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Sample records for cells displaying p-stat-3

  1. Fucoidan induces apoptosis of HepG2 cells by down-regulating p-Stat3.

    Science.gov (United States)

    Roshan, Sadia; Liu, Yun-yi; Banafa, Amal; Chen, Hui-jie; Li, Ke-xiu; Yang, Guang-xiao; He, Guang-yuan; Chen, Ming-jie

    2014-06-01

    Fucoidan is one of the main bioactive components of polysaccharides. The current study was focused on the anti-tumor effects of fucoidan on human heptoma cell line HepG2 and the possible mechanisms. Fucoidan treatment resulted in cell cycle arrest and apoptosis of HepG2 cells in a dose-dependent manner detected by MTT assay, flow cytometry and fluorescent microscopy. The results of flow cytometric analysis revealed that fucoidan induced G2/M arrest in the cell cycle progression. Hoechst 33258 and Annexin V/PI staining results showed that the apoptotic cell number was increased, which was associated with a dose-dependent up-regulation of Bax and down-regulation of Bcl-2 and p-Stat3. In parallel, the up-regulation of p53 and the increase in reactive oxygen species were also observed, which may play important roles in the inhibition of HepG2 growth by fucoidan. In the meantime, Cyclin B1 and CDK1 were down-regulated by fucoidan treatment. Down-regulation of p-Stat3 by fucoidan resulted in apoptosis and an increase in ROS in response to fucoidan exposure. We therefore concluded that fucoidan induces apoptosis through the down-regulation of p-Stat3. These results suggest that fucoidan may be used as a novel anti-cancer agent for hepatocarcinoma.

  2. 非小细胞肺癌中p-STAT3与Ki67的表达及意义%The expression and significance of phosphorylated signal transducer and activator of transcription 3 and Ki67 in non-small cell lung cancer

    Institute of Scientific and Technical Information of China (English)

    刘培杰

    2010-01-01

    Objective To study the expressions of phosphorylated signal transducer and activator of transcription 3(p-STAT3)and Ki67 in non-small cell lung cancer and their significance in tumor development and progression. Methods The expression of p-STAT3 and Ki67 were detected in 67 lung carcinoma tissues and 41 normal lung tissues by immun histochemical method. Results The positive rate of p-STAT3 in non-small cell lung cancer (67.16%,45/67) was significantly higher than that in normal lung tissue(17.07%,7/41). The positive rate of Ki67 in non-small cell lung cancer (76.12%,51/67) was significantly higher than that in normal lung tissue(7.31%,3/41).The expression of p-STAT3 and Ki67 were associated with clinical stages, lymph node transferation and histological grades(P<0.05). The expression of p-STAT3 and Ki67 were not associated with ages, sex, tumor location or size.The expression of p-STAT3 was positively correlated with Ki67 expression. Conclusions Abnormal activation of p-STAT3, can lead to excessive proliferation of tumor cells and Ki67 is a favorable indicator of proliferation of lung tumor cells, and the combined detection of p-STAT3 and Ki67 may be valuable for diagnosis and treatment of non-small cell lung cancer.%目的 研究磷酸化信号转导与转录活化因子3(p-STAT3)和 Ki67在非小细胞肺癌(NSCLC)发生、发展中的作用及相互关系.方法 采用免疫组织化学S-P法检测67例非小细胞肺癌组织和41例正常肺组织中p-STAT3与Ki67的表达.结果 非小细胞肺癌组织中p-STAT3的阳性表达率(67.16%,45/67)显著高于正常肺组织(17.07%,7/41),Ki67在非小细胞肺癌组织中的表达(76.12%,51/67)高于正常肺组织(7.31%,3/41).p-STAT3和 Ki67的表达与临床分期、有无淋巴转移和分化程度相关(P均<0.05),与患者年龄、性别、肿瘤位置及大小无关.p-STAT3的表达与Ki67表达呈正相关.结论 p-STAT3的异常活化可促进恶性肿瘤细胞的过度增殖,Ki67较好的反应了肺癌

  3. Crizotinib (PF-2341066) induces apoptosis due to downregulation of pSTAT3 and BCL-2 family proteins in NPM-ALK(+) anaplastic large cell lymphoma.

    Science.gov (United States)

    Hamedani, Farid Saei; Cinar, Munevver; Mo, Zhicheng; Cervania, Melissa A; Amin, Hesham M; Alkan, Serhan

    2014-04-01

    Nucleophosmin-anaplastic lymphoma kinase (NPM-ALK) is an aberrant fusion gene product with tyrosine kinase activity and is expressed in substantial subset of anaplastic large cell lymphomas (ALCL). It has been shown that NPM-ALK binds to and activates signal transducer and activator of transcription 3 (STAT3). Although NPM-ALK(+) ALCL overall shows a better prognosis, there is a sub-group of patients who relapses and is resistant to conventional chemotherapeutic regimens. NPM-ALK is a potential target for small molecule kinase inhibitors. Crizotinib (PF-2341066) is a small, orally bioavailable molecule that inhibits growth of tumors with ALK activity as shown in a subgroup of non-small lung cancer patients with EML4-ALK expression. In this study, we have investigated the in vitro effects of Crizotinib in ALCL cell line with NPM-ALK fusion. Crizotinib induced marked downregulation of STAT3 phosphorylation, which was associated with significant apoptotic cell death. Apoptosis induction was attributed to caspase-3 cleavage and marked downregulation of the Bcl-2 family of proteins including MCL-1. These findings implicate that Crizotinib has excellent potential to treat patients with NPM-ALK(+) ALCL through induction of apoptotic cell death and downregulation of major oncogenic proteins in this aggressive lymphoma.

  4. 白细胞介素-1β对大鼠视网膜Müller细胞磷酸化的信号转导及转录活化因子3表达的影响%Interleukin-1β promotes the expression of pSTAT3 in rat retinal Müller cells

    Institute of Scientific and Technical Information of China (English)

    沈玺; 徐格致

    2009-01-01

    目的 观察不同浓度白细胞介素-1β(IL-1β)对大鼠视网膜Müller细胞磷酸化的信号转导及转录活化因子3(pSTAT3)表达的影响.方法 将体外培养的Sprague-Dawley(SD)大鼠Müller细胞分别以0.0、0.1、1.0、5.0、10.0 ng/ml浓度的IL-1β刺激24 h后,免疫荧光法和蛋白质印迹(Western botting)检测细胞内pSTAT3的表达改变.SD大鼠32只,随机分为对照组、100、500、1000 ng/ml组,每组均为8只大鼠.分别将磷酸缓冲液(PBS)配制的浓度分别为100、500、1000 ng/ml的IL-1β注入大鼠的左眼玻璃体腔中,对照组大鼠左眼玻璃体腔注射相同容积的PBS,24 h后采用同样方法 检测pSTAT3在视网膜的表达情况.结果 免疫荧光法和Western botting检测结果 显示,IL-1β刺激24 h后,IL-1β浓度为0.0 ng/ml时,pSTAT3在细胞核内有极少量表达,当浓度达到1.0 ng/ml后,pSTAT3在细胞核内的表达随浓度增加而升高,差异有统计学意义(F=46.64,43.78;P<0.01).对照组大鼠视网膜内无明显pSTAT3的表达,当浓度达到100.0 ng/ml后,pSTAT3在视网膜内的表达随浓度增加而升高,差异有统计学意义(F=73.53,43.70;P<0.01);pSTAT3主要表达于内核层和神经节细胞层.双荧光标记显示,对照组大鼠视网膜内无pSTAT3表达,胶质纤维酸性蛋白(GFAP)主要表达于神经节细胞层,并向视网膜色素上皮层呈丝状放射;从100 ng/ml IL-1β刺激24 h后起,内核层就出现颗粒状GFAP和pSTAT3的双标染色.结论 IL-1β可以上调Müller细胞pSTAT3的表达,这可能是Müller细胞发生反应性胶质活化的通路之一.%Objective To observe the influence of interleukin-1β (IL-1β) on the expression of phosphorylated signal transducers and activators of transcription 3 (pSTAT 3) in rat retinal MOiler cells. Methods For in vitro study cultured Mailer cells were treated with IL-1β of different concentrations (0, 0.1, 1, 5 and 10 ng/ml) for 24 hours. For in vivo study, 32 Sprague-Dawley(SD)rats were

  5. Tumor suppressor PRSS8 targets Sphk1/S1P/Stat3/Akt signaling in colorectal cancer

    Science.gov (United States)

    Wang, Qian; Li, Zexin; Yang, Yiqiong; Chen, Zhiguo; Wang, Jianguo; Zhao, Weixing; Zhang, Huijuan; Chen, Jiwang; Dong, Huali; Shen, Kui; Diamond, Alan M.; Yang, Wancai

    2016-01-01

    PRSS8 is a membrane-anchored serine protease prostasin and has been shown an association with carcinogenesis. Herein we found that PRSS8 expression was significantly reduced in colorectal adenomas and adenocarcinomas. The decreased PRSS8 was well correlated with clinical stages, poor differentiation and shorter survival time of colorectal cancer. Furthermore, increase of PRSS8 led to the inhibition of colorectal cancer cell proliferation, knockdown of PRSS8 accelerated cell proliferation in vitro, and overexpressing PRSS8 retarded cancer cell growth in nude mice. Mechanistic studies revealed that PRSS8 inhibited Sphk1/S1P/Stat3/Akt signaling pathway, in terms of inverse association between PRSS8 and Sphk1 in human colorectal cancers and in Sphk1-/− mice. In conclusion, PRSS8 acts as a tumor suppressor by inhibiting Sphk1/S1P/Stat3/Akt signaling pathway, and could be used as a biomarker to monitor colorectal carcinogenesis and predict outcomes. PMID:27050145

  6. pSTAT1, pSTAT3, and T-bet as markers of disease activity in chronic inflammatory demyelinating polyradiculoneuropathy.

    Science.gov (United States)

    Madia, Francesca; Frisullo, Giovanni; Nociti, Viviana; Conte, Amelia; Luigetti, Marco; Del Grande, Alessandra; Patanella, Agata Katia; Iorio, Raffaele; Tonali, Pietro Attilio; Batocchi, Anna Paola; Sabatelli, Mario

    2009-06-01

    Chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is considered an auto-immune disorder. We evaluated expression of pSTAT1, T-bet, and pSTAT3 in circulating T-cells, B-cells, and monocytes and spontaneous production of interleukin-17 (IL17), interferon-gamma (IFN gamma), and interleukin-10 (IL10) by peripheral blood mononuclear cells (PBMCs) from 14 active CIDP patients compared with 6 patients with long-lasting remission and 20 controls. Active disease patients showed higher pSTAT1, T-bet, and pSTAT3 in CD4(+) T-cells than controls (p CIDP patients than controls (p = 0.0011, p = 0.0041, p = 0.0413, respectively) and remission patients (p = 0.0073, p = 0.0274, p = 0.0251, respectively). Moreover in CD8(+) T-cells, pSTAT3 expression was higher in active CIDP patients than in remission patients (p = 0.0345) and in controls (p = 0.0023). IL17 and IFN gamma production were significantly higher in active CIDP patients than in controls (p CIDP patients (p = 0.0073). IL10 levels were higher in active phase patients than in controls (p = 0.0334). Our data suggest that pSTAT1, T-bet, and pSTAT3 can be considered putative markers of disease activity and potential targets for specific therapies.

  7. Significance and relationship between Cripto-1 and p-STAT3 expression in gastric cancer and precancerous lesions

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    AIM:To explore the relationship between Cripto-1 (CR-1) and tyrosine phosphorylation STAT3 (p-STAT3) expressions in gastric cancer (GC) and gastric carcinogensis and metastasis.METHODS: The PV9000 immunohistochemical method was used to detect the expression of CR-1 and p-STAT3 in 178 cases of GC, 95 matched normal gastric mucosa, 40 chronic atrophic gastritis (CAG), 48 intestinal meta-plasia (IM) and 25 dysplasia (DYS). RESULTS: The positive rates of CR-1 and p-STAT3 expression were significantly higher in ...

  8. The Role of p-STAT3 as a Prognostic and Clinicopathological Marker in Colorectal Cancer: A Systematic Review and Meta-Analysis

    Science.gov (United States)

    Chu, Qi; Gan, Yong; Ren, Hui; Zhang, Liyan; Wang, Liwei; Li, Xiaoxiu; Wang, Wei

    2016-01-01

    Objective High expression of phosphorylated signal transducer and activator of transcription 3 (p-STAT3) has been detected in a variety of human tumors. However, the association of positive p-STAT3 expression with clinicopathological parameters and the prognosis of colorectal cancer patients remain controversial. To identify the relationship between p-STAT3 expression and clinicopathological parameters and prognosis in patients with colorectal cancer, a systematic review and meta-analysis were performed. Methods We performed a comprehensive literature search from PubMed, EMBASE, and SinoMed through 27 March, 2016. Hazard ratios (HRs) with 95% confidence intervals (CI) were combined to evaluate the association between p-STAT3 expression and overall survival of colorectal cancer patients. Odds ratios (ORs) with 95% CI were combined to evaluate the association between p-STAT3 expression and clinicopathological parameters in patients with colorectal cancer. Results Seventeen studies including a total of 2,346 colorectal cancer patients were included in this meta-analysis. The combined HR was 1.43 (95% CI: 1.23–1.67, P < 0.001), which suggested a positive relationship between p-STAT3 overexpression and poorer overall survival of colorectal cancer patients. In addition, the results indicated that positive p-STAT3 expression was significantly associated with the presence of lymph node metastasis (OR: 2.43, 95% CI: 1.18–5.01, P = 0.02) but was not associated with TNM stage, tumor differentiation or gender. Conclusion The meta-analysis results suggest that p-STAT3 overexpression is unfavorable for the prognosis of colorectal cancer patients, and p-STAT3 overexpression is associated with the presence of lymph node metastasis among colorectal cancer patients. PMID:27504822

  9. Dynamic Balance of pSTAT1 and pSTAT3 in C57BL/6 Mice Infected with Lethal or Nonlethal Plasmodium yoelii

    Institute of Scientific and Technical Information of China (English)

    Xibao Shi; Li Qin; Guangjie Liu; Siting Zhao; Nanzheng Peng; Xiaoping Chen

    2008-01-01

    Signal transducer and activator of transcription(STAT)proteins play an important role in cytokine signaling pathways and regulation of immune responses.The balance of the phosphorylated(activated)STAT1(pSTAT1) and STAT3 (pSTAT3)has been documented in cancer immunology.In this study,we investigated the dynamic balance of pSTAT1 and pSTAT3 in C57BL/6 mice infected with either a nonlethal(py17XNL)or lethal(ey17XL) strain of Plasmodium yoelii.Both pylNL and ey17XL infections induced a maximum activation of STAT1 and STAT3 On the first day after parasite inoculation.Additionally,the py17XNL infection induced a pSTAT1- dominant response in mice during the early stage of infection,with the resolution of parasitemia.In contrast, Py17XL infection induced a pSTAT3-dominant response during the early phase of infection,with the death of the animals.Our results indicated that maximum activation of STAT1 and STAT3 occurred much earlier than the peak levels of cytokines induced by Plasmodium yoelii infection based on previous reports and that infection with Py17XNL-and py17XL induced different dynamic patterns of pSTAT1 and pSTAT3 balance.

  10. 磷酸化STAT3检测对肺腺癌所致恶性胸腔积液的诊断价值%The significance of pSTAT3 test in the diagnosis of malignant pleural effusion induced by adenocarcinoma of lung

    Institute of Scientific and Technical Information of China (English)

    田伊卿; 朱立强; 方先勇

    2015-01-01

    Objective To investigate the diagnostic value and clinical significance of phosphorylated signal transducer and activa-tor of transcription 3(pSTAT3)in malignant pleural effusion(MPE)caused by lung adenocarcinoma.Methods 47 pleural effusion samples were collected and detected by using liquid-based cytologic test (LCT).Furthermore,the expression of pSTAT3 was detec-ted by using the method of immunocytochemistry in all of the specimens.Results Among the 47 cases of pleural effusion,40 cases of MPE were caused by lung adenocarcinoma and 7 cases were benign pleural effusion,which were confirmed by clinical manifesta-tion,biopsy and imaging data.The positive expression rate of pSTAT3 was 80.0%(32/40)in MPE caused by lung adenocarcino-ma,significantly higher than benign group(P <0.05).13 cases of MPE caused by lung adenocarcinoma and 7 cases of benign pleural effusion were diagnosed with liquid-based cytologic test alone.The other 27 cases were undetermined and needed to be differentiate between adenocarcinoma and atypical mesothelial cells.However,32 cases of lung adenocarcinoma and 7 cases could be clearly diag-nosed if liquid-based cytologic test was used combined with immunocytochemistry detection of pSTAT3 expression.Only 8 cases were undetermined.Diagnosis of grey area was significantly narrowed.There was significant statistical significance between the two methods(P <0.05).Conclusion High expression of pSTAT3 is related to the MPE caused by lung adenocarcinoma.Detection of pSTAT3 expression can be used as a new method in the diagnosis of MPE caused by lung adenocarcinoma.pSTAT3 may play an important role in the development of the MPE.%目的:探讨磷酸化信号转导与转录激活因子3(pSTAT3)蛋白表达对肺腺癌所致恶性胸腔积液的诊断价值。方法应用液基细胞学技术,对临床送检的47例胸腔积液进行检测,同时每例均利用薄层细胞涂片行免疫细胞化学染色,检测 pSTAT3蛋白的表达。结果40例

  11. Expression of p-STAT3 and vascular endothelial growth factor in MNNG-induced precancerous lesions and gastric tumors in rats

    Institute of Scientific and Technical Information of China (English)

    Xiao-Yan Wang; Lou-Lei Wang; Xuan Zheng; Li-Na Meng; Bin Lyu; Hai-Feng Jin

    2016-01-01

    AIM: To investigate the dynamic expression of p-signal transducer and activator of transcription 3(STAT3) and vascular endothelial growth factor(VEGF) in the formation of gastric tumors induced by drinking water containing N-methyl-N’-nitro-N-nitrosoguanidine(MNNG) in Wistar rats.METHODS: One hundred and twenty Wistar rats were randomly divided into two groups(60 in each group): Control group and Model group. The rats in each group were then randomly divided into three groups(20 in each group): C/M15, C/M25 and C/M40(15, 25 and 40 represent the number of feeding weeks from termination). Rats in the control group received normal drinking water and rats in the model group received drinking water containing 100 μg/m L MNNG. Stomach tissues were collected at the end of the 15 th, 25 th and 40 th week, respectively, for microscopic measurement using hematoxylin and eosin staining. The expression of p-STAT3 and VEGF in different pathological types of gastric tissue, including normal, inflammation, atrophy, hyperplasia and gastric stromal tumor, was observed by immunohistochemistry and Western blot, and the corelation between p-STAT3 and VEGF was analyzed. RESULTS:(1) The expression of p-STAT3 in tissue with gastritis, atrophy, dysplasia and gastric stromal tumor were significantly increased in the model group compared with the control group(2.5 ± 1.0, 2.75 ±0.36, 6.2 ± 0.45, 5.67 ± 0.55 vs 0.75 ± 0.36, P = 0.026, 0.035, 0.001, 0.002, respectively); the expression of p-STAT3 in tissue with dysplasia was higher than that in samples with gastritis or atrophy(6.2 ± 0.45 vs 2.5 ± 1.0, P = 0.006; 6.2 ± 0.45 vs 2.75 ± 0.36, P = 0.005, respectively); however, the expression of p-STAT3 in gastritis and atrophy was not significantly different(P > 0.05);(2) the expression of VEGF in tissue with gastritis, atrophy, dysplasia and gastric stromal tumor was significantly increased in the model group compared with normal gastric mucosa; and the expression of VEGF in tissue

  12. STAT3与P-STAT3在转基因AD小鼠脑组织中的表达及意义%Expression of STAT3 and P-STAT3 in the brain of a transgenic mouse model of Alzheimer's disease

    Institute of Scientific and Technical Information of China (English)

    田密; 侯德仁; 邓炎尧; 李维; 奉夏露

    2013-01-01

    目的:通过检测STAT3及P-STAT3在APPswe/PS△E9双转基因阿尔茨海默病(AD)小鼠脑组织中的表达,探讨其在AD发病过程中的可能作用。方法采用免疫组化法检测APPswe/PS△E9双转基因AD小鼠及对照小鼠脑组织中STAT3和P-STAT3的表达。结果STAT3和P-STAT3表达于脑组织的不同部位。STAT3在转基因AD小鼠和对照小鼠的大脑皮层、基底前脑、海马及小脑中的阳性表达率分别为93.75%、87.50%,87.50%、43.75%,81.25%、37.50%,62.50%、0.00%,其中差异有统计学意义的是基底前脑、海马及小脑中的表达(P<0.05);P-STAT3在转基因AD小鼠和对照小鼠的大脑皮层、基底前脑、海马及小脑中的阳性表达率分别为0.00%、0.00%,68.75%、0.00%,62.50%、12.50%,43.75%、0.00%,其中差异有统计学意义的是基底前脑、海马及小脑中的表达(P<0.05);且STAT3和P-STAT3呈正相关(P<0.05)。结论STAT3和P-STAT3在转基因AD小鼠的基底前脑、海马及小脑中存在高表达,其可能参与了AD发病的病理过程。%Objective To detect the expression of signal transducer and activator of transcription 3 (STAT3) and P-STAT3 in the brain of the APPswe/PS△E9 double transgenic mouse model of Alzhaimer's disease (AD) and invesitgate their possible role in AD. Methods APPswe/PS △E9 double transgenic mice and control mice were examined for cerebral STAT3 and P-STAT3 expressions using immunothistochemistry. Results STAT3 and P-STAT3 were expressed in the different regions of mouse brain. In the transgenic mice and the control mice, the positivity rates of STAT3 were 93.75%and 87.50%in the cerebral cortex, 87.50% and 43.75% in the basal forebrain, 81.25% and 37.50% in the hippocampus, and 62.50% and 0.00% in the cerebellum, respectively, showing significant differences between the mice in the STAT3 expressions in the basal forebrain, hippocampus and cerebellum (P<0.05). The positivity rates of P-STAT3 in the

  13. STAT3,pSTAT3与TFF3在甲状腺乳头状癌中的临床意义%Clinical significance of STAT3, pSTAT3 and TFF3 in human papillary thyroid carcinoma

    Institute of Scientific and Technical Information of China (English)

    张晨蕾; 李燕萍; 吴靖芳; 万佳鑫; 陈亚敏; 张文静; 薛刚

    2016-01-01

    目的 探讨甲状腺乳头状癌(Papillary Thyroid Carcinomas,PTC) STAT3及其磷酸化水平与三叶因子3(Trefoil Factor 3,TFF3)表达及相关性.方法 采用免疫组织化学检测31例组织芯片中PTC及癌旁组织STAT3、pSTAT3和TFF3阳性率与平均光密度(AOD)值;Western Blot检测6例PTC和癌旁手术标本中相应蛋白的含量,并进行相关分析.结果 (1)免疫组化:TFF3蛋白在PTC中均高表达(P<0.001),TFF3阳性率与AOD值在直径>2 cm组、淋巴结转移组和临床Ⅲ~Ⅳ病例明显高于直径≤2 cm组、无淋巴结转移组和Ⅰ~Ⅱ期者(x2=12.120,P=0.001,x2=7.517,P=0.043,x2=10.490, P=0.003;AOD值:t=2.810, P=0.0064, t=2.234, P=0.039, t=2.926, P=0.0058);PTC中STAT3阳性率和AOD值与临床资料无关.pSTAT3为胞核阳性,阳性率和AOD值在PTC组织低于癌旁组织,直径>2 cm组、淋巴结转移组和临床Ⅲ~Ⅳ病例明显低于直径≤2 cm组、无淋巴结转移组和Ⅰ~Ⅱ期者(x2=4.963, P=0.046,x2=4.882, P=0.033;AOD值:t=2.195, P=0.041, t=2.790, P=0.0068, t=2.018, P=0.042);TFF3与STAT3呈正相关,与pSTAT3呈负相关(P<0.01). (2) Westem Blot: TFF3和STAT3蛋白在PTC的表达高于癌旁组织(t=2.841,P=0.008,t=2.925,P=0.007),但pSTAT3却明显低于癌旁组织(t=2.713,P=0.009).结论 甲状腺乳头状癌TFF3和STAT3高表达、pSTAT3低表达对PTC不良预后的预测有指导意义.

  14. Matrix-addressable electrochromic display cell

    Science.gov (United States)

    Beni, G.; Schiavone, L. M.

    1981-04-01

    We report an electrochromic display cell with intrinsic matrix addressability. The cell, based on a sputtered iridium oxide film (SIROF) and a tantalum-oxide hysteretic counterelectrode, has electrochromic parameters (i.e., response times, operating voltages, and contrast) similar to those of other SIROF display devices, but in addition, has short-circuit memory and voltage threshold. Memory and threshold are sufficiently large to allow, in principle, multiplexing of electrochromic display panels of large-screen TV pixel size.

  15. Clinical Implications of Phosphorylated STAT3 Expression in de novo Diffuse Large B-cell Lymphoma

    DEFF Research Database (Denmark)

    Ok, Chi Y; Chen, Jiayu; Xu-Monette, Ziju

    2014-01-01

    of phosphorylated STAT3 (pSTAT3) on prognosis are limited. EXPERIMENTAL DESIGN: We evaluated expression of pSTAT3 in de novo DLBCL using immunohistochemistry, gene expression profiling (GEP), and gene set enrichment analysis (GSEA). Results were analyzed in correlation with cell-of-origin (COO), critical lymphoma...

  16. Effects of gabapentin on the expression of p-STAT3 in dorsal roto ganglia of diabetic neuropathological pain rats%加巴喷丁对糖尿病神经病理痛大鼠背根神经节磷酸化信号转导和转录激活因子3表达的影响

    Institute of Scientific and Technical Information of China (English)

    苗蓓; 周田田; 邵翠杰

    2014-01-01

    目的:观察加巴喷丁(gabapentin, GBP)对糖尿病神经病理痛大鼠背根神经节磷酸化信号转导和转录激活因子3(p-STAT3)表达的影响。方法80只成年健康雄性SD大鼠,随机分为正常组(Control 组)、糖尿病神经病理痛组( DNP组)、生理盐水+DNP组( SC+DNP组)和加巴喷丁+DNP组( GBP+DNP组),每组20只。采用链脲佐菌素(STZ)诱导的糖尿病神经病理痛大鼠模型;SC+DNP组和GBP+DNP组于STZ注射15天起分别腹腔注射生理盐水或GBP 50 mg/kg,1次/d,连续7天;采用行为学测试测定STZ注射前1天及注射后3、7、10、14、21、28天大鼠缩足反射机械刺激阈值(PWMT)和缩足反射热辐射潜伏期(PWTL),免疫组化方法检测大鼠背根神经节(DRG)中p-STAT3的表达。结果与Control 组比较, DNP组PWMT〔(3.8±0.84) g〕降低、PWTL〔(5.9±0.94) s〕缩短,DRG中p-STAT3蛋白的表达上调(P<0.05);与DNP组比较, SC+DNP组大鼠在腹腔注射生理盐水后,PWMT〔(4.03±0.5) g〕、PWTL〔(6.2±0.7) s〕和p-STAT3蛋白的表达无明显改变(P>0.05);与DNP组和SC+DNP组比较, GBP+DNP组PWMT〔(8.4±0.87) g〕升高,PWTL〔(10±1.2) s〕延长, DRG中p-STAT3蛋白的表达下调(P<0.05)。结论 GBP能减轻大鼠糖尿病神经病理痛,其机制可能与抑制p-STAT3的表达水平有关。%Objective To investigate the effects of gabapentin on the expression of p -STAT3 in dorsal root ganglia of a rat model of diabetic neuropathological pain ( DNP) .Methods 80 male SD rats weighing 200-220 g were randomly divided into 4groups (20 in each):a control group, a DNP group, a saline (SC+DNP) group and a gabapentin (GBP+DNP) group.Then a rat model of diabetic neuropathological pain was established through injection of streptozocin ( STZ) .Fifteen days after STZ administration , the rats of the SC

  17. Yeast surface display of dehydrogenases in microbial fuel-cells.

    Science.gov (United States)

    Gal, Idan; Schlesinger, Orr; Amir, Liron; Alfonta, Lital

    2016-12-01

    Two dehydrogenases, cellobiose dehydrogenase from Corynascus thermophilus and pyranose dehydrogenase from Agaricus meleagris, were displayed for the first time on the surface of Saccharomyces cerevisiae using the yeast surface display system. Surface displayed dehydrogenases were used in a microbial fuel cell and generated high power outputs. Surface displayed cellobiose dehydrogenase has demonstrated a midpoint potential of -28mV (vs. Ag/AgCl) at pH=6.5 and was used in a mediator-less anode compartment of a microbial fuel cell producing a power output of 3.3μWcm(-2) using lactose as fuel. Surface-displayed pyranose dehydrogenase was used in a microbial fuel cell and generated high power outputs using different substrates, the highest power output that was achieved was 3.9μWcm(-2) using d-xylose. These results demonstrate that surface displayed cellobiose dehydrogenase and pyranose dehydrogenase may successfully be used in microbial bioelectrochemical systems.

  18. Development of exosome surface display technology in living human cells.

    Science.gov (United States)

    Stickney, Zachary; Losacco, Joseph; McDevitt, Sophie; Zhang, Zhiwen; Lu, Biao

    2016-03-25

    Surface display technology is an emerging key player in presenting functional proteins for targeted drug delivery and therapy. Although a number of technologies exist, a desirable mammalian surface display system is lacking. Exosomes are extracellular vesicles that facilitate cell-cell communication and can be engineered as nano-shuttles for cell-specific delivery. In this study, we report the development of a novel exosome surface display technology by exploiting mammalian cell secreted nano-vesicles and their trans-membrane protein tetraspanins. By constructing a set of fluorescent reporters for both the inner and outer surface display on exosomes at two selected sites of tetraspanins, we demonstrated the successful exosomal display via gene transfection and monitoring fluorescence in vivo. We subsequently validated our system by demonstrating the expected intracellular partitioning of reporter protein into sub-cellular compartments and secretion of exosomes from human HEK293 cells. Lastly, we established the stable engineered cells to harness the ability of this robust system for continuous production, secretion, and uptake of displayed exosomes with minimal impact on human cell biology. In sum, our work paved the way for potential applications of exosome, including exosome tracking and imaging, targeted drug delivery, as well as exosome-mediated vaccine and therapy.

  19. Calculated characteristics of an ac plasma display panel cell

    Energy Technology Data Exchange (ETDEWEB)

    Boeuf, J.P.; Pitchford, L.C. [Universite Paul Sabatier, Toulouse (France)

    1996-02-01

    Equipotential contours and contours of constant power deposition into excitation of xenon, calculated from a two-dimensional (2-D) fluid model, are presented at times during the evolution of a single discharge event in an ac plasma display panel cell with a dielectric barrier rib geometry.

  20. OVCAR-3 spheroid-derived cells display distinct metabolic profiles.

    Directory of Open Access Journals (Sweden)

    Kathleen A Vermeersch

    Full Text Available Recently, multicellular spheroids were isolated from a well-established epithelial ovarian cancer cell line, OVCAR-3, and were propagated in vitro. These spheroid-derived cells displayed numerous hallmarks of cancer stem cells, which are chemo- and radioresistant cells thought to be a significant cause of cancer recurrence and resultant mortality. Gene set enrichment analysis of expression data from the OVCAR-3 cells and the spheroid-derived putative cancer stem cells identified several metabolic pathways enriched in differentially expressed genes. Before this, there had been little previous knowledge or investigation of systems-scale metabolic differences between cancer cells and cancer stem cells, and no knowledge of such differences in ovarian cancer stem cells.To determine if there were substantial metabolic changes corresponding with these transcriptional differences, we used two-dimensional gas chromatography coupled to mass spectrometry to measure the metabolite profiles of the two cell lines.These two cell lines exhibited significant metabolic differences in both intracellular and extracellular metabolite measurements. Principal components analysis, an unsupervised dimensional reduction technique, showed complete separation between the two cell types based on their metabolite profiles. Pathway analysis of intracellular metabolomics data revealed close overlap with metabolic pathways identified from gene expression data, with four out of six pathways found enriched in gene-level analysis also enriched in metabolite-level analysis. Some of those pathways contained multiple metabolites that were individually statistically significantly different between the two cell lines, with one of the most broadly and consistently different pathways, arginine and proline metabolism, suggesting an interesting hypothesis about cancerous and stem-like metabolic phenotypes in this pair of cell lines.Overall, we demonstrate for the first time that metabolism

  1. Effect of Wall Charge on Striation in Plasma Display Cells

    Institute of Scientific and Technical Information of China (English)

    HE Feng; OUYANG Jiting; CAO Jing; FENG Shuo; MIAO Jinsong; WANG Jianqi

    2007-01-01

    Different configurations and driving voltages have been employed to investigate the effect of the wall charge on the striations in macroscopic plasma display panel (PDP) cells.The experimental results show that a discharge channel near the dielectric layer is indispensable to striation occurring in the anode area during a discharge,while the pre-accumulated charge on the dielectric layer and the surface state are not important.The origin of the striation is related only to the physical process in the cell.The dielectric layer acts as a charge collector during a PDP discharge.

  2. Guiding plant virus particles to integrin-displaying cells

    Science.gov (United States)

    Hovlid, Marisa L.; Steinmetz, Nicole F.; Laufer, Burkhardt; Lau, Jolene L.; Kuzelka, Jane; Wang, Qian; Hyypiä, Timo; Nemerow, Glen R.; Kessler, Horst; Manchester, Marianne; Finn, M. G.

    2012-05-01

    Viral nanoparticles (VNPs) are structurally regular, highly stable, tunable nanomaterials that can be conveniently produced in high yields. Unmodified VNPs from plants and bacteria generally do not show tissue specificity or high selectivity in binding to or entry into mammalian cells. They are, however, malleable by both genetic and chemical means, making them useful scaffolds for the display of large numbers of cell- and tissue-targeting ligands, imaging moieties, and/or therapeutic agents in a well-defined manner. Capitalizing on this attribute, we modified the genetic sequence of the Cowpea mosaic virus (CPMV) coat protein to display an RGD oligopeptide sequence derived from human adenovirus type 2 (HAdV-2). Concurrently, wild-type CPMV was modified via NHS acylation and Cu(i)-catalyzed azide-alkyne cycloaddition (CuAAC) chemistry to attach an integrin-binding cyclic RGD peptide. Both types of particles showed strong and selective affinity for several different cancer cell lines that express RGD-binding integrin receptors.Viral nanoparticles (VNPs) are structurally regular, highly stable, tunable nanomaterials that can be conveniently produced in high yields. Unmodified VNPs from plants and bacteria generally do not show tissue specificity or high selectivity in binding to or entry into mammalian cells. They are, however, malleable by both genetic and chemical means, making them useful scaffolds for the display of large numbers of cell- and tissue-targeting ligands, imaging moieties, and/or therapeutic agents in a well-defined manner. Capitalizing on this attribute, we modified the genetic sequence of the Cowpea mosaic virus (CPMV) coat protein to display an RGD oligopeptide sequence derived from human adenovirus type 2 (HAdV-2). Concurrently, wild-type CPMV was modified via NHS acylation and Cu(i)-catalyzed azide-alkyne cycloaddition (CuAAC) chemistry to attach an integrin-binding cyclic RGD peptide. Both types of particles showed strong and selective affinity

  3. Plasmids and packaging cell lines for use in phage display

    Science.gov (United States)

    Bradbury, Andrew M.

    2012-07-24

    The invention relates to a novel phagemid display system for packaging phagemid DNA into phagemid particles which completely avoids the use of helper phage. The system of the invention incorporates the use of bacterial packaging cell lines which have been transformed with helper plasmids containing all required phage proteins but not the packaging signals. The absence of packaging signals in these helper plasmids prevents their DNA from being packaged in the bacterial cell, which provides a number of significant advantages over the use of both standard and modified helper phage. Packaged phagemids expressing a protein or peptide of interest, in fusion with a phage coat protein such as g3p, are generated simply by transfecting phagemid into the packaging cell line.

  4. Yeast cell surface display for lipase whole cell catalyst and its applications

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yun; Zhang, Rui; Lian, Zhongshuai; Wang, Shihui; Wright, Aaron T.

    2014-08-01

    The cell surface display technique allows for the expression of target proteins or peptides on the microbial cell surface by fusing an appropriate protein as an anchoring motif. Yeast display systems, such as Pichia pastoris, Yarowia lipolytica and Saccharomyces cerevisiae, are ideal, alternative and extensive display systems with the advantage of simple genetic manipulation and post-translational modification of expressed heterologous proteins. Engineered yeasts show high performance characteristics and variant utilizations. Herein, we comprehensively summarize the variant factors affecting lipase whole cell catalyst activity and display efficiency, including the structure and size of target proteins, screening anchor proteins, type and chain length of linkers, and the appropriate matching rules among the above-mentioned display units. Furthermore, we also address novel approaches to enhance stability and activity of recombinant lipases, such as VHb gene co-expression, multi-enzyme co-display technique, and the micro-environmental interference and self-assembly techniques. Finally, we represent the variety of applications of whole cell surface displayed lipases on yeast cells in non-aqueous phases, including synthesis of esters, PUFA enrichment, resolution of chiral drugs, organic synthesis and biofuels. We demonstrate that the lipase surface display technique is a powerful tool for functionalizing yeasts to serve as whole cell catalysts, and increasing interest is providing an impetus for broad application of this technique.

  5. Invasive oral cancer stem cells display resistance to ionising radiation.

    Science.gov (United States)

    Gemenetzidis, Emilios; Gammon, Luke; Biddle, Adrian; Emich, Helena; Mackenzie, Ian C

    2015-12-22

    There is a significant amount of evidence to suggest that human tumors are driven and maintained by a sub-population of cells, known as cancer stem cells (CSC). In the case of head and neck cancer, such cells have been characterised by high expression levels of CD44 cell surface glycoprotein, while we have previously shown the presence of two diverse oral CSC populations in vitro, with different capacities for cell migration and proliferation. Here, we examined the response of oral CSC populations to ionising radiation (IR), a front-line measure for the treatment of head and neck tumors. We show that oral CSC initially display resistance to IR-induced growth arrest as well as relative apoptotic resistance. We propose that this is a result of preferential activation of the DNA damagerepair pathway in oral CSC with increased activation of ATM and BRCA1, elevated levels of DNA repair proteins RAD52, XLF, and a significantly faster rate of DNA double-strand-breaks clearance 24 hours following IR. By visually identifying CSC sub-populations undergoing EMT, we show that EMT-CSC represent the majority of invasive cells, and are more radio-resistant than any other population in re-constructed 3D tissues. We provide evidence that IR is not sufficient to eliminate CSC in vitro, and that sensitization of CD44hi/ESAlow cells to IR, followed by secondary EMT blockade, could be critical in order to reduce primary tumor recurrence, but more importantly to be able to eradicate cells capable of invasion and distant metastasis.

