented in Z heterozygotes (30.0 vs. 16.7%: P = 0.13). Exploratory data analyses suggested a high genetic risk for extrahepatic tumour localisation (OR 3.0; p(c) = 0.016) and potentially female Z allele carriers (OR 3.37; unadjusted P = 0.022, p(c) = 0.088). These data point to a novel role of α1AT Z heterozygosity as a potential genetic susceptibility factor for cholangiocarcinoma formation and suggest a contribution of aberrant α1AT function in biliary carcinogenesis. However, given the overall low rs28929474 minor allele frequency, larger studies are warranted to confirm and extend our findings. © 2010 Blackwell Publishing Ltd.

  12. Stereological quantification of mast cells in human synovium

    DEFF Research Database (Denmark)

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


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

  13. Technical Challenges in the Derivation of Human Pluripotent Cells

    Directory of Open Access Journals (Sweden)

    Parinya Noisa


    Full Text Available It has long been discovered that human pluripotent cells could be isolated from the blastocyst state of embryos and called human embryonic stem cells (ESCs. These cells can be adapted and propagated indefinitely in culture in an undifferentiated manner as well as differentiated into cell representing the three major germ layers: endoderm, mesoderm, and ectoderm. However, the derivation of human pluripotent cells from donated embryos is limited and restricted by ethical concerns. Therefore, various approaches have been explored and proved their success. Human pluripotent cells can also be derived experimentally by the nuclear reprogramming of somatic cells. These techniques include somatic cell nuclear transfer (SCNT, cell fusion and overexpression of pluripotent genes. In this paper, we discuss the technical challenges of these approaches for nuclear reprogramming, involving their advantages and limitations. We will also highlight the possible applications of these techniques in the study of stem cell biology.

  14. Human B cell activating factor (BCAF): production by a human T cell tumor line. (United States)

    Fevrier, M; Diu, A; Mollier, P; Abadie, A; Olive, D; Mawas, C; Theze, J


    In a previous study, we demonstrated that supernatants from human T cell clones stimulated by a pair of anti-CD2 monoclonal antibodies cause resting human B cells to become activated and to proliferate in the absence of any other signals. The activity responsible for these effects was shown to be different from already characterized lymphokines and in particular from IL-2 and IL-4, and was named B Cell Activating Factor or BCAF. In this paper, we describe the production of BCAF by a human T cell tumor line T687 after phorbol myristate acetate (PMA) stimulation; this production can be potentiated by phytohemagglutinin (PHA). We further show that the stimulatory phase can be separated from the secretory phase thereby avoiding contamination of BCAF-containing supernatant by PMA and PHA. Supernatants produced under these conditions do not contain either IL-4 or IFN but contain traces of lymphotoxin and 2 to 10 ng/ml of IL-2. The T687 cell line will allow us to obtain a large volume of supernatant for biochemical study and purification of the molecule(s) responsible for BCAF activity.

  15. Trichloroethylene toxicity in a human hepatoma cell line

    Energy Technology Data Exchange (ETDEWEB)

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


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

  16. Human Immunodeficiency Syndromes Affecting Human Natural Killer Cell Cytolytic Activity


    Ham, Hyoungjun; Billadeau, Daniel D.


    Natural killer (NK) cells are lymphocytes of the innate immune system that secrete cytokines upon activation and mediate the killing of tumor cells and virus-infected cells, especially those that escape the adaptive T cell response caused by the down regulation of MHC-I. The induction of cytotoxicity requires that NK cells contact target cells through adhesion receptors, and initiate activation signaling leading to increased adhesion and accumulation of F-actin at the NK cell cytotoxic synaps...

  17. Experience with gemcitabine and cisplatin in the therapy of inoperable and metastatic cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Chaiyut Charoentum; Sumitra Thongprasert; Busyamas Chewaskulyong; Sutthirak Munprakan


    AIM:To study the activity of gemcitabine and cisplatin in a cohort of patients with inoperable or metastatic cholangiocarcinoma.METHODS: Chemotherapy-naive patients with pathologically proven cholangiocarcinoma, receiving treatment that consisted of gemcitabine at 1250 mg/m2in a 30-min infusion on d 1 and 8, and cisplatin at 75 mg/m2 at every 21-d cycle, were retrospectively analyzed.RESULTS: From June 2003 to December 2005, 42patients were evaluated. Twelve patients (28%) had unresectable disease and 30 (72%) had metastatic disease. There were 28 males and 14 females with a median age of 51 years (range 33-67) and median ECOG PS of 1 (range 0-2). A total of 171 cycles were given with a median number of cycles of 4 (range 1-6).There were 0 CR, 9 PR, 11 SD and 13 PD (response rate 21%). Grade 3-4 hematologic toxicities were: anemia in 33%, neutropenia in 22% and thrombocytopenia in 5%.Non-hematologic toxicity was generally mild. No cases of febrile neutropenia or treatment-related death were noted. The median survival was 10.8 mo (range 8.4-13mo) and progression free survival was 8.5 mo. One-year survival rate was 40%.CONCLUSION: Our results indicate that the combination of gemcitabine and cisplatin had consistent efficacy in patients with unresectable or metastatic cholangiocarcinoma.

  18. Correlation of p53 gene mutation and expression of P53 protein in cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Xiao-Fang Liu; Hao Zhang; Shi-Guang Zhu; Xian-Ting Zhou; Hai-Long Su; Zheng Xu; Shao-Jun Li


    AIM: To characterize the tumor suppressor gene p53 mutations and study the correlation of p53 gene mutation and the expression of P53 protein in cholangiocarcinoma.METHODS: A total of 36 unselected, frozen samples of cholangiocarcinoma were collected. p53 gene status(exon 5-8) and P53 protein were examined by automated sequencing and immunohistochemical staining, combined with the clinical parameters of patients.RESULTS: p53 gene mutations were found in 22 of 36 (61.1%) patients. Nineteen of 36 (52.8%) patients were positive for P53 protein expression. There were significant differences in extent of differentiation and invasion between the positive and negative expression of P53 protein. However, there were no significant differences in pathologic parameters between the mutations and non-mutations.CONCLUSION: The alterations of the p53 gene evaluated by DNA sequence analysis is relatively accurate. Expression of P53 protein could not act as an independent index to estimate the prognosis of cholangiocarcinoma.

  19. Cholangiocarcinoma presenting as a solitary epididymal metastasis: a case report and review of the literature

    Directory of Open Access Journals (Sweden)

    Bailey David M


    Full Text Available Abstract Background Solid tumor metastasis to the epididymis is a rare occurrence and is mostly discovered incidentally at autopsy or after therapeutic orchidectomy for prostate cancer. Other primary carcinomas that have been demonstrated to metastasize to the paratesticular region include those originating in the stomach, kidney, ileum, and colon. Case presentation A 72-year-old gentleman presented with a firm and tender mass involving the right epididymis. On examination, he was jaundiced. Computed tomography of the abdomen demonstrated an obstructive stricture of the extra-hepatic bile ducts, in keeping with a cholangiocarcinoma, through which a metal stent was endoscopically inserted for symptomatic relief. Subsequent right radical orchidectomy yielded a diffusely infiltrative adenocarcinoma obliterating the epididymis, extending into the rete testis, vas deferens and spermatic cord and showing widespread vascular and perineural invasion. Residual epididymal, rete, and testicular tubules showed no in situ neoplasia. Morphologically and immunohistochemically the features were in keeping with a metastasis from a primary cholangiocarcinoma. Conclusion Only two cases of bile duct carcinoma metastasising to the male genital tract have previously been reported in the literature, the testis being the main site of metastasis in both cases. To our knowledge, this is the first described case of cholangiocarcinoma metastasising primarily to the epididymis, and presenting as a solitary epididymal metastasis in the absence of disseminated disease. It serves to highlight the importance of performing a thorough examination of the male external genitalia both clinically, in the follow up of cancer patients, and at autopsy.

  20. Human immunodeficiency virus type 1 infection of human uterine epithelial cells: viral shedding and cell contact-mediated infectivity. (United States)

    Asin, Susana N; Wildt-Perinic, Dunja; Mason, Sarah I; Howell, Alexandra L; Wira, Charles R; Fanger, Michael W


    We examined the mechanism of human immunodeficiency virus (HIV) type 1 infection of human uterine epithelial cells to gain a clearer understanding of the events by which HIV-1 infects cells within the female reproductive tract. We demonstrated that these cells can be productively infected by HIV-1 and that infection is associated with viral RNA reverse transcription, DNA transcription, and secretion of infectious virus. Levels of viral DNA and secreted virus decreased gradually after infection. Moreover, virus released by the uterine epithelial cells shortly after infection was able to infect human T cell lines, but virus released later did not. In contrast, human CD4(+) T cell lines were infected after cocultivation with epithelial cells at both early and late stages of infection. These data demonstrated that HIV-1 infects human epithelial cells of upper reproductive tract origin and that productive viral infection of epithelial cells may be an important mechanism of transmission of HIV-1 infection in women.