  6. Endothelial progenitor cells display clonal restriction in multiple myeloma

    Directory of Open Access Journals (Sweden)

    Dai Kezhi

    2006-06-01

    Full Text Available Abstract Background In multiple myeloma (MM, increased neoangiogenesis contributes to tumor growth and disease progression. Increased levels of endothelial progenitor cells (EPCs contribute to neoangiogenesis in MM, and, importantly, covary with disease activity and response to treatment. In order to understand the mechanisms responsible for increased EPC levels and neoangiogenic function in MM, we investigated whether these cells were clonal by determining X-chromosome inactivation (XCI patterns in female patients by a human androgen receptor assay (HUMARA. In addition, EPCs and bone marrow cells were studied for the presence of clonotypic immunoglobulin heavy-chain (IGH gene rearrangement, which indicates clonality in B cells; thus, its presence in EPCs would indicate a close genetic link between tumor cells in MM and endothelial cells that provide tumor neovascularization. Methods A total of twenty-three consecutive patients who had not received chemotherapy were studied. Screening in 18 patients found that 11 displayed allelic AR in peripheral blood mononuclear cells, and these patients were further studied for XCI patterns in EPCs and hair root cells by HUMARA. In 2 patients whose EPCs were clonal by HUMARA, and in an additional 5 new patients, EPCs were studied for IGH gene rearrangement using PCR with family-specific primers for IGH variable genes (VH. Results In 11 patients, analysis of EPCs by HUMARA revealed significant skewing (≥ 77% expression of a single allele in 64% (n = 7. In 4 of these patients, XCI skewing was extreme (≥ 90% expression of a single allele. In contrast, XCI in hair root cells was random. Furthermore, PCR amplification with VH primers resulted in amplification of the same product in EPCs and bone marrow cells in 71% (n = 5 of 7 patients, while no IGH rearrangement was found in EPCs from healthy controls. In addition, in patients with XCI skewing in EPCs, advanced age was associated with poorer clinical status

  7. Efficient display of active lipase LipB52 with a Pichia pastoris cell surface display system and comparison with the LipB52 displayed on Saccharomyces cerevisiae cell surface

    Directory of Open Access Journals (Sweden)

    Ma Yushu

    2008-01-01

    Full Text Available Abstract Background For industrial bioconversion processes, the utilization of surface-displayed lipase in the form of whole-cell biocatalysts is more advantageous, because the enzymes are displayed on the cell surface spontaneously, regarded as immobilized enzymes. Results Two Pichia pastoris cell surface display vectors based on the flocculation functional domain of FLO with its own secretion signal sequence or the α-factor secretion signal sequence were constructed respectively. The lipase gene lipB52 fused with the FLO gene was successfully transformed into Pichia pastoris KM71. The lipase LipB52 was expressed under the control of the AOX1 promoter and displayed on Pichia pastoris KM71 cell surface with the two Pichia pastoris cell surface display vectors. Localization of the displayed LipB52 on the cell surface was confirmed by the confocal laser scanning microscopy (CLSM. The LipB52 displayed on the Pichia pastoris cell surface exhibited activity toward p-nitrophenol ester with carbon chain length ranging from C10 to C18, and the optimum substrate was p-nitrophenol-caprate (C10, which was consistent with it displayed on the Saccharomyces cerevisiae EBY100 cell surface. The hydrolysis activity of lipase LipB52 displayed on Pichia pastoris KM71-pLHJ047 and KM71-pLHJ048 cell surface reached 94 and 91 U/g dry cell, respectively. The optimum temperature of the displayed lipases was 40°C at pH8.0, they retained over 90% activity after incubation at 60°C for 2 hours at pH 7.0, and still retained 85% activity after incubation for 3 hours. Conclusion The LipB52 displayed on the Pichia pastoris cell surface exhibited better stability than the lipase LipB52 displayed on Saccharomyces cerevisiae cell surface. The displayed lipases exhibited similar transesterification activity. But the Pichia pastoris dry cell weight per liter (DCW/L ferment culture was about 5 times than Saccharomyces cerevisiae, the lipase displayed on Pichia pastoris are more

  8. Fixation and stabilization of Escherichia coli cells displaying genetically engineered cell surface proteins.

    Science.gov (United States)

    Freeman, A; Abramov, S; Georgiou, G

    1996-12-05

    A large biotechnological potential is inherent in the display of proteins (e.g., enzymes, single-chain antibodies, on the surface of bacterial cells) (Georgiou et al., 1993). Applications such as immobilized whole-cell biocatalysts or cellular adsorbents require cell fixation to prevent disintegration, stabilization of the anchored protein from leakage, denaturation or proteolysis, and total loss of cell viability, preventing medium and potential product contamination with cells. In this article we describe the adaptation of a simple two-stage chemical crosslinking procedure based on "bi-layer encagement" (Tor et al., 1989) for stabilizing Escherichia coli cells expressing an Lpp-OmpA (46-159)-beta-lactamase fusion that displays beta-lactamase on the cell surface. Bilayer crosslinking and coating the bacteria with a polymeric matrix is accomplished by treating the cells first with either glutaraldehyde or polyglutaraldehyde, followed by secondary crosslinking with polyacrylamide hydrazide. These treatments resulted in a 5- to 25-fold reduction of the thermal inactivation rate constant at 55 degrees C of surface anchored beta-lactamase and completely prevented the deterioration of the cells for at least a week of storage at 4 degrees C. The stabilization procedure developed paves the way to scalable biotechnological applications of E. coli displaying surface anchored proteins as whole-cell biocatalysts and adsorbents.

  9. Fixation and stabilization of Escherichia coli cells displaying genetically engineered cell surface proteins

    Energy Technology Data Exchange (ETDEWEB)

    Freeman, A.; Abramov, S. [Tel-Aviv Univ. (Israel); Georgiou, G. [Univ. of Texas, Austin, TX (United States). Dept. of Chemical Engineering

    1996-12-05

    A large biotechnological potential is inherent in the display of proteins. Applications such as immobilized whole-cell biocatalysts or cellular adsorbents require cell fixation to prevent disintegration, stabilization of the anchored protein from leakage, denaturation or proteolysis, and total loss of cell viability, preventing medium and potential product contamination with cells. In this article the authors describe the adaptation of a simple two-stage chemical crosslinking procedure based on bi-layer encagement for stabilizing Escherichia coli cells expressing an Lpp-OmpA-{beta}-lactamase fusion that displays {beta}-lactamase on the cell surface. Bilayer crosslinking and coating the bacteria with a polymeric matrix is accomplished by treating the cells first with either glutaraldehyde or polyglutaraldehyde, followed by secondary crosslinking with polyacrylamide hydrazide. These treatments resulted in a 5- to 25-fold reduction of the thermal inactivation rate constant at 55 C of surface anchored {beta}-lactamase and completely prevented the deterioration of the cells for at least a week of storage at 4 C. The stabilization procedure developed paves the way to scalable biotechnological applications of E. coli displaying surface anchored proteins as whole-cell biocatalysts and adsorbents.

  10. The small molecule curcumin analog FLLL32 induces apoptosis in melanoma cells via STAT3 inhibition and retains the cellular response to cytokines with anti-tumor activity

    Directory of Open Access Journals (Sweden)

    Young Gregory S

    2010-06-01

    Full Text Available Abstract Background We characterized the biologic effects of a novel small molecule STAT3 pathway inhibitor that is derived from the natural product curcumin. We hypothesized this lead compound would specifically inhibit the STAT3 signaling pathway to induce apoptosis in melanoma cells. Results FLLL32 specifically reduced STAT3 phosphorylation at Tyr705 (pSTAT3 and induced apoptosis at micromolar amounts in human melanoma cell lines and primary melanoma cultures as determined by annexin V/propidium iodide staining and immunoblot analysis. FLLL32 treatment reduced expression of STAT3-target genes, induced caspase-dependent apoptosis, and reduced mitochondrial membrane potential. FLLL32 displayed specificity for STAT3 over other homologous STAT proteins. In contrast to other STAT3 pathway inhibitors (WP1066, JSI-124, Stattic, FLLL32 did not abrogate IFN-γ-induced pSTAT1 or downstream STAT1-mediated gene expression as determined by Real Time PCR. In addition, FLLL32 did not adversely affect the function or viability of immune cells from normal donors. In peripheral blood mononuclear cells (PBMCs, FLLL32 inhibited IL-6-induced pSTAT3 but did not reduce signaling in response to immunostimulatory cytokines (IFN-γ, IL 2. Treatment of PBMCs or natural killer (NK cells with FLLL32 also did not decrease viability or granzyme b and IFN-γ production when cultured with K562 targets as compared to vehicle (DMSO. Conclusions These data suggest that FLLL32 represents a lead compound that could serve as a platform for further optimization to develop improved STAT3 specific inhibitors for melanoma therapy.

  11. Efficient method to optimize antibodies using avian leukosis virus display and eukaryotic cells.

    Science.gov (United States)

    Yu, Changming; Pike, Gennett M; Rinkoski, Tommy A; Correia, Cristina; Kaufmann, Scott H; Federspiel, Mark J

    2015-08-11

    Antibody-based therapeutics have now had success in the clinic. The affinity and specificity of the antibody for the target ligand determines the specificity of therapeutic delivery and off-target side effects. The discovery and optimization of high-affinity antibodies to important therapeutic targets could be significantly improved by the availability of a robust, eukaryotic display technology comparable to phage display that would overcome the protein translation limitations of microorganisms. The use of eukaryotic cells would improve the diversity of the displayed antibodies that can be screened and optimized as well as more seamlessly transition into a large-scale mammalian expression system for clinical production. In this study, we demonstrate that the replication and polypeptide display characteristics of a eukaryotic retrovirus, avian leukosis virus (ALV), offers a robust, eukaryotic version of bacteriophage display. The binding affinity of a model single-chain Fv antibody was optimized by using ALV display, improving affinity >2,000-fold, from micromolar to picomolar levels. We believe ALV display provides an extension to antibody display on microorganisms and offers virus and cell display platforms in a eukaryotic expression system. ALV display should enable an improvement in the diversity of properly processed and functional antibody variants that can be screened and affinity-optimized to improve promising antibody candidates.

  12. Ovarian tumor-initiating cells display a flexible metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, Angela S. [Department of Human Nutrition, Foods, and Exercise, Virginia Tech, Blacksburg, VA (United States); Roberts, Paul C. [Biomedical Science and Pathobiology, Virginia Tech, Blacksburg, VA (United States); Frisard, Madlyn I. [Department of Human Nutrition, Foods, and Exercise, Virginia Tech, Blacksburg, VA (United States); Hulver, Matthew W., E-mail: hulvermw@vt.edu [Department of Human Nutrition, Foods, and Exercise, Virginia Tech, Blacksburg, VA (United States); Schmelz, Eva M., E-mail: eschmelz@vt.edu [Department of Human Nutrition, Foods, and Exercise, Virginia Tech, Blacksburg, VA (United States)

    2014-10-15

    An altered metabolism during ovarian cancer progression allows for increased macromolecular synthesis and unrestrained growth. However, the metabolic phenotype of cancer stem or tumor-initiating cells, small tumor cell populations that are able to recapitulate the original tumor, has not been well characterized. In the present study, we compared the metabolic phenotype of the stem cell enriched cell variant, MOSE-L{sub FFLv} (TIC), derived from mouse ovarian surface epithelial (MOSE) cells, to their parental (MOSE-L) and benign precursor (MOSE-E) cells. TICs exhibit a decrease in glucose and fatty acid oxidation with a concomitant increase in lactate secretion. In contrast to MOSE-L cells, TICs can increase their rate of glycolysis to overcome the inhibition of ATP synthase by oligomycin and can increase their oxygen consumption rate to maintain proton motive force when uncoupled, similar to the benign MOSE-E cells. TICs have an increased survival rate under limiting conditions as well as an increased survival rate when treated with AICAR, but exhibit a higher sensitivity to metformin than MOSE-E and MOSE-L cells. Together, our data show that TICs have a distinct metabolic profile that may render them flexible to adapt to the specific conditions of their microenvironment. By better understanding their metabolic phenotype and external environmental conditions that support their survival, treatment interventions can be designed to extend current therapy regimens to eradicate TICs. - Highlights: • Ovarian cancer TICs exhibit a decreased glucose and fatty acid oxidation. • TICs are more glycolytic and have highly active mitochondria. • TICs are more resistant to AICAR but not metformin. • A flexible metabolism allows TICs to adapt to their microenvironment. • This flexibility requires development of specific drugs targeting TIC-specific changes to prevent recurrent TIC outgrowth.

  13. Extracellular Vesicles from Ovarian Carcinoma Cells Display Specific Glycosignatures

    Directory of Open Access Journals (Sweden)

    Joana Gomes

    2015-08-01

    Full Text Available Cells release vesicles to the extracellular environment with characteristic nucleic acid, protein, lipid, and glycan composition. Here we have isolated and characterized extracellular vesicles (EVs and total cell membranes (MBs from ovarian carcinoma OVMz cells. EVs were enriched in specific markers, including Tsg101, CD63, CD9, annexin-I, and MBs contained markers of cellular membrane compartments, including calnexin, GRASP65, GS28, LAMP-1, and L1CAM. The glycoprotein galectin-3 binding protein (LGALS3BP was strongly enriched in EVs and it contained sialylated complex N-glycans. Lectin blotting with a panel of lectins showed that EVs had specific glycosignatures relative to MBs. Furthermore, the presence of glycoproteins bearing complex N-glycans with α2,3-linked sialic acid, fucose, bisecting-GlcNAc and LacdiNAc structures, and O-glycans with the T-antigen were detected. The inhibition of N-glycosylation processing from high mannose to complex glycans using kifunensine caused changes in the composition of EVs and induced a decrease of several glycoproteins. In conclusion, the results showed that glycosignatures of EVs were specific and altered glycosylation within the cell affected the composition and/or dynamics of EVs release. Furthermore, the identified glycosignatures of EVs could provide novel biomarkers for ovarian cancer.

  14. ProtEx: a novel technology to display exogenous proteins on the cell surface for immunomodulation.

    Science.gov (United States)

    Singh, Narendra P; Yolcu, Esma S; Askenasy, Nadir; Shirwan, Haval

    2005-11-01

    Gene therapy as an immunomodulatory approach has the potential to treat various inherited and acquired immune-based human diseases. However, its clinical application has several challenges, varying from the efficiency of gene transfer, control of gene expression, cell and tissue targeting, and safety concerns associated with the introduction of exogenous DNA into cells/tissues. Gene therapy is also a time- and labor-intensive procedure. As an alternative, we recently developed a novel technology, ProtEx, that allows for rapid, efficient, and durable display of exogenous proteins on the surface of cells, tissues, and organs without detectable toxicity. This technology exploits the strong binding affinity (Kd = 10(-15) M) of streptavidin with biotin and involves generation of chimeric molecules composed of the extracellular portions of immunological proteins of interest and a modified form of streptavidin, biotinylation of biological surfaces, and decoration of the modified surface with chimeric proteins. Biotin persists on the cell surface for weeks both in vitro and in vivo, thereby providing a platform to display exogenous proteins with extended cell surface kinetics. Two chimeric proteins, rat FasL (SA-FasL) and human CD80 (CD80-SA), were generated and tested for cell surface display and immunomodulatory functions. SA-FasL and CD80-SA molecules persisted on the surface of various cell types for extended periods, varying from days to weeks in vitro and in vivo. The cell surface kinetics, however, were protein and cell type dependent. SA-FasL showed potent apoptotic activity against Fas+ cells as a soluble protein or displayed on the cell surface and effectively blocked alloreactive responses. The display of CD80-SA on the surface of tumor cells, however, converted them into antigen-presenting cells for effective stimulation of autologous and allogeneic T-cell responses. ProtEx technology, therefore, represents a practical and effective alternative to DNA

  15. Inhibition of STAT3 reduces astrocytoma cell invasion and constitutive activation of STAT3 predicts poor prognosis in human astrocytoma.

    Directory of Open Access Journals (Sweden)

    Qinchuan Liang

    Full Text Available Astrocytoma cells characteristically possess high invasion potentials. Recent studies have revealed that knockdown of signal transducers and activators of transcription 3 (STAT3 expression by RNAi induces apoptosis in astrocytoma cell. Nevertheless, the distinct roles of STAT3 in astrocytoma's invasion and recurrence have not been elucidated. In this study, we silenced STAT3 using Small interfering RNAs in two human glioblastoma multiforme (GBM cell lines (U251 and U87, and investigated the effect on GBM cell adhesion and invasion. Our results demonstrate that disruption of STAT3 inhibits GBM cell's adhesion and invasion. Knockdown of STAT3 significantly increased E-cadherin but decreased N-cadherin, vascular endothelial growth factor, matrix metalloproteinase 2 and matrix metalloproteinase 9. Additionally, expression of pSTAT3(Tyr705 correlates with astrocytoma WHO classification, Karnofsky performance status scale score, tumor recurrence and survival. Furthermore, pSTAT3(Tyr705 is a significant prognostic factor in astrocytoma. In conclusion, STAT3 may affect astrocytoma invasion, expression of pSTAT3(Tyr705 is a significant prognostic factor in tumor recurrence and overall survival in astrocytoma patients. Therefore, STAT3 may provide a potential target for molecular therapy in human astrocytoma, and pSTAT3(Tyr705could be an important biomarker for astrocytoma prognosis.

  16. A Fluid Membrane-Based Soluble Ligand Display System for Live CellAssays

    Energy Technology Data Exchange (ETDEWEB)

    Nam, Jwa-Min; Nair, Pradeep N.; Neve, Richard M.; Gray, Joe W.; Groves, Jay T.

    2005-10-14

    Cell communication modulates numerous biological processes including proliferation, apoptosis, motility, invasion and differentiation. Correspondingly, there has been significant interest in the development of surface display strategies for the presentation of signaling molecules to living cells. This effort has primarily focused on naturally surface-bound ligands, such as extracellular matrix components and cell membranes. Soluble ligands (e.g. growth factors and cytokines) play an important role in intercellular communications, and their display in a surface-bound format would be of great utility in the design of array-based live cell assays. Recently, several cell microarray systems that display cDNA, RNAi, or small molecules in a surface array format were proven to be useful in accelerating high-throughput functional genetic studies and screening therapeutic agents. These surface display methods provide a flexible platform for the systematic, combinatorial investigation of genes and small molecules affecting cellular processes and phenotypes of interest. In an analogous sense, it would be an important advance if one could display soluble signaling ligands in a surface assay format that allows for systematic, patterned presentation of soluble ligands to live cells. Such a technique would make it possible to examine cellular phenotypes of interest in a parallel format with soluble signaling ligands as one of the display parameters. Herein we report a ligand-modified fluid supported lipid bilayer (SLB) assay system that can be used to functionally display soluble ligands to cells in situ (Figure 1A). By displaying soluble ligands on a SLB surface, both solution behavior (the ability to become locally enriched by reaction-diffusion processes) and solid behavior (the ability to control the spatial location of the ligands in an open system) could be combined. The method reported herein benefits from the naturally fluid state of the supported membrane, which allows

  17. Identification of human embryonic progenitor cell targeting peptides using phage display.

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    Paola A Bignone

    Full Text Available Human pluripotent stem (hPS cells are capable of differentiation into derivatives of all three primary embryonic germ layers and can self-renew indefinitely. They therefore offer a potentially scalable source of replacement cells to treat a variety of degenerative diseases. The ability to reprogram adult cells to induced pluripotent stem (iPS cells has now enabled the possibility of patient-specific hPS cells as a source of cells for disease modeling, drug discovery, and potentially, cell replacement therapies. While reprogramming technology has dramatically increased the availability of normal and diseased hPS cell lines for basic research, a major bottleneck is the critical unmet need for more efficient methods of deriving well-defined cell populations from hPS cells. Phage display is a powerful method for selecting affinity ligands that could be used for identifying and potentially purifying a variety of cell types derived from hPS cells. However, identification of specific progenitor cell-binding peptides using phage display may be hindered by the large cellular heterogeneity present in differentiating hPS cell populations. We therefore tested the hypothesis that peptides selected for their ability to bind a clonal cell line derived from hPS cells would bind early progenitor cell types emerging from differentiating hPS cells. The human embryonic stem (hES cell-derived embryonic progenitor cell line, W10, was used and cell-targeting peptides were identified. Competition studies demonstrated specificity of peptide binding to the target cell surface. Efficient peptide targeted cell labeling was accomplished using multivalent peptide-quantum dot complexes as detected by fluorescence microscopy and flow cytometry. The cell-binding peptides were selective for differentiated hPS cells, had little or no binding on pluripotent cells, but preferential binding to certain embryonic progenitor cell lines and early endodermal hPS cell derivatives. Taken

  18. Bacterial whole-cell biocatalysts by surface display of enzymes: toward industrial application.

    Science.gov (United States)

    Schüürmann, Jan; Quehl, Paul; Festel, Gunter; Jose, Joachim

    2014-10-01

    Despite the first report on the bacterial display of a recombinant peptide appeared almost 30 years ago, industrial application of cells with surface-displayed enzymes is still limited. To display an enzyme on the surface of a living cell bears several advantages. First of all, neither the substrate nor the product of the enzymatic reaction needs to cross a membrane barrier. Second, the enzyme being linked to the cell can be separated from the reaction mixture and hence the product by simple centrifugation. Transfer to a new substrate preparation results in multiple cycles of enzymatic conversion. Finally, the anchoring in a matrix, in this case, the cell envelope stabilizes the enzyme and makes it less accessible to proteolytic degradation and material adsorption resulting in continuous higher activities. These advantages in common need to balance some disadvantages before this application can be taken into account for industrial processes, e.g., the exclusion of the enzyme from the cellular metabolome and hence from redox factors or other co-factors that need to be supplied. Therefore, this digest describes the different systems in Gram-positive and Gram-negative bacteria that have been used for the surface display of enzymes so far and focuses on examples among these which are suitable for industrial purposes or for the production of valuable resources, not least in order to encourage a broader application of whole-cell biocatalysts with surface-displayed enzymes.

  19. Selection of functional T cell receptor mutants from a yeast surface-display library.

    Science.gov (United States)

    Kieke, M C; Shusta, E V; Boder, E T; Teyton, L; Wittrup, K D; Kranz, D M

    1999-05-11

    The heterodimeric alphabeta T cell receptor (TCR) for antigen is the key determinant of T cell specificity. The structure of the TCR is very similar to that of antibodies, but the engineering of TCRs by directed evolution with combinatorial display libraries has not been accomplished to date. Here, we report that yeast surface display of a TCR was achieved only after the mutation of specific variable region residues. These residues are located in two regions of the TCR, at the interface of the alpha- and beta-chains and in the beta-chain framework region that is thought to be in proximity to the CD3 signal-transduction complex. The mutations are encoded naturally in many antibody variable regions, indicating specific functional differences that have not been appreciated between TCRs and antibodies. The identification of these residues provides an explanation for the inherent difficulties in the display of wild-type TCRs compared with antibodies. Yeast-displayed mutant TCRs bind specifically to the peptide/MHC antigen, enabling engineering of soluble T cell receptors as specific T cell antagonists. This strategy of random mutagenesis followed by selection for surface expression may be of general use in the directed evolution of other eukaryotic proteins that are refractory to display.

  20. A yeast surface display system for the discovery of ligands that trigger cell activation.

    Science.gov (United States)

    Cho, B K; Kieke, M C; Boder, E T; Wittrup, K D; Kranz, D M

    1998-11-01

    Opposing cells often communicate signalling events using multivalent interactions between receptors present on their cell surface. For example, T cells are typically activated when the T cell receptor (TCR) and its associated costimulatory molecules are multivalently engaged by the appropriate ligands present on an antigen presenting cell. In this report, yeast expressing high cell-surface levels of a TCR ligand (a recombinant antibody to the TCR Vbeta domain) were shown to act as 'pseudo' antigen presenting cells and induce T cell activation as monitored by increased levels of CD25 and CD69 and by downregulation of cell surface TCR. Similar levels of T cell activation could occur even when a 30-fold excess of irrelevant yeast was present, suggesting that such a yeast display system, by virtue of its ability to present ligands multivalently, may be used in highly sensitive procedures to identify novel polypeptides that interact multivalently with cell surface receptors and thereby trigger specific cellular responses.

  1. Display of GPI-anchored anti-EGFR nanobodies on extracellular vesicles promotes tumour cell targeting

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    Sander A. A. Kooijmans

    2016-03-01

    Full Text Available Background: Extracellular vesicles (EVs are attractive candidate drug delivery systems due to their ability to functionally transport biological cargo to recipient cells. However, the apparent lack of target cell specificity of exogenously administered EVs limits their therapeutic applicability. In this study, we propose a novel method to equip EVs with targeting properties, in order to improve their interaction with tumour cells. Methods: EV producing cells were transfected with vectors encoding for anti-epidermal growth factor receptor (EGFR nanobodies, which served as targeting ligands for tumour cells, fused to glycosylphosphatidylinositol (GPI anchor signal peptides derived from decay-accelerating factor (DAF. EVs were isolated using ultrafiltration/size-exclusion liquid chromatography and characterized using western blotting, Nanoparticle Tracking Analysis, and electron microscopy. EV–tumour cell interactions were analyzed under static conditions using flow cytometry and under flow conditions using a live-cell fluorescence microscopy-coupled perfusion system. Results: V analysis showed that GPI-linked nanobodies were successfully displayed on EV surfaces and were highly enriched in EVs compared with parent cells. Display of GPI-linked nanobodies on EVs did not alter general EV characteristics (i.e. morphology, size distribution and protein marker expression, but greatly improved EV binding to tumour cells dependent on EGFR density under static conditions. Moreover, nanobody-displaying EVs showed a significantly improved cell association to EGFR-expressing tumour cells under flow conditions. Conclusions: We show that nanobodies can be anchored on the surface of EVs via GPI, which alters their cell targeting behaviour. Furthermore, this study highlights GPI-anchoring as a new tool in the EV toolbox, which may be applied for EV display of a variety of proteins, such as antibodies, reporter proteins and signaling molecules.

  2. Display of GPI-anchored anti-EGFR nanobodies on extracellular vesicles promotes tumour cell targeting

    Science.gov (United States)

    Kooijmans, Sander A. A.; Aleza, Clara Gómez; Roffler, Steve R.; van Solinge, Wouter W.; Vader, Pieter; Schiffelers, Raymond M.

    2016-01-01

    Background Extracellular vesicles (EVs) are attractive candidate drug delivery systems due to their ability to functionally transport biological cargo to recipient cells. However, the apparent lack of target cell specificity of exogenously administered EVs limits their therapeutic applicability. In this study, we propose a novel method to equip EVs with targeting properties, in order to improve their interaction with tumour cells. Methods EV producing cells were transfected with vectors encoding for anti-epidermal growth factor receptor (EGFR) nanobodies, which served as targeting ligands for tumour cells, fused to glycosylphosphatidylinositol (GPI) anchor signal peptides derived from decay-accelerating factor (DAF). EVs were isolated using ultrafiltration/size-exclusion liquid chromatography and characterized using western blotting, Nanoparticle Tracking Analysis, and electron microscopy. EV–tumour cell interactions were analyzed under static conditions using flow cytometry and under flow conditions using a live-cell fluorescence microscopy-coupled perfusion system. Results EV analysis showed that GPI-linked nanobodies were successfully displayed on EV surfaces and were highly enriched in EVs compared with parent cells. Display of GPI-linked nanobodies on EVs did not alter general EV characteristics (i.e. morphology, size distribution and protein marker expression), but greatly improved EV binding to tumour cells dependent on EGFR density under static conditions. Moreover, nanobody-displaying EVs showed a significantly improved cell association to EGFR-expressing tumour cells under flow conditions. Conclusions We show that nanobodies can be anchored on the surface of EVs via GPI, which alters their cell targeting behaviour. Furthermore, this study highlights GPI-anchoring as a new tool in the EV toolbox, which may be applied for EV display of a variety of proteins, such as antibodies, reporter proteins and signaling molecules. PMID:26979463

  3. The use of scFv-displaying yeast in mammalian cell surface selections.

    Science.gov (United States)

    Wang, Xin Xiang; Shusta, Eric V

    2005-09-01

    Yeast surface display has proven to be a powerful tool for the directed evolution of immunological proteins when soluble ligands are available (Cho, B.K., Kieke, M.C., Boder, E.T., Wittrup, K.D., Kranz, D.M., 1998. A yeast surface display system for the discovery of ligands that trigger cell activation. J. Immunol. Methods 220, 179; Boder, E.T., Midelfort, K.S., Wittrup, K.D., 2000. Directed evolution of antibody fragments with monovalent femtomolar antigen-binding affinity. Proc. Natl. Acad. Sci. U. S. A. 97, 10701; Shusta, E.V., Holler, P.D., Kieke, M.C., Kranz, D.M., Wittrup, K.D., 2000. Directed evolution of a stable scaffold for T-cell receptor engineering. Nat. Biotechnol. 18, 754; Esteban, O., Zhao, H., 2004. Directed evolution of soluble single-chain human class II MHC molecules. J. Mol. Biol. 340, 81). This investigation extends the utility of this display platform by demonstrating its capacity for use in cell panning selections. This was accomplished by employing a model single-chain antibody (scFv)-hapten system that allowed for detailed investigation of the factors governing panning success. Yeast displaying anti-fluorescein scFv (4-4-20) exhibited specific interactions with the fluoresceinated endothelial cells and could be recovered from large backgrounds of irrelevant yeast in just three rounds. Successful selections required as few as 1700 fluorescein ligands per cell, and a three-round enrichment ratio of 10(6) was possible. These results indicate that yeast surface display is a viable option for use in cell or tissue-based selections.

  4. Impact of STAT1-siRNA transfection on the expressions of STAT3 and transforming growth factor β1 in human glomerulur mesangial cells induced by high glucose

    Institute of Scientific and Technical Information of China (English)

    郑晓玉

    2013-01-01

    Objective To observe the cell proliferation and the protein expression of STAT1,phosphorylation of STAT1(p-STAT1),STAT3,p-STAT3and transforming growth factor β1(TGF-β1) in human glomerulur mesangial

  5. Construction of a Pichia pastoris cell-surface display system using Flo1p anchor system.

    Science.gov (United States)

    Tanino, Takanori; Fukuda, Hideki; Kondo, Akihiko

    2006-01-01

    A Pichia pastoris cell-surface display system was constructed using a Flo1p anchor system, which was developed in Saccharomyces cerevisiae. The lipase from Rhizopus oryzae with a pro sequence (ProROL) was used as the model protein and was genetically fused to the anchor consisting of amino acids 1-1099 of Flo1p (FS anchor). The resulting fusion protein FSProROL was expressed under the control of the alcohol oxidase 1 promoter (pAOX1). The fluorescence microscopy of immunolabeled P. pastoris cells revealed that ProROL was displayed on the cell surface, and Western blot analysis revealed that the fusion protein FSProROL was noncovalently attached to the cell wall and highly glycosylated. The lipase activity of P. pastoris cells was affected by the methanol concentration for the induction phase. Surprisingly, the activity of lipase displayed on the cells incubated at 60 degrees C was not only stable but also increased to about 6.5 times the initial value after 4 h incubation.

  6. Isolation of anti-T cell receptor scFv mutants by yeast surface display.

    Science.gov (United States)

    Kieke, M C; Cho, B K; Boder, E T; Kranz, D M; Wittrup, K D

    1997-11-01

    Yeast surface display and sorting by flow cytometry have been used to isolate mutants of an scFv that is specific for the Vbeta8 region of the T cell receptor. Selection was based on equilibrium binding by two fluorescently labeled probes, a soluble Vbeta8 domain and an antibody to the c-myc epitope tag present at the carboxy-terminus of the scFv. The mutants that were selected in this screen included a scFv with threefold increased affinity for the Vbeta8 and scFv clones that were bound with reduced affinities by the anti-c-myc antibody. The latter finding indicates that the yeast display system may be used to map conformational epitopes, which cannot be revealed by standard peptide screens. Equilibrium antigen binding constants were estimated within the surface display format, allowing screening of isolated mutants without necessitating subcloning and soluble expression. Only a relatively small library of yeast cells (3 x 10[5]) displaying randomly mutagenized scFv was screened to identify these mutants, indicating that this system will provide a powerful tool for engineering the binding properties of eucaryotic secreted and cell surface proteins.

  7. Automated cell identification and tracking using nanoparticle moving-light-displays.

    Directory of Open Access Journals (Sweden)

    James A Tonkin

    Full Text Available An automated technique for the identification, tracking and analysis of biological cells is presented. It is based on the use of nanoparticles, enclosed within intra-cellular vesicles, to produce clusters of discrete, point-like fluorescent, light sources within the cells. Computational analysis of these light ensembles in successive time frames of a movie sequence, using k-means clustering and particle tracking algorithms, provides robust and automated discrimination of live cells and their motion and a quantitative measure of their proliferation. This approach is a cytometric version of the moving light display technique which is widely used for analyzing the biological motion of humans and animals. We use the endocytosis of CdTe/ZnS, core-shell quantum dots to produce the light displays within an A549, epithelial, lung cancer cell line, using time-lapse imaging with frame acquisition every 5 minutes over a 40 hour time period. The nanoparticle moving light displays provide simultaneous collection of cell motility data, resolution of mitotic traversal dynamics and identification of familial relationships allowing construction of multi-parameter lineage trees.

  8. Automated cell identification and tracking using nanoparticle moving-light-displays.

    Science.gov (United States)

    Tonkin, James A; Rees, Paul; Brown, Martyn R; Errington, Rachel J; Smith, Paul J; Chappell, Sally C; Summers, Huw D

    2012-01-01

    An automated technique for the identification, tracking and analysis of biological cells is presented. It is based on the use of nanoparticles, enclosed within intra-cellular vesicles, to produce clusters of discrete, point-like fluorescent, light sources within the cells. Computational analysis of these light ensembles in successive time frames of a movie sequence, using k-means clustering and particle tracking algorithms, provides robust and automated discrimination of live cells and their motion and a quantitative measure of their proliferation. This approach is a cytometric version of the moving light display technique which is widely used for analyzing the biological motion of humans and animals. We use the endocytosis of CdTe/ZnS, core-shell quantum dots to produce the light displays within an A549, epithelial, lung cancer cell line, using time-lapse imaging with frame acquisition every 5 minutes over a 40 hour time period. The nanoparticle moving light displays provide simultaneous collection of cell motility data, resolution of mitotic traversal dynamics and identification of familial relationships allowing construction of multi-parameter lineage trees.

  9. ABI domain-containing proteins contribute to surface protein display and cell division in Staphylococcus aureus.

    Science.gov (United States)

    Frankel, Matthew B; Wojcik, Brandon M; DeDent, Andrea C; Missiakas, Dominique M; Schneewind, Olaf

    2010-10-01

    The human pathogen Staphylococcus aureus requires cell wall anchored surface proteins to cause disease. During cell division, surface proteins with YSIRK signal peptides are secreted into the cross-wall, a layer of newly synthesized peptidoglycan between separating daughter cells. The molecular determinants for the trafficking of surface proteins are, however, still unknown. We screened mutants with non-redundant transposon insertions by fluorescence-activated cell sorting for reduced deposition of protein A (SpA) into the staphylococcal envelope. Three mutants, each of which harboured transposon insertions in genes for transmembrane proteins, displayed greatly reduced envelope abundance of SpA and surface proteins with YSIRK signal peptides. Characterization of the corresponding mutations identified three transmembrane proteins with abortive infectivity (ABI) domains, elements first described in lactococci for their role in phage exclusion. Mutations in genes for ABI domain proteins, designated spdA, spdB and spdC (surface protein display), diminish the expression of surface proteins with YSIRK signal peptides, but not of precursor proteins with conventional signal peptides. spdA, spdB and spdC mutants display an increase in the thickness of cross-walls and in the relative abundance of staphylococci with cross-walls, suggesting that spd mutations may represent a possible link between staphylococcal cell division and protein secretion.

  10. Cell Surface Display and Characterization of Rhizopus oryzae Lipase in Pichia pastoris Using Sed1p as an Anchor Protein.

    Science.gov (United States)

    Li, Wenqian; Shi, Hao; Ding, Huaihai; Wang, Liangliang; Zhang, Yu; Li, Xun; Wang, Fei

    2015-07-01

    It has been investigated to conduct the surface displaying of lipase from Rhizopus oryzae onto the cells of Pichia pastoris yeast using Sed1p as an anchor protein. A yeast cell surface display plasmid pPICZαA-rol-histag-sed1p was constructed by fusing rol and sed1p gene fragments into the plasmid pPICZαA, followed by introducing recombinant plasmid into P. pastoris cells. Surface display levels were monitored by Western Blot and immunofluorescence microscopy. The activity of displaying lipase obtained from recombinant mutS reached at 60 U/g-dry cell. In addition, the displaying lipase was stable in broad ranges of temperatures and pH, with the optimum temperature at 40 °C and pH 7.5. These results indicate that the P. pastoris displaying lipase may have potential in whole-cell biocatalyst.

  11. Proximity effect among cellulose-degrading enzymes displayed on the Saccharomyces cerevisiae cell surface.

    Science.gov (United States)

    Bae, Jungu; Kuroda, Kouichi; Ueda, Mitsuyoshi

    2015-01-01

    Proximity effect is a form of synergistic effect exhibited when cellulases work within a short distance from each other, and this effect can be a key factor in enhancing saccharification efficiency. In this study, we evaluated the proximity effect between 3 cellulose-degrading enzymes displayed on the Saccharomyces cerevisiae cell surface, that is, endoglucanase, cellobiohydrolase, and β-glucosidase. We constructed 2 kinds of arming yeasts through genome integration: ALL-yeast, which simultaneously displayed the 3 cellulases (thus, the different cellulases were near each other), and MIX-yeast, a mixture of 3 kinds of single-cellulase-displaying yeasts (the cellulases were far apart). The cellulases were tagged with a fluorescence protein or polypeptide to visualize and quantify their display. To evaluate the proximity effect, we compared the activities of ALL-yeast and MIX-yeast with respect to degrading phosphoric acid-swollen cellulose after adjusting for the cellulase amounts. ALL-yeast exhibited 1.25-fold or 2.22-fold higher activity than MIX-yeast did at a yeast concentration equal to the yeast cell number in 1 ml of yeast suspension with an optical density (OD) at 600 nm of 10 (OD10) or OD0.1. At OD0.1, the distance between the 3 cellulases was greater than that at OD10 in MIX-yeast, but the distance remained the same in ALL-yeast; thus, the difference between the cellulose-degrading activities of ALL-yeast and MIX-yeast increased (to 2.22-fold) at OD0.1, which strongly supports the proximity effect between the displayed cellulases. A proximity effect was also observed for crystalline cellulose (Avicel). We expect the proximity effect to further increase when enzyme display efficiency is enhanced, which would further increase cellulose-degrading activity. This arming yeast technology can also be applied to examine proximity effects in other diverse fields.

  12. In vitro Fab display: a cell-free system for IgG discovery.

    Science.gov (United States)

    Stafford, Ryan L; Matsumoto, Marissa L; Yin, Gang; Cai, Qi; Fung, Juan Jose; Stephenson, Heather; Gill, Avinash; You, Monica; Lin, Shwu-Hwa; Wang, Willie D; Masikat, Mary Rose; Li, Xiaofan; Penta, Kalyani; Steiner, Alex R; Baliga, Ramesh; Murray, Christopher J; Thanos, Christopher D; Hallam, Trevor J; Sato, Aaron K

    2014-04-01

    Selection technologies such as ribosome display enable the rapid discovery of novel antibody fragments entirely in vitro. It has been assumed that the open nature of the cell-free reactions used in these technologies limits selections to single-chain protein fragments. We present a simple approach for the selection of multi-chain proteins, such as antibody Fab fragments, using ribosome display. Specifically, we show that a two-chain trastuzumab (Herceptin) Fab domain can be displayed in a format which tethers either the heavy or light chain to the ribosome while retaining functional antigen binding. Then, we constructed synthetic Fab HC and LC libraries and performed test selections against carcinoembryonic antigen (CEA) and vascular endothelial growth factor (VEGF). The Fab selection output was reformatted into full-length immunoglobulin Gs (IgGs) and directly expressed at high levels in an optimized cell-free system for immediate screening, purification and characterization. Several novel IgGs were identified using this cell-free platform that bind to purified CEA, CEA positive cells and VEGF.

  13. Exploiting bacterial peptide display technology to engineer biomaterials for neural stem cell culture.

    Science.gov (United States)

    Little, Lauren E; Dane, Karen Y; Daugherty, Patrick S; Healy, Kevin E; Schaffer, David V

    2011-02-01

    Stem cells are often cultured on substrates that present extracellular matrix (ECM) proteins; however, the heterogeneous and poorly defined nature of ECM proteins presents challenges both for basic biological investigation of cell-matrix investigations and translational applications of stem cells. Therefore, fully synthetic, defined materials conjugated with bioactive ligands, such as adhesive peptides, are preferable for stem cell biology and engineering. However, identifying novel ligands that engage cellular receptors can be challenging, and we have thus developed a high throughput approach to identify new adhesive ligands. We selected an unbiased bacterial peptide display library for the ability to bind adult neural stem cells (NSCs), and 44 bacterial clones expressing peptides were identified and found to bind to NSCs with high avidity. Of these clones, four contained RGD motifs commonly found in integrin binding domains, and three exhibited homology to ECM proteins. Three peptide clones were chosen for further analysis, and their synthetic analogs were adsorbed on tissue culture polystyrene (TCPS) or grafted onto an interpenetrating polymer network (IPN) for cell culture. These three peptides were found to support neural stem cell self-renewal in defined medium as well as multi-lineage differentiation. Therefore, bacterial peptide display offers unique advantages to isolate bioactive peptides from large, unbiased libraries for applications in biomaterials engineering.

  14. STAT3 can be activated through paracrine signaling in breast epithelial cells

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    Sasser A Kate

    2008-10-01

    Full Text Available Abstract Background Many cancers, including breast cancer, have been identified with increased levels of phosphorylated or the active form of Signal Transducers and Activators of Transcription 3 (STAT3 protein. However, whether the tumor microenvironment plays a role in this activation is still poorly understood. Methods Conditioned media, which contains soluble factors from MDA-MB-231 and MDA-MB-468 breast cancer cells and breast cancer associated fibroblasts, was added to MCF-10A breast epithelial and MDA-MB-453 breast cancer cells. The stimulation of phosphorylated STAT3 (p-STAT3 levels by conditioned media was assayed by Western blot in the presence or absence of neutralized IL-6 antibody, or a JAK/STAT3 inhibitor, JSI-124. The stimulation of cell proliferation in MCF-10A cells by conditioned media in the presence or absence of JSI-124 was subjected to MTT analysis. IL-6, IL-10, and VEGF levels were determined by ELISA analysis. Results Our results demonstrated that conditioned media from cell lines with constitutively active STAT3 are sufficient to induce p-STAT3 levels in various recipients that do not possess elevated p-STAT3 levels. This signaling occurs through the JAK/STAT3 pathway, leading to STAT3 phosphorylation as early as 30 minutes and is persistent for at least 24 hours. ELISA analysis confirmed a correlation between elevated levels of IL-6 production and p-STAT3. Neutralization of the IL-6 ligand or gp130 was sufficient to block increased levels of p-STAT3 (Y705 in treated cells. Furthermore, soluble factors within the MDA-MB-231 conditioned media were also sufficient to stimulate an increase in IL-6 production from MCF-10A cells. Conclusion These results demonstrate STAT3 phosphorylation in breast epithelial cells can be stimulated by paracrine signaling through soluble factors from both breast cancer cells and breast cancer associated fibroblasts with elevated STAT3 phosphorylation. The induction of STAT3 phosphorylation is

  15. Ophiobolin A, a sesterterpenoid fungal phytotoxin, displays higher in vitro growth-inhibitory effects in mammalian than in plant cells and displays in vivo antitumor activity.

    Science.gov (United States)

    Bury, Marina; Novo-Uzal, Esther; Andolfi, Anna; Cimini, Sara; Wauthoz, Nathalie; Heffeter, Petra; Lallemand, Benjamin; Avolio, Fabiana; Delporte, Cédric; Cimmino, Alessio; Dubois, Jacques; Van Antwerpen, Pierre; Zonno, Maria Chiara; Vurro, Maurizio; Poumay, Yves; Berger, Walter; Evidente, Antonio; De Gara, Laura; Kiss, Robert; Locato, Vittoria

    2013-08-01

    Ophiobolin A, a sesterterpenoid produced by plant pathogenic fungi, was purified from the culture extract of Drechslera gigantea and tested for its growth-inhibitory activity in both plant and mammalian cells. Ophiobolin A induced cell death in Nicotiana tabacum L. cv. Bright Yellow 2 (TBY-2) cells at concentrations ≥10 µM, with the TBY-2 cells showing typical features of apoptosis-like cell death. At a concentration of 5 µM, ophiobolin A did not affect plant cell viability but prevented cell proliferation. When tested on eight cancer cell lines, concentrations <1 µM of ophiobolin A inhibited growth by 50% after 3 days of culture irrespective of their multidrug resistance (MDR) phenotypes and their resistance levels to pro-apoptotic stimuli. It is, thus, unlikely that ophiobolin A exerts these in vitro growth-inhibitory effects in cancer cells by activating pro-apoptotic processes. Highly proliferative human keratinocytes appeared more sensitive to the growth-inhibitory effects of ophiobolin A than slowly proliferating ones. Ophiobolin A also displayed significant antitumor activity at the level of mouse survival when assayed at 10 mg/kg in the B16F10 mouse melanoma model with lung pseudometastases. Ophiobolin A could, thus, represent a novel scaffold to combat cancer types that display various levels of resistance to pro-apoptotic stimuli and/or various MDR phenotypes.

  16. Targeting Phosphatidylserine on Apoptotic Cells with Phages and Peptides Selected from a Bacteriophage Display Library

    Directory of Open Access Journals (Sweden)

    Ruping Shao

    2007-11-01

    Full Text Available Phosphatidylserine (PS is a well-characterized biomarker for apoptosis. Ligands that bind to PS can be used for noninvasive imaging of therapy-induced cell death, particularly apoptosis. In this study, we screened a random 12-mer peptide phage library on liposomes prepared from PS. One clone displaying the peptide SVSVGMKPSPRP (designated as PS3-10 bound to PS approximately 4-fold better than its binding to phosphatidylcholine and 18-fold better than to bovine serum albumin in a solid-phase binding assay. In addition, the binding of the corresponding PS3-10 peptide to PS was significantly higher than that of a scrambled peptide. PS3-10 phages, but not a control 4-2-2 phage, bound to aged red blood cells that had PS exposed on their surface. Binding of PS3-10 phages and PS3-10 peptide to TRAIL-induced apoptotic DLD1 cells was 3.2 and 5.4 times higher than their binding to untreated viable cells, respectively. Significantly, immunohistochemical staining confirmed selective binding of PS3-10 phages to apoptotic cells. Our data suggest that panning of phage display libraries may allow the selection of suitable peptide ligands for apoptotic cells and that PS3-10 peptide may serve as a template for further development of molecular probes for in vitro and in vivo imaging of apoptosis.

  17. Parametric studies of AC plasma display panel cells in complex geometries

    Energy Technology Data Exchange (ETDEWEB)

    Punset, C.; Boeuf, J.P.; Pitchford, L.C. [Univ. Paul Sabatier, Toulouse (France). CPAT

    1996-12-31

    The authors have developed a two-dimensional fluid model of AC plasma panel cells. This model was used to study the operation of a plasma panel cell in a barrier rib geometry in single pulse operation. In this communication, they report results from calculations in several different geometries and for a series of applied voltage pulses. The purpose of this work is to study the effect of geometry on the electrical characteristics of AC plasma display panel cells. The calculations reported here are representative of actual device operation. They have also used the model to study the effect of the geometrical parameters of the cell, to optimize the discharge efficiency in producing UV photons and to study cross-talk, i.e., interaction between adjacent cells. The conclusions of these parametric studies will be presented.

  18. Spatial uniformity inspection apparatus for solar cells using a projection display.

    Science.gov (United States)

    Yoo, Jae-Keun; Kim, Seung Kwan; Lee, Dong-Hoon; Park, Seung-Nam

    2012-07-10

    We demonstrate a measurement apparatus to inspect spatial uniformity of quantum efficiency of solar cells using a beam projector. Deviation of irradiance from the used beam projector over the area of 1.5×0.8 m on the cell plane was flattened within ±2.6% through gray scale adjustment, which was originally about 200%. Scanning a small square image with an area of 3×3 mm over a square-shaped photovoltaic cell with an area of 15.6×15.6 cm, we could identify the locations according to efficiency level and showed that the cell had quantum efficiency deviation of more than 10%. Utilizing the advantageous feature of a projection display, we also demonstrated that this apparatus can inspect the spatial uniformity of solar modules and panels consisting of multiple solar cells.

  19. Liquid crystal cell design of VGA field sequential color LCoS display

    Science.gov (United States)

    Liu, Yanyan; Geng, Weidong; Dai, Yongping

    2009-07-01

    The design of liquid crystal cell is an important factor to determine the display quality of LCoS display device. The goal of this paper is to gain VGA field sequential color (FSC) LCoS device used for near-to-eye system. The characteristics of optics and electrooptics for the twist nematic liquid crystal material and the material requirements of the FSC LCoS were studied. The LCOS liquid crystal cell optimized by dynamic parameter space method had an uniform reflectivity (about 90%) for the light with wave length from 450nm to 650nm. Both considering the electrooptic response curve of liquid crystal and the relationship between the contrast ratio and pixel size, we determined to use high speed twist nematic liquid crystal working in normally white mode. The liquid crystal cell gap and the pixel size were determined as 2.5um and 12um, respectively. The VGA FSC LCoS device was fabricated with SMIC 0.35um CMOS process and filled with LC-A liquid crystal of Merck in Varitronix. The measurement showed that the response time of liquid crystal from light to dark was 1.8ms and from dark to light was 4.4ms. The contrast ratio is bigger than 50:1. The LCoS displays well.

  20. Radioresistant Sf9 insect cells display moderate resistance against cumene hydroperoxide.

    Science.gov (United States)

    Kumar, Jyoti Swaroop; Suman, Shubhankar; Singh, Vijaypal; Chandna, Sudhir

    2012-08-01

    Lepidopteran insect cells serve as excellent model to study stress responses and are known to display resistance against DNA damaging agents including ionizing radiation; however, limited information is available on the effects of membrane damaging agents in these cells. In this study, we investigated the response of Sf9 cells (derived from ovaries of Spodoptera frugiperda; order Lepidoptera) to cumene hydroperoxide (CHPx), compared to human BMG-1 cells. CHPx treatment at doses lethal for human cells also caused typical necrosis in Sf9. Severe necrosis in human BMG-1 cells was observed at 125 μM, whereas similar effect in Sf9 cells was observed at 250 μM. In Sf9 cells, CHPx (250 μM) induced negligible changes in mitochondrial membrane potential and intracellular reactive oxygen species, while moderate effect was observed on intracellular calcium distribution. Reduced DNA damage and lipid (including cardiolipin) oxidation was observed in Sf9 cells that could be due to moderate total antioxidant status and constitutive/induced glutathione S-transferase activity. This study importantly demonstrates that Lepidopteran insect cells having extensive resistance towards DNA damaging agents show only moderately higher resistance to membrane damaging agents. A stronger reducing environment involving efficient antioxidant system seems to contribute significantly in this response.

  1. Mimotopes for alloreactive and conventional T cells in a peptide-MHC display library.

    Directory of Open Access Journals (Sweden)

    Frances Crawford

    2004-04-01

    Full Text Available The use of peptide libraries for the identification and characterization of T cell antigen peptide epitopes and mimotopes has been hampered by the need to form complexes between the peptides and an appropriate MHC molecule in order to construct a complete T cell ligand. We have developed a baculovirus-based peptide library method in which the sequence encoding the peptide is embedded within the genes for the MHC molecule in the viral DNA, such that insect cells infected with virus encoding a library of different peptides each displays a unique peptide-MHC complex on its surface. We have fished in such a library with two different fluorescent soluble T cell receptors (TCRs, one highly peptide specific and the other broadly allo-MHC specific and hypothesized to be much less focused on the peptide portion of the ligand. A single peptide sequence was selected by the former alphabetaTCR that, not unexpectedly, was highly related to the immunizing peptide. As hypothesized, the other alphabetaTCR selected a large family of peptides, related only by a similarity to the immunizing peptide at the p5 position. These findings have implications for the relative importance of peptide and MHC in TCR ligand recognition. This display method has broad applications in T cell epitope identification and manipulation and should be useful in general in studying interactions between complex proteins.

  2. Prazosin Displays Anticancer Activity against Human Prostate Cancers: Targeting DNA, Cell Cycle

    Directory of Open Access Journals (Sweden)

    Ssu-Chia Lin

    2007-10-01

    Full Text Available Quinazoline-based α1,-adrenoceptor antagonists, in particular doxazosin, terazosin, are suggested to display antineoplastic activity against prostate cancers. However, there are few studies elucidating the effect of prazosin. In this study, prazosin displayed antiproliferative activity superior to that of other α1-blockers, including doxazosin, terazosin, tamsulosin, phentolamine. Prazosin induced G2 checkpoint arrest, subsequent apoptosis in prostate cancer PC-3, DU-145, LNCaP cells. In p53-null PC-3 cells, prazosin induced an increase in DNA str, breaks, ATM/ATR checkpoint pathways, leading to the activation of downstream signaling cascades, including Cdc25c phosphorylation at Ser216, nuclear export of Cdc25c, cyclin-dependent kinase (Cdk 1 phosphorylation at Tyr15. The data, together with sustained elevated cyclin A levels (other than cyclin B1 levels, suggested that Cdki activity was inactivated by prazosin. Moreover, prazosin triggered mitochondria-mediated, caspaseexecuted apoptotic pathways in PC-3 cells. The oral administration of prazosin significantly reduced tumor mass in PC-3-derived cancer xenografts in nude mice. In summary, we suggest that prazosin is a potential antitumor agent that induces cell apoptosis through the induction of DNA damage stress, leading to Cdki inactivation, G2 checkpoint arrest. Subsequently, mitochondriamediated caspase cascades are triggered to induce apoptosis in PC-3 cells.

  3. Collecting duct-derived cells display mesenchymal stem cell properties and retain selective in vitro and in vivo epithelial capacity.

    Science.gov (United States)

    Li, Joan; Ariunbold, Usukhbayar; Suhaimi, Norseha; Sunn, Nana; Guo, Jinjin; McMahon, Jill A; McMahon, Andrew P; Little, Melissa

    2015-01-01

    We previously described a mesenchymal stem cell (MSC)-like population within the adult mouse kidney that displays long-term colony-forming efficiency, clonogenicity, immunosuppression, and panmesodermal potential. Although phenotypically similar to bone marrow (BM)-MSCs, kidney MSC-like cells display a distinct expression profile. FACS sorting from Hoxb7/enhanced green fluorescent protein (GFP) mice identified the collecting duct as a source of kidney MSC-like cells, with these cells undergoing an epithelial-to-mesenchymal transition to form clonogenic, long-term, self-renewing MSC-like cells. Notably, after extensive passage, kidney MSC-like cells selectively integrated into the aquaporin 2-positive medullary collecting duct when microinjected into the kidneys of neonatal mice. No epithelial integration was observed after injection of BM-MSCs. Indeed, kidney MSC-like cells retained a capacity to form epithelial structures in vitro and in vivo, and conditioned media from these cells supported epithelial repair in vitro. To investigate the origin of kidney MSC-like cells, we further examined Hoxb7(+) fractions within the kidney across postnatal development, identifying a neonatal interstitial GFP(lo) (Hoxb7(lo)) population displaying an expression profile intermediate between epithelium and interstitium. Temporal analyses with Wnt4(GCE/+):R26(tdTomato/+) mice revealed evidence for the intercalation of a Wnt4-expressing interstitial population into the neonatal collecting duct, suggesting that such intercalation may represent a normal developmental mechanism giving rise to a distinct collecting duct subpopulation. These results extend previous observations of papillary stem cell activity and collecting duct plasticity and imply a role for such cells in collecting duct formation and, possibly, repair.

  4. Amaryllidaceae alkaloids belonging to different structural subgroups display activity against apoptosis-resistant cancer cells.

    Science.gov (United States)

    Van Goietsenoven, Gwendoline; Andolfi, Anna; Lallemand, Benjamin; Cimmino, Alessio; Lamoral-Theys, Delphine; Gras, Thierry; Abou-Donia, Amina; Dubois, Jacques; Lefranc, Florence; Mathieu, Véronique; Kornienko, Alexander; Kiss, Robert; Evidente, Antonio

    2010-07-23

    Fifteen Amaryllidaceae alkaloids (1-15) were evaluated for their antiproliferative activities against six distinct cancer cell lines. Several of these natural products were found to have low micromolar antiproliferative potencies. The log P values of these compounds did not influence their observed activity. When active, the compounds displayed cytostatic, not cytotoxic activity, with the exception of pseudolycorine (3), which exhibited cytotoxic profiles. The active compounds showed similar efficacies toward cancer cells irrespective of whether the cell lines were responsive or resistant to proapoptotic stimuli. Altogether, the data from the present study revealed that lycorine (1), amarbellisine (6), haemanthamine (14), and haemanthidine (15) are potentially useful chemical scaffolds to generate further compounds to combat cancers associated with poor prognoses, especially those naturally resistant to apoptosis, such as glioblastoma, melanoma, non-small-cell lung, and metastatic cancers.

  5. Bioadsorption of Rare Earth Elements through Cell Surface Display of Lanthanide Binding Tags.

    Science.gov (United States)

    Park, Dan M; Reed, David W; Yung, Mimi C; Eslamimanesh, Ali; Lencka, Malgorzata M; Anderko, Andrzej; Fujita, Yoshiko; Riman, Richard E; Navrotsky, Alexandra; Jiao, Yongqin

    2016-03-01

    With the increasing demand for rare earth elements (REEs) in many emerging clean energy technologies, there is an urgent need for the development of new approaches for efficient REE extraction and recovery. As a step toward this goal, we genetically engineered the aerobic bacterium Caulobacter crescentus for REE adsorption through high-density cell surface display of lanthanide binding tags (LBTs) on its S-layer. The LBT-displayed strains exhibited enhanced adsorption of REEs compared to cells lacking LBT, high specificity for REEs, and an adsorption preference for REEs with small atomic radii. Adsorbed Tb(3+) could be effectively recovered using citrate, consistent with thermodynamic speciation calculations that predicted strong complexation of Tb(3+) by citrate. No reduction in Tb(3+) adsorption capacity was observed following citrate elution, enabling consecutive adsorption/desorption cycles. The LBT-displayed strain was effective for extracting REEs from the acid leachate of core samples collected at a prospective rare earth mine. Our collective results demonstrate a rapid, efficient, and reversible process for REE adsorption with potential industrial application for REE enrichment and separation.

  6. NK cells, displaying early activation, cytotoxicity and adhesion molecules, are associated with mild dengue disease.

    Science.gov (United States)

    Azeredo, E L; De Oliveira-Pinto, L M; Zagne, S M; Cerqueira, D I S; Nogueira, R M R; Kubelka, C F

    2006-02-01

    During the innate immune response against infections, Natural Killer (NK) cells are as important effector cells as are Cytotoxic T lymphocytes (CTL) generated after antigenic stimulation in the adaptative response. NK cells increase in numbers, after viral infection or vaccination. We investigated the NK cell and CD8 T lymphocyte status in 55 dengue infected patients. The NK (CD56+CD3-) and CD56+ T cell (CD56+CD3+) rates rise during the acute phase of disease. The majority of NK cells from dengue patients display early markers for activation (CD69, HLA-DR, and CD38) and cell adhesion molecules (CD44, CD11a) during the acute phase of disease. The intracellular cytotoxic granule, TIA-1, is also up-regulated early in NK cells. Most of these markers appear also on CD8+ T lymphocytes but during the late acute phase. Circulating IL-15 is elevated in a significant number of patients during early acute infection and its values were statistically correlated with NK frequencies and cytotoxic markers on NKs. We have therefore shown that dengue virus infection is very likely stimulating a cytotoxic response that may be efficient in controlling the virus in synergism with CD8+ T lymphocytes. Interestingly, the heightened CD56+CD3-, CD56+CD3+, CD56+TIA-1+ and CD56+CD11a+ cell rates are associated with mild dengue clinical manifestations and might indicate a good prognosis of the disease.

  7. NK cells, displaying early activation, cytotoxicity and adhesion molecules, are associated with mild dengue disease

    Science.gov (United States)

    Azeredo, E L; De Oliveira-Pinto, L M; Zagne, S M; Cerqueira, D I S; Nogueira, R M R; Kubelka, C F

    2006-01-01

    During the innate immune response against infections, Natural Killer (NK) cells are as important effector cells as are Cytotoxic T lymphocytes (CTL) generated after antigenic stimulation in the adaptative response. NK cells increase in numbers, after viral infection or vaccination. We investigated the NK cell and CD8 T lymphocyte status in 55 dengue infected patients. The NK (CD56+CD3−) and CD56+ T cell (CD56+CD3+) rates rise during the acute phase of disease. The majority of NK cells from dengue patients display early markers for activation (CD69, HLA-DR, and CD38) and cell adhesion molecules (CD44, CD11a) during the acute phase of disease. The intracellular cytotoxic granule, TIA-1, is also up-regulated early in NK cells. Most of these markers appear also on CD8+ T lymphocytes but during the late acute phase. Circulating IL-15 is elevated in a significant number of patients during early acute infection and its values were statistically correlated with NK frequencies and cytotoxic markers on NKs. We have therefore shown that dengue virus infection is very likely stimulating a cytotoxic response that may be efficient in controlling the virus in synergism with CD8+ T lymphocytes. Interestingly, the heightened CD56+CD3−, CD56+CD3+, CD56+TIA-1+ and CD56+CD11a+ cell rates are associated with mild dengue clinical manifestations and might indicate a good prognosis of the disease. PMID:16412060

  8. Display of wasp venom allergens on the cell surface of Saccharomyces cerevisiae

    Directory of Open Access Journals (Sweden)

    Poulsen Lars K

    2010-09-01

    Full Text Available Abstract Background Yeast surface display is a technique, where the proteins of interest are expressed as fusions with yeast surface proteins and thus remain attached to the yeast cell wall after expression. Our purpose was to study whether allergens expressed on the cell surface of baker's yeast Saccharomyces cerevisiae preserve their native allergenic properties and whether the yeast native surface glycoproteins interfere with IgE binding. We chose to use the major allergens from the common wasp Vespula vulgaris venom: phospholipase A1, hyaluronidase and antigen 5 as the model. Results The proteins were expressed on the surface as fusions with a-agglutinin complex protein AGA2. The expression was confirmed by fluorescent cytometry (FACS after staining the cells with antibody against a C-tag attached to the C-terminal end of the allergens. Phospholipase A1 and hyaluronidase retained their enzymatic activities. Phospholipase A1 severely inhibited the growth of the yeast cells. Antigen 5 - expressing yeast cells bound IgE antibodies from wasp venom allergic patient sera but not from control sera as demonstrated by FACS. Moreover, antigen 5 - expressing yeast cells were capable of mediating allergen-specific histamine release from human basophils. Conclusions All the three major wasp venom allergens were expressed on the yeast surface. A high-level expression, which was observed only for antigen 5, was needed for detection of IgE binding by FACS and for induction of histamine release. The non-modified S. cerevisiae cells did not cause any unspecific reaction in FACS or histamine release assay despite the expression of high-mannose oligosaccharides. In perspective the yeast surface display may be used for allergen discovery from cDNA libraries and possibly for sublingual immunotherapy as the cells can serve as good adjuvant and can be produced in large amounts at a low price.

  9. Individually programmable cell stretching microwell arrays actuated by a Braille display.

    Science.gov (United States)

    Kamotani, Yoko; Bersano-Begey, Tommaso; Kato, Nobuhiro; Tung, Yi-Chung; Huh, Dongeun; Song, Jonathan W; Takayama, Shuichi

    2008-06-01

    Cell culture systems are often static and are therefore nonphysiological. In vivo, many cells are exposed to dynamic surroundings that stimulate cellular responses in a process known as mechanotransduction. To recreate this environment, stretchable cell culture substrate systems have been developed, however, these systems are limited by being macroscopic and low throughput. We have developed a device consisting of 24 miniature cell stretching chambers with flexible bottom membranes that are deformed using the computer-controlled, piezoelectrically actuated pins of a Braille display. We have also developed efficient image capture and analysis protocols to quantify morphological responses of the cells to applied strain. Human dermal microvascular endothelial cells (HDMECs) were found to show increasing degrees of alignment and elongation perpendicular to the radial strain in response to cyclic stretch at increasing frequencies of 0.2, 1, and 5 Hz, after 2, 4, and 12h. Mouse myogenic C2C12 cells were also found to align in response to the stretch, while A549 human lung adenocarcinoma epithelial cells did not respond to stretch.

  10. Development of RI-based real-time display technology of apoptotic cell death

    Energy Technology Data Exchange (ETDEWEB)

    Park, Sang Hyun; Jagn, Beom Su; Hayu, Tyas Utami

    2012-01-15

    Apoptosis, or the programmed cell death, is the generally normal death of a cell in living organisms. Inappropriate apoptosis (either too little or too much) is a factor in many human disease including neurodegenerative diseases, autoimmune disorders and many types of cancer. Therefore, it is one of the most challenging and widely studied topics currently. Development of RI-based real-time display technology of apoptosis can be provided invaluable analysis data for diagnosis and treatment of various diseases. In this study, bifunctional chelator (BFC) for Tc-99m tricarbonyl was synthesized for ML-10 derivative radiolabeling. The formation of complexation of apoptotic cells was developed by combining the ML-10 moiety with the BFC for {sup 99m}Tc-tricarbonyl precursor. The results of this project will be utilized for the development of RI-Biomics Center-based Total Analysis System (TAS) through the optimization of equipment in the RI-Biomics Center.

  11. Display of wasp venom allergens on the cell surface of Saccharomyces cerevisiae

    DEFF Research Database (Denmark)

    Borodina, Irina; Jensen, B. M.; Søndergaard, Ib;

    2010-01-01

    Background: Yeast surface display is a technique, where the proteins of interest are expressed as fusions with yeast surface proteins and thus remain attached to the yeast cell wall after expression. Our purpose was to study whether allergens expressed on the cell surface of baker's yeast...... Saccharomyces cerevisiae preserve their native allergenic properties and whether the yeast native surface glycoproteins interfere with IgE binding. We chose to use the major allergens from the common wasp Vespula vulgaris venom: phospholipase A1, hyaluronidase and antigen 5 as the model. Results: The proteins...... were expressed on the surface as fusions with a-agglutinin complex protein AGA2. The expression was confirmed by fluorescent cytometry (FACS) after staining the cells with antibody against a C-tag attached to the C-terminal end of the allergens. Phospholipase A1 and hyaluronidase retained...

  12. Generation and selection of immunized Fab phage display library against human B cell lymphoma

    Institute of Scientific and Technical Information of China (English)

    Yongmei Shen; Xiaochun Yang; Ningzheng Dong; Xiaofang Xie; Xia Bai; Yizhen Shi

    2007-01-01

    The approval of using monoclonal antibodies as a targeted therapy in the management of patients with B cell lymphoma has led to new treatment options for this group of patients. Production of monoclonal antibodies by the traditional hybridoma technology is costly, and the resulting murine antibodies often have the disadvantage of triggering human anti-mouse antibody (HAMA) response. Therefore recombinant Fab antibodies generated by the phage display technology can be a suitable alternative in managing B cell lymphoma. In this study, we extracted total RNA from spleen cells of BALB/c mice immunized with human B lymphoma cells, and used RT-PCR to amplify cDNAs coding for the K light chains and Fd fragments of heavy chains. After appropriate restriction digests, these cDNA fragments were successively inserted into the phagemid vector pComb3H-SS to construct an immunized Fab phage display library. The diversity of the constructed library was approximately 1.94×107. Following five rounds of biopanning, soluble Fab antibodies were produced from positive clones identified by ELISA. From eight positive clones, FabC06, FabC21, FabC43 and FabC59 were selected for sequence analysis. At the level of amino acid sequences, the variable heavy domains (VH) and variable light domains (VL) were found to share 88-92% and 89-94% homology with sequences coded by the corresponding murine germline genes respectively. Furthermore, reactivity with membrane proteins of the B cell lymphoma was demonstrated by immunohistochemistry and western blotting. These immunized Fab antibodies may provide a valuable tool for further study of B cell lymphoma and could also contribute to the improvement of disease therapy.

  13. Thin film polycrystalline silicon: Promise and problems in displays and solar cells

    Energy Technology Data Exchange (ETDEWEB)

    Fonash, S.J. [Pennsylvania State Univ., University Park, PA (United States)

    1995-08-01

    Thin film polycrystalline Si (poly-Si) with its carrier mobilities, potentially good stability, low intragrain defect density, compatibility with silicon processing, and ease of doping activation is an interesting material for {open_quotes}macroelectronics{close_quotes} applications such as TFTs for displays and solar cells. The poly-Si films needed for these applications can be ultra-thin-in the 500{Angstrom} to 1000{Angstrom} thickness range for flat panel display TFTs and in the 4{mu}m to 10{mu}m thickness range for solar cells. Because the films needed for these microelectronics applications can be so thin, an effective approach to producing the films is that of crystallizing a-Si precursor material. Unlike cast materials, poly-Si films made this way can be produced using low temperature processing. Unlike deposited poly-Si films, these crystallized poly-Si films can have grain widths that are much larger than the film thickness and almost atomically smooth surfaces. This thin film poly-Si crystallized from a-Si precursor films, and its promise and problems for TFTs and solar cells, is the focus of this discussion.

  14. Antigen 43-mediated autotransporter display, a versatile bacterial cell surface presentation system

    DEFF Research Database (Denmark)

    Kjærgaard, Kristian; Hasman, Henrik; Schembri, Mark;

    2002-01-01

    Antigen 43 (Ag43), a self-recognizing outer membrane protein of Escherichia coli, has been converted into an efficient and versatile tool for surface display of foreign protein segments. Ag43 is an autotransporter protein characterized by the feature that all information required for transport...... to the outer membrane and secretion through the cell envelope is contained within the protein itself. Ag43 consists of two subunits (alpha and beta), where the beta-subunit forms an integral outer membrane translocator to which the alpha-subunit is noncovalently attached. The simplicity of the Ag43 system...

  15. Display of Clostridium cellulovorans xylose isomerase on the cell surface of Saccharomyces cerevisiae and its direct application to xylose fermentation.

    Science.gov (United States)

    Ota, Miki; Sakuragi, Hiroshi; Morisaka, Hironobu; Kuroda, Kouichi; Miyake, Hideo; Tamaru, Yutaka; Ueda, Mitsuyoshi

    2013-01-01

    Xylose isomerase (XI) is a key enzyme in the conversion of D-xylose, which is a major component of lignocellulosic biomass, to D-xylulose. Genomic analysis of the bacterium Clostridium cellulovorans revealed the presence of XI-related genes. In this study, XI derived from C. cellulovorans was produced and displayed using the yeast cell-surface display system, and the xylose assimilation and fermentation properties of this XI-displaying yeast were examined. XI-displaying yeast grew well in medium containing xylose as the sole carbon source and directly produced ethanol from xylose under anaerobic conditions.

  16. Large-scale differential display analysis of T helper cell differentiation.

    Science.gov (United States)

    Ojala, Pekka; Virtanen, Eveliina; Chen, Zhi

    2007-03-01

    We have developed a novel large-scale multicapillary fluorescent differential display (FDD) platform amenable to further automation. The power of the method is demonstrated by the analysis of T helper cell differentiation. Eight RNA samples from wild type, Stat4 knockout and Stat6 knockout mice were analyzed with 16 anchoring primers and 24 arbitrary primers, resulting in 285 294 sample peaks. Visually selected patterns of differential expression suggest two major regulatory mechanisms: activation and Stat4 genotype. A subset of the findings is reproduced in the confirmatory differential display (DD) that included technical and biological replicates. In a small fragment identification pilot study, we identify Ifi27 and Cct8 to be up-regulated by T cell activation. We present a method for the analysis of electropherogram similarity across large datasets, based on correlation of low-resolution representations of electrophoretic data. We show how it can be applied to analyze experimental and technical variables. Using this method, we demonstrate the effect of activation and genotype. In addition, agreement of our real experimental data to the theoretical basis of DD, as well as issues in anchoring primer selectivity, are studied.

  17. Differential display of vincristine-resistance-related genes in gastric cancer SGC7901 cells

    Institute of Scientific and Technical Information of China (English)

    Xin Wang; Bo-Rong Pan; Jian-Ping Jin; Dai-Ming Fan; Mei Lan; Yong-Quan Shi; Ju Lu; Yue-Xia Zhong; Han-Ping Wu; Hui-Hong Zai; Jie Ding; Kai-Cun Wu

    2002-01-01

    AIM: To isolate and clone the vincristine-resistine-relatedgenes in gastric cancer SGC7901 cell line and to clarify themultidrug-resistant molecular mechanism of gastric cancercells.METHODS: The modified differential-display polymerasechain reaction (DD-PCR) was used to examine thedifferences in the mRNA composition of Vincristine-resistantgastric cancer SGC 7901 cells (SGC7901/VCR), induced byvincristine sulfate versus SGC7901cells. The differentiallyexpressed cDNA fragments were confirmed byreverseNorthern analysis, sequencing, BLAST analysis andNorthern bolt analysis.RESULTS: DD-PCR identified that 54 cDNA fragments werepreferentially expressed in SGC 7901/VCR cells. When thesecDNA fragments were analyzed by reverseNorthern blot, 20were reproducibly expressed at a high level in SGC7901/VCR. Sequencing and BLAST analysis revealed that sevenof the genes were known genes: ADP-ribosylation factor 4,cytochrorne oxidase subunit Ⅱ, Ss-A/Ro ribonucleoprteinautoantigen 60kd subunit, ribosomal protein S13, galaectin-8 gene, oligophrenin 1 mRNA, and ribosomal protein L23mRNA; and thirteen of the genes were unknown genes. Thelength and abundance of the four unknown genes mRNAwere further confirmed by Northern blot analysis.CONCLUSION: The twenty differential known and unknowngenes may be related to the vincristine-resistant mechanismin human gastric cancer SGC7901 cell line.

  18. Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst

    Science.gov (United States)

    Moura, Marcelo Victor Holanda; da Silva, Giulia Pontes; Machado, Antônio Carlos de Oliveira; Torres, Fernando Araripe Gonçalves; Freire, Denise Maria Guimarães; Almeida, Rodrigo Volcan

    2015-01-01

    Yeast Surface Display (YSD) is a strategy to anchor proteins on the yeast cell wall which has been employed to increase enzyme stability thus decreasing production costs. Lipase B from Candida antarctica (LipB) is one of the most studied enzymes in the context of industrial biotechnology. This study aimed to assess the biochemical features of this important biocatalyst when immobilized on the cell surface of the methylotrophic yeast Pichia pastoris using the YSD approach. For that purpose, two anchors were tested. The first (Flo9) was identified after a prospection of the P. pastoris genome being related to the family of flocculins similar to Flo1 but significantly smaller. The second is the Protein with Internal Repeats (Pir1) from P. pastoris. An immunolocalization assay showed that both anchor proteins were able to display the reporter protein EGFP in the yeast outer cell wall. LipB was expressed in P. pastoris fused either to Flo9 (FLOLIPB) or Pir1 (PIRLIPB). Both constructions showed hydrolytic activity towards tributyrin (>100 U/mgdcw and >80 U/mgdcw, respectively), optimal hydrolytic activity around 45°C and pH 7.0, higher thermostability at 45°C and stability in organic solvents when compared to a free lipase. PMID:26510006

  19. Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst.

    Directory of Open Access Journals (Sweden)

    Marcelo Victor Holanda Moura

    Full Text Available Yeast Surface Display (YSD is a strategy to anchor proteins on the yeast cell wall which has been employed to increase enzyme stability thus decreasing production costs. Lipase B from Candida antarctica (LipB is one of the most studied enzymes in the context of industrial biotechnology. This study aimed to assess the biochemical features of this important biocatalyst when immobilized on the cell surface of the methylotrophic yeast Pichia pastoris using the YSD approach. For that purpose, two anchors were tested. The first (Flo9 was identified after a prospection of the P. pastoris genome being related to the family of flocculins similar to Flo1 but significantly smaller. The second is the Protein with Internal Repeats (Pir1 from P. pastoris. An immunolocalization assay showed that both anchor proteins were able to display the reporter protein EGFP in the yeast outer cell wall. LipB was expressed in P. pastoris fused either to Flo9 (FLOLIPB or Pir1 (PIRLIPB. Both constructions showed hydrolytic activity towards tributyrin (>100 U/mgdcw and >80 U/mgdcw, respectively, optimal hydrolytic activity around 45°C and pH 7.0, higher thermostability at 45°C and stability in organic solvents when compared to a free lipase.