  1. Effect of human neural progenitor cells on injured spinal cord

    Institute of Scientific and Technical Information of China (English)

    XU Guang-hui; BAI Jin-zhu; CAI Qin-lin; LI Xiao-xia; LI Ling-song; SHEN Li


    Objective: To study whether human neural progenitor cells can differentiate into neural cells in vivo and improve the recovery of injured spinal cord in rats.Methods: Human neural progenitor cells were transplanted into the injured spinal cord and the functional recovery of the rats with spinal cord contusion injury was evaluated with Basso-Beattie-Bresnahan (BBB) locomotor scale and motor evoked potentials. Additionally, the differentiation of human neural progenitor cells was shown by immunocytochemistry.Results: Human neural progenitor cells developed into functional cells in the injured spinal cord and improved the recovery of injured spinal cord in both locomotor scores and electrophysiological parameters in rats.Conclusions: Human neural progenitor cells can treat injured spinal cord, which may provide a new cell source for research of clinical application.

  2. Generation of induced pluripotent stem cells from human blood. (United States)

    Loh, Yuin-Han; Agarwal, Suneet; Park, In-Hyun; Urbach, Achia; Huo, Hongguang; Heffner, Garrett C; Kim, Kitai; Miller, Justine D; Ng, Kitwa; Daley, George Q


    Human dermal fibroblasts obtained by skin biopsy can be reprogrammed directly to pluripotency by the ectopic expression of defined transcription factors. Here, we describe the derivation of induced pluripotent stem cells from CD34+ mobilized human peripheral blood cells using retroviral transduction of OCT4/SOX2/KLF4/MYC. Blood-derived human induced pluripotent stem cells are indistinguishable from human embryonic stem cells with respect to morphology, expression of surface antigens, and pluripotency-associated transcription factors, DNA methylation status at pluripotent cell-specific genes, and the capacity to differentiate in vitro and in teratomas. The ability to reprogram cells from human blood will allow the generation of patient-specific stem cells for diseases in which the disease-causing somatic mutations are restricted to cells of the hematopoietic lineage.

  3. A Chemical Probe that Labels Human Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Nao Hirata


    Full Text Available A small-molecule fluorescent probe specific for human pluripotent stem cells would serve as a useful tool for basic cell biology research and stem cell therapy. Screening of fluorescent chemical libraries with human induced pluripotent stem cells (iPSCs and subsequent evaluation of hit molecules identified a fluorescent compound (Kyoto probe 1 [KP-1] that selectively labels human pluripotent stem cells. Our analyses indicated that the selectivity results primarily from a distinct expression pattern of ABC transporters in human pluripotent stem cells and from the transporter selectivity of KP-1. Expression of ABCB1 (MDR1 and ABCG2 (BCRP, both of which cause the efflux of KP-1, is repressed in human pluripotent stem cells. Although KP-1, like other pluripotent markers, is not absolutely specific for pluripotent stem cells, the identified chemical probe may be used in conjunction with other reagents.

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

    DEFF Research Database (Denmark)

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


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


    NARCIS (Netherlands)



    In the present study the cytotoxicity of 16 proanthocyanidins was evaluated in GLC(4), a human small cell lung carcinoma cell line, and in COLO 320, a human colorectal cancer cell line, using the microculture tetrazolium (MTT) assay. With IC50 values ranging from 18 to >200 mu m following continuous

  6. Derivation and differentiation of haploid human embryonic stem cells. (United States)

    Sagi, Ido; Chia, Gloryn; Golan-Lev, Tamar; Peretz, Mordecai; Weissbein, Uri; Sui, Lina; Sauer, Mark V; Yanuka, Ofra; Egli, Dieter; Benvenisty, Nissim


    Diploidy is a fundamental genetic feature in mammals, in which haploid cells normally arise only as post-meiotic germ cells that serve to ensure a diploid genome upon fertilization. Gamete manipulation has yielded haploid embryonic stem (ES) cells from several mammalian species, but haploid human ES cells have yet to be reported. Here we generated and analysed a collection of human parthenogenetic ES cell lines originating from haploid oocytes, leading to the successful isolation and maintenance of human ES cell lines with a normal haploid karyotype. Haploid human ES cells exhibited typical pluripotent stem cell characteristics, such as self-renewal capacity and a pluripotency-specific molecular signature. Moreover, we demonstrated the utility of these cells as a platform for loss-of-function genetic screening. Although haploid human ES cells resembled their diploid counterparts, they also displayed distinct properties including differential regulation of X chromosome inactivation and of genes involved in oxidative phosphorylation, alongside reduction in absolute gene expression levels and cell size. Surprisingly, we found that a haploid human genome is compatible not only with the undifferentiated pluripotent state, but also with differentiated somatic fates representing all three embryonic germ layers both in vitro and in vivo, despite a persistent dosage imbalance between the autosomes and X chromosome. We expect that haploid human ES cells will provide novel means for studying human functional genomics and development.

  7. Human embryonic stem cells and respect for life


    Meyer, J.(CERN, Geneva, Switzerland)


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

  8. Abnormalities in human pluripotent cells due to reprogramming mechanisms. (United States)

    Ma, Hong; Morey, Robert; O'Neil, Ryan C; He, Yupeng; Daughtry, Brittany; Schultz, Matthew D; Hariharan, Manoj; Nery, Joseph R; Castanon, Rosa; Sabatini, Karen; Thiagarajan, Rathi D; Tachibana, Masahito; Kang, Eunju; Tippner-Hedges, Rebecca; Ahmed, Riffat; Gutierrez, Nuria Marti; Van Dyken, Crystal; Polat, Alim; Sugawara, Atsushi; Sparman, Michelle; Gokhale, Sumita; Amato, Paula; Wolf, Don P; Ecker, Joseph R; Laurent, Louise C; Mitalipov, Shoukhrat


    Human pluripotent stem cells hold potential for regenerative medicine, but available cell types have significant limitations. Although embryonic stem cells (ES cells) from in vitro fertilized embryos (IVF ES cells) represent the 'gold standard', they are allogeneic to patients. Autologous induced pluripotent stem cells (iPS cells) are prone to epigenetic and transcriptional aberrations. To determine whether such abnormalities are intrinsic to somatic cell reprogramming or secondary to the reprogramming method, genetically matched sets of human IVF ES cells, iPS cells and nuclear transfer ES cells (NT ES cells) derived by somatic cell nuclear transfer (SCNT) were subjected to genome-wide analyses. Both NT ES cells and iPS cells derived from the same somatic cells contained comparable numbers of de novo copy number variations. In contrast, DNA methylation and transcriptome profiles of NT ES cells corresponded closely to those of IVF ES cells, whereas iPS cells differed and retained residual DNA methylation patterns typical of parental somatic cells. Thus, human somatic cells can be faithfully reprogrammed to pluripotency by SCNT and are therefore ideal for cell replacement therapies.

  9. Proteomic analysis of human blastocoel fluid and blastocyst cells

    DEFF Research Database (Denmark)

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


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


    Institute of Scientific and Technical Information of China (English)

    张建军; 丁尔迅; 王强; 陈学云; 付志仁


    To investigate the expression of Fas ligand in human colon carcinoma cell lines. Methods: A total of six human colon cancer cell lines were examined for the expression of Fas ligand mRNA and cell surface protein by using RT-PCR and flow cytometry respectively. Results: The results showed that Fas ligand mRNA was expressed in all of the six cancer cell lines and Fas ligand cell surface protein was expressed in part of them. Conclusion: These data suggest that Fas ligand was expressed, at least in part, in human colon cancer cell lines and might facilitate to escape from immune surveillance of the host.

  11. Derivation and characterization of human embryonic stem cells on human amnion epithelial cells. (United States)

    Lai, Dongmei; Wang, Yongwei; Sun, Jian; Chen, Yifei; Li, Ting; Wu, Yi; Guo, Lihe; Wei, Chunsheng


    Culture conditions that support the growth of undifferentiated human embryonic stem cells (hESCs) have already been established using primary human amnion epithelial cells (hAECs) as an alternative to traditional mitotically inactivated mouse embryonic fibroblasts (MEFs). In the present work, inner cell masses (ICM) were isolated from frozen embryos obtained as donations from couples undergoing in vitro fertilization (IVF) treatment and four new hESC lines were derived using hAECs as feeder cells. This feeder system was able to support continuous growth of what were, according to their domed shape and markers, undifferentiated naïve-like hESCs. Their pluripotent potential were also demonstrated by embryoid bodies developing to the expected three germ layers in vitro and the productions of teratoma in vivo. The cell lines retained their karyotypic integrity for over 35 passages. Transmission electron microscopy (TEM) indicated that these newly derived hESCs consisted mostly of undifferentiated cells with large nuclei and scanty cytoplasm. The new hESCs cultured on hAECs showed distinct undifferentiated characteristics in comparison to hESCs of the same passage maintained on MEFs. This type of optimized culture system may provide a useful platform for establishing clinical-grade hESCs and assessing the undifferentiated potential of hESCs.

  12. Melatonin inhibits cholangiocarcinoma and reduces liver injury in Opisthorchis viverrini-infected and N-nitrosodimethylamine-treated hamsters. (United States)

    Laothong, Umawadee; Pinlaor, Porntip; Boonsiri, Patcharee; Pairojkul, Chawalit; Priprem, Aroonsri; Johns, Nutjaree Pratheepawanit; Charoensuk, Lakhanawan; Intuyod, Kitti; Pinlaor, Somchai


    The human liver fluke Opisthorchis viverrini infection and N-nitrosodimethylamine (NDMA) administration induce cholangiocarcinoma (CCA) and liver injury in hamsters. Melatonin protects against liver injury and reduces the alteration of mitochondrial structure, mitochondrial membrane potential, and mitochondrial pro- and anti-apoptotic pathways in various cancer types. To investigate the chemopreventive effect of melatonin on CCA genesis and liver injury, hamsters were treated with a combination of O. viverrini infection and NDMA concurrently administered with melatonin (10 mg/kg and 50 mg/kg) for 120 days. Melatonin treatment at 50 mg/kg caused a significant reduction in liver/body weight ratios and decreased tumor volumes leading to an increase in the survival of animals. In the tumorous tissues, the high-dose melatonin reduced DNA fragmentation and mitochondrial apoptosis by inducing anti-apoptotic protein (Bcl-2) in the mitochondrial fraction and down-regulating cytochrome c, pro-apoptotic protein (Bax), and caspase-3 in tumor cytosol. Moreover, a high-dose melatonin treatment significantly increased mitochondrial antioxidant enzymes and prevented mitochondrial ultrastructure changes in the tumor. Overall, melatonin has potent chemopreventive effects in inhibiting CCA genesis and also reduces liver injury in hamster CCA, which, in part, might involve in the suppression of CCA by reducing tumor mitochondria alteration.

  13. Dynamic behaviour of human neuroepithelial cells in the developing forebrain (United States)

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


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

  14. Derivation of a Homozygous Human Androgenetic Embryonic Stem Cell Line. (United States)

    Ding, Chenhui; Huang, Sunxing; Qi, Quan; Fu, Rui; Zhu, Wanwan; Cai, Bing; Hong, Pingping; Liu, Zhengxin; Gu, Tiantian; Zeng, Yanhong; Wang, Jing; Xu, Yanwen; Zhao, Xiaoyang; Zhou, Qi; Zhou, Canquan


    Human embryonic stem cells (hESCs) have long been considered as a promising source for cell replacement therapy. However, one major obstacle for the use of these cells is immune compatibility. Histocompatible human parthenogenetic ESCs have been reported as a new method for generating human leukocyte antigen (HLA)-matched hESCs. To further investigate the possibility of obtaining histocompatible stem cells from uniparental embryos, we tried to produce androgenetic haploid human embryos by injecting a single spermatozoon into enucleated human oocyte, and establish human androgenetic embryonic stem (hAGES) cell lines from androgenetic embryos. In the present study, a diploid hAGES cell line has been established, which exhibits typical features of human ESCs, including the expression of pluripotency markers, having differentiation potential in vitro and in vivo, and stable propagation in an undifferentiated state (>P40). Bisulfite sequencing of the H19, Snrpn, Meg3, and Kv imprinting control regions suggested that hAGES cells maintained to a certain extent a sperm methylation pattern. Genome-wide single nucleotide polymorphism, short tandem repeat, and HLA analyses revealed that the hAGES cell genome was highly homozygous. These results suggest that hAGES cells from spermatozoon could serve as a useful tool for studying the mechanisms underlying genomic imprinting in humans. It might also be used as a potential resource for cell replacement therapy as parthenogenetic stem cells.

  15. Human Cell and Tissue Establishment Registration Public Query (United States)

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

  16. Human Cell and Tissue Establishment Registration Public Query (United States)

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

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

    African Journals Online (AJOL)

    Human tissue legislation in South Africa: Focus on stem cell research and therapy. ... Related Substances Act, the Consumer Protection Act, the Children's Act and ... human tissue legislation in SA, the legislator has an opportunity to mirror the ...

  18. Human Stem Cell Derived Cardiomyocytes: An Alternative ... (United States)

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

  19. Orotate phosphoribosyl transferase mRNA expression and the response of cholangiocarcinoma to 5-fluorouracil

    Institute of Scientific and Technical Information of China (English)

    Chariya Hahnvajanawong; Jariya Chaiyagool; Wunchana Seubwai; Vajarabhongsa Bhudhisawasdi; Nisana Namwat; Narong Khuntikeo; Banchob Sripa


    AIM:To determine whether expression of certain enzymes related to 5-fluorouracil (5-FU) metabolism predicts 5-FU chemosensitivity in cholangiocarcinoma (CCA).METHODS:The histoculture drug response assay (HDRA) was performed using surgically resected CCA tissues.Tumor cell viability was determined morphologically with hematoxylin and eosin-and terminal deoxynucleotide transferase-mediated dUTP nick-end labeling-stained tissues.The mRNA expression of thymidine phosphorylase (TP),orotate phosphoribosyl transferase (OPRT),thymidylate synthase (TS),and dihydropyrimidine dehydrogenase (DPD) was determined with realtime reverse transcriptase-polymerase chain reaction.The levels of gene expression and the sensitivity to 5-FU were evaluated.RESULTS:Twenty-three CCA tissues were obtained from patients who had been diagnosed with intrahepatic CCA and who underwent surgical resection at Srinagarind Hospital,Khon Kaen University from 2007 to 2009.HDRA was used to determine the response of these CCA tissues to 5-FU.Based on the dose-response curve,200 μg/mL 5-FU was selected as the test concentration.The percentage of inhibition index at the median point was selected as the cut-off point to differentiate the responding and non-responding tumors to 5-FU.When the relationship between TP,OPRT,TS and DPD mRNA expression levels and the sensitivity of CCA tissues to 5-FU was examined,only OPRT mRNA expression was significantly correlated with the response to 5-FU.The mean expression level of OPRT was significantly higher in the responder group compared to the non-responder group (0.41 ± 0.25 vs 0.22 ± 0.12,P < 0.05).CONCLUSION:OPRT mRNA expression may be a useful predictor of 5-FU chemosensitivity of CCA.Whether OPRT mRNA could be used to predict the success of 5-FU chemotherapy in CCA patients requires confirmation in patients.

  20. Preoperative assessment of hilar cholangiocarcinoma:combination of cholangiographyandCTangiography

    Institute of Scientific and Technical Information of China (English)

    Shi-An Yu; Cheng Zhang; Jia-Min Zhang; Gen-Jun Mao; Long-Tang Xu; Xiao-Kang Wu; Jin-Er Shu; Guang-Hong Lv; Zhang-Dong Zheng


    BACKGROUND: Hilar cholangiocarcinoma is one of the most dififcult carcinomas to manage because of the location of the main tumor at the hepatic hilus and the complex anatomy of the biliary, arterial, and portal systems. To plan an operation, it is important to acquire accurate information about the relationship between hilar cholangiocarcinoma and adjacent vessels. This study aimed to evaluate the clinical value of cholangiography combined with spiral CT three-dimensional (3D) angiography for a preoperative assessment of hilar cholangiocarcinoma. METHODS: From March 2007 to August 2009, cholangiography was performed in 13 patients with hilar cholangiocarcinoma. Meanwhile, contrast-enhanced abdominal scanning was performed using 16-slice spiral CT, and the 3D images of the hepatic artery and portal vein were acquired. The level and range of invasion of the hepatic artery, the portal vein, and the bile duct, the preoperative Bismuth classiifcation, and T-staging were recorded and compared with those after surgical exploration. RESULTS: The hepatic artery and portal vein were recon-structed successfully in all these patients. Percutaneous transhepatic cholangiography was performed in 9 patients, endoscopic retrograde cholangiopancreatography in 1, and magnetic resonance cholangiopancreatography in 3. The CT angiography records of invasion of the hepatic artery were consistent with the results of explorations in these patients. The data from 5 of the 13 patients were consistent with those on invasion of the portal vein. The results of the Bismuth classiifcation and the T-staging system were consistent with those of surgical exploration in 12 of the 13 patients. Seven of 8 patients who were estimated to be suitable for operation based on images were curatively treated and 5 who were judged to be unsuitable for curative operation by cholangiography and CT angiography were conifrmed intraoperatively and underwent palliative procedures. CONCLUSIONS

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


    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.

  2. Identification of molecules derived from human fibroblast feeder cells that support the proliferation of human embryonic stem cells

    DEFF Research Database (Denmark)

    Anisimov, Sergey V.; Christophersen, Nicolaj S.; Correia, Ana S.


    the proliferation and pluripotency of these cells. Importantly, feeder cells generally lose their capacity to support human embryonic stem cell proliferation in vitro following long-term culture. In this study, we performed large-scale gene expression profiles of human foreskin fibroblasts during early...