  20. Displaying Lipase B from Candida antarctica in Pichia pastoris Using the Yeast Surface Display Approach: Prospection of a New Anchor and Characterization of the Whole Cell Biocatalyst.

    Science.gov (United States)

    Moura, Marcelo Victor Holanda; da Silva, Giulia Pontes; Machado, Antônio Carlos de Oliveira; Torres, Fernando Araripe Gonçalves; Freire, Denise Maria Guimarães; Almeida, Rodrigo Volcan

    2015-01-01

    Yeast Surface Display (YSD) is a strategy to anchor proteins on the yeast cell wall which has been employed to increase enzyme stability thus decreasing production costs. Lipase B from Candida antarctica (LipB) is one of the most studied enzymes in the context of industrial biotechnology. This study aimed to assess the biochemical features of this important biocatalyst when immobilized on the cell surface of the methylotrophic yeast Pichia pastoris using the YSD approach. For that purpose, two anchors were tested. The first (Flo9) was identified after a prospection of the P. pastoris genome being related to the family of flocculins similar to Flo1 but significantly smaller. The second is the Protein with Internal Repeats (Pir1) from P. pastoris. An immunolocalization assay showed that both anchor proteins were able to display the reporter protein EGFP in the yeast outer cell wall. LipB was expressed in P. pastoris fused either to Flo9 (FLOLIPB) or Pir1 (PIRLIPB). Both constructions showed hydrolytic activity towards tributyrin (>100 U/mgdcw and >80 U/mgdcw, respectively), optimal hydrolytic activity around 45°C and pH 7.0, higher thermostability at 45°C and stability in organic solvents when compared to a free lipase.

  1. Affibody-displaying bionanocapsules for specific drug delivery to HER2-expressing cancer cells.

    Science.gov (United States)

    Shishido, Takuya; Mieda, Hiroaki; Hwang, Sang Youn; Nishimura, Yuya; Tanaka, Tsutomu; Ogino, Chiaki; Fukuda, Hideki; Kondo, Akihiko

    2010-10-01

    A novel HER2-targeted carrier was developed using bionanocapsules (BNCs). Bionanocapsules (BNCs) are 100-nm hollow nanoparticles composed of the L-protein of hepatitis B virus surface antigen. An affibody of HER2 was genetically displayed on the BNC surface (Z(HER2)-BNC). For the investigation of binding affinity, Z(HER2)-BNC was incubated with the cancer cell lines SK-BR-3 (HER2 positive), and MDA-MB-231 (HER2 negative). For analysis of HER2 targeting specificity, Z(HER2)-BNC or Z(WT)-BNC (without affibody) was incubated with both SK-BR-3 and MDA-MB-231 cells by time lapse and concentration. For the delivery of encapsulated molecules (calcein), fluorescence of Z(HER2)-BNC mixed with liposomes was also compared with that of Z(WT)-BNC and nude liposomes by incubation with SK-BR-3 cells. As a result, Z(HER2)-BNC-liposome complex demonstrated the delivery to HER2-expressing cells (SK-BR-3) with a high degree of specificity. This indicates that genetically engineered BNCs are promising carrier for cancer treatment.

  2. Screening of a specific peptide binding to esophageal squamous carcinoma cells from phage displayed peptide library.

    Science.gov (United States)

    Ma, Caixia; Li, Chunyan; Jiang, Dongliang; Gao, Xiaojie; Han, Juanjuan; Xu, Nan; Wu, Qiong; Nie, Guochao; Chen, Wei; Lin, Fenghuei; Hou, Yingchun

    2015-06-01

    To select a specifically binding peptide for imaging detection of human esophageal squamous cell carcinoma (ESCC), a phage-displayed 12-mer peptide library was used to screen the peptide that bind to ESCC cells specifically. After four rounds of bio-panning, the phage recovery rate gradually increased, and specific phage clones were effectively enriched. The 60 randomly selected phage clones were tested using cellular enzyme-linked immunosorbent assay (ELISA), and 41 phage clones were identified as positive clones with the over 2.10 ratio of absorbance higher than other clones, IRP and PBS controls. From the sequencing results of the positive clones, 14 peptide sequences were obtained and ESCP9 consensus sequence was identified as the peptide with best affinity to ESCC cells via competitive inhibition, fluorescence microscopy, and flow cytometry. The results indicate that the peptide ESCP9 can bind to ESCC cells specifically and sensitively, and it is a potential candidate to be developed as an useful molecule to the imaging detection and targeting therapy for ESCC.

  3. Conditionally reprogrammed normal and transformed mouse mammary epithelial cells display a progenitor-cell-like phenotype.

    Directory of Open Access Journals (Sweden)

    Francisco R Saenz

    Full Text Available Mammary epithelial (ME cells cultured under conventional conditions senesce after several passages. Here, we demonstrate that mouse ME cells isolated from normal mammary glands or from mouse mammary tumor virus (MMTV-Neu-induced mammary tumors, can be cultured indefinitely as conditionally reprogrammed cells (CRCs on irradiated fibroblasts in the presence of the Rho kinase inhibitor Y-27632. Cell surface progenitor-associated markers are rapidly induced in normal mouse ME-CRCs relative to ME cells. However, the expression of certain mammary progenitor subpopulations, such as CD49f+ ESA+ CD44+, drops significantly in later passages. Nevertheless, mouse ME-CRCs grown in a three-dimensional extracellular matrix gave rise to mammary acinar structures. ME-CRCs isolated from MMTV-Neu transgenic mouse mammary tumors express high levels of HER2/neu, as well as tumor-initiating cell markers, such as CD44+, CD49f+, and ESA+ (EpCam. These patterns of expression are sustained in later CRC passages. Early and late passage ME-CRCs from MMTV-Neu tumors that were implanted in the mammary fat pads of syngeneic or nude mice developed vascular tumors that metastasized within 6 weeks of transplantation. Importantly, the histopathology of these tumors was indistinguishable from that of the parental tumors that develop in the MMTV-Neu mice. Application of the CRC system to mouse mammary epithelial cells provides an attractive model system to study the genetics and phenotype of normal and transformed mouse epithelium in a defined culture environment and in vivo transplant studies.

  4. Novel cell parameter determination of a twisted-nematic liquid crystal display

    Institute of Scientific and Technical Information of China (English)

    Huang Xia; Jing Hai; Fu Guo-Zhu

    2008-01-01

    In this paper a novel method is proposed to determine the cell parameters including the twist angle, optic retardation and rubbing direction of twisted-nematic liquid crystal displays (TNLCD) by rotating the TNLCD. It is a single-wavelength method. Because using subtraction equation of transmittance as curve fitting equation, the influence of the light from environment and the absorption by polarizer, the sample of TNLCD and analyser on the transmittance is eliminated. Accurate results can also be obtained in imperfect darkness. By large numbers of experiments, we found that not only the experimental setup is quite simple and can be easily adopted to be carried out, but also the results are accurate.

  5. iNKT cells from patients with secondary progressive multiple sclerosis display pro-inflammatory profiles

    Directory of Open Access Journals (Sweden)

    Sara De Biasi

    2016-11-01

    Full Text Available Background. Multiple Sclerosis (MS, an autoimmune disease with neurodegeneration and inflammation, is characterized by several alterations of different T cell subsets. However, few data exist on the role of iNKT lymphocytes. Objective. To identify possible changes in the phenotype of iNKT cells in patients with different clinical forms of MS, and find alterations in their polyfunctionality (i.e., ability to produce simultaneously up to 4 cytokines such as IL-17, TNF-α, IFN-γ, IL-4.Methods. We studied a total of 165 patients, 91 with a Relapsing Remitting form RR; 31 were treated with interferon (IFN1-β, 25 with natalizumab (Nat, 29 with glatiramer acetate (Gla; 17 were newly-diagnosed RR without treatment, 19 not active RR without treatment. Forty-four patients had a Progressive MS: 20 Primary Progressive (PP, 24 Secondary Progressive (SP. A total of 55 age- and sex-matched subjects represented healthy controls (CTR. Among fresh peripheral blood mononuclear cells (PBMC iNKT cells were identified by flow cytometry. Moreover, the capability of iNKT cells to produce different cytokines (IL-17, TNF-α, IFN-γ, and IL-4 after in vitro stimulation were evaluated in 18 RR (11 treated with Nat and 7 with IFN, 4 PP, 6 SP and 16 CTR.Results. No main differences were found in iNKT cell phenotype among MS patients with different MS forms, or during different treatments. However, the polyfunctional response of iNKT cells showed Th1 and Th17 profiles. This was well evident in patients with secondary progressive form, who are characterized by high levels of inflammation and neurodegeneration, and exhibited a sustained increase in the production of Th17 cytokines. Patients treated with natalizumab displayed lower levels of iNKT cells producing IL-17, TNF-α and IFN-γ.Conclusion. Our data suggest that the progressive phase of the disease is characterized by permanent iNKT activation and a skewing towards an inflammatory phenotype. Compared to other

  6. Screening and Identification of a Novel Hepatocellular Carcinoma Cell Binding Peptide by Using a Phage Display Library

    Institute of Scientific and Technical Information of China (English)

    Xiaohua ZHU; Hua WU; Sha LUO; Zhiqun XIANYU; Dan ZHU

    2008-01-01

    The purpose of this study was to screen peptides that can specifically bind to human hepatocellular carcinoma (hHCC) cells using phage display of random peptide library in order to develope a peptide-based carrier for the diagnosis or therapy of hHCC. A peptide 12-mer phage display library was employed and 4 rounds of subtractive panning were performed using the hHCC cell line HepG2 as the target. After panning, the phages that specifically bound to and internalized in hHCC cells were selected. The selected phages demonstrated highly specific affinity to HepG2 cells analyzed by ELISA and immunofluorescence analysis. 57.3% of the selected phage clones displayed repeated sequence FLLEPHLMDTSM, and 4 amino acid residues, FLEP were extremely conservative. Based on the sequencing results, a 16-mer peptide (WH-16) was synthesized. The competitive ELISA showed that the binding of the phage clones displayed sequence FLLEPHLMDTSM to HepG2 cells was efficiently inhibited by WH-16. Our findings indicate that cellular binding of phage is mediated via its displayed peptide and the synthesized 16-mer peptide may have the potential to be a delivery Carrier in target diagnosis or therapy for hHCC.

  7. Mesenchymal Cell Reprogramming in Experimental MPLW515L Mouse Model of Myelofibrosis

    Science.gov (United States)

    Wei, Max; Ren, Xiubao; Shao, Zonghong; Zhang, Ling; Levine, Ross L.; Epling-Burnette, Pearlie K.

    2017-01-01

    Myelofibrosis is an indicator of poor prognosis in myeloproliferative neoplasms (MPNs), but the precise mechanism(s) contributing to extracellular matrix remodeling and collagen deposition in the bone marrow (BM) niche remains unanswered. In this study, we isolated mesenchymal stromal cells (MSCs) from mice transplanted with wild-type thrombopoietin receptor (MPLWT) and MPLW515L retroviral-transduced bone marrow. Using MSCs derived from MPLW515-transplant recipients, excessive collagen deposition was maintained in the absence of the virus and neoplastic hematopoietic cells suggested that the MSCs were reprogrammed in vivo. TGFβ production by malignant megakaryocytes plays a definitive role promoting myelofibrosis in MPNs. However, TGFβ was equally expressed by MSCs derived from MPLWT and MPLW515L expressing mice and the addition of neutralizing anti-TGFβ antibody only partially reduced collagen secretion in vitro. Interestingly, profibrotic MSCs displayed increased levels of pSmad3 and pSTAT3 suggesting that inflammatory mediators cooperating with the TGFβ-receptor signaling may maintain the aberrant phenotype ex vivo. FGFb is a known suppressor of TGFβ signaling. Reduced collagen deposition by FGFb-treated MSCs derived from MPLW515L mice suggests that the activating pathway is vulnerable to this suppressive mediator. Therefore, our findings have implications for the future investigation of therapies to reverse fibrosis in MPNs. PMID:28135282

  8. Pulmonary Large Cell Carcinoma Displays High Expression of EMMPRIN and VEGF

    Institute of Scientific and Technical Information of China (English)

    Yushuang Zheng; Miao Yu; Huachuan Zheng; Yifu Guan; Yasuo Takano

    2008-01-01

    OBJECTIVE To investigate the expression of extracellular matrix metalloproteinase inducer (EMMPRIN) and vascular endothelial growth factor (VEGF) in lung carcinomas,and to clarify their roles in carcinoma progression.METHODS Expression of EMMPRIN and VEGF was examined with tissue microarrays (TMAs) of lung carcinomas (n = 181),and their suppression in adjacent normal lung samples (n = 40) were determined by immunohistochemistry.The results were compared with clinicopathological findings for the same tumors.RESULTS Both EMMPRIN and VEGF were occasionally expressed in pseudostratified columnar epithelium and frequently in lung carcinomas.Histologically,EMMPRIN and VEGF displayed higher levels in large (LCC) cell carcinomas than adenocarcinoma (AD),squamous (SQ) and small cell carcinomas (SCC) (P < 0.05).EMMPRIN was more highly expressed in SQ as compared with AD (P < 0.05),while the converse was true for VEGF (P < 0.05).Binding was generally more intense for EMMPRIN in samples from male compared to female patients (P < 0.05),whereas the latter tended to exhibit more VEGF expression (P < 0.05).Positive associations of VEGF expression with the TNM stage and amounts of EMMPRIN were noted in the lung carcinomas (P < 0.05).CONCLUSION EMMPRIN and VEGF possibly contribute to physiological repair of normal lung and histogenesis of lung carcinoma.Both proteins might be involved in the molecular basis for differences in the incidence of lung carcinoma between men and women.

  9. Neolignans from Nectandra megapotamica (Lauraceae Display in vitro Cytotoxic Activity and Induce Apoptosis in Leukemia Cells

    Directory of Open Access Journals (Sweden)

    Vitor Ponci

    2015-07-01

    Full Text Available Nectandra megapotamica (Spreng. Mez. (Lauraceae is a well-known Brazilian medicinal plant that has been used in folk medicine to treat several diseases. In continuation of our ongoing efforts to discover new bioactive natural products from the Brazilian flora, this study describes the identification of cytotoxic compounds from the MeOH extract of N. megapotamica (Lauraceae leaves using bioactivity-guided fractionation. This approach resulted in the isolation and characterization of eight tetrahydrofuran neolignans: calopeptin (1, machilin-G (2, machilin-I (3, aristolignin (4, nectandrin A (5, veraguensin (6, ganschisandrin (7, and galgravin (8. Different assays were conducted to evaluate their cytotoxic activities and to determine the possible mechanism(s related to the activity displayed against human leukemia cells. The most active compounds 4, 5 and 8 gave IC50 values of 14.2 ± 0.7, 16.9 ± 0.8 and 16.5 ± 0.8 µg/mL, respectively, against human leukemia (HL-60 tumor cells. Moreover, these compounds induced specific apoptotic hallmarks, such as plasma membrane bleb formation, nuclear DNA condensation, specific chromatin fragmentation, phosphatidyl-serine exposure on the external leaflet of the plasma membrane, cleavage of PARP as well as mitochondrial damage, which as a whole could be related to the intrinsic apoptotic pathway.

  10. Noncoding flavivirus RNA displays RNA interference suppressor activity in insect and Mammalian cells.

    Science.gov (United States)

    Schnettler, Esther; Sterken, Mark G; Leung, Jason Y; Metz, Stefan W; Geertsema, Corinne; Goldbach, Rob W; Vlak, Just M; Kohl, Alain; Khromykh, Alexander A; Pijlman, Gorben P

    2012-12-01

    West Nile virus (WNV) and dengue virus (DENV) are highly pathogenic, mosquito-borne flaviviruses (family Flaviviridae) that cause severe disease and death in humans. WNV and DENV actively replicate in mosquitoes and human hosts and thus encounter different host immune responses. RNA interference (RNAi) is the predominant antiviral response against invading RNA viruses in insects and plants. As a countermeasure, plant and insect RNA viruses encode RNA silencing suppressor (RSS) proteins to block the generation/activity of small interfering RNA (siRNA). Enhanced flavivirus replication in mosquitoes depleted for RNAi factors suggests an important biological role for RNAi in restricting virus replication, but it has remained unclear whether or not flaviviruses counteract RNAi via expression of an RSS. First, we established that flaviviral RNA replication suppressed siRNA-induced gene silencing in WNV and DENV replicon-expressing cells. Next, we showed that none of the WNV encoded proteins displayed RSS activity in mammalian and insect cells and in plants by using robust RNAi suppressor assays. In contrast, we found that the 3'-untranslated region-derived RNA molecule known as subgenomic flavivirus RNA (sfRNA) efficiently suppressed siRNA- and miRNA-induced RNAi pathways in both mammalian and insect cells. We also showed that WNV sfRNA inhibits in vitro cleavage of double-stranded RNA by Dicer. The results of the present study suggest a novel role for sfRNA, i.e., as a nucleic acid-based regulator of RNAi pathways, a strategy that may be conserved among flaviviruses.

  11. Phage display-derived inhibitor of the essential cell wall biosynthesis enzyme MurF

    Directory of Open Access Journals (Sweden)

    Blewett Ann

    2008-12-01

    Full Text Available Abstract Background To develop antibacterial agents having novel modes of action against bacterial cell wall biosynthesis, we targeted the essential MurF enzyme of the antibiotic resistant pathogen Pseudomonas aeruginosa. MurF catalyzes the formation of a peptide bond between D-Alanyl-D-Alanine (D-Ala-D-Ala and the cell wall precursor uridine 5'-diphosphoryl N-acetylmuramoyl-L-alanyl-D-glutamyl-meso-diaminopimelic acid (UDP-MurNAc-Ala-Glu-meso-A2pm with the concomitant hydrolysis of ATP to ADP and inorganic phosphate, yielding UDP-N-acetylmuramyl-pentapeptide. As MurF acts on a dipeptide, we exploited a phage display approach to identify peptide ligands having high binding affinities for the enzyme. Results Screening of a phage display 12-mer library using purified P. aeruginosa MurF yielded to the identification of the MurFp1 peptide. The MurF substrate UDP-MurNAc-Ala-Glumeso-A2pm was synthesized and used to develop a sensitive spectrophotometric assay to quantify MurF kinetics and inhibition. MurFp1 acted as a weak, time-dependent inhibitor of MurF activity but was a potent inhibitor when MurF was pre-incubated with UDP-MurNAc-Ala-Glu-meso-A2pm or ATP. In contrast, adding the substrate D-Ala-D-Ala during the pre-incubation nullified the inhibition. The IC50 value of MurFp1 was evaluated at 250 μM, and the Ki was established at 420 μM with respect to the mixed type of inhibition against D-Ala-D-Ala. Conclusion MurFp1 exerts its inhibitory action by interfering with the utilization of D-Ala-D-Ala by the MurF amide ligase enzyme. We propose that MurFp1 exploits UDP-MurNAc-Ala-Glu-meso-A2pm-induced structural changes for better interaction with the enzyme. We present the first peptide inhibitor of MurF, an enzyme that should be exploited as a target for antimicrobial drug development.

  12. REAL-Select: full-length antibody display and library screening by surface capture on yeast cells.

    Science.gov (United States)

    Rhiel, Laura; Krah, Simon; Günther, Ralf; Becker, Stefan; Kolmar, Harald; Hock, Björn

    2014-01-01

    We describe a novel approach named REAL-Select for the non-covalent display of IgG-molecules on the surface of yeast cells for the purpose of antibody engineering and selection. It relies on the capture of secreted native full-length antibodies on the cell surface via binding to an externally immobilized ZZ domain, which tightly binds antibody Fc. It is beneficial for high-throughput screening of yeast-displayed IgG-libraries during antibody discovery and development. In a model experiment, antibody-displaying yeast cells were isolated from a 1:1,000,000 mixture with control cells confirming the maintenance of genotype-phenotype linkage. Antibodies with improved binding characteristics were obtained by affinity maturation using REAL-Select, demonstrating the ability of this system to display antibodies in their native form and to detect subtle changes in affinity by flow cytometry. The biotinylation of the cell surface followed by functionalization with a streptavidin-ZZ fusion protein is an approach that is independent of the genetic background of the antibody-producing host and therefore can be expected to be compatible with other eukaryotic expression hosts such as P. pastoris or mammalian cells.

  13. REAL-Select: full-length antibody display and library screening by surface capture on yeast cells.

    Directory of Open Access Journals (Sweden)

    Laura Rhiel

    Full Text Available We describe a novel approach named REAL-Select for the non-covalent display of IgG-molecules on the surface of yeast cells for the purpose of antibody engineering and selection. It relies on the capture of secreted native full-length antibodies on the cell surface via binding to an externally immobilized ZZ domain, which tightly binds antibody Fc. It is beneficial for high-throughput screening of yeast-displayed IgG-libraries during antibody discovery and development. In a model experiment, antibody-displaying yeast cells were isolated from a 1:1,000,000 mixture with control cells confirming the maintenance of genotype-phenotype linkage. Antibodies with improved binding characteristics were obtained by affinity maturation using REAL-Select, demonstrating the ability of this system to display antibodies in their native form and to detect subtle changes in affinity by flow cytometry. The biotinylation of the cell surface followed by functionalization with a streptavidin-ZZ fusion protein is an approach that is independent of the genetic background of the antibody-producing host and therefore can be expected to be compatible with other eukaryotic expression hosts such as P. pastoris or mammalian cells.

  14. Effector and naturally occurring regulatory T cells display no abnormalities in activation induced cell death in NOD mice.

    Directory of Open Access Journals (Sweden)

    Ayelet Kaminitz

    Full Text Available BACKGROUND: Disturbed peripheral negative regulation might contribute to evolution of autoimmune insulitis in type 1 diabetes. This study evaluates the sensitivity of naïve/effector (Teff and regulatory T cells (Treg to activation-induced cell death mediated by Fas cross-linking in NOD and wild-type mice. PRINCIPAL FINDINGS: Both effector (CD25(-, FoxP3(- and suppressor (CD25(+, FoxP3(+ CD4(+ T cells are negatively regulated by Fas cross-linking in mixed splenocyte populations of NOD, wild type mice and FoxP3-GFP trangeneess. Proliferation rates and sensitivity to Fas cross-linking are dissociated in Treg cells: fast cycling induced by IL-2 and CD3/CD28 stimulation improve Treg resistance to Fas-ligand (FasL in both strains. The effector and suppressor CD4(+ subsets display balanced sensitivity to negative regulation under baseline conditions, IL-2 and CD3/CD28 stimulation, indicating that stimulation does not perturb immune homeostasis in NOD mice. Effective autocrine apoptosis of diabetogenic cells was evident from delayed onset and reduced incidence of adoptive disease transfer into NOD.SCID by CD4(+CD25(- T cells decorated with FasL protein. Treg resistant to Fas-mediated apoptosis retain suppressive activity in vitro. The only detectable differential response was reduced Teff proliferation and upregulation of CD25 following CD3-activation in NOD mice. CONCLUSION: These data document negative regulation of effector and suppressor cells by Fas cross-linking and dissociation between sensitivity to apoptosis and proliferation in stimulated Treg. There is no evidence that perturbed AICD in NOD mice initiates or promotes autoimmune insulitis.

  15. Sorafenib analogue SC-60 induces apoptosis through the SHP-1/STAT3 pathway and enhances docetaxel cytotoxicity in triple-negative breast cancer cells.

    Science.gov (United States)

    Liu, Chun-Yu; Su, Jung-Chen; Huang, Tzu-Ting; Chu, Pei-Yi; Huang, Chun-Teng; Wang, Wan-Lun; Lee, Chia-Han; Lau, Ka-Yi; Tsai, Wen-Chun; Yang, Hsiu-Ping; Shiau, Chung-Wai; Tseng, Ling-Ming; Chen, Kuen-Feng

    2017-03-01

    Recurrent triple-negative breast cancer (TNBC) needs new therapeutic targets. Src homology region 2 domain-containing phosphatase-1 (SHP-1) can act as a tumor suppressor by dephosphorylating oncogenic kinases. One major target of SHP-1 is STAT3, which is highly activated in TNBC. In this study, we tested a sorafenib analogue SC-60, which lacks angiokinase inhibition activity, but acts as a SHP-1 agonist, in TNBC cells. SC-60 inhibited proliferation and induced apoptosis by dephosphorylating STAT3 in both a dose- and time-dependent manner in TNBC cells (MDA-MB-231, MDA-MB-468, and HCC1937). By contrast, ectopic expression of STAT3 rescued the anticancer effect induced by SC-60. SC-60 also increased the SHP-1 activity, but this effect was inhibited when the N-SH2 domain (DN1) was deleted or with SHP-1 point mutation (D61A), implying that SHP-1 is the major target of SC-60 in TNBC. The use of SC-60 in combination with docetaxel synergized the anticancer effect induced by SC-60 through the SHP-1/STAT3 pathway in TNBC cells. Importantly, SC-60 also displayed a significant antitumor effect in an MDA-MB-468 xenograft model by modulating the SHP-1/STAT3 axis, indicating the anticancer potential of SC-60 in TNBC treatment. Targeting SHP-1/p-STAT3 and the potential combination of SHP-1 agonist with chemotherapeutic docetaxel is a feasible therapeutic strategy for TNBC.

  16. RGD peptide-displaying M13 bacteriophage/PLGA nanofibers as cell-adhesive matrices for smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Shin, Yong Cheol; Lee, Jong Ho; Jin, Oh Seong; Lee, Eun Ji; Jin, Lin Hua; Kim, Chang Seok; Hong, Suck Won; Han, Dong Wook; Kim, Chun Tae; Oh, Jin Woo [Pusan National University, Busan (Korea, Republic of)

    2015-01-15

    Extracellular matrices (ECMs) are network structures that play an essential role in regulating cellular growth and differentiation. In this study, novel nanofibrous matrices were fabricated by electrospinning M13 bacteriophage and poly(lactic-co-glycolic acid) (PLGA) and were shown to be structurally and functionally similar to natural ECMs. A genetically-engineered M13 bacteriophage was constructed to display Arg-Gly-Asp (RGD) peptides on its surface. The physicochemical properties of RGD peptide-displaying M13 bacteriophage (RGD-M13 phage)/PLGA nanofibers were characterized by using scanning electron microscopy and Fourier-transform infrared spectroscopy. We used immunofluorescence staining to confirm that M13 bacteriophages were homogenously distributed in RGD-M13 phage/PLGA matrices. Furthermore, RGD-M13 phage/PLGA nanofibrous matrices, having excellent biocompatibility, can enhance the behaviors of vascular smooth muscle cells. This result suggests that RGD-M13 phage/PLGA nanofibrous matrices have potentials to serve as tissue engineering scaffolds.

  17. Amperometric L-glutamate biosensor based on bacterial cell-surface displayed glutamate dehydrogenase

    Energy Technology Data Exchange (ETDEWEB)

    Liang, Bo [Laboratory for Biosensing, Key Laboratory of Biofuels, and Shandong Provinicial Key Laboratory of Energy Genetics, Qingdao Institute of Bioenergy & Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao 266101 (China); University of Chinese Academy of Sciences, 19A Yuquan Road, Beijing 100049 (China); Zhang, Shu [Laboratory for Biosensing, Key Laboratory of Biofuels, and Shandong Provinicial Key Laboratory of Energy Genetics, Qingdao Institute of Bioenergy & Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao 266101 (China); Key Laboratory of Marine Chemistry Theory and Technology of Ministry of Education, Ocean University of China, 238 Songling Road, Qingdao 266100 (China); Lang, Qiaolin [Laboratory for Biosensing, Key Laboratory of Biofuels, and Shandong Provinicial Key Laboratory of Energy Genetics, Qingdao Institute of Bioenergy & Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao 266101 (China); Song, Jianxia; Han, Lihui [Key Laboratory of Marine Chemistry Theory and Technology of Ministry of Education, Ocean University of China, 238 Songling Road, Qingdao 266100 (China); Liu, Aihua, E-mail: liuah@qibebt.ac.cn [Laboratory for Biosensing, Key Laboratory of Biofuels, and Shandong Provinicial Key Laboratory of Energy Genetics, Qingdao Institute of Bioenergy & Bioprocess Technology, Chinese Academy of Sciences, 189 Songling Road, Qingdao 266101 (China); University of Chinese Academy of Sciences, 19A Yuquan Road, Beijing 100049 (China)

    2015-07-16

    Highlights: • E. coli surface-dispalyed Gldh exhibiting excellent enzyme activity and stability. • Sensitive amperometric biosensor for glutamate using Gldh-bacteria and MWNTs. • The glutamate biosensor exhibited high specificity and stability. - Abstract: A novel L-glutamate biosensor was fabricated using bacteria surface-displayed glutamate dehydrogenase (Gldh-bacteria). Here the cofactor NADP{sup +}-specific dependent Gldh was expressed on the surface of Escherichia coli using N-terminal region of ice nucleation protein (INP) as the anchoring motif. The cell fractionation assay and SDS-PAGE analysis indicated that the majority of INP-Gldh fusion proteins were located on the surface of cells. The biosensor was fabricated by successively casting polyethyleneimine (PEI)-dispersed multi-walled carbon nanotubes (MWNTs), Gldh-bacteria and Nafion onto the glassy carbon electrode (Nafion/Gldh-bacteria/PEI-MWNTs/GCE). The MWNTs could not only significantly lower the oxidation overpotential towards NAPDH, which was the product of NADP{sup +} involving in the oxidation of glutamate by Gldh, but also enhanced the current response. Under the optimized experimental conditions, the current–time curve of the Nafion/Gldh-bacteria/PEI-MWNTs/GCE was performed at +0.52 V (vs. SCE) by amperometry varying glutamate concentration. The current response was linear with glutamate concentration in two ranges (10 μM–1 mM and 2–10 mM). The low limit of detection was estimated to be 2 μM glutamate (S/N = 3). Moreover, the proposed biosensor is stable, specific, reproducible and simple, which can be applied to real samples detection.

  18. Selection of scFvs specific for the HepG2 cell line using ribosome display

    Indian Academy of Sciences (India)

    Lei Zhou; Wei-Ping Mao; Juan Fen; Hong-Yun Liu; Chuan-Jing Wei; Wen-Xiu Li; Feng-Yun Zhou

    2009-06-01

    The aim of this study was to construct a ribosome display library of single chain variable fragments (scFvs) associated with hepatocarcinoma and screen such a library for hepatocarcinoma-binding scFvs. mRNA was isolated from the spleens of mice immunized with hepatocellular carcinoma cell line HepG2. Heavy and k chain genes (VH and k) were amplified separately by RT-PCR, and an anti-HepG2 VH/k chain ribosome display library was constructed by assembling VH and k into the VH/k chain with a specially constructed linker by SOE-PCR. The VH/k chain library was transcribed and translated in vitro using a rabbit reticulocyte lysate system. In order to isolate specific scFvs, recognizing HepG2 negative selection on a normal hepatocyte line WRL-68 was carried out before three rounds of positive selection on HepG2. After three rounds of panning, cell enzyme-linked immunosorbent assay (ELISA) showed that one of the scFvs had high affinity for the HepG2 cell and lower affinity for the WRL-68 cell. In this study, we successfully constructed a native ribosome display library. Such a library would prove useful for direct intact cell panning using ribosome display technology. The selected scFv had a potential value for hepatocarcinoma treatment.

  19. Site-protected fixation and immobilization of Escherichia coli cells displaying surface-anchored beta-lactamase.

    Science.gov (United States)

    Freeman, A; Abramov, S; Georgiou, G

    1999-01-20

    Bacteria displaying heterologous receptors or enzymes on their surface hold great potential as whole-cell adsorbents and biocatalysts, respectively. For industrial applications, such surface-engineered cells need to be killed and chemically fixed to prevent disintegration and leakage of the displayed proteins under process conditions. It is also highly desirable to couple the chemically stabilized cells onto a solid support matrix for additional mechanical stability, flexibility in reactor choice, and easy separation from processed medium. Recently, we described the development of a readily scalable methodology for cell killing, fixation, and outer membrane stabilization via glutaraldehyde fixation followed by secondary crosslinking (Freeman, A., Abramov, S. and Georgiou, G. 1996. Biotechnol. Bioeng. 52: 625-630). Glutaraldehyde treatment was also found, however, to reduce the specific activity of a model enzyme, beta-lactamase displayed on the surface of E. coli. Here, we show that crosslinking carried out in the presence of beta-lactamase inhibitors, namely phenyl boronic acid or sodium borate, protects the active site from chemical modification resulting in up to threefold higher specific activities without affecting the cell-stabilizing effect of the glutaraldehyde treatment. To prepare an immobilized whole cell biocatalyst, residual unreacted surface aldehyde groups were employed to immobilize covalently the fixed bacteria onto chitosan-coated cellulose powder. The binding of the bacteria onto chitosan-coated cellulose was quantitative up to cell loading of 83 mg dry cell weight/g of support. Cell immobilization did not introduce mass transfer limitations and created only a modest reduction in Vmax. Thus, chemical crosslinking, affected in presence of reversible active-site inhibitors and coupled with cell immobilization on chitosan-coated cellulose represents a widely useful methodology for the process application of recombinant bacteria displaying surface

  20. Cataloging altered gene expression in young and senescent cells using enhanced differential display

    NARCIS (Netherlands)

    Linskens, Maarten H.K.; Feng, Junli; Andrews, William H.; Enlow, Brett E.; Saati, Shahin M.; Tonkin, Leath A.; Funk, Walter D.; Villeponteau, Bryant

    1995-01-01

    Recently, a novel PCR-based technique, differential display (DD), has facilitated the study of differentially expressed genes at the mRNA level. We report here an improved version of DD, which we call Enhanced Differential Display (EDD). We have modified the technique to enhance reproducibility and

  1. RGD Peptide Cell-Surface Display Enhances the Targeting and Therapeutic Efficacy of Attenuated Salmonella-mediated Cancer Therapy.

    Science.gov (United States)

    Park, Seung-Hwan; Zheng, Jin Hai; Nguyen, Vu Hong; Jiang, Sheng-Nan; Kim, Dong-Yeon; Szardenings, Michael; Min, Jung Hyun; Hong, Yeongjin; Choy, Hyon E; Min, Jung-Joon

    2016-01-01

    Bacteria-based anticancer therapies aim to overcome the limitations of current cancer therapy by actively targeting and efficiently removing cancer. To achieve this goal, new approaches that target and maintain bacterial drugs at sufficient concentrations during the therapeutic window are essential. Here, we examined the tumor tropism of attenuated Salmonella typhimurium displaying the RGD peptide sequence (ACDCRGDCFCG) on the external loop of outer membrane protein A (OmpA). RGD-displaying Salmonella strongly bound to cancer cells overexpressing αvβ3, but weakly bound to αvβ3-negative cancer cells, suggesting the feasibility of displaying a preferential homing peptide on the bacterial surface. In vivo studies revealed that RGD-displaying Salmonellae showed strong targeting efficiency, resulting in the regression in αvβ3-overexpressing cancer xenografts, and prolonged survival of mouse models of human breast cancer (MDA-MB-231) and human melanoma (MDA-MB-435). Thus, surface engineering of Salmonellae to display RGD peptides increases both their targeting efficiency and therapeutic effect.

  2. Long-Term Cultured Human Term Placenta-Derived Mesenchymal Stem Cells of Maternal Origin Displays Plasticity

    Directory of Open Access Journals (Sweden)

    Vikram Sabapathy

    2012-01-01

    Flow analysis established bonafied MSCs phenotypic characteristics, staining positively for CD29, CD73, CD90, CD105 and negatively for CD14, CD34, CD45 markers. Pluripotency of the cultured MSCs was assessed by in vitro differentiation towards not only intralineage cells like adipocytes, osteocytes, chondrocytes, and myotubules cells but also translineage differentiated towards pancreatic progenitor cells, neural cells, and retinal cells displaying plasticity. These cells did not significantly alter cell cycle or apoptosis pattern while maintaining the normal karyotype; they also have limited expression of MHC-II antigens and are Naive for stimulatory factors CD80 and CD 86. Further soft agar assays revealed that placental MSCs do not have the ability to form invasive colonies. Taking together all these characteristics into consideration, it indicates that placental MSCs could serve as good candidates for development and progress of stem-cell based therapeutics.

  3. Cell membrane modification for rapid display of proteins as a novel means of immunomodulation: FasL-decorated cells prevent islet graft rejection.

    Science.gov (United States)

    Yolcu, Esma S; Askenasy, Nadir; Singh, Narendra P; Cherradi, Salah-Eddine Lamhamedi; Shirwan, Haval

    2002-12-01

    Long-term display of exogenous proteins on the cell surface may have important research and therapeutic implications. We report a novel method for the cell-surface display of proteins that involves generation of a chimeric protein with core streptavidin, biotinylation of cells, and "decoration" with the protein. A chimeric protein with the extracellular portions of FasL (SA-FasL) was efficiently displayed on the cell surface within 2 hr without detectable cellular toxicity. Biotin and SA-FasL persisted on the cell surface for weeks in vitro and in vivo. Immunomodulation with SA-FasL-decorated splenocytes effectively blocked alloreactive responses in naive and presensitized rodents and prevented the rejection of allogeneic pancreatic islets. This approach may serve as an alternative to gene transfer-based expression with broad research and therapeutic applications.

  4. Structurally modified curcumin analogs inhibit STAT3 phosphorylation and promote apoptosis of human renal cell carcinoma and melanoma cell lines.

    Directory of Open Access Journals (Sweden)

    Matthew A Bill

    Full Text Available The Janus kinase-2 (Jak2-signal transducer and activator of transcription-3 (STAT3 pathway is critical for promoting an oncogenic and metastatic phenotype in several types of cancer including renal cell carcinoma (RCC and melanoma. This study describes two small molecule inhibitors of the Jak2-STAT3 pathway, FLLL32 and its more soluble analog, FLLL62. These compounds are structurally distinct curcumin analogs that bind selectively to the SH2 domain of STAT3 to inhibit its phosphorylation and dimerization. We hypothesized that FLLL32 and FLLL62 would induce apoptosis in RCC and melanoma cells and display specificity for the Jak2-STAT3 pathway. FLLL32 and FLLL62 could inhibit STAT3 dimerization in vitro. These compounds reduced basal STAT3 phosphorylation (pSTAT3, and induced apoptosis in four separate human RCC cell lines and in human melanoma cell lines as determined by Annexin V/PI staining. Apoptosis was also confirmed by immunoblot analysis of caspase-3 processing and PARP cleavage. Pre-treatment of RCC and melanoma cell lines with FLLL32/62 did not inhibit IFN-γ-induced pSTAT1. In contrast to FLLL32, curcumin and FLLL62 reduced downstream STAT1-mediated gene expression of IRF1 as determined by Real Time PCR. FLLL32 and FLLL62 significantly reduced secretion of VEGF from RCC cell lines in a dose-dependent manner as determined by ELISA. Finally, each of these compounds inhibited in vitro generation of myeloid-derived suppressor cells. These data support further investigation of FLLL32 and FLLL62 as lead compounds for STAT3 inhibition in RCC and melanoma.