  3. Cholangiocarcinoma Secondary to Primary Sclerosing Cholangitis in Explanted Livers: A Single-Center Study in the South of Iran

    Directory of Open Access Journals (Sweden)



    Full Text Available Background Primary sclerosing cholangitis (PSC is a chronic disease, characterized by chronic inflammation and fibrosis of bile duct epithelial cells. This is a significant contributory factor to the development of malignancy, most commonly cholangiocarcinoma (CCA, which is the second most common malignant liver tumor. Objectives For the first time in Iran, we intend to describe our experience with cases of PSC, with and without CCA, in explanted livers, and compare our results with those found in other areas of the world. Patients and Methods The study population comprised 181 individuals with a diagnosis of PSC who had undergone liver transplantation in the main liver transplant center of Iran, the largest center of hepatobiliary surgery in the south of that country, over a 3-year period between 2012 and 2014. All explanted livers, with and without CCA, were evaluated. Results Of the 181 patients, 16 were found to have CCA, two of whom had been diagnosed after pathologic study of the explanted livers. Therefore it appeared that 8.8% of the patients with PSC in our center had developed CCA before liver transplantation. Conclusions A comparison of our results with those obtained from other centers in both Western and Asian countries (which reported CCA in 3.6% - 36.5% of patients with PSC, shows that the incidence of CCA in the patients we studied is intermediate.

  4. Predictors of long term survival after hepatic resection for hilar cholangiocarcinoma:A retrospective study of 5-year survivors

    Institute of Scientific and Technical Information of China (English)

    Mohamed Abd El Wahab; Ayman El Nakeeb; Ehab El Hanafy; Ahmad M Sultan; Ahmed Elghawalby; Waleed Askr; Mahmoud Ali; Mohamed Abd El Gawad; Tarek Salah


    AIM:To determine predictors of long term survival after resection of hilar cholangiocarcinoma(HC) by comparing patients surviving > 5 years with those who survived 5 years.RESULTS:There were 34(14%) long term survivors(5 year survivors) among the 243 patients.Fiveyear survivors were younger at diagnosis than those surviving less than 5 years(mean age,50.47 ± 4.45 vs 54.59 ± 4.98,p = 0.001).Gender,clinical presentation,preoperative drainage,preoperative serum bilirubin,albumin and serum glutamic-pyruvic transaminase were similar between the two groups.The level of CA 19-9 was significantly higher in patients surviving 5 year,includingyoung age(p = 0.001),serum CA19-9(p = 0.0001),non-cirrhotic liver(p = 0.02),major hepatic resection(p = 0.001),caudate lobe resection(p = 0.006),well differentiated tumour(p = 0.03),lymph node status(0.008),R0 resection margin(p = 0.0001) and early postoperative liver cell failure(p = 0.02).CONCLUSION:Liver status,resection of caudate lobe,lymph node status,R0 resection and CA19-9 were demonstrated to be independent risk factors for long term survival.

  5. microRNA与胆管癌关系的研究进展%MicroRNAs in cholangiocarcinoma: recent progress

    Institute of Scientific and Technical Information of China (English)



    microRNA(miRNA)是一组内源性的非编码RNA,其通过转录后水平调节靶基因的表达,在细胞增殖、分化、凋亡及肿瘤的发生、发展中起重要作用.笔者就近年来microRNA在胆管癌研究领域的最新进展进行综述,主要介绍miR-21,miR-370,let-7家族,miR-373,miR-200家族,miR-29b,miR-204,miR-320等与胆管癌的关系.%MicroRNAs (miRNAs) are tiny, endogenously expressed noncoding RNAs that act as the posttranscriptional regulators of gene expression, and play very important roles in cell proliferation, differentiation and apoptosis as well as in cancer occurrence and development. In this paper, the authors overview the recent advances of miRNAs involvement in cholangiocarcinoma, focusing on miR-21, miR-370, let-7 family, miR-373, miR-200 family, miR-29b, miR-204 and miR-320.

  6. Human embryonic stem cell research: ethical and legal issues. (United States)

    Robertson, J A


    The use of human embryonic stem cells to replace damaged cells and tissues promises future hope for the treatment of many diseases. However, many countries now face complex ethical and legal questions as a result of the research needed to develop these cell-replacement therapies. The challenge that must be met is how to permit research on human embryonic tissue to occur while maintaining respect for human life generally.

  7. The diagnostic value of joint detection of serum CA19-9, CA125 and CA242 for cholangiocarcinoma

    Institute of Scientific and Technical Information of China (English)

    Cui Da-Peng; Han Lei; Liu Zhen-Xian; Yang He; Zhang Ying-Chun


    Objective:To study the diagnostic value of joint detection of serum CA19-9, CA125 and CA242 for cholangiocarcinoma.Methods:A total of 35 patients with cholangiocarcinoma who received surgical resection in our hospital were selected as malignant group, 30 patients with cholelithiasis who received surgical resection in our hospital during the same period were selected as benign group, serum samples were collected before surgery to determine CA19-9, CA125 and CA242 content, and cholangiocarcinoma tissue and normal bile duct tissue were collected after surgery to determine the content of proliferation and invasion molecules. Results:Serum CA19-9, CA125 and CA242 levels of malignant group were significantly higher than those of control group; PROX-1, Ki-67, Bcl-2, Bad, Gab1, LOXL2, TRPM7 and CXCL12 levels in cholangiocarcinoma tissue were higher than those in benign bile duct tissue, and E-cadherin level was lower than that in benign bile duct tissue; serum CA19-9, CA125 and CA242 levels were positively correlated with PROX-1, Ki-67, Bcl-2, Bad, Gab1, LOXL2, TRPM7 and CXCL12 levels, and negatively correlated with E-cadherin level.Conclusion:Joint detection of serum CA19-9, CA125 and CA242 can not only provide reference for the diagnosis of cholangiocarcinoma, but can also provide basis for the evaluation of proliferation, invasion and other malignant biological behaviors.

  8. Intrahepatic cholangiocarcinoma in a worker at an offset color proof-printing company: An autopsy case report. (United States)

    Tomimaru, Yoshito; Kobayashi, Shogo; Wada, Hiroshi; Hama, Naoki; Kawamoto, Koichi; Eguchi, Hidetoshi; Kira, Toshihiko; Morii, Eiichi; Doki, Yuichiro; Mori, Masaki; Nagano, Hiroaki


    A 40-year-old Japanese man visited our hospital after test results indicated elevated hepatobiliary enzymes. He had worked at a printing plant for 8 years and been exposed to organic solvents, including 1,2-dichloropropane (1,2-DCP) and dichloromethane (DCM). Abdominal computed tomography (CT) showed an intrahepatic tumor with dilation of the intrahepatic bile duct. He was diagnosed with intrahepatic cholangiocarcinoma. He had no known risk factors for cholangiocarcinoma. Extended left hepatectomy with lymph node dissection was performed and the tumor was histologically diagnosed as well-differentiated adenocarcinoma. A histological examination also showed biliary intraepithelial preneoplastic lesions in non-cancerous liver areas. Two years after surgery, the patient developed jaundice, esophageal varices and ascites. A CT examination showed liver cirrhosis without recurrence of the cholangiocarcinoma. Although a liver transplantation was planned as a therapeutic option for his liver cirrhosis, his liver failure progressed rapidly and he died before transplantation could be performed. At autopsy, fibrosis was found in the whole liver, especially in the wall of the bile duct and periductal area suggesting chronic bile duct injury due to exposure to organic solvents. Taken together, the current case may suggest that exposure to organic solvents, including 1,2-DCP and DCM, is a risk factor for cholangiocarcinoma. Identifying risk factors for cholangiocarcinoma will help identify the mechanism and help prevent development of the disease.

  9. Epigenetic Regulation of Human Embryonic Stem Cells

    Directory of Open Access Journals (Sweden)

    Qidong eHu


    Full Text Available Recently, there has been tremendous progress in characterizing the transcriptional network regulating hESCs (MacArthur et al., 2009; Loh et al., 2011, including those signaling events mediated by Oct4, Nanog and Sox2. There is growing interest in the epigenetic machinery involved in hESC self-renewal and differentiation. In general, epigenetic regulation includeschromatin reorganization, DNA modification and histone modification, which are not directly related to alterations in DNA sequences. Various protein complexes, includingPolycomb, trithorax, NuRD, SWI/SNF andOct4, have been shown to play critical roles in epigenetic control of hESC maintenance and differentiation. Hence, we will formally review recent advances in unraveling the multifaceted role of epigenetic regulation in hESC self-renewal and induced differentiation, particularly with respect to chromatin remodeling and DNA methylation events. Unraveling the molecular mechanisms underlying the maintenance/differentiation of hESCs and reprogramming of somatic cells will greatly strengthen our capacity to generate various types of cells to treat human diseases.

  10. Mast cells and human hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Fabio Grizzi; Barbara Franceschini; Maurizio Chiriva-Internati; Young Liu; Paul L. Hermonat; Nicola Dioguardi


    AIM: To investigate the density of mast cells (MCs) in human hepatocellular carcinoma (HCC), and to determine whether the MCs density has any correlations with histopathological grading, staging or some baseline patient characteristics.METHODS: Tissue sections of 22 primary HCCs were histochemically stained with toluidine blue, in order to be able to quantify the MCs in and around the neoplasm using a computer-assisted image analysis system. HCC was staged and graded by two independent pathologists. To identify the sinusoidal capillarisation of each specimen 3μm thick sections were histochemically stained with sirius red, and semi-quantitatively evaluated by two independent observers. The data were statistically analysed using Spearman′s correlation and Student′s t-test when appropriate.RESULTS: MCs density did not correlate with the age or sex of the patients, the serum alanine aminotransferase (ALT) or aspartate aminotransferase (AST) levels, or the stage or grade of the HCC. No significant differences were found between the MCs density of the patients with and without hepatitis C virus infection, but they were significantly higher in the specimens showing marked sinusoidal capillarisation.CONCLUSION: The lack of any significant correlation between MCs density and the stage or grade of the neoplastic lesions suggests that there is no causal relationship between MCs recruitment and HCC. However, as capillarisation proceeds concurrently with arterial blood supply during hepatocarcinogenesis, MCs may be considered of primary importance in the transition from sinusoidal to capillary-type endothelial cells and the HCC growth.

  11. Derivation, propagation and differentiation of human embryonic stem cells. (United States)

    Conley, Brock J; Young, Julia C; Trounson, Alan O; Mollard, Richard


    Embryonic stem (ES) cells are in vitro cultivated pluripotent cells derived from the inner cell mass (ICM) of the embryonic blastocyst. Attesting to their pluripotency, ES cells can be differentiated into representative derivatives of all three embryonic germ layers (endoderm, ectoderm and mesoderm) both in vitro and in vivo. Although mouse ES cells have been studied for many years, human ES cells have only more recently been derived and successfully propagated. Many biochemical differences and culture requirements between mouse and human ES cells have been described, yet despite these differences the study of murine ES cells has provided important insights into methodologies aimed at generating a greater and more in depth understanding of human ES cell biology. One common feature of both mouse and human ES cells is their capacity to undergo controlled differentiation into spheroid structures termed embryoid bodies (EBs). EBs recapitulate several aspects of early development, displaying regional-specific differentiation programs into derivatives of all three embryonic germ layers. For this reason, EB formation has been utilised as an initial step in a wide range of studies aimed at differentiating both mouse and human ES cells into a specific and desired cell type. Recent reports utilising specific growth factor combinations and cell-cell induction systems have provided alternative strategies for the directed differentiation of cells into a desired lineage. According to each one of these strategies, however, a relatively high cell lineage heterogeneity remains, necessitating subsequent purification steps including mechanical dissection, selective media or fluorescent or magnetic activated cell sorting (FACS and MACS, respectively). In the future, the ability to specifically direct differentiation of human ES cells at 100% efficiency into a desired lineage will allow us to fully explore the potential of these cells in the analysis of early human development, drug

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

    Directory of Open Access Journals (Sweden)

    Nigel G. Kooreman


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

  13. New frontiers in human cell biology and medicine: can pluripotent stem cells deliver? (United States)

    Goldstein, Lawrence S B


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

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

    Directory of Open Access Journals (Sweden)

    Xiaoti Xu


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

  15. CD56 marks human dendritic cell subsets with cytotoxic potential

    NARCIS (Netherlands)

    Roothans, D.; Smits, E.; Lion, E.; Tel, J.; Anguille, S.


    Human plasmacytoid and myeloid dendritic cells (DCs), when appropriately stimulated, can express the archetypal natural killer (NK)-cell surface marker CD56. In addition to classical DC functions, CD56+ DCs are endowed with an unconventional cytotoxic capacity.

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

    Directory of Open Access Journals (Sweden)

    Yajing Huang


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

  17. Stem Cells: A Renaissance in Human Biology Research. (United States)

    Wu, Jun; Izpisua Belmonte, Juan Carlos


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

  18. Neoplastic human embryonic stem cells as a model of radiation resistance of human cancer stem cells. (United States)

    Dingwall, Steve; Lee, Jung Bok; Guezguez, Borhane; Fiebig, Aline; McNicol, Jamie; Boreham, Douglas; Collins, Tony J; Bhatia, Mick


    Studies have implicated that a small sub-population of cells within a tumour, termed cancer stem cells (CSCs), have an enhanced capacity for tumour formation in multiple cancers and may be responsible for recurrence of the disease after treatment, including radiation. Although comparisons have been made between CSCs and bulk-tumour, the more important comparison with respect to therapy is between tumour-sustaining CSC versus normal stem cells that maintain the healthy tissue. However, the absence of normal known counterparts for many CSCs has made it difficult to compare the radiation responses of CSCs with the normal stem cells required for post-radiotherapy tissue regeneration and the maintenance of tissue homeostasis. Here we demonstrate that transformed human embryonic stem cells (t-hESCs), showing features of neoplastic progression produce tumours resistant to radiation relative to their normal counterpart upon injection into immune compromised mice. We reveal that t-hESCs have a reduced capacity for radiation induced cell death via apoptosis and exhibit altered cell cycle arrest relative to hESCs in vitro. t-hESCs have an increased expression of BclXL in comparison to their normal counterparts and re-sensitization of t-hESCs to radiation upon addition of BH3-only mimetic ABT737, suggesting that overexpression of BclXL underpins t-hESC radiation insensitivity. Using this novel discovery platform to investigate radiation resistance in human CSCs, our study indicates that chemotherapy targeting Bcl2-family members may prove to be an adjuvant to radiotherapy capable of targeting CSCs.

  19. Molecular aging and rejuvenation of human muscle stem cells

    DEFF Research Database (Denmark)

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


    Very little remains known about the regulation of human organ stem cells (in general, and during the aging process), and most previous data were collected in short-lived rodents. We examined whether stem cell aging in rodents could be extrapolated to genetically and environmentally variable humans....... Our findings establish key evolutionarily conserved mechanisms of human stem cell aging. We find that satellite cells are maintained in aged human skeletal muscle, but fail to activate in response to muscle attrition, due to diminished activation of Notch compounded by elevated transforming growth...... factor beta (TGF-beta)/phospho Smad3 (pSmad3). Furthermore, this work reveals that mitogen-activated protein kinase (MAPK)/phosphate extracellular signal-regulated kinase (pERK) signalling declines in human muscle with age, and is important for activating Notch in human muscle stem cells. This molecular...

  20. Ultrastructure of interstitial cells in subserosa of human colon

    DEFF Research Database (Denmark)

    Rumessen, Jüri Johannes; Vanderwinden, Jean-Marie; Hansen, Alastair;


    We studied the ultrastructure of interstitial cells in the subserosal/adventitial layer in human colon. An interstitial cell type with an ultrastructure intermediate between fibroblast-like cells (FLC) and interstitial cells of Cajal was identified (IC-SS). IC-SS had thin and flattened branching...

  1. Differentiation of neuroepithelia from human embryonic stem cells



    We describe the method for in vitro differentiation of neuroepithelial cells from human embryonic stem cells under a chemically defined condition. The protocol is established following the fundamental principle of in vivo neuroectodermal specification. The primitive neuroepithelial cells generated by this protocol can be further induced into neuronal and glia cells with forebrain, mid/hind brain, and spinal cord identities.

  2. Identification of a candidate stem cell in human gallbladder. (United States)

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


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

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

    Energy Technology Data Exchange (ETDEWEB)

    Thilly, W.G.


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

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

    Energy Technology Data Exchange (ETDEWEB)

    Thilly, W.G.


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

  5. Training human mesenchymal stromal cells for bone tissue engineering applications

    NARCIS (Netherlands)

    Doorn, J.


    Human mesenchymal stromal cells (hMSCs) are an interesting source for cell therapies and tissue engineering applications, because these cells are able to differentiate into various target tissues, such as bone, cartilage, fat and endothelial cells. In addition, they secrete a wide array of growth fa

  6. 77 FR 5489 - Identification of Human Cell Lines Project (United States)


    ... working with cells derived from one individual or animal species, only to eventually learn that the cells..., morphology, pathologic or disease-state, hybrid or mixed culture, feeder cells, date of origin, etc), the STR... National Institute of Standards and Technology Identification of Human Cell Lines Project AGENCY: National...

  7. The evolution of human cells in terms of protein innovation. (United States)

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


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

  8. Adult human brain cell culture for neuroscience research. (United States)

    Gibbons, Hannah M; Dragunow, Mike


    Studies of the brain have progressed enormously through the use of in vivo and in vitro non-human models. However, it is unlikely such studies alone will unravel the complexities of the human brain and so far no neuroprotective treatment developed in animals has worked in humans. In this review we discuss the use of adult human brain cell culture methods in brain research to unravel the biology of the normal and diseased human brain. The advantages of using adult human brain cells as tools to study human brain function from both historical and future perspectives are discussed. In particular, studies using dissociated cultures of adult human microglia, astrocytes, oligodendrocytes and neurons are described and the applications of these types of study are evaluated. Alternative sources of human brain cells such as adult neural stem cells, induced pluripotent stem cells and slice cultures of adult human brain tissue are also reviewed. These adult human brain cell culture methods could benefit basic research and more importantly, facilitate the translation of basic neuroscience research to the clinic for the treatment of brain disorders. Copyright 2009 Elsevier Ltd. All rights reserved.

  9. 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


    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.