  5. Engineering the cell surface display of cohesins for assembly of cellulosome-inspired enzyme complexes on Lactococcus lactis

    Directory of Open Access Journals (Sweden)

    Wieczorek Andrew S

    2010-09-01

    Full Text Available Abstract Background The assembly and spatial organization of enzymes in naturally occurring multi-protein complexes is of paramount importance for the efficient degradation of complex polymers and biosynthesis of valuable products. The degradation of cellulose into fermentable sugars by Clostridium thermocellum is achieved by means of a multi-protein "cellulosome" complex. Assembled via dockerin-cohesin interactions, the cellulosome is associated with the cell surface during cellulose hydrolysis, forming ternary cellulose-enzyme-microbe complexes for enhanced activity and synergy. The assembly of recombinant cell surface displayed cellulosome-inspired complexes in surrogate microbes is highly desirable. The model organism Lactococcus lactis is of particular interest as it has been metabolically engineered to produce a variety of commodity chemicals including lactic acid and bioactive compounds, and can efficiently secrete an array of recombinant proteins and enzymes of varying sizes. Results Fragments of the scaffoldin protein CipA were functionally displayed on the cell surface of Lactococcus lactis. Scaffolds were engineered to contain a single cohesin module, two cohesin modules, one cohesin and a cellulose-binding module, or only a cellulose-binding module. Cell toxicity from over-expression of the proteins was circumvented by use of the nisA inducible promoter, and incorporation of the C-terminal anchor motif of the streptococcal M6 protein resulted in the successful surface-display of the scaffolds. The facilitated detection of successfully secreted scaffolds was achieved by fusion with the export-specific reporter staphylococcal nuclease (NucA. Scaffolds retained their ability to associate in vivo with an engineered hybrid reporter enzyme, E. coli β-glucuronidase fused to the type 1 dockerin motif of the cellulosomal enzyme CelS. Surface-anchored complexes exhibited dual enzyme activities (nuclease and β-glucuronidase, and were

  6. Two-dimensional model of an AC plasma display panel cell in a neon-xenon mixture

    Energy Technology Data Exchange (ETDEWEB)

    Boeuf, J.P.; Pitchford, L.C. [Universite Paul Sabatier, Toulouse (France)

    1995-12-31

    We present a 2-D fluid model of an AC plasma display panel cell. Plasma Display Panels (PDP) are flat display devices where the light of each picture element is emitted from a plasma created by an electric discharge. In the simplest electrode configuration, AC plasma display panels consist of two glass plates, each with parallel electrodes deposited on their surfaces. The electrodes are covered with a dielectric film above which a protective MgO layer is deposited. The plates are sealed together with their electrodes at right angles, and the cap between the plates is filled with a rare gas mixture. An electric discharge can be initiated in the gas cap by applying a voltage pulse between a line electrode and a column electrode. This discharge is transient due to the dielectric layers covering the electrode: the charges deposited on the dielectric surfaces induce in the gas cap a voltage which opposes the electrode voltage. Since the electrode voltage is AC (frequency in the 10-100 kHz range), a discharge is initiated each time the electrode voltage changes sign (the voltage across the dielectric layers adds to the electrode voltage when it changes sign). The {open_quotes}ON{close_quotes} state of a picture element is therefore a succession of transient discharges. In color displays, the UV light of the discharge is used to excite phosphors in the three fundamental colors (at least three discharge cells are used for one pixel of the screen). Neon-xenon or helium-xenon mixtures are generally used in color displays where photons emitted by excited atomic (147 nm) and molecular (150 nm and 173 nm) xenon are used to excite the phosphors.

  7. Affinity isolation of antigen-specific circulating B cells for generation of phage display-derived human monoclonal antibodies

    DEFF Research Database (Denmark)

    Ditzel, Henrik

    2009-01-01

    A method is described for affinity isolation of antigen-specific circulating B cells of interest for subsequent generation of immune antibody phage display libraries. This approach should overcome the problem of low yields of monoclonal antibodies of interest in the libraries generated from...... peripheral blood lymphocytes caused by the low abundance of antigen-specific B cells in the circulation. The preselection of B cells is based on the specificity of the surface Ig receptor and is accomplished using the antigen of interest conjugated to magnetic beads. This method should significantly increase...

  8. Production of flavor esters catalyzed by CALB-displaying Pichia pastoris whole-cells in a batch reactor.

    Science.gov (United States)

    Jin, Zi; Ntwali, Janvier; Han, Shuang-Yan; Zheng, Sui-Ping; Lin, Ying

    2012-05-31

    Candida antarctica lipase B (CALB) has been employed as an efficient catalyst in the preparation of many flavor esters. A CALB-displaying yeast whole-cell biocatalyst could be an attractive alternative to commercial immobilized CALB because of its low-cost preparation and high enzymatic activity. We investigated the potential application of CALB-displaying Pichia pastoris cells for the production of flavor esters. The optimal conditions for flavor esters synthesis by this biocatalyst were determined in 50-ml shake flasks. Under optimized conditions, the synthesis of 12 kinds flavor esters were scaled up in a 5-l batch stirred reactor. Among these, the mole conversions of 10 exceeded 95% after reactions for 4h. In addition, this biocatalyst showed good tolerance for high substrates concentration and excellent operational stability. Repeated use of the cells in 10 batches resulted in an activity loss of less than 10%. Thus, CALB-displaying P. pastoris whole cells are robust biocatalysts with potential commercial application in the large-scale production of flavor esters in non-aqueous media.

  9. Identification of Peptides Inhibiting Adhesion of Monocytes to the Injured Vascular Endothelial Cells through Phage-displaying Screening

    Institute of Scientific and Technical Information of China (English)

    Yu GUO; Jia ZHANG; Ji-Cheng WANG; Feng-Xiang YAN; Bing-Yang ZHU; Hong-Lin HUANG; Duan-Fang LIAO

    2005-01-01

    Using oxidized low-density lipoprotein (LDL)-injured vascular endothelial cells (ECs) as target cells, peptides specifically binding to the injured ECs were screened from a phage-displaying peptide library by using the whole-cell screening technique after three cycles of the "adsorption-elution-amplification"procedure. Positive phage clones were identified by ELISA, and the inserted amino acid sequences in the displaying peptides were deduced from confirmation with DNA sequencing. The adhesion rate of ECs to monocytes was evaluated by cell counting. The activity of endothelial nitric oxide synthase (eNOS), and the expression levels of caveolin- 1 and intercellular adhesion molecule- 1 (ICAM- 1) were determined by Western blotting. Six positive clones specifically binding to injured ECV304 endothelial cells were selected from fourteen clones. Interestingly, four phages had peptides with tandem leucine, and two of these even shared an identical sequence. Functional analysis demonstrated that the YCPRYVRRKLENELLVL peptide shared by two clones inhibited the expression of ICAM-1, increased nitric oxide concentration in the culture media, and upregulated the expression of caveolin-1 and eNOS. As a result, the adhesion rate of monocytes to ECV304 cells was significantly reduced by 12.1%. These data suggest that the anti-adhesion effect of these novel peptides is related to the regulation of the caveolin-1/nitric oxide signal transduction pathway, and could be of use in potential therapeutic agents against certain cardiovascular diseases initiated by vascular endothelial cell damage.

  10. Microbial surface displayed enzymes based biofuel cell utilizing degradation products of lignocellulosic biomass for direct electrical energy.

    Science.gov (United States)

    Fan, Shuqin; Hou, Chuantao; Liang, Bo; Feng, Ruirui; Liu, Aihua

    2015-09-01

    In this work, a bacterial surface displaying enzyme based two-compartment biofuel cell for the direct electrical energy conversion from degradation products of lignocellulosic biomass is reported. Considering that the main degradation products of the lignocellulose are glucose and xylose, xylose dehydrogenase (XDH) displayed bacteria (XDH-bacteria) and glucose dehydrogenase (GDH) displayed bacteria (GDH-bacteria) were used as anode catalysts in anode chamber with methylene blue as electron transfer mediator. While the cathode chamber was constructed with laccase/multi-walled-carbon nanotube/glassy-carbon-electrode. XDH-bacteria exhibited 1.75 times higher catalytic efficiency than GDH-bacteria. This assembled enzymatic fuel cell exhibited a high open-circuit potential of 0.80 V, acceptable stability and energy conversion efficiency. Moreover, the maximum power density of the cell could reach 53 μW cm(-2) when fueled with degradation products of corn stalk. Thus, this finding holds great potential to directly convert degradation products of biomass into electrical energy.

  11. Power generating reflective-type liquid crystal displays using a reflective polariser and a polymer solar cell

    Science.gov (United States)

    Ho Huh, Yoon; Park, Byoungchoo

    2015-06-01

    We herein report the results of a study of a power generating reflective-type liquid crystal display (LCD), composed of a 90° twisted nematic (TN) LC cell attached to the top of a light-absorbing polymer solar cell (PSC), i.e., a Solar-LCD. The PSC consisted of a polymer bulk-heterojunction photovoltaic (PV) layer of poly[[9-(1-octylnonyl)-9H-carbazole-2,7-diyl]-2,5-thiophenediyl-2,1,3-benzothiadiazole-4,7-diyl-2,5-thiophenediyl] and [6,6]-phenyl C71 butyric acid methyl ester (PCDTBT:PCBM70), and showed a high power conversion efficiency of about 5%. In order to improve the visibility of the Solar-LCD, between the TN-LC and the PV cells we inserted a reflective polariser of a giant birefringent optical (GBO) film. The reflectivity from the Solar-LCD was observed to be considerably increased by more than 13-15% under illumination by visible light. The Solar-LCD also exhibited a significantly improved contrast ratio of more than 17-19. We believe there is a clear case for using such Solar-LCDs in new power-generating reflective-type displays; taken as a whole these results also demonstrate the possibility of their application in a number of energy-harvesting opto-electrical display devices.

  12. A novel peptide, selected from phage display library of random peptides, can efficiently target into human breast cancer cell

    Institute of Scientific and Technical Information of China (English)

    DONG Jian; LIU WeiQing; JIANG AiMei; ZHANG KeJian; CHEN MingQing

    2008-01-01

    To develop a targeting vector for breast cancer biotherapy, MDA-MB-231 cell, a human breast cancer cell line, was co-cultured with pC89 (9 aa) phage display library of random peptides. In multiple inde-pendent peptide-presenting phage screening trials, subtilisin was used as a protease to inactivate ex-tra-cellular phages. The internalized phages were collected by cell lysising and amplified in E. coli XLI-Blue. Through five rounds of selection, the peptide-presenting phages which could be internalized in MDA-MB-231 cells were isolated. A comparison was made between internalization capacities of pep-tide-presenting phages isolated from MDA-MB-231 cells and RGD-integrin binding phage by cocultur-ing them with other human tumor cell lines and normal cells. The nucleotide sequences of isolated peptide-presenting phages were then determined by DNA sequencing. To uncover whether phage coat protein or amino acid order was required for the character of the peptide to MDA-MB-231 cells, three peptides were synthesized. They are CASPSGALRSC, ASPSGALRS and CGVIFDHSVPC (the shifted sequence of CASPSGALRSC), and after coculturing them with different cell lines, their targeting ca-pacities to MDA-MB-231 cells were detected. These data suggested that the internalization process was highly selective, and capable of capturing a specific peptide from parent peptide variants. Moreover, the targeting internalization event of peptides was an amino acid sequence dependent manner. The results demonstrated the feasibility of using phage display library of random peptides to develop new targeting system for intracellular delivery of macromolecules, and the peptide we obtained might be modified as a targeting vector for breast cancer gene therapy.

  13. Carcinoma in situ testis displays permissive chromatin modifications similar to immature foetal germ cells

    DEFF Research Database (Denmark)

    Almstrup, K; Nielsen, J E; Mlynarska, O;

    2010-01-01

    The majority of testicular germ cell cancers develop through a pre-invasive carcinoma in situ (CIS) stage. The CIS cell is a neoplastic counterpart of foetal germ cells. During their development, foetal germ cells undergo extensive and essential epigenetic modifications, but little is known about...

  14. Display of GPI-anchored anti-EGFR nanobodies on extracellular vesicles promotes tumour cell targeting

    NARCIS (Netherlands)

    Kooijmans, Sander A A; Aleza, Clara Gómez; Roffler, Steve R; van Solinge, Wouter W; Vader, Pieter; Schiffelers, Raymond M

    2016-01-01

    BACKGROUND: Extracellular vesicles (EVs) are attractive candidate drug delivery systems due to their ability to functionally transport biological cargo to recipient cells. However, the apparent lack of target cell specificity of exogenously administered EVs limits their therapeutic applicability. In

  15. Enhancing thermostability of a Rhizomucor miehei lipase by engineering a disulfide bond and displaying on the yeast cell surface.

    Science.gov (United States)

    Han, Zhen-lin; Han, Shuang-yan; Zheng, Sui-ping; Lin, Ying

    2009-11-01

    To increase the thermostability of Rhizomucor miehei lipase, the software Disulfide by Design was used to engineer a novel disulfide bond between residues 96 and 106, and the corresponding double cysteine mutants were constructed. The R. miehei lipase mutant could be expressed by Pichia pastoris in a free secreted form or could be displayed on the cell surface. The new disulfide bond spontaneously formed in the mutant R. miehei lipase. Thermostability was examined by measuring of hydrolysis activity using 4-nitrophenyl caprylate as a substrate. The engineered disulfide bond contributed to thermostability in the free form of the R. miehei lipase variant. The variant displayed on the yeast cell surface had significantly increased residual hydrolytic activity in aqueous solution after incubation at 60 degrees C for 5 h and increased synthetic activity in organic solvent at 60 degrees C. These results indicated that yeast surface display might improve the stability of R. miehei lipase, as well as amplifying the thermostability through the engineered disulfide bond.

  16. Cells Lacking mtDNA Display Increased dNTP Pools upon DNA Damage

    DEFF Research Database (Denmark)

    Skovgaard, Tine; Rasmussen, Lene Juel; Munch-Petersen, Birgitte

    and mitochondrial function we have examined the effect of DNA damage on dNTP pools in cells deficient of mtDNA. We show that DNA damage induced by UV irradiation, in a dose corresponding to LD50, induces cell cycle synchronization in different human osteosarcoma cell lines. The UV pulse also has a destabilizing...

  17. An experimental and numerical investigation of flat panel display cell using magnetic fluid

    CERN Document Server

    Seo, J W; Park, S J; Lee, H S

    2002-01-01

    Optical and fluid dynamical properties of magnetic fluid have been studied experimentally and numerically using a test device with a water-base magnetite magnetic fluid. It has been found that the 3.5 mu m thick fluid film absorbs most of the incoming visible light and can be actuated fast enough to realize display devices. The computational simulation shows that the surface tension of the liquid plays the most dominant roles for the test device, and a device that can actuate the magnetic fluid magnetically is proposed.

  18. Transfected Cell Microarrays for the Expression of Membrane-Displayed Single-Chain Antibodies

    Science.gov (United States)

    2011-01-01

    T4 DNA ligase (1 U/μL) and 5× ligation buffer (Invitro- gen) were stored at –20◦C for up to 6 months (see Note 6). 10. Ligation reaction buffer...the need to purify the dephosphory- lated vector. 6. T4 DNA ligase is unstable on ice for long periods of time. It is best to keep the enzyme at –20◦C...Delehanty 10. The digested, cleaned PCR products (7 μl) were ligated to 50 ng of pDisplay vector also digested with SfiI and SalI with 1U T4 DNA

  19. A tri-copper(II) complex displaying DNA-cleaving properties and antiproliferative activity against cancer cells.

    Science.gov (United States)

    Suntharalingam, Kogularamanan; Hunt, Douglas J; Duarte, Alexandra A; White, Andrew J P; Mann, David J; Vilar, Ramon

    2012-11-19

    A new disubstituted terpyridine ligand and the corresponding tri-copper(II) complex have been prepared and characterised. The binding affinity and binding mode of this tri-copper complex (as well as the previously reported mono- and di-copper analogues) towards duplex DNA were determined by using UV/Vis spectroscopic titrations and fluorescent indicator displacement (FID) assays. These studies showed the three complexes to bind moderately (in the order of 10(4)  M(-1)) to duplex DNA (ct-DNA and a 26-mer sequence). Furthermore, the number of copper centres and the nature of the substituents were found to play a significant role in defining the binding mode (intercalative or groove binding). The nuclease potential of the three complexes was investigated by using circular plasmid DNA as a substrate and analysing the products by agarose-gel electrophoresis. The cleaving activity was found to be dependent on the number of copper centres present (cleaving potency was in the order: tri-copper>di-copper>mono-copper). Interestingly, the tri-copper complex was able to cleave DNA without the need of external co-reductants. As this complex displayed the most promising nuclease properties, cell-based studies were carried out to establish if there was a direct link between DNA cleavage and cellular toxicity. The tri-copper complex displayed high cytotoxicity against four cancer cell lines. Of particular interest was that it displayed high cytotoxicity against the cisplatin-resistant MOLT-4 leukaemia cell line. Cellular uptake studies showed that the tri-copper complex was able to enter the cell and more importantly localise in the nucleus. Immunoblotting analysis (used to monitor changes in protein levels related to the DNA damage response pathway) and DNA-flow cytometric studies suggested that this tri-copper(II) complex is able to induce cellular DNA damage.

  20. Simple display system of mechanical properties of cells and their dispersion.

    Directory of Open Access Journals (Sweden)

    Yuji Shimizu

    Full Text Available The mechanical properties of cells are unique indicators of their states and functions. Though, it is difficult to recognize the degrees of mechanical properties, due to small size of the cell and broad distribution of the mechanical properties. Here, we developed a simple virtual reality system for presenting the mechanical properties of cells and their dispersion using a haptic device and a PC. This system simulates atomic force microscopy (AFM nanoindentation experiments for floating cells in virtual environments. An operator can virtually position the AFM spherical probe over a round cell with the haptic handle on the PC monitor and feel the force interaction. The Young's modulus of mesenchymal stem cells and HEK293 cells in the floating state was measured by AFM. The distribution of the Young's modulus of these cells was broad, and the distribution complied with a log-normal pattern. To represent the mechanical properties together with the cell variance, we used log-normal distribution-dependent random number determined by the mode and variance values of the Young's modulus of these cells. The represented Young's modulus was determined for each touching event of the probe surface and the cell object, and the haptic device-generating force was calculated using a Hertz model corresponding to the indentation depth and the fixed Young's modulus value. Using this system, we can feel the mechanical properties and their dispersion in each cell type in real time. This system will help us not only recognize the degrees of mechanical properties of diverse cells but also share them with others.

  1. The integration of novel EAP-based Braille cells for use in a refreshable tactile display

    Science.gov (United States)

    Di Spigna, N.; Chakraborti, P.; Winick, D.; Yang, P.; Ghosh, T.; Franzon, P.

    2010-04-01

    Structures demonstrating the viability of both the hydraulic and latching Braille dot, and the dielectric elastomer fiber Braille dot have been fabricated and characterized. A hydraulic proof-of-concept structure has achieved the necessary volumetric change required to lift a Braille dot over 0.5mm at voltages under 1000V and at speeds under 100ms. Long bimorphs have been fabricated that demonstrate large tip displacements over 2mm that could be used to mechanically latch the Braille rod in the 'up' position to achieve the force requirement. The addition of radial prestrain in dielectric elastomer tubes has reduced the wall thickness and directed the strain in the axial direction which has had a dramatic impact on their resulting characteristics. The required bias voltage for the dielectric elastomer fiber Braille dot has been reduced from 15.5kV to 8.75kV while the Braille head tip displacement of a fabricated prototype has almost tripled on average and now also exceeds the required displacement for a refreshable Braille display. Finally, potential solutions to the current shortcomings of both designs in meeting all of the requirements for such a display are discussed.

  2. HCMV Displays a Unique Transcriptome of Immunomodulatory Genes in Primary Monocyte-Derived Cell Types

    Science.gov (United States)

    Van Damme, Ellen; Thys, Kim; Tuefferd, Marianne; Van Hove, Carl; Aerssens, Jeroen; Van Loock, Marnix

    2016-01-01

    Human cytomegalovirus (HCMV) is a betaherpesvirus which rarely presents problems in healthy individuals, yet may result in severe morbidity in immunocompromised patients and in immune-naïve neonates. HCMV has a large 235 kb genome with a coding capacity of at least 165 open reading frames (ORFs). This large genome allows complex gene regulation resulting in different sets of transcripts during lytic and latent infection. While latent virus mainly resides within monocytes and CD34+ progenitor cells, reactivation to lytic infection is driven by differentiation towards terminally differentiated myeloid dendritic cells and macrophages. Consequently, it has been suggested that macrophages and dendritic cells contribute to viral spread in vivo. Thus far only limited knowledge is available on the expression of HCMV genes in terminally differentiated myeloid primary cells and whether or not the virus exhibits a different set of lytic genes in primary cells compared with lytic infection in NHDF fibroblasts. To address these questions, we used Illumina next generation sequencing to determine the HCMV transcriptome in macrophages and dendritic cells during lytic infection and compared it to the transcriptome in NHDF fibroblasts. Here, we demonstrate unique expression profiles in macrophages and dendritic cells which significantly differ from the transcriptome in fibroblasts mainly by modulating the expression of viral transcripts involved in immune modulation, cell tropism and viral spread. In a head to head comparison between macrophages and dendritic cells, we observed that factors involved in viral spread and virion composition are differentially regulated suggesting that the plasticity of the virion facilitates the infection of surrounding cells. Taken together, this study provides the full transcript expression analysis of lytic HCMV genes in monocyte-derived type 1 and type 2 macrophages as well as in monocyte-derived dendritic cells. Thereby underlining the potential

  3. Displaying the researched body: Growing cell portraits in a medical museum

    DEFF Research Database (Denmark)

    Whiteley, Louise Emma; Tybjerg, Karin; Pedersen, Bente Vinge

    2016-01-01

    In Heirloom, artist Gina Czarnecki and scientist John Hunt grow portraits of the artist’s daughters from their own cells, onto glass casts of their faces. This required the development of novel scien- tific techniques to allow the growth of human cells in a gallery. Heirloom was exhibited...

  4. Cells Lacking mtDNA Display Increased dNTP Pools upon DNA Damage

    DEFF Research Database (Denmark)

    Skovgaard, Tine; Rasmussen, Lene Juel; Munch-Petersen, Birgitte

    and mitochondrial function we have examined the effect of DNA damage on dNTP pools in cells deficient of mtDNA. We show that DNA damage induced by UV irradiation, in a dose corresponding to LD50, induces an S phase delay in different human osteosarcoma cell lines. The UV pulse also has a destabilizing effect...

  5. Cardiomyocytes display low mitochondrial priming and are highly resistant toward cytotoxic T‐cell killing

    Science.gov (United States)

    Zheng, Xiang; Halle, Stephan; Yu, Kai; Mishra, Pooja; Scherr, Michaela; Pietzsch, Stefan; Willenzon, Stefanie; Janssen, Anika; Boelter, Jasmin; Hilfiker‐Kleiner, Denise; Eder, Matthias

    2016-01-01

    Following heart transplantation, alloimmune responses can cause graft rejection by damaging donor vascular and parenchymal cells. However, it remains unclear whether cardiomyocytes are also directly killed by immune cells. Here, we used two‐photon microscopy to investigate how graft‐specific effector CD8+ T cells interact with cardiomyocytes in a mouse heart transplantation model. Surprisingly, we observed that CD8+ T cells are completely impaired in killing cardiomyocytes. Even after virus‐mediated preactivation, antigen‐specific CD8+ T cells largely fail to lyse these cells although both cell types engage in dynamic interactions. Furthermore, we established a two‐photon microscopy‐based assay using intact myocardium to determine the susceptibility of cardiomyocytes to undergo apoptosis. This feature, also known as mitochondrial priming reveals an unexpected weak predisposition of cardiomyocytes to undergo apoptosis in situ. These observations together with the early exhaustion phenotype of graft‐infiltrating specific T cells provide an explanation why cardiomyocytes are largely protected from direct CD8+ T‐cell‐mediated killing. PMID:26970349

  6. Flexoelectric effect in an in-plane switching (IPS) liquid crystal cell for low-power consumption display devices

    Science.gov (United States)

    Kim, Min Su; Bos, Philip J.; Kim, Dong-Woo; Yang, Deng-Ke; Lee, Joong Hee; Lee, Seung Hee

    2016-10-01

    Technology of displaying static images in portable displays, advertising panels and price tags pursues significant reduction in power consumption and in product cost. Driving at a low-frequency electric field in fringe-field switching (FFS) mode can be one of the efficient ways to save powers of the recent portable devices, but a serious drop of image-quality, so-called image-flickering, has been found in terms of the coupling of elastic deformation to not only quadratic dielectric effect but linear flexoelectric effect. Despite of the urgent requirement of solving the issue, understanding of such a phenomenon is yet vague. Here, we thoroughly analyze and firstly report the flexoelectric effect in in-plane switching (IPS) liquid crystal cell. The effect takes place on the area above electrodes due to splay and bend deformations of nematic liquid crystal along oblique electric fields, so that the obvious spatial shift of the optical transmittance is experimentally observed and is clearly demonstrated based on the relation between direction of flexoelectric polarization and electric field polarity. In addition, we report that the IPS mode has inherent characteristics to solve the image-flickering issue in the low-power consumption display in terms of the physical property of liquid crystal material and the electrode structure.

  7. The Anoikis Effector Bit1 Displays Tumor Suppressive Function in Lung Cancer Cells

    OpenAIRE

    Xin Yao; Scott Jennings; Shubha Kale Ireland; Tri Pham; Brandi Temple; Mya Davis; Renwei Chen; Ian Davenport; Hector Biliran

    2014-01-01

    The mitochondrial Bit1 (Bcl-2 inhibitor of transcription 1) protein is a part of an apoptotic pathway that is uniquely regulated by integrin-mediated attachment. As an anoikis effector, Bit1 is released into the cytoplasm following loss of cell attachment and induces a caspase-independent form of apoptosis. Considering that anoikis resistance is a critical determinant of transformation, we hypothesized that cancer cells may circumvent the Bit1 apoptotic pathway to attain anchorage-independenc...

  8. Green tea polyphenol tailors cell adhesivity of RGD displaying surfaces: multicomponent models monitored optically

    Science.gov (United States)

    Peter, Beatrix; Farkas, Eniko; Forgacs, Eniko; Saftics, Andras; Kovacs, Boglarka; Kurunczi, Sandor; Szekacs, Inna; Csampai, Antal; Bosze, Szilvia; Horvath, Robert

    2017-01-01

    The interaction of the anti-adhesive coating, poly(L-lysine)-graft-poly(ethylene glycol) (PLL-g-PEG) and its Arg-Gly-Asp (RGD) functionalized form, PLL-g-PEG-RGD, with the green tea polyphenol, epigallocatechin-gallate (EGCg) was in situ monitored. After, the kinetics of cellular adhesion on the EGCg exposed coatings were recorded in real-time. The employed plate-based waveguide biosensor is applicable to monitor small molecule binding and sensitive to sub-nanometer scale changes in cell membrane position and cell mass distribution; while detecting the signals of thousands of adhering cells. The combination of this remarkable sensitivity and throughput opens up new avenues in testing complicated models of cell-surface interactions. The systematic studies revealed that, despite the reported excellent antifouling properties of the coatings, EGCg strongly interacted with them, and affected their cell adhesivity in a concentration dependent manner. Moreover, the differences between the effects of the fresh and oxidized EGCg solutions were first demonstrated. Using a semiempirical quantumchemical method we showed that EGCg binds to the PEG chains of PLL-g-PEG-RGD and effectively blocks the RGD sites by hydrogen bonds. The calculations supported the experimental finding that the binding is stronger for the oxidative products. Our work lead to a new model of polyphenol action on cell adhesion ligand accessibility and matrix rigidity. PMID:28186133

  9. Adipose mesenchymal stem cells isolated after manual or water jet-assisted liposuction display similar properties

    Directory of Open Access Journals (Sweden)

    Claire eBony

    2016-01-01

    Full Text Available Mesenchymal stem or stromal cells (MSC are under investigation in many clinical trials for their therapeutic potential in a variety of diseases, including autoimmune and inflammatory disorders. One of the main sources of MSCs is the adipose tissue, which is mainly obtained by manual liposuction using a cannula linked to a syringe. However, in the last years, a number of devices for fat liposuction intended for clinical use have been commercialized but few papers have compared these procedures in terms of stromal vascular fraction (SVF or adipose stromal cells (ASC. The objective of the present study was to compare and qualify for clinical use the adipose stromal cells (ASC obtained from fat isolated with the manual or the Bodyjet® waterjet-assisted procedure. Although the initial number of cells after collagenase digestion was higher with the manual procedure, both the percentage of dead cells, the number of CFU-F and the phenotype of cells were identical in the SVF at isolation and in the ASC populations at day 14. We also showed that the osteogenic and adipogenic differentiation potentials of ASCs were identical between preparations while a slight but significant higher in vitro immunosuppressive effect was observed with ASCs isolated from fat removed with a cannula. The difference in the immunomodulatory effect between ASC populations was however not observed in vivo using the delayed-type hypersensitivity model. Our data therefore indicate that the procedure for fat liposuction does not impact the characteristics or the therapeutic function of ASCs.

  10. Coxsackievirus B exits the host cell in shed microvesicles displaying autophagosomal markers.

    Directory of Open Access Journals (Sweden)

    Scott M Robinson

    2014-04-01

    Full Text Available Coxsackievirus B3 (CVB3, a member of the picornavirus family and enterovirus genus, causes viral myocarditis, aseptic meningitis, and pancreatitis in humans. We genetically engineered a unique molecular marker, "fluorescent timer" protein, within our infectious CVB3 clone and isolated a high-titer recombinant viral stock (Timer-CVB3 following transfection in HeLa cells. "Fluorescent timer" protein undergoes slow conversion of fluorescence from green to red over time, and Timer-CVB3 can be utilized to track virus infection and dissemination in real time. Upon infection with Timer-CVB3, HeLa cells, neural progenitor and stem cells (NPSCs, and C2C12 myoblast cells slowly changed fluorescence from green to red over 72 hours as determined by fluorescence microscopy or flow cytometric analysis. The conversion of "fluorescent timer" protein in HeLa cells infected with Timer-CVB3 could be interrupted by fixation, suggesting that the fluorophore was stabilized by formaldehyde cross-linking reactions. Induction of a type I interferon response or ribavirin treatment reduced the progression of cell-to-cell virus spread in HeLa cells or NPSCs infected with Timer-CVB3. Time lapse photography of partially differentiated NPSCs infected with Timer-CVB3 revealed substantial intracellular membrane remodeling and the assembly of discrete virus replication organelles which changed fluorescence color in an asynchronous fashion within the cell. "Fluorescent timer" protein colocalized closely with viral 3A protein within virus replication organelles. Intriguingly, infection of partially differentiated NPSCs or C2C12 myoblast cells induced the release of abundant extracellular microvesicles (EMVs containing matured "fluorescent timer" protein and infectious virus representing a novel route of virus dissemination. CVB3 virions were readily observed within purified EMVs by transmission electron microscopy, and infectious virus was identified within low-density isopycnic

  11. The anoikis effector Bit1 displays tumor suppressive function in lung cancer cells.

    Directory of Open Access Journals (Sweden)

    Xin Yao

    Full Text Available The mitochondrial Bit1 (Bcl-2 inhibitor of transcription 1 protein is a part of an apoptotic pathway that is uniquely regulated by integrin-mediated attachment. As an anoikis effector, Bit1 is released into the cytoplasm following loss of cell attachment and induces a caspase-independent form of apoptosis. Considering that anoikis resistance is a critical determinant of transformation, we hypothesized that cancer cells may circumvent the Bit1 apoptotic pathway to attain anchorage-independence and tumorigenic potential. Here, we provide the first evidence of the tumor suppressive effect of Bit1 through a mechanism involving anoikis induction in human lung adenocarcinoma derived A549 cells. Restitution of Bit1 in anoikis resistant A549 cells is sufficient to induce detachment induced-apoptosis despite defect in caspase activation and impairs their anchorage-independent growth. Conversely, stable downregulation of Bit1 in these cells significantly enhances their anoikis resistance and anchorage-independent growth. The Bit1 knockdown cells exhibit significantly enhanced tumorigenecity in vivo. It has been previously shown that the nuclear TLE1 corepressor is a putative oncogene in lung cancer, and we show here that TLE1 blocks Bit1 mediated anoikis in part by sequestering the pro-apoptotic partner of Bit1, the Amino-terminal Enhancer of Split (AES protein, in the nucleus. Taken together, these findings suggest a tumor suppressive role of the caspase-independent anoikis effector Bit1 in lung cancer. Consistent with its role as a tumor suppressor, we have found that Bit1 is downregulated in human non-small cell lung cancer (NSCLC tissues.

  12. The anoikis effector Bit1 displays tumor suppressive function in lung cancer cells.

    Science.gov (United States)

    Yao, Xin; Jennings, Scott; Ireland, Shubha Kale; Pham, Tri; Temple, Brandi; Davis, Mya; Chen, Renwei; Davenport, Ian; Biliran, Hector

    2014-01-01

    The mitochondrial Bit1 (Bcl-2 inhibitor of transcription 1) protein is a part of an apoptotic pathway that is uniquely regulated by integrin-mediated attachment. As an anoikis effector, Bit1 is released into the cytoplasm following loss of cell attachment and induces a caspase-independent form of apoptosis. Considering that anoikis resistance is a critical determinant of transformation, we hypothesized that cancer cells may circumvent the Bit1 apoptotic pathway to attain anchorage-independence and tumorigenic potential. Here, we provide the first evidence of the tumor suppressive effect of Bit1 through a mechanism involving anoikis induction in human lung adenocarcinoma derived A549 cells. Restitution of Bit1 in anoikis resistant A549 cells is sufficient to induce detachment induced-apoptosis despite defect in caspase activation and impairs their anchorage-independent growth. Conversely, stable downregulation of Bit1 in these cells significantly enhances their anoikis resistance and anchorage-independent growth. The Bit1 knockdown cells exhibit significantly enhanced tumorigenecity in vivo. It has been previously shown that the nuclear TLE1 corepressor is a putative oncogene in lung cancer, and we show here that TLE1 blocks Bit1 mediated anoikis in part by sequestering the pro-apoptotic partner of Bit1, the Amino-terminal Enhancer of Split (AES) protein, in the nucleus. Taken together, these findings suggest a tumor suppressive role of the caspase-independent anoikis effector Bit1 in lung cancer. Consistent with its role as a tumor suppressor, we have found that Bit1 is downregulated in human non-small cell lung cancer (NSCLC) tissues.

  13. Display of phytase on the cell surface of Saccharomyces cerevisiae to degrade phytate phosphorus and improve bioethanol production.

    Science.gov (United States)

    Chen, Xianzhong; Xiao, Yan; Shen, Wei; Govender, Algasan; Zhang, Liang; Fan, You; Wang, Zhengxiang

    2016-03-01

    Currently, development of biofuels as an alternative fuel has gained much attention due to resource and environmental challenges. Bioethanol is one of most important and dominant biofuels, and production using corn or cassava as raw materials has become a prominent technology. However, phytate contained in the raw material not only decreases the efficiency of ethanol production, but also leads to an increase in the discharge of phosphorus, thus impacting on the environment. In this study, to decrease phytate and its phosphorus content in an ethanol fermentation process, Saccharomyces cerevisiae was engineered through a surface-displaying system utilizing the C-terminal half of the yeast α-agglutinin protein. The recombinant yeast strain, PHY, was constructed by successfully displaying phytase on the surface of cells, and enzyme activity reached 6.4 U/g wet biomass weight. Ethanol productions using various strains were compared, and the results demonstrated that the specific growth rate and average fermentation rate of the PHY strain were higher 20 and 18 %, respectively, compared to the control strain S. cerevisiae CICIMY0086, in a 5-L bioreactor process by simultaneous saccharification and fermentation. More importantly, the phytate phosphorus concentration decreased by 89.8 % and free phosphorus concentration increased by 142.9 % in dry vinasse compared to the control in a 5-L bioreactor. In summary, we constructed a recombinant S. cerevisiae strain displaying phytase on the cell surface, which could improve ethanol production performance and effectively reduce the discharge of phosphorus. The strain reported here represents a useful novel engineering platform for developing an environment-friendly system for bioethanol production from a corn substrate.

  14. Autosomal mutants of proton-exposed kidney cells display frequent loss of heterozygosity on nonselected chromosomes.

    Science.gov (United States)

    Grygoryev, Dmytro; Dan, Cristian; Gauny, Stacey; Eckelmann, Bradley; Ohlrich, Anna P; Connolly, Marissa; Lasarev, Michael; Grossi, Gianfranco; Kronenberg, Amy; Turker, Mitchell S

    2014-05-01

    High-energy protons found in the space environment can induce mutations and cancer, which are inextricably linked. We hypothesized that some mutants isolated from proton-exposed kidneys arose through a genome-wide incident that causes loss of heterozygosity (LOH)-generating mutations on multiple chromosomes (termed here genomic LOH). To test this hypothesis, we examined 11 pairs of nonselected chromosomes for LOH events in mutant cells isolated from the kidneys of mice exposed to 4 or 5 Gy of 1 GeV protons. The mutant kidney cells were selected for loss of expression of the chromosome 8-encoded Aprt gene. Genomic LOH events were also assessed in Aprt mutants isolated from isogenic cultured kidney epithelial cells exposed to 5 Gy of protons in vitro. Control groups were spontaneous Aprt mutants and clones isolated without selection from the proton-exposed kidneys or cultures. The in vivo results showed significant increases in genomic LOH events in the Aprt mutants from proton-exposed kidneys when compared with spontaneous Aprt mutants and when compared with nonmutant (i.e., nonselected) clones from the proton-exposed kidneys. A bias for LOH events affecting chromosome 14 was observed in the proton-induced Aprt mutants, though LOH for this chromosome did not confer increased radiation resistance. Genomic LOH events were observed in Aprt mutants isolated from proton-exposed cultured kidney cells; however the incidence was fivefold lower than in Aprt mutants isolated from exposed intact kidneys, suggesting a more permissive environment in the intact organ and/or the evolution of kidney clones prior to their isolation from the tissue. We conclude that proton exposure creates a subset of viable cells with LOH events on multiple chromosomes, that these cells form and persist in vivo, and that they can be isolated from an intact tissue by selection for a mutation on a single chromosome.

  15. Ferrocenyl chalcone difluoridoborates inhibit HIV-1 integrase and display low activity towards cancer and endothelial cells.

    Science.gov (United States)

    Monserrat, Jean-Philippe; Al-Safi, Rasha I; Tiwari, Keshri Nath; Quentin, Lionel; Chabot, Guy G; Vessières, Anne; Jaouen, Gérard; Neamati, Nouri; Hillard, Elizabeth A

    2011-10-15

    We report here the discovery of a potent series of HIV-1 integrase (IN) inhibitors based on the ferrocenyl chalcone difluoridoborate structure. Ten new compounds have been synthesized and were generally found to have similar inhibitory activities against the IN 3' processing and strand transfer (ST) processes. IC(50) values were found to be in the low micromolar range, and significantly lower than those found for the non-coordinated ferrocenyl chalcones and other ferrocene molecules. The ferrocenyl chalcone difluoridoborates furthermore exhibited low cytotoxicity against cancer cells and low morphological activity against epithelial cells.

  16. Multivalent display of quinic acid based ligands for targeting E-selectin expressing cells.

    Science.gov (United States)

    Shamay, Yosi; Paulin, Denise; Ashkenasy, Gonen; David, Ayelet

    2009-10-08

    The site-specific expression of molecular markers on endothelial cells of blood vessels during inflammatory response and angiogenesis provides an opportunity to target drugs and imaging molecules to the vascular endothelium of diseased tissues. This paper describes an innovative strategy for selective delivery of polymer conjugates to E- and P-selectin expressing cells using a series of quinic acid (Qa) based non-carbohydrate analogues of the natural ligand sialyl Lewis(x) (sLe(x)) as targeting moieties. We demonstrate that such analogues antagonize the adhesion of sLe(x) expressing HL-60 cells to both E- and P-selectin. Significantly, the apparent avidity of polymer conjugates carrying multiple Qa copies has increased by 3 orders of magnitude relative to their monomeric forms. Furthermore, we found that the major mechanism of copolymer entry and delivery into E-selectin expressing cells is endocytosis. These selectin-targetable copolymers provide the foundation to support controlled delivery of anticancer drugs and imaging agents to tumor vasculature for therapeutic and diagnostic applications.

  17. Phenotypically Dormant and Immature Leukaemia Cells Display Increased Ribosomal Protein S6 Phosphorylation.

    Directory of Open Access Journals (Sweden)

    Monica Pallis

    Full Text Available Mechanistic/mammalian target of rapamycin (mTOR activity drives a number of key metabolic processes including growth and protein synthesis. Inhibition of the mTOR pathway promotes cellular dormancy. Since cells from patients with acute myeloid leukaemia (AML can be phenotypically dormant (quiescent, we examined biomarkers of their mTOR pathway activity concurrently with Ki-67 and CD71 (indicators of cycling cells by quantitative flow cytometry. Using antibodies to phosphorylated epitopes of mTOR (S2448 and its downstream targets ribosomal protein S6 (rpS6, S235/236 and 4E-BP1 (T36/45, we documented that these phosphorylations were negligible in lymphocytes, but evident in dormant as well as proliferating subsets of both mobilised normal stem cell harvest CD34+ cells and AML blasts. Although mTOR phosphorylation in AML blasts was lower than that of the normal CD34+ cells, p-4E-BP1 was 2.6-fold higher and p-rpS6 was 22-fold higher. Moreover, in contrast to 4E-BP1, rpS6 phosphorylation was higher in dormant than proliferating AML blasts, and was also higher in the immature CD34+CD38- blast subset. Data from the Cancer Genome Atlas show that rpS6 expression is associated with that of respiratory chain enzymes in AML. We conclude that phenotypic quiescence markers do not necessarily predict metabolic dormancy and that elevated rpS6 ser235/236 phosphorylation is characteristic of AML.

  18. Pluripotent and Multipotent Stem Cells Display Distinct Hypoxic miRNA Expression Profiles

    Science.gov (United States)

    Agrawal, Rahul; Dale, Tina P.; Al-Zubaidi, Mohammed A.; Benny Malgulwar, Prit; Forsyth, Nicholas R.; Kulshreshtha, Ritu

    2016-01-01

    MicroRNAs are reported to have a crucial role in the regulation of self-renewal and differentiation of stem cells. Hypoxia has been identified as a key biophysical element of the stem cell culture milieu however, the link between hypoxia and miRNA expression in stem cells remains poorly understood. We therefore explored miRNA expression in hypoxic human embryonic and mesenchymal stem cells (hESCs and hMSCs). A total of 50 and 76 miRNAs were differentially regulated by hypoxia (2% O2) in hESCs and hMSCs, respectively, with a negligible overlap of only three miRNAs. We found coordinate regulation of precursor and mature miRNAs under hypoxia suggesting their regulation mainly at transcriptional level. Hypoxia response elements were located upstream of 97% of upregulated hypoxia regulated miRNAs (HRMs) suggesting hypoxia-inducible-factor (HIF) driven transcription. HIF binding to the candidate cis-elements of specific miRNAs under hypoxia was confirmed by Chromatin immunoprecipitation coupled with qPCR. Role analysis of a subset of upregulated HRMs identified linkage to reported inhibition of differentiation while a downregulated subset of HRMs had a putative role in the promotion of differentiation. MiRNA-target prediction correlation with published hypoxic hESC and hMSC gene expression profiles revealed HRM target genes enriched in the cytokine:cytokine receptor, HIF signalling and pathways in cancer. Overall, our study reveals, novel and distinct hypoxia-driven miRNA signatures in hESCs and hMSCs with the potential for application in optimised culture and differentiation models for both therapeutic application and improved understanding of stem cell biology. PMID:27783707

  19. Adipose Mesenchymal Stem Cells Isolated after Manual or Water-jet-Assisted Liposuction Display Similar Properties

    Science.gov (United States)

    Bony, Claire; Cren, Mailys; Domergue, Sophie; Toupet, Karine; Jorgensen, Christian; Noël, Danièle

    2016-01-01

    Mesenchymal stem or stromal cells (MSC) are under investigation in many clinical trials for their therapeutic potential in a variety of diseases, including autoimmune and inflammatory disorders. One of the main sources of MSCs is the adipose tissue, which is mainly obtained by manual liposuction using a cannula linked to a syringe. However, in the past years, a number of devices for fat liposuction intended for clinical use have been commercialized but few papers have compared these procedures in terms of stromal vascular fraction (SVF) or adipose mesenchymal stromal cells (ASC). The objective of the present study was to compare and qualify for clinical use the ASC obtained from fat isolated with the manual or the Bodyjet® water-jet-assisted procedure. Although the initial number of cells obtained after collagenase digestion was higher with the manual procedure, the percentage of dead cells, the number of colony forming unit-fibroblast and the phenotype of cells were identical in the SVF at isolation (day 0) and in the ASC populations at day 14. We also showed that the osteogenic and adipogenic differentiation potentials of ASCs were identical between preparations while a slight but significant higher in vitro immunosuppressive effect was observed with ASCs isolated from fat removed with a cannula. The difference in the immunomodulatory effect between ASC populations was, however, not observed in vivo using the delayed-type hypersensitivity (DTH) model. Our data, therefore, indicate that the procedure for fat liposuction does not impact the characteristics or the therapeutic function of ASCs. PMID:26834736

  20. Post-storage cell wall metabolism in two sweet cherry (Prunus avium L.) cultivars displaying different postharvest performance.

    Science.gov (United States)

    Belge, Burcu; Comabella, Eva; Graell, Jordi; Lara, Isabel

    2015-09-01

    The biochemical processes underlying firmness loss of sweet cherry (Prunus avium L.) fruit are poorly understood. Studies on cell wall metabolism of sweet cherry have been generally undertaken during on-tree development or at harvest maturity, while published reports on postharvest changes are scarce and fragmentary. In this work, cell wall modifications after storage at 0 ℃ were studied in two cherry cultivars ('Celeste' and 'Somerset') displaying different postharvest potential. Firmness was largely determined by the yields of the Na2CO3- and KOH-soluble fractions, enriched in covalently-bound pectins and in matrix glycans, respectively, and correlated well with ascorbic acid contents. The yields of these two cell wall fractions were correlated inversely with pectinmethylesterase and endo-1,4-β-d-glucanase activities, indicating a relevant role of these two enzymes in postharvest firmness changes in sweet cherry. The amount of solubilised cell wall materials was closely associated to the contents of dehydroascorbic acid, suggesting the possible involvement of oxidative mechanisms in cell wall disassembly. These data may help understanding the evolution of fruit quality during the marketing period, and give hints for the design of suitable management strategies to preserve key attributes.

  1. Zika Virus Strains Potentially Display Different Infectious Profiles in Human Neural Cells

    Directory of Open Access Journals (Sweden)

    Yannick Simonin

    2016-10-01

    Full Text Available The recent Zika virus (ZIKV epidemic has highlighted the poor knowledge on its physiopathology. Recent studies showed that ZIKV of the Asian lineage, responsible for this international outbreak, causes neuropathology in vitro and in vivo. However, two African lineages exist and the virus is currently found circulating in Africa. The original African strain was also suggested to be neurovirulent but its laboratory usage has been criticized due to its multiple passages. In this study, we compared the French Polynesian (Asian ZIKV strain to an African strain isolated in Central African Republic and show a difference in infectivity and cellular response between both strains in human neural stem cells and astrocytes. Consistently, this African strain led to a higher infection rate and viral production, as well as stronger cell death and anti-viral response. Our results highlight the need to better characterize the physiopathology and predict neurological impairment associated with African ZIKV.

  2. Gaucher Disease-Induced Pluripotent Stem Cells Display Decreased Erythroid Potential and Aberrant Myelopoiesis.

    Science.gov (United States)

    Sgambato, Judi A; Park, Tea Soon; Miller, Diana; Panicker, Leelamma M; Sidransky, Ellen; Lun, Yu; Awad, Ola; Bentzen, Søren M; Zambidis, Elias T; Feldman, Ricardo A

    2015-08-01

    Gaucher disease (GD) is the most common lysosomal storage disease resulting from mutations in the lysosomal enzyme glucocerebrosidase (GCase). The hematopoietic abnormalities in GD include the presence of characteristic Gaucher macrophages that infiltrate patient tissues and cytopenias. At present, it is not clear whether these cytopenias are secondary to the pathological activity of Gaucher cells or a direct effect of GCase deficiency on hematopoietic development. To address this question, we differentiated induced pluripotent stem cells (iPSCs) derived from patients with types 1, 2, and 3 GD to CD34(+)/CD45(+)/CD43(+)/CD143(+) hematopoietic progenitor cells (HPCs) and examined their developmental potential. The formation of GD-HPCs was unaffected. However, these progenitors demonstrated a skewed lineage commitment, with increased myeloid differentiation and decreased erythroid differentiation and maturation. Interestingly, myeloid colony-formation assays revealed that GD-HPCs, but not control-HPCs, gave rise to adherent, macrophage-like cells, another indication of abnormal myelopoiesis. The extent of these hematologic abnormalities correlated with the severity of the GCase mutations. All the phenotypic abnormalities of GD-HPCs observed were reversed by incubation with recombinant GCase, indicating that these developmental defects were caused by the mutated GCase. Our results show that GCase deficiency directly impairs hematopoietic development. Additionally, our results suggest that aberrant myelopoiesis might contribute to the pathological properties of Gaucher macrophages, which are central to GD manifestations. The hematopoietic developmental defects we observed reflect hematologic abnormalities in patients with GD, demonstrating the utility of GD-iPSCs for modeling this disease.

  3. Human Muscle Progenitor Cells Displayed Immunosuppressive Effect through Galectin-1 and Semaphorin-3A

    Directory of Open Access Journals (Sweden)

    Séverine Lecourt

    2012-01-01

    Full Text Available In human skeletal muscle, myoblasts represent the main population of myogenic progenitors. We previously showed that, beside their myogenic differentiation capacities, myoblasts also differentiate towards osteogenic and chondrogenic lineages, some properties generally considered being hallmarks of mesenchymal stem cells (MSCs. MSCs are also characterized by their immunosuppressive potential, through cell-cell contacts and soluble factors, including prostaglandin E-2 (PGE-2, transforming growth factor-β1 (TGF-β1, interleukine-10, or indoleamine 2,3-dioxygenase. We and others also reported that Galectin-1 (Gal-1 and Semaphorin-3A (Sema-3A were involved in MSCs-mediated immunosuppression. Here, we show that human myoblasts induce a significant and dose-dependant proliferation inhibition, independently of PGE-2 and TGF-β1. Our experiments revealed that myoblasts, in culture or in situ in human muscles, expressed and secreted Gal-1 and Sema-3A. Furthermore, myoblasts immunosuppressive functions were reverted by using blocking antibodies against Gal-1 or Sema-3A. Together, these results demonstrate an unsuspected immunosuppressive effect of myoblasts that may open new therapeutic perspectives.

  4. K562 cells display different vulnerability to H₂O₂ induced oxidative stress in differing cell cycle phases.

    Science.gov (United States)

    Akcakaya, Handan; Dal, Fulya; Tok, Sabiha; Cinar, Suzan-Adin; Nurten, Rustem

    2015-02-01

    Oxidative stress can be defined as the increase of oxidizing agents like reactive oxygen and nitrogen species, or the imbalance between the antioxidative defense mechanism and oxidants. Cell cycle checkpoint response can be defined as the arrest of the cell cycle functioning after damaging chemical exposure. This temporary arrest may be a period of time given to the cells to repair the DNA damage before entering the cycle again and completing mitosis. In order to determine the effects of oxidative stress on several cell cycle phases, human erytroleukemia cell line (K562) was synchronized with mimosine and genistein, and cell cycle analysis carried out. Synchronized cells were exposed to oxidative stress with hydrogen peroxide (H2O2) at several concentrations and different times. Changes on mitochondria membrane potential (ΔΨm) of K562 cells were analyzed in G1, S, and G2 /M using Rhodamine 123 (Rho 123). To determine apoptosis and necrosis, stressed cells were stained with Annexin V (AnnV) and propidium iodide (PI) for flow cytometry. Changes were observed in the ΔΨm of synchronized and asynchronized cells that were exposed to oxidative stress. Synchronized cells in S phase proved resistant to the effects of oxidative stress and synchronized cells at G2 /M phase were sensitive to the effects of H2O2 -induced oxidative stress at 500 μM and above.

  5. Selection of single domain antibodies from immune libraries displayed on the surface of E. coli cells with two β-domains of opposite topologies.

    Directory of Open Access Journals (Sweden)

    Valencio Salema

    Full Text Available Screening of antibody (Ab libraries by direct display on the surface of E. coli cells is hampered by the presence of the outer membrane (OM. In this work we demonstrate that the native β-domains of EhaA autotransporter and intimin, two proteins from enterohemorrhagic E. coli O157:H7 (EHEC with opposite topologies in the OM, are effective systems for the display of immune libraries of single domain Abs (sdAbs from camelids (nanobodies or VHH on the surface of E. coli K-12 cells and for the selection of high affinity sdAbs using magnetic cell sorting (MACS. We analyzed the capacity of EhaA and intimin β-domains to display individual sdAbs and sdAb libraries obtained after immunization with the extracellular domain of the translocated intimin receptor from EHEC (TirM(EHEC. We demonstrated that both systems displayed functional sdAbs on the surface of E. coli cells with little proteolysis and cellular toxicity, although E. coli cells displaying sdAbs with the β-domain of intimin showed higher antigen-binding capacity. Both E. coli display libraries were screened for TirM(EHEC binding clones by MACS. High affinity binders were selected by both display systems, although more efficiently with the intimin β-domain. The specificity of the selected clones against TirM(EHEC was demonstrated by flow cytometry of E. coli cells, along with ELISA and surface plasmon resonance with purified sdAbs. Finally, we employed the E. coli cell display systems to provide an estimation of the affinity of the selected sdAb by flow cytometry analysis under equilibrium conditions.

  6. Menstrual blood cells display stem cell-like phenotypic markers and exert neuroprotection following transplantation in experimental stroke.

    Science.gov (United States)

    Borlongan, Cesar V; Kaneko, Yuji; Maki, Mina; Yu, Seong-Jin; Ali, Mohammed; Allickson, Julie G; Sanberg, Cyndy D; Kuzmin-Nichols, Nicole; Sanberg, Paul R

    2010-04-01

    Cell therapy remains an experimental treatment for neurological disorders. A major obstacle in pursuing the clinical application of this therapy is finding the optimal cell type that will allow benefit to a large patient population with minimal complications. A cell type that is a complete match of the transplant recipient appears as an optimal scenario. Here, we report that menstrual blood may be an important source of autologous stem cells. Immunocytochemical assays of cultured menstrual blood reveal that they express embryonic-like stem cell phenotypic markers (Oct4, SSEA, Nanog), and when grown in appropriate conditioned media, express neuronal phenotypic markers (Nestin, MAP2). In order to test the therapeutic potential of these cells, we used the in vitro stroke model of oxygen glucose deprivation (OGD) and found that OGD-exposed primary rat neurons that were co-cultured with menstrual blood-derived stem cells or exposed to the media collected from cultured menstrual blood exhibited significantly reduced cell death. Trophic factors, such as VEGF, BDNF, and NT-3, were up-regulated in the media of OGD-exposed cultured menstrual blood-derived stem cells. Transplantation of menstrual blood-derived stem cells, either intracerebrally or intravenously and without immunosuppression, after experimentally induced ischemic stroke in adult rats also significantly reduced behavioral and histological impairments compared to vehicle-infused rats. Menstrual blood-derived cells exemplify a source of "individually tailored" donor cells that completely match the transplant recipient, at least in women. The present neurostructural and behavioral benefits afforded by transplanted menstrual blood-derived cells support their use as a stem cell source for cell therapy in stroke.

  7. Combined EGFR- and notch inhibition display additive inhibitory effect on glioblastoma cell viability and glioblastoma-induced endothelial cell sprouting in vitro

    DEFF Research Database (Denmark)

    Staberg, Mikkel; Michaelsen, Signe Regner; Stobbe Olsen, Marie-Louise;

    2016-01-01

    aimed at testing a combination treatment strategy using inhibitors targeting the notch and EGFR pathways. METHODS: For evaluation of cell viability a standard MTT assay was used. Western blotting (WB) and Q-RT-PCR were employed in order to assess the protein- and mRNA expression levels, respectively....... In order to determine angiogenic processes, we used an endothelial spheroid sprouting assay. For assessment of secreted VEGF from GBM cells we performed a VEGF-quantikine ELISA. RESULTS: GBM cells were confirmed to express EGFR and Notch and to have the capacity to induce endothelial cell sprouting....... Inhibition of EGFR and Notch signaling was achieved using either Iressa (gefitinib) or the gamma-secretase inhibitor DAPT. Our data showed that DAPT combined with Iressa treatment displayed increased inhibitory effect on cell viability and abrogated expression and activation of major pro-survival pathways...

  8. Stat3 promotes invasion of esophageal squamous cell carcinoma through up-regulation of MMP2.

    Science.gov (United States)

    Xuan, Xaioyan; Li, Shanshan; Lou, Xi; Zheng, Xianzhao; Li, Yunyun; Wang, Feng; Gao, Yuan; Zhang, Hongyan; He, Hongliu; Zeng, Qingru

    2015-05-01

    Stat3 alters the expression of its downstream genes and is associated with tumor invasion and metastasis in several human cancers. Its role in esophageal squamous cell carcinoma (ESCC) has not been well characterized. We examined the tumor sections of 100 cases of ESCC by immunohistochemistry and observed significant overexpression of Stat3 in the cytoplasm of 89% of ESCC cells and of phosphorylated Stat3 (p-Stat3) in the nuclei of 71% of ESCC when compare with normal esophageal mucosa (72%, p = 0.02; and 31%, p = 0.001). Overexpression of Stat3 and p-Stat3 positively correlated with that of matrix metalloproteinase-2 (MMP2), a known regulator for cell migration, in 65% of ESCC while only 26% shown in benign esophageal mucosa. To further investigate the association of Stat3 with tumor metastasis in vitro, invasion of EC-1 cells (a human ESCC cell line) were investigated with Boyden chambers. The results showed that transfection of Stat3 not only promoted invasion of EC-1 cells but also significantly induced MMP2 expression in a dose-dependent manner. In contrast, suppressing expression of endogenous Stat3 mRNA and protein by Stat3 siRNA significantly reduced EC-1 cell invasion and MMP2 expression. A high-affinity Stat3-binding element was localized to the positions of 648-641 bp (TTCTCGAA) in the MMP2 promoter with electrophoretic mobility shift assay. Our results suggest that Stat3, p-Stat3, and MMP2 were overexpressed in ESCC and associated with invasion of ESCC; and Stat3 up-regulated expression of MMP2 in ESCC through directly binding to the MMP2 promoter.

  9. Alpha-1 proteinase inhibitor M358R reduces thrombin generation when displayed on the surface of cells expressing tissue factor.

    Science.gov (United States)

    Gierczak, Richard F; Pepler, Laura; Bhagirath, Vinai; Liaw, Patricia C; Sheffield, William P

    2014-11-01

    The M358R variant of alpha-1-proteinase inhibitor (API) is a potent soluble inhibitor of thrombin. Previously we engineered AR-API M358R, a membrane-bound form of this protein and showed that it inhibited exogenous thrombin when expressed on transfected cells lacking tissue factor (TF). To determine the suitability of AR-API M358R for gene transfer to vascular cells to limit thrombogenicity, we tested the ability of AR-API M358R to inhibit endogenous thrombin generated in plasma via co-expression co-expressing it on the surface of cells expressing TF. Transfected AR-API M358R formed inhibitory complexes with thrombin following exposure of recalcified, defibrinated plasma to TF on T24/83 cells, but discontinuously monitored thrombin generation was unaffected. Similarly, AR-API M358R expression did not reduce continuously monitored thrombin generation by T24/83 cell suspensions exposed to recalcified normal plasma in a Thrombogram-Thrombinoscope-type thrombin generation assay (TGA); in contrast, 1 μM hirudin variant 3 or soluble API M358R abolished thrombin generation. Gene transfer of TF to HEK 293 conferred the ability to support TF-dependent thrombin generation on HEK 293 cells. Co-transfection of HEK 293 cells with a 9:1 excess of DNA encoding AR-API M358R to that encoding TF reduced peak thrombin generation approximately 3-fold compared to controls. These in vitro results suggest that surface display of API M358R inhibits thrombin generation when the tethered serpin is expressed in excess of TF, and suggest its potential to limit thrombosis in appropriate vascular beds in animal models.

  10. Targeting essential Eimeria ninakohlyakimovae sporozoite ligands for caprine host endothelial cell invasion with a phage display peptide library.

    Science.gov (United States)

    Ruiz, A; Pérez, D; Muñoz, M C; Molina, J M; Taubert, A; Jacobs-Lorena, M; Vega-Rodríguez, J; López, A M; Hermosilla, C

    2015-11-01

    Eimeria ninakohlyakimovae is an important coccidian parasite of goats which causes severe diarrhoea in young animals. Specific molecules that mediate E. ninakohlyakimovae host interactions and molecular mechanisms involved in the pathogenesis are still unknown. Although strong circumstantial evidence indicates that E. ninakohlyakimovae sporozoite interactions with caprine endothelial host cells (ECs) are specific, hardly any information is available about the interacting molecules that confer host cell specificity. In this study, we describe a novel method to identify surface proteins of caprine umbilical vein endothelial cells (CUVEC) using a phage display library. After several panning rounds, we identified a number of peptides that specifically bind to the surface of CUVEC. Importantly, caprine endothelial cell peptide 2 (PCEC2) and PCEC5 selectively reduced the infection rate by E. ninakohlyakimovae sporozoites. These preliminary data give new insight for the molecular identification of ligands involved in the interaction between E. ninakohlyakimovae sporozoites and host ECs. Further studies using this phage approach might be useful to identify new potential target molecules for the development of anti-coccidial drugs or even new vaccine strategies.

  11. Increased micronucleated cell frequency related to exposure to radiation emitted by computer cathode ray tube video display monitors

    Directory of Open Access Journals (Sweden)

    Carbonari Karina

    2005-01-01

    Full Text Available It is well recognized that electromagnetic fields can affect the biological functions of living organisms at both cellular and molecular level. The potential damaging effects of electromagnetic fields and very low frequency and extremely low frequency radiation emitted by computer cathode ray tube video display monitors (VDMs has become a concern within the scientific community. We studied the effects of occupational exposure to VDMs in 10 males and 10 females occupationally exposed to VDMs and 20 unexposed control subjects matched for age and sex. Genetic damage was assessed by examining the frequency of micronuclei in exfoliated buccal cells and the frequency of other nuclear abnormalities such as binucleated and broken egg cells. Although there were no differences regarding binucleated cells between exposed and control individuals our analysis revealed a significantly higher frequency of micronuclei (p < 0.001 and broken egg cells (p < 0.05 in individuals exposed to VDMs as compared to unexposed. We also found that the differences between individuals exposed to VDMs were significantly related to the sex of the individuals and that there was an increase in skin, central nervous system and ocular disease in the exposed individuals. These preliminary results indicate that microcomputer workers exposed to VDMs are at risk of significant cytogenetic damage and should periodically undergo biological monitoring.

  12. Alkamides from the fruits of Piper longum and Piper nigrum displaying potent cell adhesion inhibition.

    Science.gov (United States)

    Lee, Seung Woong; Kim, Young Kook; Kim, Koanhoi; Lee, Hyun Sun; Choi, Jung Ho; Lee, Woo Song; Jun, Chang-Duk; Park, Jee Hun; Lee, Jeong Min; Rho, Mun-Chual

    2008-08-15

    Eight alkamides 1-8 were isolated by bioassay-guided isolation of EtOH extracts of the fruits of Piper longum and Piper nigum (Piperaceae). Their structures were elucidated by spectroscopic analysis ((1)H, (13)C NMR, and ESI-MS) as follows: guineensine (1), retrofracamide C (2), (2E,4Z,8E)-N-[9-(3,4-methylenedioxyphenyl)-2,4,8-nonatrienoyl]piperidine (3), pipernonaline (4), piperrolein B (5), piperchabamide D (6), pellitorin (7), and dehydropipernonaline (8). Their compounds 3-5, 7, and 8 inhibited potently the direct binding between sICAM-1 and LFA-1 of THP-1 cells in a dose-dependent manner, with IC(50) values of 10.7, 8.8, 13.4, 13.5, and 6.0 microg/mL, respectively.

  13. MR1-restricted MAIT cells display ligand discrimination and pathogen selectivity through distinct T cell receptor usage

    DEFF Research Database (Denmark)

    Gold, Marielle C.; McLaren, James E.; Reistetter, Joseph A.

    2015-01-01

    Mucosal-associated invariant T (MAIT) cells express a semi-invariant T cell receptor (TCR) that detects microbial metabolites presented by the nonpolymorphic major histocompatibility complex (MHC)-like molecule MR1. The highly conserved nature of MR1 in conjunction with biased MAIT TCRα chain usage...... is widely thought to indicate limited ligand presentation and discrimination within a pattern-like recognition system. Here, we evaluated the TCR repertoire of MAIT cells responsive to three classes of microbes. Substantial diversity and heterogeneity were apparent across the functional MAIT cell repertoire...... with this interpretation, MAIT cell clones with distinct TCRs responded differentially to a riboflavin metabolite. These results suggest that MAIT cells can discriminate between pathogen-derived ligands in a clonotype-dependent manner, providing a basis for adaptive memory via recruitment of specific repertoires shaped...

  14. MR1-restricted MAIT cells display ligand discrimination and pathogen selectivity through distinct T cell receptor usage

    DEFF Research Database (Denmark)

    Gold, Marielle C.; McLaren, James E.; Reistetter, Joseph A.

    2014-01-01

    Mucosal-associated invariant T (MAIT) cells express a semi-invariant T cell receptor (TCR) that detects microbial metabolites presented by the nonpolymorphic major histocompatibility complex (MHC)-like molecule MR1. The highly conserved nature of MR1 in conjunction with biased MAIT TCRα chain usage...... is widely thought to indicate limited ligand presentation and discrimination within a pattern-like recognition system. Here, we evaluated the TCR repertoire of MAIT cells responsive to three classes of microbes. Substantial diversity and heterogeneity were apparent across the functional MAIT cell repertoire...... with this interpretation, MAIT cell clones with distinct TCRs responded differentially to a riboflavin metabolite. These results suggest that MAIT cells can discriminate between pathogen-derived ligands in a clonotype-dependent manner, providing a basis for adaptive memory via recruitment of specific repertoires shaped...

  15. Stimulation of the B-cell receptor activates the JAK2/STAT3 signaling pathway in chronic lymphocytic leukemia cells.

    Science.gov (United States)

    Rozovski, Uri; Wu, Ji Yuan; Harris, David M; Liu, Zhiming; Li, Ping; Hazan-Halevy, Inbal; Ferrajoli, Alessandra; Burger, Jan A; O'Brien, Susan; Jain, Nitin; Verstovsek, Srdan; Wierda, William G; Keating, Michael J; Estrov, Zeev

    2014-06-12

    In chronic lymphocytic leukemia (CLL), stimulation of the B-cell receptor (BCR) triggers survival signals. Because in various cells activation of the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway provides cells with survival advantage, we wondered whether BCR stimulation activates the JAK/STAT pathway in CLL cells. To stimulate the BCR we incubated CLL cells with anti-IgM antibodies. Anti-IgM antibodies induced transient tyrosine phosphorylation and nuclear localization of phosphorylated (p) STAT3. Immunoprecipitation studies revealed that anti-JAK2 antibodies coimmunoprecipitated pSTAT3 and pJAK2 in IgM-stimulated but not unstimulated CLL cells, suggesting that activation of the BCR induces activation of JAK2, which phosphorylates STAT3. Incubation of CLL cells with the JAK1/2 inhibitor ruxolitinib inhibited IgM-induced STAT3 phosphorylation and induced apoptosis of IgM-stimulated but not unstimulated CLL cells in a dose- and time-dependent manner. Whether ruxolitinib treatment would benefit patients with CLL remains to be determined.

  16. Production, secretion, and cell surface display of recombinant Sporosarcina ureae S-layer fusion proteins in Bacillus megaterium.

    Science.gov (United States)

    Knobloch, Denise; Ostermann, Kai; Rödel, Gerhard

    2012-01-01

    Monomolecular crystalline bacterial cell surface layers (S-layers) have broad application potential in nanobiotechnology due to their ability to generate functional supramolecular structures. Here, we report that Bacillus megaterium is an excellent host organism for the heterologous expression and efficient secretion of hemagglutinin (HA) epitope-tagged versions of the S-layer protein SslA from Sporosarcina ureae ATCC 13881. Three chimeric proteins were constructed, comprising the precursor, C-terminally truncated, and N- and C-terminally truncated forms of the S-layer SslA protein tagged with the human influenza hemagglutinin epitope. For secretion of fusion proteins, the open reading frames were cloned into the Escherichia coli-Bacillus megaterium shuttle vector pHIS1525. After transformation of the respective plasmids into Bacillus megaterium protoplasts, the recombinant genes were successfully expressed and the proteins were secreted into the growth medium. The isolated S-layer proteins are able to assemble in vitro into highly ordered, crystalline, sheetlike structures with the fused HA tag accessible to antibody. We further show by fluorescent labeling that the secreted S-layer fusion proteins are also clustered on the cell envelope of Bacillus megaterium, indicating that the cell surface can serve in vivo as a nucleation point for crystallization. Thus, this system can be used as a display system that allows the dense and periodic presentation of S-layer proteins or the fused tags.

  17. Mouse mutants for the nicotinic acetylcholine receptor ß2 subunit display changes in cell adhesion and neurodegeneration response genes.

    Directory of Open Access Journals (Sweden)

    Carol M Rubin

    Full Text Available Mice lacking expression of the ß2 subunit of the neuronal nicotinic acetylcholine receptor (CHRNB2 display abnormal retinal waves and a dispersed projection of retinal ganglion cell (RGC axons to their dorsal lateral geniculate nuclei (dLGNs. Transcriptomes of LGN tissue from two independently generated Chrnb2-/- mutants and from wildtype mice were obtained at postnatal day 4 (P4, during the normal period of segregation of eye-specific afferents to the LGN. Microarray analysis reveals reduced expression of genes located on the cell membrane or in extracellular space, and of genes active in cell adhesion and calcium signaling. In particular, mRNA for cadherin 1 (Cdh1, a known axon growth regulator, is reduced to nearly undetectable levels in the LGN of P4 mutant mice and Lypd2 mRNA is similarly suppressed. Similar analysis of retinal tissue shows increased expression of crumbs 1 (Crb1 and chemokine (C-C motif ligand 21 (Ccl21 mRNAs in Chrnb2-/- mutant animals. Mutations in these genes are associated with retinal neuronal degeneration. The retinas of Chrnb2-/- mutants are normal in appearance, but the increased expression of these genes may also be involved in the abnormal projection patterns of RGC to the LGN. These data may provide the tools to distinguish the interplay between neural activity and molecular expression. Finally, comparison of the transcriptomes of the two different Chrnb2-/- mutant strains reveals the effects of genetic background upon gene expression.

  18. Phage display selection on whole cells yields a small peptide specific for HCV receptor human CD81

    Institute of Scientific and Technical Information of China (English)

    JIE CAO; PING ZHAO; X1AO HUI MIAO; LAN JUAN ZHAO; LI JUN XUE; ZHONG TIAN QI

    2003-01-01

    The human CD81(hCD81),the most recently proposed receptor of hepatitis C virus(HCV),can especifically bind to HCV envelope glycoprotein 2(E2).In this study,hCD81-expressing murine NIH/3T3 cells were used to select hCD81-binding peptides from a phage displayed nonapeptide library(PVIII9aaCys).Eighteen of the 75clones selected from the library showed specific binding to the hCD81-expressing NIH/3T3 cells by enzyme linked immunosorbent assay(ELISA)and competitive inhibition test.Twelve out of the 18 clones shared the amino acid motif SPQYWTGPA.Sequence comparison of the motif showed no amino acid homology with the native HCV E2.The motif-containing phages could competitively inhibit the ability of HCV E2 binding to native hCD81-expressing MOLT-4 cells,and induce HCV E2 specific immune response in vivo.These results suggest that the selected motif SPQYWTGPA should be a mimotope of HCV E2 to bind to hCD81 molecules.Our findings cast new light on developing HCV receptor antagonists.

  19. Mammalian Cell Surface Display as a Novel Method for Developing Engineered Lectins with Novel Characteristics

    Directory of Open Access Journals (Sweden)

    Keisuke Soga

    2015-07-01

    Full Text Available Leguminous lectins have a conserved carbohydrate recognition site comprising four loops (A–D. Here, we randomly mutated the sequence and length of loops C and D of peanut agglutinin (PNA and expressed the proteins on the surface of mouse green fluorescent protein (GFP-reporter cells. Flow cytometry, limiting dilution, and cDNA cloning were used to screen for several mutated PNAs with distinct properties. The mutated PNA clones obtained using NeuAcα2-6(Galβ1-3GalNAc as a ligand showed preference for NeuAcα2-6(Galβ1-3GalNAc rather than non-sialylated Galβ1-3GlcNAc, whereas wild-type PNA binds to Galβ1-3GlcNAc but not sialylated Galβ1-3GalNAc. Sequence analyses revealed that for all of the glycan-reactive mutated PNA clones, (i loop C was eight amino acids in length, (ii loop D was identical to that of wild-type PNA, (iii residue 127 was asparagine, (iv residue 125 was tryptophan, and (v residue 130 was hydrophobic tyrosine, phenylalanine, or histidine. The sugar-binding ability of wild-type PNA was increased nine-fold when Tyr125 was mutated to tryptophan, and that of mutated clone C was increased more than 30-fold after His130 was changed to tyrosine. These results provide an insight into the relationship between the amino acid sequences of the carbohydrate recognition site and sugar-binding abilities of leguminous lectins.

  20. An integrated microfluidic system for screening of phage-displayed peptides specific to colon cancer cells and colon cancer stem cells.

    Science.gov (United States)

    Che, Yu-Jui; Wu, Huei-Wen; Hung, Lien-Yu; Liu, Ching-Ann; Chang, Hwan-You; Wang, Kuan; Lee, Gwo-Bin

    2015-09-01

    Affinity reagents recognizing biomarkers specifically are essential components of clinical diagnostics and target therapeutics. However, conventional methods for screening of these reagents often have drawbacks such as large reagent consumption, the labor-intensive or time-consuming procedures, and the involvement of bulky or expensive equipment. Alternatively, microfluidic platforms could potentially automate the screening process within a shorter period of time and reduce reagent and sample consumption dramatically. It has been demonstrated recently that a subpopulation of tumor cells known as cancer stem cells possess high drug resistance and proliferation potential and are regarded as the main cause of metastasis. Therefore, a peptide that recognizes cancer stem cells and differentiates them from other cancer cells will be extremely useful in early diagnosis and target therapy. This study utilized M13 phage display technology to identify peptides that bind, respectively, to colon cancer cells and colon cancer stem cells using an integrated microfluidic system. In addition to positive selection, a negative selection process was integrated on the chip to achieve the selection of peptides of high affinity and specificity. We successfully screened three peptides specific to colon cancer cells and colon cancer stem cells, namely, HOLC-1, HOLC-2, and COLC-1, respectively, and their specificity was measured by the capture rate between target, control, and other cell lines. The capture rates are 43.40 ± 7.23%, 45.16 ± 7.12%, and 49.79 ± 5.34% for colon cancer cells and colon cancer stem cells, respectively, showing a higher specificity on target cells than on control and other cell lines. The developed technique may be promising for early diagnosis of cancer cells and target therapeutics.

  1. MR1-restricted MAIT cells display ligand discrimination and pathogen selectivity through distinct T cell receptor usage.

    Science.gov (United States)

    Gold, Marielle C; McLaren, James E; Reistetter, Joseph A; Smyk-Pearson, Sue; Ladell, Kristin; Swarbrick, Gwendolyn M; Yu, Yik Y L; Hansen, Ted H; Lund, Ole; Nielsen, Morten; Gerritsen, Bram; Kesmir, Can; Miles, John J; Lewinsohn, Deborah A; Price, David A; Lewinsohn, David M

    2014-07-28

    Mucosal-associated invariant T (MAIT) cells express a semi-invariant T cell receptor (TCR) that detects microbial metabolites presented by the nonpolymorphic major histocompatibility complex (MHC)-like molecule MR1. The highly conserved nature of MR1 in conjunction with biased MAIT TCRα chain usage is widely thought to indicate limited ligand presentation and discrimination within a pattern-like recognition system. Here, we evaluated the TCR repertoire of MAIT cells responsive to three classes of microbes. Substantial diversity and heterogeneity were apparent across the functional MAIT cell repertoire as a whole, especially for TCRβ chain sequences. Moreover, different pathogen-specific responses were characterized by distinct TCR usage, both between and within individuals, suggesting that MAIT cell adaptation was a direct consequence of exposure to various exogenous MR1-restricted epitopes. In line with this interpretation, MAIT cell clones with distinct TCRs responded differentially to a riboflavin metabolite. These results suggest that MAIT cells can discriminate between pathogen-derived ligands in a clonotype-dependent manner, providing a basis for adaptive memory via recruitment of specific repertoires shaped by microbial exposure.