  10. Nanoscale Mechanical Stimulation of Human Mesenchymal Stem Cells

    Directory of Open Access Journals (Sweden)

    H Nikukar


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

  11. Activation of GPR119 Stimulates Human β-Cell Replication and Neogenesis in Humanized Mice with Functional Human Islets (United States)

    Ansarullah; Free, Colette; Christopherson, Jenica; Chen, Quanhai; Gao, Jie; Liu, Chengyang; Naji, Ali; Rabinovitch, Alex; Guo, Zhiguang


    Using humanized mice with functional human islets, we investigated whether activating GPR119 by PSN632408, a small molecular agonist, can stimulate human β-cell regeneration in vivo. Human islets were transplanted under the left kidney capsule of immunodeficient mice with streptozotocin- (STZ-) induced diabetes. The recipient mice were treated with PSN632408 or vehicle and BrdU daily. Human islet graft function in the mice was evaluated by nonfasting glucose levels, oral glucose tolerance, and removal of the grafts. Immunostaining for insulin, glucagon, and BrdU or Ki67 was performed in islet grafts to evaluate α- and β-cell replication. Insulin and CK19 immunostaining was performed to evaluate β-cell neogenesis. Four weeks after human islet transplantation, 71% of PSN632408-treated mice achieved normoglycaemia compared with 24% of vehicle-treated mice. Also, oral glucose tolerance was significantly improved in the PSN632408-treated mice. PSN632408 treatment significantly increased both human α- and β-cell areas in islet grafts and stimulated α- and β-cell replication. In addition, β-cell neogenesis was induced from pancreatic duct cells in the islet grafts. Our results demonstrated that activation of GPR119 increases β-cell mass by stimulating human β-cell replication and neogenesis. Therefore, GPR119 activators may qualify as therapeutic agents to increase human β-cell mass in patients with diabetes. PMID:27413754

  12. Analysis of lead toxicity in human cells

    Directory of Open Access Journals (Sweden)

    Gillis Bruce S


    Full Text Available Abstract Background Lead is a metal with many recognized adverse health side effects, and yet the molecular processes underlying lead toxicity are still poorly understood. Quantifying the injurious effects of lead is also difficult because of the diagnostic limitations that exist when analyzing human blood and urine specimens for lead toxicity. Results We analyzed the deleterious impact of lead on human cells by measuring its effects on cytokine production and gene expression in peripheral blood mononuclear cells. Lead activates the secretion of the chemokine IL-8 and impacts mitogen-dependent activation by increasing the secretion of the proinflammatory cytokines IL-6 and TNF-α and of the chemokines IL-8 and MIP1-α in the presence of phytohemagglutinin. The recorded changes in gene expression affected major cellular functions, including metallothionein expression, and the expression of cellular metabolic enzymes and protein kinase activity. The expression of 31 genes remained elevated after the removal of lead from the testing medium thereby allowing for the measurement of adverse health effects of lead poisoning. These included thirteen metallothionein transcripts, three endothelial receptor B transcripts and a number of transcripts which encode cellular metabolic enzymes. Cellular responses to lead correlated with blood lead levels and were significantly altered in individuals with higher lead content resultantly affecting the nervous system, the negative regulation of transcription and the induction of apoptosis. In addition, we identified changes in gene expression in individuals with elevated zinc protoporphyrin blood levels and found that genes regulating the transmission of nerve impulses were affected in these individuals. The affected pathways were G-protein mediated signaling, gap junction signaling, synaptic long-term potentiation, neuropathic pain signaling as well as CREB signaling in neurons. Cellular responses to lead were

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

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    Sabine Rutti

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

  14. Extracellular protonation modulates cell-cell interaction mechanics and tissue invasion in human melanoma cells (United States)

    Hofschröer, Verena; Koch, Kevin Alexander; Ludwig, Florian Timo; Friedl, Peter; Oberleithner, Hans; Stock, Christian; Schwab, Albrecht


    Detachment of cells from the primary tumour precedes metastatic progression by facilitating cell release into the tissue. Solid tumours exhibit altered pH homeostasis with extracellular acidification. In human melanoma, the Na+/H+ exchanger NHE1 is an important modifier of the tumour nanoenvironment. Here we tested the modulation of cell-cell-adhesion by extracellular pH and NHE1. MV3 tumour spheroids embedded in a collagen matrix unravelled the efficacy of cell-cell contact loosening and 3D emigration into an environment mimicking physiological confinement. Adhesive interaction strength between individual MV3 cells was quantified using atomic force microscopy and validated by multicellular aggregation assays. Extracellular acidification from pHe7.4 to 6.4 decreases cell migration and invasion but increases single cell detachment from the spheroids. Acidification and NHE1 overexpression both reduce cell-cell adhesion strength, indicated by reduced maximum pulling forces and adhesion energies. Multicellular aggregation and spheroid formation are strongly impaired under acidification or NHE1 overexpression. We show a clear dependence of melanoma cell-cell adhesion on pHe and NHE1 as a modulator. These effects are opposite to cell-matrix interactions that are strengthened by protons extruded via NHE1. We conclude that these opposite effects of NHE1 act synergistically during the metastatic cascade. PMID:28205573

  15. Generation of Corneal Keratocytes from Human Embryonic Stem Cells. (United States)

    Hertsenberg, Andrew J; Funderburgh, James L


    Human Embryonic Stem Cells (hESC) offer an important resource as a limitless supply of any differentiated cell type of the human body. Keratocytes, cells from the corneal stroma, may have the potential for restoration of vision in cell therapy and biomedical engineering applications, but these specialized cells are not readily expanded in vitro. Here we describe a two-part method to produce keratocytes from the H1 hESC cell line. The hESC cells, maintained and expanded in feeder-free culture medium are first differentiated to neural crest cells using the stromal-derived inducing activity (SDIA) of the PA6 mouse embryonic fibroblast cell line. The resulting neural crest cells are selected by their expression of cell-surface CD271 and subsequently cultured as 3D pellets in a defined differentiation medium to induce a keratocyte phenotype.

  16. Derivation of multipotent mesenchymal precursors from human embryonic stem cells.

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    Full Text Available BACKGROUND: Human embryonic stem cells provide access to the earliest stages of human development and may serve as a source of specialized cells for regenerative medicine. Thus, it becomes crucial to develop protocols for the directed differentiation of embryonic stem cells into tissue-restricted precursors. METHODS AND FINDINGS: Here, we present culture conditions for the derivation of unlimited numbers of pure mesenchymal precursors from human embryonic stem cells and demonstrate multilineage differentiation into fat, cartilage, bone, and skeletal muscle cells. CONCLUSION: Our findings will help to elucidate the mechanism of mesoderm specification during embryonic stem cell differentiation and provide a platform to efficiently generate specialized human mesenchymal cell types for future clinical applications.

  17. Nucleosome Organization in Human Embryonic Stem Cells.

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    Puya G Yazdi

    Full Text Available The fundamental repeating unit of eukaryotic chromatin is the nucleosome. Besides being involved in packaging DNA, nucleosome organization plays an important role in transcriptional regulation and cellular identity. Currently, there is much debate about the major determinants of the nucleosome architecture of a genome and its significance with little being known about its role in stem cells. To address these questions, we performed ultra-deep sequencing of nucleosomal DNA in two human embryonic stem cell lines and integrated our data with numerous epigenomic maps. Our analyses have revealed that the genome is a determinant of nucleosome organization with transcriptionally inactive regions characterized by a "ground state" of nucleosome profiles driven by underlying DNA sequences. DNA sequence preferences are associated with heterogeneous chromatin organization around transcription start sites. Transcription, histone modifications, and DNA methylation alter this "ground state" by having distinct effects on both nucleosome positioning and occupancy. As the transcriptional rate increases, nucleosomes become better positioned. Exons transcribed and included in the final spliced mRNA have distinct nucleosome profiles in comparison to exons not included at exon-exon junctions. Genes marked by the active modification H3K4m3 are characterized by lower nucleosome occupancy before the transcription start site compared to genes marked by the inactive modification H3K27m3, while bivalent domains, genes associated with both marks, lie exactly in the middle. Combinatorial patterns of epigenetic marks (chromatin states are associated with unique nucleosome profiles. Nucleosome organization varies around transcription factor binding in enhancers versus promoters. DNA methylation is associated with increasing nucleosome occupancy and different types of methylations have distinct location preferences within the nucleosome core particle. Finally, computational

  18. Nucleosome Organization in Human Embryonic Stem Cells. (United States)

    Yazdi, Puya G; Pedersen, Brian A; Taylor, Jared F; Khattab, Omar S; Chen, Yu-Han; Chen, Yumay; Jacobsen, Steven E; Wang, Ping H


    The fundamental repeating unit of eukaryotic chromatin is the nucleosome. Besides being involved in packaging DNA, nucleosome organization plays an important role in transcriptional regulation and cellular identity. Currently, there is much debate about the major determinants of the nucleosome architecture of a genome and its significance with little being known about its role in stem cells. To address these questions, we performed ultra-deep sequencing of nucleosomal DNA in two human embryonic stem cell lines and integrated our data with numerous epigenomic maps. Our analyses have revealed that the genome is a determinant of nucleosome organization with transcriptionally inactive regions characterized by a "ground state" of nucleosome profiles driven by underlying DNA sequences. DNA sequence preferences are associated with heterogeneous chromatin organization around transcription start sites. Transcription, histone modifications, and DNA methylation alter this "ground state" by having distinct effects on both nucleosome positioning and occupancy. As the transcriptional rate increases, nucleosomes become better positioned. Exons transcribed and included in the final spliced mRNA have distinct nucleosome profiles in comparison to exons not included at exon-exon junctions. Genes marked by the active modification H3K4m3 are characterized by lower nucleosome occupancy before the transcription start site compared to genes marked by the inactive modification H3K27m3, while bivalent domains, genes associated with both marks, lie exactly in the middle. Combinatorial patterns of epigenetic marks (chromatin states) are associated with unique nucleosome profiles. Nucleosome organization varies around transcription factor binding in enhancers versus promoters. DNA methylation is associated with increasing nucleosome occupancy and different types of methylations have distinct location preferences within the nucleosome core particle. Finally, computational analysis of nucleosome