  2. Remarkable heterogeneity displayed by oval cells in rat and mouse models of stem cell-mediated liver regeneration

    DEFF Research Database (Denmark)

    Jelnes, Peter; Santoni-Rugiu, Eric; Rasmussen, Morten

    2007-01-01

    -collidin (DDC) diet; and N-acetyl-paraaminophen (APAP). Reproducibly, oval cells showing reactivity for cytokeratins (CKs), muscle pyruvate kinase (MPK), the adenosine triphosphate-binding cassette transporter ABCG2/BCRP1 (ABCG2), alpha-fetoprotein (AFP), and delta-like protein 1/preadipocyte factor 1 (Dlk....../Pref-1) were induced in rat liver treated according to the AAF/PHx and CDE but not the DDC protocol. In mouse liver, the CDE, DDC, and APAP protocols all induced CKs and ABCG2-positive oval cells. However, AFP and Dlk/Pref-1 expression was rarely detected in oval cells. CONCLUSION: Our results delineate...

  3. Effector and Naturally Occurring Regulatory T Cells Display No Abnormalities in Activation Induced Cell Death in NOD Mice

    OpenAIRE

    Ayelet Kaminitz; Esma S Yolcu; Askenasy, Enosh M.; Jerry Stein; Isaac Yaniv; Haval Shirwan; Nadir Askenasy

    2011-01-01

    BACKGROUND: Disturbed peripheral negative regulation might contribute to evolution of autoimmune insulitis in type 1 diabetes. This study evaluates the sensitivity of naïve/effector (Teff) and regulatory T cells (Treg) to activation-induced cell death mediated by Fas cross-linking in NOD and wild-type mice. PRINCIPAL FINDINGS: Both effector (CD25(-), FoxP3(-)) and suppressor (CD25(+), FoxP3(+)) CD4(+) T cells are negatively regulated by Fas cross-linking in mixed splenocyte populations of NOD...

  4. Heterologous protein display on the cell surface of lactic acid bacteria mediated by the s-layer protein

    Directory of Open Access Journals (Sweden)

    Han Lanlan

    2011-10-01

    Full Text Available Abstract Background Previous studies have revealed that the C-terminal region of the S-layer protein from Lactobacillus is responsible for the cell wall anchoring, which provide an approach for targeting heterologous proteins to the cell wall of lactic acid bacteria (LAB. In this study, we developed a new surface display system in lactic acid bacteria with the C-terminal region of S-layer protein SlpB of Lactobacillus crispatus K2-4-3 isolated from chicken intestine. Results Multiple sequence alignment revealed that the C-terminal region (LcsB of Lb. crispatus K2-4-3 SlpB had a high similarity with the cell wall binding domains SA and CbsA of Lactobacillus acidophilus and Lb. crispatus. To evaluate the potential application as an anchoring protein, the green fluorescent protein (GFP or beta-galactosidase (Gal was fused to the N-terminus of the LcsB region, and the fused proteins were successfully produced in Escherichia coli, respectively. After mixing them with the non-genetically modified lactic acid bacteria cells, the fused GFP-LcsB and Gal-LcsB were functionally associated with the cell surface of various lactic acid bacteria tested. In addition, the binding capacity could be improved by SDS pretreatment. Moreover, both of the fused proteins could simultaneously bind to the surface of a single cell. Furthermore, when the fused DNA fragment of gfp:lcsB was inserted into the Lactococcus lactis expression vector pSec:Leiss:Nuc, the GFP could not be secreted into the medium under the control of the nisA promoter. Western blot, in-gel fluorescence assay, immunofluorescence microscopy and SDS sensitivity analysis confirmed that the GFP was successfully expressed onto the cell surface of L. lactis with the aid of the LcsB anchor. Conclusion The LcsB region can be used as a functional scaffold to target the heterologous proteins to the cell surfaces of lactic acid bacteria in vitro and in vivo, and has also the potential for biotechnological

  5. An anti-tumor protein produced by Trichinella spiralis and identified by screening a T7 phage display library, induces apoptosis in human hepatoma H7402 cells

    Science.gov (United States)

    Trichinella spiralis infection confers effective resistance to tumor cell expansion. In this study, a T7 phage cDNA display library was constructed to express genes encoded by T. spiralis. Organic phase multi-cell screening was used to sort through candidate proteins in a transfected human chronic m...

  6. IL-6/STAT3 signaling pathway is activated in plasma cell mastitis.

    Science.gov (United States)

    Liu, Yang; Zhang, Jian; Zhou, Yu-Hui; Jiang, Yi-Na; Zhang, Wei; Tang, Xiao-Jiang; Ren, Yu; Han, Shui-Ping; Liu, Pei-Jun; Xu, Jing; He, Jian-Jun

    2015-01-01

    Plasma cell mastitis (PCM), a particular type of mastitis, mainly occurs in females at nonpregnant and nonlactating stages. The infiltration of abundant plasma cells and lymphocytes is the hallmark of the disease. The incidence rate of PCM increased gradually and its pathogenesis remained unclear. In this study, we investigated the expression of IL-6/STAT3 signaling pathway, which is vital not only for the differentiation of plasma cells but also for survival of plasma cells and T lymphocytes, in 30 PCM cases, 10 acute mastitis cases and 10 normal breast tissues by immunohistochemical analysis. IL-6 level was significantly higher in PCM patients than in acute mastitis patients or normal group. The positive rate of IL-6 and p-STAT3 staining in PCM samples was 93.3% (28/30) and 70% (21/30), respectively, and there was a significant positive association between IL-6 and p-STAT3 staining (r=0.408, P=0.025). In PCM group, the rate of nipple retraction was 40% (12/30). Significantly higher IL-6 expression was found in PCM patients with nipple retraction than in other PCM patients. However, no significant difference in IL-6 or p-STAT3 staining was detected between PCM patients experiencing recurrence and other PCM patients. In addition, Bcl-2 level was higher in PCM patients than in acute mastitis patients or normal group, but there was no difference in Bcl-2 immunostaining between PCM patients experiencing recurrence and other PCM patients. These indicate that IL-6/STAT3 signaling is activated in PCM and may play an important role in the pathogenesis of PCM.

  7. Human Renal Normal, Tumoral, and Cancer Stem Cells Express Membrane-Bound Interleukin-15 Isoforms Displaying Different Functions

    Directory of Open Access Journals (Sweden)

    Sandy Azzi

    2015-06-01

    Full Text Available Intrarenal interleukin-15 (IL-15 participates to renal pathophysiology, but the role of its different membrane-bound isoforms remains to be elucidated. In this study, we reassess the biology of membrane-bound IL-15 (mb-IL-15 isoforms by comparing primary cultures of human renal proximal tubular epithelial cells (RPTEC to peritumoral (ptumTEC, tumoral (RCC, and cancer stem cells (CSC/CD105+. RPTEC express a 14 to 16 kDa mb-IL-15, whose existence has been assumed but never formally demonstrated and likely represents the isoform anchored at the cell membrane through the IL-15 receptor α (IL-15Rα chain, because it is sensitive to acidic treatment and is not competent to deliver a reverse signal. By contrast, ptumTEC, RCC, and CSC express a novel N-hyperglycosylated, short-lived transmembrane mb-IL-15 (tmb-IL-15 isoform around 27 kDa, resistant to acidic shock, delivering a reverse signal in response to its soluble receptor (sIL-15Rα. This reverse signal triggers the down-regulation of the tumor suppressor gene E-cadherin in ptumTEC and RCC but not in CSC/CD105+, where it promotes survival. Indeed, through the AKT pathway, tmb-IL-15 protects CSC/CD105+ from non-programmed cell death induced by serum starvation. Finally, both mb-IL-15 and tmb-IL-15 are sensitive to metalloproteases, and the cleaved tmb-IL-15 (25 kDa displays a powerful anti-apoptotic effect on human hematopoietic cells. Overall, our data indicate that both mb-IL-15 and tmb-IL-15 isoforms play a complex role in renal pathophysiology downregulating E-cadherin and favoring cell survival. Moreover, “apparently normal” ptumTEC cells, sharing different properties with RCC, could contribute to organize an enlarged peritumoral “preneoplastic” environment committed to favor tumor progression.

  8. JAK2 inhibitor TG101348 overcomes erlotinib-resistance in non-small cell lung carcinoma cells with mutated EGF receptor.

    Science.gov (United States)

    Zhang, Fu-quan; Yang, Wen-tao; Duan, Shan-zhou; Xia, Ying-chen; Zhu, Rong-ying; Chen, Yong-bing

    2015-06-10

    Non-small cell lung cancer (NSCLC) patients with epidermal growth factor receptor (EGFR) mutations are responsive to EGFR-tyrosine kinase inhibitor (EGFR-TKI). However, NSCLC patients with secondary somatic EGFR mutations are resistant to EGFR-TKI treatment. In this study, we investigated the effect of TG101348 (a JAK2 inhibitor) on the tumor growth of erlotinib-resistant NSCLC cells. Cell proliferation, apoptosis, gene expression and tumor growth were evaluated by diphenyltetrazolium bromide (MTT) assay, flow cytometry, terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) staining, Western Blot and a xenograft mouse model, respectively. Results showed that erlotinib had a stronger impact on the induction of apoptosis in erlotinib-sensitive PC-9 cells but had a weaker effect on erlotinib-resistant H1975 and H1650 cells than TG101348. TG101348 significantly enhanced the cytotoxicity of erlotinib to erlotinib-resistant NSCLC cells, stimulated erlotinib-induced apoptosis and downregulated the expressions of EGFR, p-EGFR, p-STAT3, Bcl-xL and survivin in erlotinib-resistant NSCLC cells. Moreover, the combined treatment of TG101348 and erlotinib induced apoptosis, inhibited the activation of p-EGFR and p-STAT3, and inhibited tumor growth of erlotinib-resistant NSCLC cells in vivo. Our results indicate that TG101348 is a potential adjuvant for NSCLC patients during erlotinib treatment.

  9. Generation and characterisation of cisplatin-resistant non-small cell lung cancer cell lines displaying a stem-like signature.

    Directory of Open Access Journals (Sweden)

    Martin P Barr

    Full Text Available INTRODUCTION: Inherent and acquired cisplatin resistance reduces the effectiveness of this agent in the management of non-small cell lung cancer (NSCLC. Understanding the molecular mechanisms underlying this process may result in the development of novel agents to enhance the sensitivity of cisplatin. METHODS: An isogenic model of cisplatin resistance was generated in a panel of NSCLC cell lines (A549, SKMES-1, MOR, H460. Over a period of twelve months, cisplatin resistant (CisR cell lines were derived from original, age-matched parent cells (PT and subsequently characterized. Proliferation (MTT and clonogenic survival assays (crystal violet were carried out between PT and CisR cells. Cellular response to cisplatin-induced apoptosis and cell cycle distribution were examined by FACS analysis. A panel of cancer stem cell and pluripotent markers was examined in addition to the EMT proteins, c-Met and β-catenin. Cisplatin-DNA adduct formation, DNA damage (γH2AX and cellular platinum uptake (ICP-MS was also assessed. RESULTS: Characterisation studies demonstrated a decreased proliferative capacity of lung tumour cells in response to cisplatin, increased resistance to cisplatin-induced cell death, accumulation of resistant cells in the G0/G1 phase of the cell cycle and enhanced clonogenic survival ability. Moreover, resistant cells displayed a putative stem-like signature with increased expression of CD133+/CD44+cells and increased ALDH activity relative to their corresponding parental cells. The stem cell markers, Nanog, Oct-4 and SOX-2, were significantly upregulated as were the EMT markers, c-Met and β-catenin. While resistant sublines demonstrated decreased uptake of cisplatin in response to treatment, reduced cisplatin-GpG DNA adduct formation and significantly decreased γH2AX foci were observed compared to parental cell lines. CONCLUSION: Our results identified cisplatin resistant subpopulations of NSCLC cells with a putative stem

  10. Quantitative evaluation of Candia antarctica lipase B displayed on the cell surface of a Pichia pastoris based on an FS anchor system.

    Science.gov (United States)

    Liang, Xing-xiang; Wang, Bei-bei; Sun, Yu-fei; Lin, Ying; Han, Shuang-yan; Zheng, Sui-ping; Cui, Tang-bing

    2013-03-01

    A new approach is described to quantify the number of enzyme molecules, such as Candia antarctica lipase B, that are displayed on the cell surface of Pichia pastoris. Enhanced green fluorescent protein (EGFP) and Candida antarctica lipase B (CALB) were fused and displayed on the surface of P. pastoris by linking to the anchor flocculation functional domain of FLO1p from Saccharomyces cerevisiae. Confocal laser scanning microscopy, flow cytometry, and fluorescence spectrophotometry were used to monitor the fluorescence intensity of fused EGFP. Combined with the corresponding protein concentration detected in the medium, a standard curve describing the relationship between the fusion protein concentration and fluorescence intensity were obtained and could be used to number CALB displayed on the cell surface. The results showed that approx. 10(4) molecules of CALB molecules were immobilized on the single P. pastoris cell wall based on FS anchor system.

  11. Characterization of the Discoidin Domain Receptor 2 Kinase as a Novel Therapeutic Target for Squamous Cell Lung Cancer

    Science.gov (United States)

    2012-09-01

    Immunoblots were performed using the Nupage System (Invitrogen) per the manufacturer’s protocol. Cells were lysed in 1% Nonidet P - 40 with protease (Roche...based on genetic lesions. J Clin Invest 2009;119:1727– 40 . 33. Du J, Bernasconi P , Clauser KR, Mani DR, Finn SP, Beroukhim R, et al. Bead-based...DDR2 mutations. DDR2 knockdown was associated with a decrease in both cell proliferation and in markers of DDR2 signaling including p -Src and p -STAT3

  12. Efficient cell surface display of Lip2 lipase using C-domains of glycosylphosphatidylinositol-anchored cell wall proteins of Yarrowia lipolytica.

    Science.gov (United States)

    Yuzbasheva, Evgeniya Y; Yuzbashev, Tigran V; Laptev, Ivan A; Konstantinova, Tatiana K; Sineoky, Sergey P

    2011-08-01

    The cell surface display of enzymes is of great interest because of its simplified purification stage and the possibility for recycling in industrial processes. In this study, we have focused on the cell wall immobilization of Yarrowia lipolytica Lip2 protein--an enzyme that has a wide technological application. By genome analysis of Y. lipolytica in addition to already characterized Ylcwp1, we identified five putative open reading frames encoding glycosylphosphatidylinositol-anchored proteins. Lip2 translation fusion with the carboxyl termini of these proteins revealed that all proteins were capable of immobilizing lipase in active form on the cell surface. The highest level of cell-bound lipase activity was achieved using C-domains encoded by YlCWP1, YlCWP3 (YALI0D27214g) and YlCWP6 (YALI0F18282g) comprising 16,173 ± 1,800, 18,785 ± 1,130 and 17,700 ± 2,101 U/g dry cells, respectively. To the best of our knowledge, these results significantly exceed the highest cell-bound lipase activity previously reported for engineered Saccharomyces cerevisiae and Pichia pastoris strains. Furthermore, the lyophilized biomass retained the activity and was robust to collecting/resuspending procedures. Nevertheless, in most cases, a substantial amount of lipase activity was also found in the growth medium. Further work will be necessary to better understand the nature of this phenomenon.

  13. Primary clear cell renal carcinoma cells display minimal mitochondrial respiratory capacity resulting in pronounced sensitivity to glycolytic inhibition by 3-Bromopyruvate.

    Science.gov (United States)

    Nilsson, H; Lindgren, D; Mandahl Forsberg, A; Mulder, H; Axelson, H; Johansson, M E

    2015-01-08

    Changes of cellular metabolism are an integral property of the malignant potential of most cancer cells. Already in the 1930s, Otto Warburg observed that tumor cells preferably utilize glycolysis and lactate fermentation for energy production, rather than the mitochondrial oxidative phosphorylation dominating in normal cells, a phenomenon today known as the Warburg effect. Even though many tumor types display a high degree of aerobic glycolysis, they still retain the activity of other energy-producing metabolic pathways. One exception seems to be the clear cell variant of renal cell carcinoma, ccRCC, where the activity of most other pathways than that of glycolysis has been shown to be reduced. This makes ccRCC a promising candidate for the use of glycolytic inhibitors in treatment of the disease. However, few studies have so far addressed this issue. In this report, we show a strikingly reduced mitochondrial respiratory capacity of primary human ccRCC cells, resulting in enhanced sensitivity to glycolytic inhibition by 3-Bromopyruvate (3BrPA). This effect was largely absent in established ccRCC cell lines, a finding that highlights the importance of using biologically relevant models in the search for new candidate cancer therapies. 3BrPA markedly reduced ATP production in primary ccRCC cells, followed by cell death. Our data suggest that glycolytic inhibitors such as 3BrPA, that has been shown to be well tolerated in vivo, should be further analyzed for the possible development of selective treatment strategies for patients with ccRCC.

  14. Deep mutational scanning of an antibody against epidermal growth factor receptor using mammalian cell display and massively parallel pyrosequencing.

    Science.gov (United States)

    Forsyth, Charles M; Juan, Veronica; Akamatsu, Yoshiko; DuBridge, Robert B; Doan, Minhtam; Ivanov, Alexander V; Ma, Zhiyuan; Polakoff, Dixie; Razo, Jennifer; Wilson, Keith; Powers, David B

    2013-01-01

    We developed a method for deep mutational scanning of antibody complementarity-determining regions (CDRs) that can determine in parallel the effect of every possible single amino acid CDR substitution on antigen binding. The method uses libraries of full length IgGs containing more than 1000 CDR point mutations displayed on mammalian cells, sorted by flow cytometry into subpopulations based on antigen affinity and analyzed by massively parallel pyrosequencing. Higher, lower and neutral affinity mutations are identified by their enrichment or depletion in the FACS subpopulations. We applied this method to a humanized version of the anti-epidermal growth factor receptor antibody cetuximab, generated a near comprehensive data set for 1060 point mutations that recapitulates previously determined structural and mutational data for these CDRs and identified 67 point mutations that increase affinity. The large-scale, comprehensive sequence-function data sets generated by this method should have broad utility for engineering properties such as antibody affinity and specificity and may advance theoretical understanding of antibody-antigen recognition.

  15. Ultra-high-throughput screening of an in vitro-synthesized horseradish peroxidase displayed on microbeads using cell sorter.

    Directory of Open Access Journals (Sweden)

    Bo Zhu

    Full Text Available The C1a isoenzyme of horseradish peroxidase (HRP is an industrially important heme-containing enzyme that utilizes hydrogen peroxide to oxidize a wide variety of inorganic and organic compounds for practical applications, including synthesis of fine chemicals, medical diagnostics, and bioremediation. To develop a ultra-high-throughput screening system for HRP, we successfully produced active HRP in an Escherichia coli cell-free protein synthesis system, by adding disulfide bond isomerase DsbC and optimizing the concentrations of hemin and calcium ions and the temperature. The biosynthesized HRP was fused with a single-chain Cro (scCro DNA-binding tag at its N-terminal and C-terminal sites. The addition of the scCro-tag at both ends increased the solubility of the protein. Next, HRP and its fusion proteins were successfully synthesized in a water droplet emulsion by using hexadecane as the oil phase and SunSoft No. 818SK as the surfactant. HRP fusion proteins were displayed on microbeads attached with double-stranded DNA (containing the scCro binding sequence via scCro-DNA interactions. The activities of the immobilized HRP fusion proteins were detected with a tyramide-based fluorogenic assay using flow cytometry. Moreover, a model microbead library containing wild type hrp (WT and inactive mutant (MUT genes was screened using fluorescence-activated cell-sorting, thus efficiently enriching the WT gene from the 1:100 (WT:MUT library. The technique described here could serve as a novel platform for the ultra-high-throughput discovery of more useful HRP mutants and other heme-containing peroxidases.

  16. Ultra-high-throughput screening of an in vitro-synthesized horseradish peroxidase displayed on microbeads using cell sorter.

    Science.gov (United States)

    Zhu, Bo; Mizoguchi, Takuro; Kojima, Takaaki; Nakano, Hideo

    2015-01-01

    The C1a isoenzyme of horseradish peroxidase (HRP) is an industrially important heme-containing enzyme that utilizes hydrogen peroxide to oxidize a wide variety of inorganic and organic compounds for practical applications, including synthesis of fine chemicals, medical diagnostics, and bioremediation. To develop a ultra-high-throughput screening system for HRP, we successfully produced active HRP in an Escherichia coli cell-free protein synthesis system, by adding disulfide bond isomerase DsbC and optimizing the concentrations of hemin and calcium ions and the temperature. The biosynthesized HRP was fused with a single-chain Cro (scCro) DNA-binding tag at its N-terminal and C-terminal sites. The addition of the scCro-tag at both ends increased the solubility of the protein. Next, HRP and its fusion proteins were successfully synthesized in a water droplet emulsion by using hexadecane as the oil phase and SunSoft No. 818SK as the surfactant. HRP fusion proteins were displayed on microbeads attached with double-stranded DNA (containing the scCro binding sequence) via scCro-DNA interactions. The activities of the immobilized HRP fusion proteins were detected with a tyramide-based fluorogenic assay using flow cytometry. Moreover, a model microbead library containing wild type hrp (WT) and inactive mutant (MUT) genes was screened using fluorescence-activated cell-sorting, thus efficiently enriching the WT gene from the 1:100 (WT:MUT) library. The technique described here could serve as a novel platform for the ultra-high-throughput discovery of more useful HRP mutants and other heme-containing peroxidases.

  17. Combined utilization of lipase-displaying Pichia pastoris whole-cell biocatalysts to improve biodiesel production in co-solvent media.

    Science.gov (United States)

    Jin, Zi; Han, Shuang-Yan; Zhang, Li; Zheng, Sui-Ping; Wang, Yong; Lin, Ying

    2013-02-01

    Lipase-displaying whole cells appear to be efficient biocatalysts because of their low preparation costs and simple recycling procedure. The combined utilization of Candida antarctica lipase B (CALB) and Rhizomucor miehei lipase (RML), separately displayed on Pichia pastoris whole cells, to produce biodiesel in co-solvent media was investigated. A response surface methodology incorporating a D-optimal design was employed to obtain the optimum reaction conditions for methyl ester (ME) synthesis. The synergistic effect of the two displayed lipases and the use of tert-butanol and isooctane as the co-solvent media were found to significantly improve the transesterification reaction. Scaled-up reactions using various types of feedstock were carried out in a 0.5-l stirred reactor under optimum conditions, affording ME yields over 90% in 12h. Moreover, the ME yields remained above 85% after 20 repeated batch cycles. In conclusion, this biocatalyst affords a promising route to efficient biodiesel production.

  18. Epitope mapping porcine reproductive and respiratory syndrome virus by phage display: the nsp2 fragment of the replicase polyprotein contains a cluster of B-cell epitopes

    DEFF Research Database (Denmark)

    Oleksiewicz, M.B.; Bøtner, Anette; Toft, P.;

    2001-01-01

    We screened phage display libraries of porcine reproductive and respiratory syndrome virus (PRRSV) protein fragments with sera from experimentally infected pigs to identify linear B-cell epitopes that are commonly recognized during infection in vivo. We identified 10 linear epitope sites (ES) 11...

  19. Coupling of Insulin Secretion and Display of a Granule-resident Zinc Transporter ZnT8 on the Surface of Pancreatic Beta Cells.

    Science.gov (United States)

    Huang, Qiong; Merriman, Chengfeng; Zhang, Hao; Fu, Dax

    2017-03-10

    The islet-specific zinc transporter ZnT8 mediates zinc enrichment in the insulin secretory granules of the pancreatic beta cell. This granular zinc transporter is also a major self-antigen found in type 1 diabetes patients. It is not clear whether ZnT8 can be displayed on the cell surface and how insulin secretion may regulate the level of ZnT8 exposure to extracellular immune surveillance. Here we report specific antibody binding to the extracellular surface of rat insulinoma INS-1E cells that stably expressed a tagged human zinc transporter ZnT8. Flow cytometry analysis after fluorescent antibody labeling revealed strong correlations among the levels of ZnT8 expression, its display on the cell surface, and glucose-stimulated insulin secretion (GSIS). Glucose stimulation increased the surface display of endogenous ZnT8 from a basal level to 32.5% of the housekeeping Na(+)/K(+) ATPase on the cell surface, thereby providing direct evidence for a GSIS-dependent surface exposure of the ZnT8 self-antigen. Moreover, the variation in tagged-ZnT8 expression and surface labeling enabled sorting of heterogeneous beta cells to subpopulations that exhibited marked differences in GSIS with parallel changes in endogenous ZnT8 expression. The abundant surface display of endogenous ZnT8 and its coupling to GSIS demonstrated the potential of ZnT8 as a surface biomarker for tracking and isolating functional beta cells in mixed cell populations.

  20. Myeloid clusters are associated with a pro-metastatic environment and poor prognosis in smoking-related early stage non-small cell lung cancer.

    Directory of Open Access Journals (Sweden)

    Wang Zhang

    Full Text Available BACKGROUND: This study aimed to understand the role of myeloid cell clusters in uninvolved regional lymph nodes from early stage non-small cell lung cancer patients. METHODS: Uninvolved regional lymph node sections from 67 patients with stage I-III resected non-small cell lung cancer were immunostained to detect myeloid clusters, STAT3 activity and occult metastasis. Anthracosis intensity, myeloid cluster infiltration associated with anthracosis and pSTAT3 level were scored and correlated with patient survival. Multivariate Cox regression analysis was performed with prognostic variables. Human macrophages were used for in vitro nicotine treatment. RESULTS: CD68+ myeloid clusters associated with anthracosis and with an immunosuppressive and metastasis-promoting phenotype and elevated overall STAT3 activity were observed in uninvolved lymph nodes. In patients with a smoking history, myeloid cluster score significantly correlated with anthracosis intensity and pSTAT3 level (P<0.01. Nicotine activated STAT3 in macrophages in long-term culture. CD68+ myeloid clusters correlated and colocalized with occult metastasis. Myeloid cluster score was an independent prognostic factor (P = 0.049 and was associated with survival by Kaplan-Maier estimate in patients with a history of smoking (P = 0.055. The combination of myeloid cluster score with either lymph node stage or pSTAT3 level defined two populations with a significant difference in survival (P = 0.024 and P = 0.004, respectively. CONCLUSIONS: Myeloid clusters facilitate a pro-metastatic microenvironment in uninvolved regional lymph nodes and associate with occult metastasis in early stage non-small cell lung cancer. Myeloid cluster score is an independent prognostic factor for survival in patients with a history of smoking, and may present a novel method to inform therapy choices in the adjuvant setting. Further validation studies are warranted.

  1. Type 1 cannabinoid receptor ligands display functional selectivity in a cell culture model of striatal medium spiny projection neurons.

    Science.gov (United States)

    Laprairie, Robert B; Bagher, Amina M; Kelly, Melanie E M; Dupré, Denis J; Denovan-Wright, Eileen M

    2014-09-05

    Modulation of type 1 cannabinoid receptor (CB1) activity has been touted as a potential means of treating addiction, anxiety, depression, and neurodegeneration. Different agonists of CB1 are known to evoke varied responses in vivo. Functional selectivity is the ligand-specific activation of certain signal transduction pathways at a receptor that can signal through multiple pathways. To understand cannabinoid-specific functional selectivity, different groups have examined the effect of individual cannabinoids on various signaling pathways in heterologous expression systems. In the current study, we compared the functional selectivity of six cannabinoids, including two endocannabinoids (2-arachidonyl glycerol (2-AG) and anandamide (AEA)), two synthetic cannabinoids (WIN55,212-2 and CP55,940), and two phytocannabinoids (cannabidiol (CBD) and Δ(9)-tetrahydrocannabinol (THC)) on arrestin2-, Gα(i/o)-, Gβγ-, Gα(s)-, and Gα(q)-mediated intracellular signaling in the mouse STHdh(Q7/Q7) cell culture model of striatal medium spiny projection neurons that endogenously express CB1. In this system, 2-AG, THC, and CP55,940 were more potent mediators of arrestin2 recruitment than other cannabinoids tested. 2-AG, AEA, and WIN55,212-2, enhanced Gα(i/o) and Gβγ signaling, with 2-AG and AEA treatment leading to increased total CB1 levels. 2-AG, AEA, THC, and WIN55,212-2 also activated Gα(q)-dependent pathways. CP55,940 and CBD both signaled through Gα(s). CP55,940, but not CBD, activated downstream Gα(s) pathways via CB1 targets. THC and CP55,940 promoted CB1 internalization and decreased CB1 protein levels over an 18-h period. These data demonstrate that individual cannabinoids display functional selectivity at CB1 leading to activation of distinct signaling pathways. To effectively match cannabinoids with therapeutic goals, these compounds must be screened for their signaling bias.

  2. Construction of a novel cell-surface display system for heterologous gene expression in Escherichia coli by using an outer membrane protein of Zymomonas mobilis as anchor motif.

    Science.gov (United States)

    He, Ming-Xiong; Feng, Hong; Zhang, Yi-Zheng

    2008-12-01

    A novel bacterial cell-surface display system was developed in Escherichia coli using omp1, a hypothetical outer membrane protein of Zymomonas mobilis. By using this system, we successfully expressed beta-amylase gene of sweet potato in E. coli. The display of enzyme on the membrane surface was also confirmed. The recombinant beta-amylase showed to significantly increase hydrolytic activity toward soluble starch. Our results provide a basis for constructing an engineered Z. mobilis strain directly fermenting raw starch to produce ethanol.

  3. Universal Numeric Segmented Display

    CERN Document Server

    Azad, Md Abul kalam; Kamruzzaman, S M

    2010-01-01

    Segmentation display plays a vital role to display numerals. But in today's world matrix display is also used in displaying numerals. Because numerals has lots of curve edges which is better supported by matrix display. But as matrix display is costly and complex to implement and also needs more memory, segment display is generally used to display numerals. But as there is yet no proposed compact display architecture to display multiple language numerals at a time, this paper proposes uniform display architecture to display multiple language digits and general mathematical expressions with higher accuracy and simplicity by using a 18-segment display, which is an improvement over the 16 segment display.

  4. Cell-surface area codes: mobile-element related gene switches generate precise and heritable cell-surface displays of address molecules that are used for constructing embryos.

    Science.gov (United States)

    Dreyer, W J; Roman-Dreyer, J

    1999-01-01

    We present an updated area code hypothesis supporting the proposal that cell surface display of seven-transmembrane olfactory receptors, protocadherins and other cell surface receptors provide codes that enable cells to find their correct partners as they sculpture embryos. The genetic mechanisms that program the expression of such displays have been largely unknown until very recently. However, increasing evidence now suggests that precise developmental control of the expression of these genes during embryogenesis is achieved in part by permanent and heritable changes in DNA. Using the developing immune system as a model, we discuss two different types of developmentally programmed genetic switches, each of which relies on recombination mechanisms related to mobile elements. We review new evidence suggesting the involvement of mobile element related switch mechanisms in the generation of protocadherin molecules, and their possible involvement in the control of expressions of olfactory receptors. As both recombinase and reverse transcriptase mechanisms play a role in the switching of the immunoglobulin genes, we searched the databases of expressed sequence tags (dbEST) for expression of related genes in other tissues. We present data revealing that transposases and reverse transcriptases are widely expressed in most tissues. We also searched these databases for expression of env (envelope) gene products, stimulated by provocative results suggesting that these molecules might function as cellular address receptors. We found that env genes are also expressed in large numbers in normal human tissues. One must assume that these three different types of mobile-element-related messenger RNA molecules (transposases, reverse transcriptases, and env proteins) are expressed for use in functions of value in the various tissues and have been preserved in the genome because of their selective advantages. We conclude that it is possible that many specific cell lineage decisions

  5. Eltrombopag, a thrombopoietin receptor agonist, enhances human umbilical cord blood hematopoietic stem/primitive progenitor cell expansion and promotes multi-lineage hematopoiesis.

    Science.gov (United States)

    Sun, Hongliang; Tsai, Ying; Nowak, Irena; Liesveld, Jane; Chen, Yuhchyau

    2012-09-01

    Umbilical cord blood (UCB) transplantation has emerged as a promising therapy, but it is challenged by scarcity of stem cells. Eltrombopag is a non-peptide, thrombopoietin (TPO) receptor agonist, which selectively activates c-Mpl in humans and chimpanzees. We investigated eltrombopag's effects on human UCB hematopoietic stem cell (HSC) and hematopoietic progenitor cell (HPC) expansion, and its effects on hematopoiesis in vivo. Eltrombopag selectively augmented the expansion of human CD45+, CD34+, and CD41+ cells in bone marrow compartment without effects on mouse bone marrow cells in the NOD/SCID mice xenotransplant model. Consequently, eltrombopag increased peripheral human platelets and white blood cells. We further examined effects in the STAT and AKT signaling pathways in serum-free cultures. Eltrombopag expanded human CD34+ CD38-, CD34+, and CD41+ cells. Both eltrombopag and recombinant human TPO (rhTPO) induced phosphorylation of STAT5 of CD34+ CD41-, CD34- CD41+, and CD34- CD41- cells. rhTPO preferentially induced pSTAT3, pAKT, and more pSTAT5 in CD34- C41+ cells, while eltrombopag had no effects on pSTAT3. In conclusion, eltrombopag enhanced expansion of HSCs/HPCs of human UCB in vivo and in vitro, and promoted multi-lineage hematopoiesis through the expansion of bone marrow HSCs/HPCs of human UCB in vivo. Eltrombopag differed somewhat from rhTPO in the signal transduction pathways by favoring earlier HSC/HPC populations.

  6. A receptor tyrosine kinase, UFO/Axl, and other genes isolated by a modified differential display PCR are overexpressed in metastatic prostatic carcinoma cell line DU145.

    Science.gov (United States)

    Jacob, A N; Kalapurakal, J; Davidson, W R; Kandpal, G; Dunson, N; Prashar, Y; Kandpal, R P

    1999-01-01

    We have used a modified differential display PCR protocol for isolating 3' restriction fragments of cDNAs specifically expressed or overexpressed in metastatic prostate carcinoma cell line DU145. Several cDNA fragments were identified that matched to milk fat globule protein, UFO/Axl, a receptor tyrosine kinase, human homologue of a Xenopus maternal transcript, laminin and laminin receptor, human carcinoma-associated antigen, and some expressed sequence tags. The transcript for milk fat globule protein, a marker protein shown to be overexpressed in breast tumors, was elevated in DU145 cells. The expression of UFO/Axl, a receptor tyrosine kinase, was considerably higher in DU145 cells as compared to normal prostate cells and prostatic carcinoma cell line PC-3. The overexpression of UFO oncogene in DU145 cells is discussed in the context of prostate cancer metastasis.

  7. Cell surface display of cold-active esterase EstPc with the use of a new autotransporter from Psychrobacter cryohalolentis K5(T).

    Science.gov (United States)

    Petrovskaya, L E; Novototskaya-Vlasova, K A; Kryukova, E A; Rivkina, E M; Dolgikh, D A; Kirpichnikov, M P

    2015-01-01

    We have cloned the gene coding for AT877-a new predicted member of the autotransporter protein family with an esterase passenger domain from permafrost bacterium Psychrobacter cryohalolentis K5(T). Expression of AT877 gene in Escherichia coli resulted in accumulation of the recombinant autotransporter in the outer membrane fraction and at the surface of the induced cells. AT877 displayed maximum hydrolytic activity toward medium-chain p-nitrophenyl esters (C8-C10) at 50 °C and was resistant to the presence of several metal ions, organic solvents and detergents. Previously, we have described a cold-active esterase EstPc from the same bacterium which possesses high activity at low temperatures and relatively high thermal stability. To construct a cell surface display system for EstPc, the hybrid autotransporter gene coding for EstPc with the α-helical linker and the translocator domain from AT877 was constructed and expressed in E. coli. According to the results of the cell fractionation studies and esterase activity measurements, the EstPc passenger was successfully displayed at the surface of the induced cells. It demonstrated a temperature optimum at 15-25 °C and a substrate preference toward p-nitrophenyl butyrate (C4). Obtained results provide a new example of the biotechnologically relevant enzyme from the permafrost microbial community with potential applications for the conversion of short- and medium-chain ester substrates and a basis for the construction of a new cell surface display platform.

  8. Reconfigurable Full-Page Braille Displays

    Science.gov (United States)

    Garner, H. Douglas

    1994-01-01

    Electrically actuated braille display cells of proposed type arrayed together to form full-page braille displays. Like other braille display cells, these provide changeable patterns of bumps driven by digitally recorded text stored on magnetic tapes or in solid-state electronic memories. Proposed cells contain electrorheological fluid. Viscosity of such fluid increases in strong electrostatic field.

  9. Refreshing Refreshable Braille Displays.

    Science.gov (United States)

    Russomanno, Alexander; O'Modhrain, Sile; Gillespie, R Brent; Rodger, Matthew W M

    2015-01-01

    The increased access to books afforded to blind people via e-publishing has given them long-sought independence for both recreational and educational reading. In most cases, blind readers access materials using speech output. For some content such as highly technical texts, music, and graphics, speech is not an appropriate access modality as it does not promote deep understanding. Therefore blind braille readers often prefer electronic braille displays. But, these are prohibitively expensive. The search is on, therefore, for a low-cost refreshable display that would go beyond current technologies and deliver graphical content as well as text. And many solutions have been proposed, some of which reduce costs by restricting the number of characters that can be displayed, even down to a single braille cell. In this paper, we demonstrate that restricting tactile cues during braille reading leads to poorer performance in a letter recognition task. In particular, we show that lack of sliding contact between the fingertip and the braille reading surface results in more errors and that the number of errors increases as a function of presentation speed. These findings suggest that single cell displays which do not incorporate sliding contact are likely to be less effective for braille reading.

  10. Cell lines, Md108 and Md66, from the hemocytes of Malacosoma disstria (Lepidoptera) display aspects of plasma-free innate non-self activities.