  19. Immunosurveillance function of human mast cell?

    Institute of Scientific and Technical Information of China (English)

    (O)ner (O)zdemir


    Mast cell (MC) is so widely recognized as a critical effector in allergic disorders that it can be difficult to think of MC in any other context. Indeed, MCs are multifunctional and recently shown that MCs can also act as antigen presenters as well as effector elements of human immune system. First observations of their possible role as anti-tumor cells in peri- or intra-tumoral tissue were mentioned five decades ago and a high content of MCs is considered as a favorable prognosis,consistent with this study. Believers of this hypothesis assumed them to be inhibitors of tumor development through their pro-apoptotic and -necrolytic granules e.g.,granzymes and TNF-α. However, some still postulate them to be enhancers of tumor development through their effects on angiogenesis due to mostly tryptase.There are also some data suggesting increased MC density causes tumor development and indicates bad prognosis. Furthermore, since MC-associated mediators have shown to influence various aspects of tumor biology, the net effect of MCs on the development/progression of tumors has been difficult to evaluate. For instance, chymase induces apoptosis in targets; yet,tryptase, another MC protease, is a well-known mitogen.MCs with these various enzyme expression patterns may mediate different functions and the predominant MC type in tissues may be determined by the environmental needs. The coexistence of tryptase-expressing MCs(MCT) and chymase and tryptase-expressing MCs (MCTC)in physiological conditions reflects a naturally occurring balance that contributes to tissue homeostasis. We have recently discussed the role and relevance of MC serine proteases in different bone marrow diseases.

  20. Biological characteristics of cell lines of human dental alveolus

    Institute of Scientific and Technical Information of China (English)

    陈世璋; 黄靖香; 孙明学; 赵斌


    Objective To investigate the biological characteristics of cell lines of healthy and diseased human dental alveoli. Methods Primary cell lines from either healthy or diseased human dental alveoli were obtained. Two cell lines, H-258 and H-171 derived from healthy and diseased human tissues respectively, were selected for morphological study and research on their growth and aging, using cell counting, and histochemical and immunohistochemical staining. Results Primary cell lines were successfully established from innormal dental alveoli. After freezing and thawing for three times, cell growth was continued and no morphological alterations were observed. The doubling time was 53.4 hours and mean division index (MDI) was 4‰. Cells were kept normal after twenty generations with no obvious reduction of doubling time and MDI. Of twenty-six primary cell lines derived from healthy human dental alveoli, only three cell lines achieved generation. After freezing and thawing for twice, cultured cells were still alive at a decreased growth speed, with doubling time of 85.9 hours and MDI of 3‰. Both cell lines, H-171 and H-258, shared the characteristics of osteoblast. Conclusions Primary cell lines of diseased human dental alveoli show greater growth potential. All cell lines of dental alveoli share characteristics of osteoblast. The technique we developed may be put into practice for the treatment of abnormal dental alveoli.

  1. Cryopreservation of human embryonic stem cells by vitrification

    Institute of Scientific and Technical Information of China (English)

    周灿权; 麦庆云; 李涛; 庄广伦


    Background The efficiency of traditional cryopreservation of human embryonic stem (ES) cells is low, and there have been few attempts to prove new cryopreservation methods effective. This study was designed to evaluate the efficiency of cryopreservation of human ES cells using vitrification method.Methods Human ES cells clumped from an identical cell line were randomly allocated to be cryopreserved by vitrification or by slow freezing. The recovery rates, the growth and differentiation potential of thawed human ES cells were compared between these two groups. The pluripotency of human ES cells after thawing was identified.Results Eighty-one point nine percent (59/72) of human ES cell clumps were recovered after vitrification, while only 22.8% (16/70) were recovered after slow freezing (P<0.01). The colonies after vitrification manifested have not only faster growth but also a lower level of differentiation when compared to colonies subjected to the slow freezing protocol. However, the rates of growth and differentiation in undifferentiated colonies from both groups were identical to the rates in those of non-cryopreserved stem cells after a prolonged culture period. Passage 6 of vitrified human ES cells retained the properties of pluripotent cells, a normal karyotype and expressed the transcription factor OCT-4, stage specific expressed antigen-4 (SSEA-4) and SSEA-3. Teratoma growth of these cells demonstrated the ability to develop into all three germ layers.Conclusions Vitrification is effective in cryopreserving human ES cells. During a prolonged culture, human ES cells retain their pluripotency after cryopreservation.

  2. Identification of skin immune cells in non-human primates. (United States)

    Adam, Lucille; Rosenbaum, Pierre; Cosma, Antonio; Le Grand, Roger; Martinon, Frédéric


    The skin is a valuable target for vaccine delivery because it contains many immune cell populations, notably antigen presenting cells. Skin immune cells have been extensively described in mice and humans but not in non-human primates, which are pertinent models for immunological research in vaccination. The aim of this work was to describe immune cell populations in the epidermis, dermis and skin draining lymph nodes in cynomolgus macaques by a single 12-parameter flow cytometry protocol. Given that skin cells share several markers, we defined a gating strategy to identify accurately immune cells and to limit contamination of one immune cell population by another. The epidermis contained CD1a(+)CD1c(-) Langerhans cells (LCs), CD3(+) T cells and putative NK cells. The dermis contained CD1a(+)CD1c(-) cells, which were similar to LCs, CD1a(+)CD1c(+) dermal dendritic cells (DDCs), CD163(high)CD11b(+) resident macrophages, CD3(+) T cells and putative NK cells. The skin also contained CD66(+) polymorphonuclear cells in some animals. Thus, immune cell populations in the macaque are similar to those in humans despite some differences in phenotype. In skin draining lymph nodes, we identified migratory LCs, CD1a(+)CD1c(+) DDCs and macrophages. The simultaneous identification of these different immune cells with one panel of markers avoids the use of large amounts of precious sample and may improve the understanding of immune mechanisms in the skin after treatment or vaccination.

  3. Beclin 1 deficiency correlated with lymph node metastasis, predicts a distinct outcome in intrahepatic and extrahepatic cholangiocarcinoma.

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    Tian-Tian Wang

    Full Text Available Autophagy can be tumor suppressive as well as promotive in regulation of tumorigenesis and disease progression. Accordingly, the prognostic significance of autophagy key regulator Beclin 1 was varied among different tumors. Here, we detected the clinicopathological and prognostic effect of Beclin 1 in the subtypes of intrahepatic cholangiocarcinoma (ICC and extrahepatic cholangiocarcinoma (ECC. Beclin 1 expression level was detected by immunohistochemistry staining in 106 ICC and 74 ECC patients. We found that Beclin 1 was lowly expressed in 126 (70% cholangiocarcinoma patients, consist of 72 ICC and 54 ECC. Moreover, the cholangiocarcinoma patients with lymph node metastasis (N1 had a lower Beclin 1 level than that of N0 subgroup (P=0.012. However, we did not detect any correlations between Beclin 1 and other clinicopathological features, including tumor subtypes, vascular invasion, HBV infection, liver cirrhosis, cholecystolithiasis and TNM stage. Survival analysis showed that, compared with the high expression subset, Beclin 1 low expression was correlated with a poorer 3-year progression-free survival (PFS, 69.1% VS 46.8%, P=041 for cholangiocarcinoma. Importantly, our stratified univariate and multivariate analysis confirmed that Beclin 1 lowly expressed ICC had an inferior PFS as well as overall survival than ECC, particularly than that of Beclin 1 highly expressed ECC patients. Thus, our study demonstrated that Beclin 1low expression, correlated with lymph node metastasis, and might be a negative prognostic biomarker for cholangiocarcinoma. Combined Beclin 1 level with the anatomical location might lead to refined prognosis for the subtypes of ICC and ECC.

  4. [In vitro strategies for human gametes production from stem cells]. (United States)

    Tosca, Lucie; Courtot, Anne-Marie; Bennaceur-Griscelli, Annelise; Tachdjian, Gérard


    Embryonic stem cells (ESC) are self-renewal and pluripotent cells that are able to differentiate in vitro into several cell types in favourable conditions. Technical protocols for in vitro gametes production have been developed in mice and human species. The functionality of such differentiated cells is not always analysed and an early meiotic arrest is a current observation. These kinds of experimentations have also been tested from human induced pluripotent stem cells (IPSC). However, differentiation ends shortly at the primordial germ cell stage.

  5. Growth in agarose of human cells infected with cytomegalovirus. (United States)

    Lang, D J; Montagnier, L; Latarjet, R


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

  6. Cytotoxinic Mechanism of Hydroxyapatite Nanoparticles on Human Hepatoma Cell Lines

    Institute of Scientific and Technical Information of China (English)

    CAO Xian-ying; QI Zhi-tao; DAI Hong-lian; YAN Yu-hua; LI Shi-pu


    Stable and single-dispersed HAP nanoparticles were synthesized with chemical method assisted by ultrasonic treatment.HAP nanoparticles were surveyed by AFM and Zataplus.The effect on the Bel-7402 human hepatoma cell lines treated with HAP nanoparticles was investigated by the MTT methods and observation of morphology,and the mechanism was studied in changes of cell cycle and ultrastructure.The result shows that inhibition of HAP nanoparticles on the Bel-7402 human hepatoma cell lines is obviously in vitro.HAP nanoparticles the entered cancer cytoplasm,and cell proliferation is stopped at G1 phase of cell cycle,thus,cancer cells die directly.

  7. Canthin-6-one induces cell death, cell cycle arrest and differentiation in human myeloid leukemia cells. (United States)

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


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

  8. Human organomics: a fresh approach to understanding human development using single-cell transcriptomics. (United States)

    Camp, J Gray; Treutlein, Barbara


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

  9. Human dental pulp stem cells: Applications in future regenerative medicine. (United States)

    Potdar, Pravin D; Jethmalani, Yogita D


    Stem cells are pluripotent cells, having a property of differentiating into various types of cells of human body. Several studies have developed mesenchymal stem cells (MSCs) from various human tissues, peripheral blood and body fluids. These cells are then characterized by cellular and molecular markers to understand their specific phenotypes. Dental pulp stem cells (DPSCs) are having a MSCs phenotype and they are differentiated into neuron, cardiomyocytes, chondrocytes, osteoblasts, liver cells and β cells of islet of pancreas. Thus, DPSCs have shown great potentiality to use in regenerative medicine for treatment of various human diseases including dental related problems. These cells can also be developed into induced pluripotent stem cells by incorporation of pluripotency markers and use for regenerative therapies of various diseases. The DPSCs are derived from various dental tissues such as human exfoliated deciduous teeth, apical papilla, periodontal ligament and dental follicle tissue. This review will overview the information about isolation, cellular and molecular characterization and differentiation of DPSCs into various types of human cells and thus these cells have important applications in regenerative therapies for various diseases. This review will be most useful for postgraduate dental students as well as scientists working in the field of oral pathology and oral medicine.

  10. Role of human mast cells and basophils in bronchial asthma. (United States)

    Marone, Gianni; Triggiani, Massimo; Genovese, Arturo; De Paulis, Amato


    Mast cells and basophils are the only cells expressing the tetrameric (alphabetagamma2) structure of the high affinity receptor for IgE (FcepsilonRI) and synthesizing histamine in humans. Human FcepsilonRI+ cells are conventionally considered primary effector cells of bronchial asthma. There is now compelling evidence that these cells differ immunologically, biochemically, and pharmacologically, which suggests that they might play distinct roles in the appearance and fluctuation of the asthma phenotype. Recent data have revealed the complexity of the involvement of human mast cells and basophils in asthma and have shed light on the control of recruitment and activation of these cells in different lung compartments. Preliminary evidence suggests that these cells might not always be detrimental in asthma but, under some circumstances, they might exert a protective effect by modulating certain aspects of innate and acquired immunity and allergic inflammation.

  11. Identification of human fibroblast cell lines as a feeder layer for human corneal epithelial regeneration.

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    Rong Lu

    Full Text Available There is a great interest in using epithelium generated in vitro for tissue bioengineering. Mouse 3T3 fibroblasts have been used as a feeder layer to cultivate human epithelia including corneal epithelial cells for more than 3 decades. To avoid the use of xeno-components, we evaluated human fibroblasts as an alternative feeder supporting human corneal epithelial regeneration. Five human fibroblast cell lines were used for evaluation with mouse 3T3 fibroblasts as a control. Human epithelial cells isolated from fresh corneal limbal tissue were seeded on these feeders. Colony forming efficiency (CFE and cell growth capacity were evaluated on days 5-14. The phenotype of the regenerated epithelia was evaluated by morphology and immunostaining with epithelial markers. cDNA microarray was used to analyze the gene expression profile of the supportive human fibroblasts. Among 5 strains of human fibroblasts evaluated, two newborn foreskin fibroblast cell lines, Hs68 and CCD1112Sk, were identified to strongly support human corneal epithelial growth. Tested for 10 passages, these fibroblasts continually showed a comparative efficiency to the 3T3 feeder layer for CFE and growth capacity of human corneal epithelial cells. Limbal epithelial cells seeded at 1 × 10(4 in a 35-mm dish (9.6 cm(2 grew to confluence (about 1.87-2.41 × 10(6 cells in 12-14 days, representing 187-241 fold expansion with over 7-8 doublings on these human feeders. The regenerated epithelia expressed K3, K12, connexin 43, p63, EGFR and integrin β1, resembling the phenotype of human corneal epithelium. DNA microarray revealed 3 up-regulated and 10 down-regulated genes, which may be involved in the functions of human fibroblast feeders. These findings demonstrate that commercial human fibroblast cell lines support human corneal epithelial regeneration, and have potential use in tissue bioengineering for corneal reconstruction.

  12. Recurrent Amplification at 13q34 Targets at CUL4A, IRS2, and TFDP1 As an Independent Adverse Prognosticator in Intrahepatic Cholangiocarcinoma.

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    Ting-Ting Liu

    Full Text Available Amplification of genes at 13q34 has been reported to be associated with tumor proliferation and progression in diverse types of cancers. However, its role in intrahepatic cholangiocarcinoma (iCCA has yet to be explored. We examined two iCCA cell lines and 86 cases of intrahepatic cholangiocarcinoma to analyze copy number of three target genes, including cullin 4A (CUL4A, insulin receptor substrate 2 (IRS2, and transcription factor Dp-1 (TFDP1 at 13q34 by quantitative real-time polymerase chain reaction. The cell lines and all tumor samples were used to test the relationship between copy number (CN alterations and protein expression by western blotting and immunohistochemical assays, respectively. IRS2 was introduced, and each target gene was silenced in cell lines. The mobility potential of cells was compared in the basal condition and after manipulation using cell migration and invasion assays. CN alterations correlated with protein expression levels. The SNU1079 cell line containing deletions of the target genes demonstrated decreased protein expression levels and significantly lower numbers of migratory and invasive cells, as opposed to the RBE cell line, which does not contain CN alterations. Overexpression of IRS2 by introducing IRS2 in SUN1079 cells increased the mobility potential. In contrast, silencing each target gene showed a trend or statistical significance toward inhibition of migratory and invasive capacities in RBE cells. In tumor samples, the amplification of each of these genes was associated with poor disease-free survival. Twelve cases (13.9% demonstrated copy numbers > 4 for all three genes tested (CUL4A, IRS2, and TFDP1, and showed a significant difference in disease-free survival by both univariate and multivariate survival analyses (hazard ratio, 2.69; 95% confidence interval, 1.23 to 5.88; P = 0.013. Our data demonstrate that amplification of genes at 13q34 plays an oncogenic role in iCCA featuring adverse disease

  13. Genetic engineering of human pluripotent cells using TALE nucleases. (United States)

    Hockemeyer, Dirk; Wang, Haoyi; Kiani, Samira; Lai, Christine S; Gao, Qing; Cassady, John P; Cost, Gregory J; Zhang, Lei; Santiago, Yolanda; Miller, Jeffrey C; Zeitler, Bryan; Cherone, Jennifer M; Meng, Xiangdong; Hinkley, Sarah J; Rebar, Edward J; Gregory, Philip D; Urnov, Fyodor D; Jaenisch, Rudolf


    Targeted genetic engineering of human pluripotent cells is a prerequisite for exploiting their full potential. Such genetic manipulations can be achieved using site-specific nucleases. Here we engineered transcription activator-like effector nucleases (TALENs) for five distinct genomic loci. At all loci tested we obtained human embryonic stem cell (ESC) and induced pluripotent stem cell (iPSC) clones carrying transgenic cassettes solely at the TALEN-specified location. Our data suggest that TALENs employing the specific architectures described here mediate site-specific genome modification in human pluripotent cells with similar efficiency and precision as do zinc-finger nucleases (ZFNs).

  14. Purification and cultivation of human pituitary growth hormone secreting cells (United States)

    Hymer, W. C.


    The maintainance of actively secreting human pituitary growth hormone cells (somatotrophs) in vitro was studied. The primary approach was the testing of agents which may be expected to increase the release of the human growth hormone (hGH). A procedure for tissue procurement is described along with the methodologies used to dissociate human pituitary tissue (obtained either at autopsy or surgery) into single cell suspensions. The validity of the Biogel cell column perfusion system for studying the dynamics of GH release was developed and documented using a rat pituitary cell system.

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

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    Zhumur Ghosh

    Full Text Available Human induced pluripotent stem cells (hiPSCs generated by de-differentiation of adult somatic cells offer potential solutions for the ethical issues surrounding human embryonic stem cells (hESCs, as well