    Science.gov (United States)

    Lapointe, Jason F; Dunphy, Gary B; Giannoulis, Paschalis; Mandato, Craig A; Nardi, James B; Gharib, Osama H; Niven, Donald F

    2011-11-01

    The innate non-self response systems of the deciduous tree pest, the forest tent caterpillar, Malacosoma disstria has been documented by us in terms of in vitro and in vivo reactions towards the Gram-positive nonpathogenic bacterium, Bacillus subtilis and Gram-negative pathogenic microbe, Xenorhabdus nematophila and their respective surface antigens, lipopoteichoic acids (LTA) and lipopolysaccharides (LPS). These studies, often conducted in whole and diluted hemolymph, preclude examination of plasma-free cellular (hemocyte) responses. Plasma-free hemocytes as primary cultures are difficult to obtain. The floating cell line Md66 and attached cell line Md108 from M. disstria hemocytes were examined as a model for plasma-free M. disstria hemocyte non-self responses. Herein, it was established that although both lines differed from each other and from the primary hemocyte cultures of M. disstria in growth parameters, cell composition and sizes both cell lines displayed granular cell-like (GL) cells and plasmatocyte-like (PL) cells according to morphological criteria and to some extent antigenic similarities based on labeling with anti-Chrysodeixis includens hemocyte monoclonal antibodies. Hemocyte-specific neuroglian-like protein was detected on cells of both cell lines and in the primary hemocyte cultures albeit with staining patterns differing according to culture and cell types, confluency levels and cell-cell adhesion. Both cell lines bound B. subtilis and X. nematophila, the reaction extent varying with the cell line and its cell types. LPS damaged both cell types in the two cell lines whereas LTA enhanced the adhesion of Md66 GL cells to flask surfaces followed by PL cell adhesion. PL cells of both lines, like the primary cultures, phagocytosed FITC-labeled B. subtilis; only Md108 GL cells phagocytosed B. subtilis. In either case phagocytosis was always less in frequency and intensity than the primary cultures. Proteins released from the cell lines differed in

  11. Epithelial cells derived from human embryonic stem cells display p16INK4A senescence, hypermotility, and differentiation properties shared by many P63+ somatic cell types

    DEFF Research Database (Denmark)

    Dabelsteen, Sally; Hercule, Paula; Barron, Patricia

    2009-01-01

    Human embryonic stem (hES) cells can generate cells expressing p63, K14, and involucrin, which have been proposed to be keratinocytes. Although these hES-derived, keratinocyte-like (hESderK) cells form epithelioid colonies when cultured in a fibroblast feeder system optimal for normal tissue......(+)/K14(+) urothelial and tracheobronchial epithelial cells. Primary and immortalized lines of these cell types had growth requirements and hypermotility responses similar to keratinocytes and bmi1 expression facilitated their immortalization by engineering to express the catalytic subunit of telomerase...

  12. DNA display selection of peptide ligands for a full-length human G protein-coupled receptor on CHO-K1 cells.

    Directory of Open Access Journals (Sweden)

    Nobuhide Doi

    Full Text Available The G protein-coupled receptors (GPCRs, which form the largest group of transmembrane proteins involved in signal transduction, are major targets of currently available drugs. Thus, the search for cognate and surrogate peptide ligands for GPCRs is of both basic and therapeutic interest. Here we describe the application of an in vitro DNA display technology to screening libraries of peptide ligands for full-length GPCRs expressed on whole cells. We used human angiotensin II (Ang II type-1 receptor (hAT1R as a model GPCR. Under improved selection conditions using hAT1R-expressing Chinese hamster ovary (CHO-K1 cells as bait, we confirmed that Ang II gene could be enriched more than 10,000-fold after four rounds of selection. Further, we successfully selected diverse Ang II-like peptides from randomized peptide libraries. The results provide more precise information on the sequence-function relationships of hAT1R ligands than can be obtained by conventional alanine-scanning mutagenesis. Completely in vitro DNA display can overcome the limitations of current display technologies and is expected to prove widely useful for screening diverse libraries of mutant peptide and protein ligands for receptors that can be expressed functionally on the surface of CHO-K1 cells.

  13. The normal counterpart of IgD myeloma cells in germinal center displays extensively mutated IgVH gene, Cmu-Cdelta switch, and lambda light chain expression.

    Science.gov (United States)

    Arpin, C; de Bouteiller, O; Razanajaona, D; Fugier-Vivier, I; Brière, F; Banchereau, J; Lebecque, S; Liu, Y J

    1998-04-20

    Human myeloma are incurable hematologic cancers of immunoglobulin-secreting plasma cells in bone marrow. Although malignant plasma cells can be almost eradicated from the patient's bone marrow by chemotherapy, drug-resistant myeloma precursor cells persist in an apparently cryptic compartment. Controversy exists as to whether myeloma precursor cells are hematopoietic stem cells, pre-B cells, germinal center (GC) B cells, circulating memory cells, or plasma blasts. This situation reflects what has been a general problem in cancer research for years: how to compare a tumor with its normal counterpart. Although several studies have demonstrated somatically mutated immunoglobulin variable region genes in multiple myeloma, it is unclear if myeloma cells are derived from GCs or post-GC memory B cells. Immunoglobulin (Ig)D-secreting myeloma have two unique immunoglobulin features, including a biased lambda light chain expression and a Cmu-Cdelta isotype switch. Using surface markers, we have previously isolated a population of surface IgM-IgD+CD38+ GC B cells that carry the most impressive somatic mutation in their IgV genes. Here we show that this population of GC B cells displays the two molecular features of IgD-secreting myeloma cells: a biased lambda light chain expression and a C&mu-Cdelta isotype switch. The demonstration of these peculiar GC B cells to differentiate into IgD-secreting plasma cells but not memory B cells both in vivo and in vitro suggests that IgD-secreting plasma and myeloma cells are derived from GCs.

  14. Flexible Bistable Cholesteric Reflective Displays

    Science.gov (United States)

    Yang, Deng-Ke

    2006-03-01

    Cholesteric liquid crystals (ChLCs) exhibit two stable states at zero field condition-the reflecting planar state and the nonreflecting focal conic state. ChLCs are an excellent candidate for inexpensive and rugged electronic books and papers. This paper will review the display cell structure,materials and drive schemes for flexible bistable cholesteric (Ch) reflective displays.

  15. Efficient display of active Geotrichum sp. lipase on Pichia pastoris cell wall and its application as a whole-cell biocatalyst to enrich EPA and DHA in fish oil.

    Science.gov (United States)

    Pan, Xiao-Xing; Xu, Li; Zhang, Yan; Xiao, Xiao; Wang, Xiao-Feng; Liu, Yun; Zhang, Hou-jin; Yan, Yun-Jun

    2012-09-26

    Geotrichum sp. lipase (GSL) was first displayed on the cell wall of Pichia pastoris on the basis of the a-agglutinin anchor system developed in Saccharomyces cerevisiae . Surface display levels were monitored using Western blotting, immunofluorescence miscroscopy, and fluorescence-activated cell sorting analysis. Lipase activity of the yeast whole cells reached a maximum at 273 ± 2.4 U/g of dry cells toward olive oil after 96 h of culture at 30 °C, with optimal pH and temperature at 7.5 and 45 °C, respectively. Displayed GSL exhibited relatively high stability between pH 6.0 and 8.0 and retained >70% of the maximum activity. The surface-displayed lipase retained 80% of its original activity after incubation at 45 °C for 4 h. Moreover, the GSL-displaying yeast whole cells were then used as a biocatalyst to enrich eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) from fish oil on the basis of selective hydrolysis. As a result, EPA and DHA increased from 1.53 and 24.1% in the original fish oil to 1.85 and 30.86%, which were increases of 1.21- and 1.29-fold, respectively. The total yield of EPA and DHA reached 46.62%.

  16. Berberine and Curcumin Target Survivin and STAT3 in Gastric Cancer Cells and Synergize Actions of Standard Chemotherapeutic 5-Fluorouracil.

    Science.gov (United States)

    Pandey, Arvind; Vishnoi, Kanchan; Mahata, Sutapa; Tripathi, Satyendra Chandra; Misra, Sri Prakash; Misra, Vatsala; Mehrotra, Ravi; Dwivedi, Manisha; Bharti, Alok C

    2015-01-01

    Aberrantly expressed survivin and STAT3 signaling have emerged as major determinants of chemoresistance in gastric cancer. We evaluated effects of potent herbal derivatives curcumin, berberine, and quercetin on STAT3 signaling, survivin expression, and response to 5-fluorouracil (5-FU) treatment in gastric cancer cells (AGS). Cytotoxic and inhibitory effects of berberine, curcumin, and quercetin alone or in combination with 5-FU were examined by MTT assay, and their effect on survivin, STAT3, and the phosphorylated active STAT3 (pSTAT3) expression was examined by western blotting. Effect of these herbal derivatives on STAT3 DNA binding activity was measured by electrophoretic mobility shift assay. Curcumin, berberine, and quercetin effectively downregulated pSTAT3 levels, survivin expression, and gastric cancer cells viability in a dose-dependent manner (with corresponding IC50 values of 40.3μM, 29.2μM and 37.5μM, respectively). Berberine was more effective in inhibiting survivin expression as compared to other herbal agents. 5-FU in combination with berberine or curcumin showed a synergistic inhibition of survivin and STAT3 level resulting in enhanced cell death in gastric cancer cells. Overall, our data suggest use of berberine and curcumin as adjunct therapeutics to overcome chemoresistance during treatment of gastric malignancies.

  17. Effects of Kupffer cell inactivation on graft survival and liver regeneration after partial liver transplantation in rats

    Institute of Scientific and Technical Information of China (English)

    Hang-Yu Luo; Shan-Fang Ma; Ji-Fu Qu; De-Hu Tian

    2015-01-01

    BACKGROUND: Gadolinium chloride (GdCl3) selectively in-activates Kupffer cells and protects against ischemia/reperfu-sion and endotoxin injury. However, the effect of Kupffer cell inactivation on liver regeneration after partial liver transplan-tation (PLTx) is not clear. This study was to investigate the role of GdCl3 pretreatment in graft function after PLTx, and to explore the potential mechanism involved in this process. METHODS: PLTx (30% partial liver transplantation) was per-formed using Kamada's cuff technique, without hepatic artery reconstruction. Rats were randomly divided into the control low-dose (5 mg/kg) and high-dose (10 mg/kg) GdCl3 groups. Liver injury was determined by the plasma levels of alanine aminotransferase and aspartate aminotransferase, liver regen-eration by PCNA staining and BrdU uptake, apoptosis by TU-NEL assay. IL-6 and p-STAT3 levels were measured by ELISA and Western blotting. RESULTS: GdCl3 depleted Kupffer cells and decreased animal survival rates, but did not significantly affect alanine amino-transferase and aspartate aminotransferase (P>0.05). GdCl3 pretreatment induced apoptosis and inhibited IL-6 overex-pression and STAT3 phosphorylation after PLTx in graft tissues. CONCLUSION: Kupffer cells may contribute to the liver re-generation after PLTx through inhibition of apoptosis and activation of the IL-6/p-STAT3 signal pathway.

  18. Increased micronucleated cell frequency related to exposure to radiation emitted by computer cathode ray tube video display monitors

    OpenAIRE

    Karina Carbonari; Luciane Gonçalves; Daniela Roth; Patrick Moreira; Ricardo Fernández; Maria da Graça Martino-Roth

    2005-01-01

    It is well recognized that electromagnetic fields can affect the biological functions of living organisms at both cellular and molecular level. The potential damaging effects of electromagnetic fields and very low frequency and extremely low frequency radiation emitted by computer cathode ray tube video display monitors (VDMs) has become a concern within the scientific community. We studied the effects of occupational exposure to VDMs in 10 males and 10 females occupationally exposed to VDMs ...

  19. Nisin production in a chitin-including continuous fermentation system with Lactococcus lactis displaying a cell wall chitin-binding domain.

    Science.gov (United States)

    Şimşek, Ömer

    2014-03-01

    The limiting factors in the continuous production of nisin are high amount of biomass loss and low dilution rate application. In this study, a chitin-including continuous nisin fermentation system (CICON-FER) was constructed for high volumetric nisin production using nisin producer L. lactis displaying cell wall chitin-binding domain (ChBD) together with chitin in the reactor. In this respect, the highest binding conditions of relevant L. lactis cells to chitin were determined. Then the chitin flakes carrying nisin-producing L. lactis cells were used within the CICON-FER system at different dilution rates (0.1-0.9 h⁻¹) and initial glucose concentrations (20-60 g l⁻¹). The results revealed that the pH 7 conditions and the use of 100 mM sodium phosphate buffer with 0.1 % Tween 20 and Triton X-100 significantly increased the binding capacity of ChBD displaying L. lactis cells to chitin. The constructed CICON-FER system maintained the presence of the ChBD surface displaying L. lactis cells in the reactor system until 0.9 h⁻¹ dilution rate that resulted in a considerably high level of volumetric nisin production and productivity (10,500 IU ml⁻¹ and 9,450 IU ml⁻¹ h⁻¹, respectively) with the combination of a 0.9-h⁻¹ dilution rate and a 40-g l⁻¹ initial glucose concentration. In conclusion, an innovative nisin fermentation system that yielded the highest nisin production thus far and that was feasible for industrial application was created.

  20. Identification of a conserved B-cell epitope on reticuloendotheliosis virus envelope protein by screening a phage-displayed random peptide library.

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    Mei Xue

    Full Text Available BACKGROUND: The gp90 protein of avian reticuloendotheliosis-associated virus (REV-A is an important envelope glycoprotein, which is responsible for inducing protective antibody immune responses in animals. B-cell epitopes on the gp90 protein of REV have not been well studied and reported. METHODS AND RESULTS: This study describes the identification of a linear B-cell epitope on the gp90 protein by screening a phage-displayed 12-mer random peptide library with the neutralizing monoclonal antibody (mAb A9E8 directed against the gp90. The mAb A9E8 recognized phages displaying peptides with the consensus motif SVQYHPL. Amino acid sequence of the motif exactly matched (213SVQYHPL(219 of the gp90. Further identification of the displayed B cell epitope was conducted using a set of truncated peptides expressed as GST fusion proteins and the Western blot results indicated that (213SVQYHPL(219 was the minimal determinant of the linear B cell epitope recognized by the mAb A9E8. Moreover, an eight amino acid peptide SVQYHPLA was proven to be the minimal unit of the epitope with the maximal binding activity to mAb A9E8. The REV-A-positive chicken serum reacted with the minimal linear epitopes in Western blot, revealing the importance of the eight amino acids of the epitope in antibody-epitope binding activity. Furthermore, we found that the epitope is a common motif shared among REV-A and other members of REV group. CONCLUSIONS AND SIGNIFICANCE: We identified (213SVQYHPL(219 as a gp90-specific linear B-cell epitope recognized by the neutralizing mAb A9E8. The results in this study may have potential applications in development of diagnostic techniques and epitope-based marker vaccines against REV-A and other viruses of the REV group.

  1. Cord blood Vα24-Vβ11 natural killer T cells display a Th2-chemokine receptor profile and cytokine responses.

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    Susanne Harner

    Full Text Available BACKGROUND: The fetal immune system is characterized by a Th2 bias but it is unclear how the Th2 predominance is established. Natural killer T (NKT cells are a rare subset of T cells with immune regulatory functions and are already activated in utero. To test the hypothesis that NKT cells are part of the regulatory network that sets the fetal Th2 predominance, percentages of Vα24(+Vβ11(+ NKT cells expressing Th1/Th2-related chemokine receptors (CKR were assessed in cord blood. Furthermore, IL-4 and IFN-γ secreting NKT cells were quantified within the single CKR(+ subsets. RESULTS: Cord blood NKT cells expressed the Th2-related CCR4 and CCR8 at significantly higher frequencies compared to peripheral blood NKT cells from adults, while CXCR3(+ and CCR5(+ cord blood NKT cells (Th1-related were present at lower percentages. Within CD4(negCD8(neg (DN NKT cells, the frequency of IL-4 producing NKT cells was significantly higher in cord blood, while frequencies of IFN-γ secreting DN NKT cells tended to be lower. A further subanalysis showed that the higher percentage of IL-4 secreting DN NKT cells was restricted to CCR3(+, CCR4(+, CCR5(+, CCR6(+, CCR7(+, CCR8(+ and CXCR4(+ DN subsets in cord blood. This resulted in significantly decreased IFN-γ /IL-4 ratios of CCR3(+, CCR6(+ and CCR8(+ cord blood DN NKT cells. Sequencing of VA24AJ18 T cell receptor (TCR transcripts in sorted cord blood Vα24Vβ11 cells confirmed the invariant TCR alpha-chain ruling out the possibility that these cells represent an unusual subset of conventional T cells. CONCLUSIONS: Despite the heterogeneity of cord blood NKT cells, we observed a clear Th2-bias at the phenotypic and functional level which was mainly found in the DN subset. Therefore, we speculate that NKT cells are important for the initiation and control of the fetal Th2 environment which is needed to maintain tolerance towards self-antigens as well as non-inherited maternal antigens.

  2. Non-pungent long chain capsaicin-analogs arvanil and olvanil display better anti-invasive activity than capsaicin in human small cell lung cancers.

    Science.gov (United States)

    Hurley, John D; Akers, Austin T; Friedman, Jamie R; Nolan, Nicholas A; Brown, Kathleen C; Dasgupta, Piyali

    2017-01-02

    The nutritional compound capsaicin inhibits the invasion of many types of human cancers. The clinical development of capsaicin as an anti-cancer drug is limited due to its unfavorable side effects like burning sensation, stomach cramps, gut pain and nausea. This study compared the anti-invasive activity of capsaicin to non-pungent long chain capsaicin analogs, namely arvanil and olvanil, in human small cell lung cancer cells. Boyden chamber invasion assays revealed that arvanil and olvanil displayed improved anti-invasive activity relative to capsaicin in human SCLC cells. The results of the Boyden chamber assay were confirmed by the spherical invasion assay, and similar results were obtained. The anti-invasive activity of arvanil, olvanil and capsaicin were independent of TRPV and CB1 receptors. Furthermore, the anti-invasive activity of arvanil, olvanil and capsaicin was mediated by the AMPK pathway. Depletion of AMPK levels by siRNA methodology abrogated the anti-invasive activity of arvanil, olvanil and capsaicin. The non-pungent capsaicin analogs arvanil and olvanil display improved anti-invasive activity relative to capsaicin in human SCLC cells. These agents may represent the second generation of capsaicin-like compounds which are more potent than the parent molecule and have a better side effect profile.

  3. HIV-infected viremic long-term non-progressors and controllers display different immunological mechanisms for preserved CD4+cell counts

    DEFF Research Database (Denmark)

    Gaardbo, J; Ronit, A; Hartling, H

    2012-01-01

    (viral load, VL>5000 copies/ml, CD4+ cell count>350 cells/ul, infected>10 years), 30 VC (VL350 cells/ul), and 25 progressors (PR) (VL>5.000 copies/ml, CD4 count>350 cells/ul) were included. Immune activation (CD4+ and CD8+cells co-expressing CD38+HLA-DR+), apoptosis (CD8+CD28-CD95+), Th17 cells (CD4+CD.......4% vs. 1.6%, P=0.007, 21.7% vs. 12.0%, P=0.051) and similar levels to PR (4.2%, 22.4%, P>0.05). Likewise, LTNP had higher frequency of apoptotic cells compared to VC (63.7% vs. 48.6%, P=0.0408) and similar levels to PR (63.8%, P>0.05). Interestingly, borderline significant trends towards lower Th17/Treg...... ratio in LTNP compared to VC were found (3.8 vs. 5.5, P=0.068) while ratios in LTNP and PR were similar (4.0, P>0.05). Conclusion: LTNP displayed high levels of immune activation, apoptotic cells and reduced Th17/Treg ratio compared to VC, while LTNP were similar to PR. Thus, the immunological mechanism...

  4. Development of an Ussuri catfish Pseudobagrus ussuriensis skin cell line displaying differential cytopathic effects to three aquatic animal viruses.

    Science.gov (United States)

    Ou, Tong; Lei, Xiao-Ying; He, Li-Bo; Zhou, Feng-Jian; Zhang, Qi-Ya

    2014-08-30

    An Ussuri catfish Pseudobagrus ussuriensis skin (UCS) cell line was developed and subcultured for more than 60 passages. UCS cells consisted of mostly epithelial-like cells and multiplied well in TC199 medium supplemented with 10% fetal bovine serum at 25°C. Chromosome analysis revealed that most UCS cells had a normal diploid karyotype with 2n=52. UCS cells showed differential cytopathic effects (CPEs) after inoculation of spring viremia of carp virus (SVCV, a negative-strand RNA virus), grass carp reovirus (GCRV, a multi-segmented double-stranded RNA virus) and Rana grylio virus (RGV, a large double-stranded DNA virus), and were indicative of high sensitivities to these three aquatic animal viruses by a virus titration study. The CPE caused by SVCV appeared as rounded and granular cells, grape-like clusters and small lytic plaques. Characteristic CPE containing plaque-like syncytia was induced by GCRV. RGV-infected cells produced typical CPE characterized by cells shrinkage and aggregation, formation of clear plaques and cell sheet detachment. Furthermore, significant fluorescent signals were observed after UCS cells were transfected with green fluorescent protein reporter plasmids, and the development of CPE induced by a recombinant RGV, ΔTK-RGV, in UCS cells was illustrated using a combination of light and fluorescence microscopy. The data from this study suggested that UCS cell line can potentially serve as a useful tool for the comparison study of different aquatic animal viruses and the isolation of some newly emerging viruses in Ussuri catfish farming.

  5. Curcumol induces apoptosis in SPC-A-1 human lung adenocarcinoma cells and displays anti-neoplastic effects in tumor bearing mice.

    Science.gov (United States)

    Tang, Qi-Ling; Guo, Ji-Quan; Wang, Qi-You; Lin, Hai-Shu; Yang, Zhou-Ping; Peng, Tong; Pan, Xue-Diao; Liu, Bing; Wang, Su-Jun; Zang, Lin-Quan

    2015-01-01

    Curcumol is a sesquiterpene originally isolated from curcuma rhizomes, a component of herbal remedies commonly used in oriental medicine. Its beneficial pharmacological activities have attract significant interest recently. In this study, anti-cancer activity of curcumol was examined with both in vitro and in vivo models. It was found that curcumol exhibited time- and concentration-dependent anti-proliferative effects in SPC-A-1 human lung adenocarcinoma cells with cell cycle arrest in the G0/G1 phase while apoptosis-induction was also confirmed with flow cytometry and morphological analyses. Interestingly, curcumol did not display growth inhibition in MRC-5 human embryonic lung fibroblasts, suggesting the anti-proliferative effects of curcumol were specific to cancer cells. Anti-neoplastic effects of curcumol were also confirmed in tumor bearing mice. Curcumol (60 mg/kg daily) significantly reduced tumor size without causing notable toxicity. In conclusion, curcumol appears a favorable anti-cancer candidate for further development.

  6. Cell surface display of minor pilin adhesins in the form of a simple heterodimeric assembly in Corynebacterium diphtheriae.

    Science.gov (United States)

    Chang, Chungyu; Mandlik, Anjali; Das, Asis; Ton-That, Hung

    2011-03-01

    Pilus assembly in Gram-positive bacteria occurs by a two-step mechanism, whereby pilins are polymerized and then covalently anchored to the cell wall. In Corynebacterium diphtheriae, the pilin-specific sortase SrtA catalyses polymerization of the SpaA-type pilus, consisting of the shaft pilin SpaA, tip pilin SpaC and minor pilin SpaB. Cell wall anchoring of the SpaA polymers is triggered when SrtA incorporates SpaB into the pilus base via lysine-mediated transpeptidation; anchoring to the cell wall peptidoglycan is subsequently catalysed by the housekeeping sortase SrtF. Here we show that SpaB and SpaC formed a heterodimer independent of SpaA polymerization. SrtA was absolutely required for the formation of the SpaBC heterodimer, while SrtF facilitated the optimal cell wall anchoring of this heterodimer. Alanine substitution of the SpaB lysine residue K139 or truncation of the SpaB cell wall-sorting signal (CWSS) abolished assembly of the SpaBC heterodimer, hence underscoring SpaB function in transpeptidation and cell wall linkage. Importantly, sortase specificity for the cell wall-anchoring step was found to be dependent on the LAFTG motif within the SpaB CWSS. Thus, C. diphtheriae employs a common sortase-catalysed mechanism involving lysine-mediated transpeptidation to generate both adhesive pilus and simple heterodimeric structures on the bacterial the cell wall.

  7. Cytotoxic Compounds from Juglans sinensis Dode Display Anti-Proliferative Activity by Inducing Apoptosis in Human Cancer Cells.

    Science.gov (United States)

    Lee, Yoo Jin; Cui, Jun; Lee, Jun; Han, Ah-Reum; Lee, Eun Byul; Jang, Ho Hee; Seo, Eun Kyoung

    2016-01-01

    Phytochemical investigation of the bark of Juglans sinensis Dode (Juglandaceae) led to the isolation of two active compounds, 8-hydroxy-2-methoxy-1,4-naphthoquinone (1) and 5-hydroxy-2-methoxy-1,4-naphthoquinone (2), together with 15 known compounds 3-17. All compounds were isolated from this plant for the first time. The structures of 1 and 2 were elucidated by spectroscopic data analysis, including 1D and 2D NMR experiments. Compounds 1-17 were tested for their cytotoxicity against the A549 human lung cancer cell line; compounds 1 and 2 exhibited significant cytotoxicity and additionally had potent cytotoxicity against six human cancer cell lines, MCF7 (breast cancer), SNU423 (liver cancer), SH-SY5Y (neuroblastoma), HeLa (cervical cancer), HCT116 (colorectal cancer), and A549 (lung cancer). In particular, breast, colon, and lung cancer cells were more sensitive to the treatment using compound 1. In addition, compounds 1 and 2 showed strong cytotoxic activity towards human breast cancer cells MCF7, HS578T, and T47D, but not towards MCF10A normal-like breast cells. They also inhibited the colony formation of MCF7, A549, and HCT116 cells in a dose-dependent manner. Flow cytometry analysis revealed that the percentage of apoptotic cells significantly increased in MCF7 cells upon the treatment with compounds 1 and 2. The mechanism of cell death caused by compounds 1 and 2 may be attributed to the upregulation of Bax and downregulation of Bcl2. These findings suggest that compounds 1 and 2 may be regarded as potential therapeutic agents against cancer.

  8. Human decidual macrophages and NK cells differentially express Toll-like receptors and display distinct cytokine profiles upon TLR stimulation.

    Directory of Open Access Journals (Sweden)

    Marion eDuriez

    2014-07-01

    Full Text Available Maternofetal pathogen transmission is partially controlled at the level of the maternal uterine mucosa at the fetal implantation site (the decidua basalis, where maternal and fetal cells are in close contact. Toll-like receptors (TLRs may play an important role in initiating rapid immune responses against pathogens in the decidua basalis, however the tolerant microenvironment should be preserved in order to allow fetal development. Here we investigated the expression and functionality of TLRs expressed by decidual macrophages (dMs and NK cells (dNKs, the major decidual immune cell populations.We report for the first time that both human dMs and dNK cells express mRNAs encoding TLRs 1-9, albeit with a higher expression level in dMs. TLR2, TLR3 and TLR4 protein expression checked by flow cytometry was positive for both dMs and dNK cells. In vitro treatment of primary dMs and dNK cells with specific TLR2, TLR3, TLR4, TLR7/8 and TLR9 agonists enhanced their secretion of pro- and anti-inflammatory cytokines, as well as cytokines and chemokines involved in immune cell crosstalk. Only dNK cells released IFN-γ, whereas only dMs released IL-1β, IL-10 and IL-12. TLR9 activation of dMs resulted in a distinct pattern of cytokine expression compared to the other TLRs. The cytokine profiles expressed by dMs and dNK cells upon TLR activation are compatible with maintenance of the fetotolerant immune environment during initiation of immune responses to pathogens at the maternofetal interface.

  9. STAT3 activation is associated with cerebrospinal fluid interleukin-10 (IL-10) in primary central nervous system diffuse large B cell lymphoma.

    Science.gov (United States)

    Mizowaki, Takashi; Sasayama, Takashi; Tanaka, Kazuhiro; Mizukawa, Katsu; Takata, Kumi; Nakamizo, Satoshi; Tanaka, Hirotomo; Nagashima, Hiroaki; Nishihara, Masamitsu; Hirose, Takanori; Itoh, Tomoo; Kohmura, Eiji

    2015-09-01

    Signal transducers and activators of transcription 3 (STAT3) are activated by various cytokines and oncogenes; however, the activity and pathogenesis of STAT3 in diffuse large B cell lymphoma of the central nervous system have not been thoroughly elucidated. We investigated the phosphorylation levels of STAT3 in 40 specimens of primary central nervous system diffuse large B-cell lymphoma (PCNS DLBCL) and analyzed the association between phsopho-STAT3 (pSTAT3) expression and cerebrospinal fluid (CSF) concentration of interleukin-10 (IL-10) or IL-6. Immunohistochemistry and Western blot analysis revealed that most of the specimens in PCNS DLBCL expressed pSTST3 protein, and a strong phosphorylation levels of STAT3 was statistically associated with high CSF IL-10 levels, but not with CSF IL-6 levels. Next, we demonstrated that recombinant IL-10 and CSF containing IL-10 induced the phosphorylation of STAT3 in PCNS DLBCL cells. Furthermore, molecular subtype classified by Hans' algorithm was correlated with pSTAT3 expression levels and CSF IL-10 levels. These results suggest that the STAT3 activity is correlated with CSF IL-10 level, which is a useful marker for STAT3 activity in PCNS DLBCLs.

  10. Primary T-cells from human CD4/CCR5-transgenic rats support all early steps of HIV-1 replication including integration, but display impaired viral gene expression

    Directory of Open Access Journals (Sweden)

    Hermann Volker

    2007-07-01

    Full Text Available Abstract Background In vivo studies on HIV-1 pathogenesis and testing of antiviral strategies have been hampered by the lack of an immunocompetent small animal model that is highly susceptible to HIV-1 infection. Since native rodents are non-permissive, we developed transgenic rats that selectively express the HIV-1 receptor complex, hCD4 and hCCR5, on relevant target cells. These animals display a transient low-level plasma viremia after HIV-1YU-2 infection, demonstrating HIV-1 susceptibility in vivo. However, unlike macrophages, primary CD4 T-cells from double-transgenic animals fail to support viral spread ex vivo. To identify quantitative limitations or absolute blocks in this rodent species, we quantitatively assessed the efficiency of key steps in the early phase of the viral replication cycle in a side-by-side comparison in infected cell lines and primary T-cells from hCD4/hCCR5-transgenic rats and human donors. Results Levels of virus entry, HIV-1 cDNA synthesis, nuclear import, and integration into the host genome were shown to be remarkably similar in cell lines and, where technically accessible, in primary T-cells from both species. In contrast, a profound impairment at the level of early HIV gene expression was disclosed at the single-cell level in primary rat T-cells and most other rat-derived cells. Macrophages were a notable exception, possibly reflecting the unique transcriptional milieu in this evolutionarily conserved target cell of all lentiviruses. Importantly, transient trans-complementation by ex vivo nucleofection with the Tat-interacting protein Cyclin T1 of human origin markedly elevated HIV gene expression in primary rat T-cells. Conclusion This is the first study that has quantitatively determined the efficiency of consecutive steps in the HIV-1 replication cycle in infected primary HIV target cells from a candidate transgenic small animal and compared it to human cells. Unlike cells derived from mice or rabbits, rat

  11. Human adipose stromal cells (ASC for the regeneration of injured cartilage display genetic stability after in vitro culture expansion.

    Directory of Open Access Journals (Sweden)

    Simona Neri

    Full Text Available Mesenchymal stromal cells are emerging as an extremely promising therapeutic agent for tissue regeneration due to their multi-potency, immune-modulation and secretome activities, but safety remains one of the main concerns, particularly when in vitro manipulation, such as cell expansion, is performed before clinical application. Indeed, it is well documented that in vitro expansion reduces replicative potential and some multi-potency and promotes cell senescence. Furthermore, during in vitro aging there is a decrease in DNA synthesis and repair efficiency thus leading to DNA damage accumulation and possibly inducing genomic instability. The European Research Project ADIPOA aims at validating an innovative cell-based therapy where autologous adipose stromal cells (ASCs are injected in the diseased articulation to activate regeneration of the cartilage. The primary objective of this paper was to assess the safety of cultured ASCs. The maintenance of genetic integrity was evaluated during in vitro culture by karyotype and microsatellite instability analysis. In addition, RT-PCR array-based evaluation of the expression of genes related to DNA damage signaling pathways was performed. Finally, the senescence and replicative potential of cultured cells was evaluated by telomere length and telomerase activity assessment, whereas anchorage-independent clone development was tested in vitro by soft agar growth. We found that cultured ASCs do not show genetic alterations and replicative senescence during the period of observation, nor anchorage-independent growth, supporting an argument for the safety of ASCs for clinical use.

  12. A novel DNMT3B splice variant expressed in tumor and pluripotent cells modulates genomic DNA methylation patterns and displays altered DNA binding.

    Science.gov (United States)

    Gopalakrishnan, Suhasni; Van Emburgh, Beth O; Shan, Jixiu; Su, Zhen; Fields, C Robert; Vieweg, Johannes; Hamazaki, Takashi; Schwartz, Philip H; Terada, Naohiro; Robertson, Keith D

    2009-10-01

    DNA methylation is an epigenetic mark essential for mammalian development, genomic stability, and imprinting. DNA methylation patterns are established and maintained by three DNA methyltransferases: DNMT1, DNMT3A, and DNMT3B. Interestingly, all three DNMTs make use of alternative splicing. DNMT3B has nearly 40 known splice variants expressed in a tissue- and disease-specific manner, but very little is known about the role of these splice variants in modulating DNMT3B function. We describe here the identification and characterization of a novel alternatively spliced form of DNMT3B lacking exon 5 within the NH(2)-terminal regulatory domain. This variant, which we term DNMT3B3Delta5 because it is closely related in structure to the ubiquitously expressed DNMT3B3 isoform, is highly expressed in pluripotent cells and brain tissue, is downregulated during differentiation, and is conserved in the mouse. Creation of pluripotent iPS cells from fibroblasts results in marked induction of DNMT3B3Delta5. DNMT3B3Delta5 expression is also altered in human disease, with tumor cell lines displaying elevated or reduced expression depending on their tissue of origin. We then compared the DNA binding and subcellular localization of DNMT3B3Delta5 versus DNMT3B3, revealing that DNMT3B3Delta5 possessed significantly enhanced DNA binding affinity and displayed an altered nuclear distribution. Finally, ectopic overexpression of DNMT3B3Delta5 resulted in repetitive element hypomethylation and enhanced cell growth in a colony formation assay. Taken together, these results show that DNMT3B3Delta5 may play an important role in stem cell maintenance or differentiation and suggest that sequences encoded by exon 5 influence the functional properties of DNMT3B.

  13. Cell surface-engineered yeast displaying a histidine oligopeptide (hexa-His) has enhanced adsorption of and tolerance to heavy metal ions.

    Science.gov (United States)

    Kuroda, K; Shibasaki, S; Ueda, M; Tanaka, A

    2001-12-01

    A histidine oligopeptide (hexa-His) with the ability to chelate divalent heavy metal ions was displayed on the yeast cell surface for the purpose of enhanced adsorption of heavy metal ions. We genetically fused a hexa-His-encoding gene with the gene encoding the C-terminal half of alpha-agglutinin that includes a glycosylphosphatidylinositol anchor attachment signal sequence and attached the hexa-His peptide on the cell wall of Saccharomyces cerevisiae. This surface-engineered yeast adsorbed three to eight times more copper ions than the parent strain and was more resistant to copper (4 mM) than the parent (below 1 mM at pH 7.8). It was possible to recover about a half of the copper ions adsorbed by whole cells with EDTA treatment without disintegrating the cells. Thus, we succeeded in constructing a novel yeast cell with both tolerance to toxic contaminants and enhanced adsorption of metal ions onto the cell surface.

  14. Tumor cell-collagen interactions: Identification and semi-quantitative evaluation of selectively-expressed genes by combination of differential display- and multiplex-PCR

    Directory of Open Access Journals (Sweden)

    Sirchia Rosalia

    2003-01-01

    Full Text Available It is widely acknowledged that the presence of extracellular matrix components as substrates can drastically modulate the phenotype and gene expression of cultured cells, including tumor cells. A number of published reports indicated that substrates made from two peculiar collagen species, i.e. type V and OF/LB, which are abnormally deposited in the stroma of primary ductal infiltrating carcinoma (d.i.c. of the breast “in vivo,” were able to exert marked and opposite effects on “in vitro” viability, growth and invasiveness of the 8701-BC cell line, isolated from d.i.c.-affected breast epithelium. To complement such functional data on the effect of cell-collagen interactions with information at molecular level, we have utilized a combination of differential display- and semi-quantitative multiplex-PCR techniques with the aim of detecting variations in the expression levels of selected genes by cells maintained in either culture condition. Here we report some prototypical data on the identification and semi-quantitation of three of the differentially-amplified PCR products found, i.e. HSP2A and MSF-B which are up-regulated in cells grown onto OF/LB collagen substrate, and SRCAP which is prominently down-regulated in the presence of type V collagen substrate. This protocol represents a powerful tool for evaluating changes in the levels and patterns of gene expression which can be theoretically adapted to any experimental model system.

  15. Smooth Muscle-Like Cells Generated from Human Mesenchymal Stromal Cells Display Marker Gene Expression and Electrophysiological Competence Comparable to Bladder Smooth Muscle Cells.

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    Juliane Brun

    Full Text Available The use of mesenchymal stromal cells (MSCs differentiated toward a smooth muscle cell (SMC phenotype may provide an alternative for investigators interested in regenerating urinary tract organs such as the bladder where autologous smooth muscle cells cannot be used or are unavailable. In this study we measured the effects of good manufacturing practice (GMP-compliant expansion followed by myogenic differentiation of human MSCs on the expression of a range of contractile (from early to late myogenic markers in relation to the electrophysiological parameters to assess the functional role of the differentiated MSCs and found that differentiation of MSCs associated with electrophysiological competence comparable to bladder SMCs. Within 1-2 weeks of myogenic differentiation, differentiating MSCs significantly expressed alpha smooth muscle actin (αSMA; ACTA2, transgelin (TAGLN, calponin (CNN1, and smooth muscle myosin heavy chain (SM-MHC; MYH11 according to qRT-PCR and/or immunofluorescence and Western blot. Voltage-gated Na+ current levels also increased within the same time period following myogenic differentiation. In contrast to undifferentiated MSCs, differentiated MSCs and bladder SMCs exhibited elevated cytosolic Ca2+ transients in response to K+-induced depolarization and contracted in response to K+ indicating functional maturation of differentiated MSCs. Depolarization was suppressed by Cd2+, an inhibitor of voltage-gated Ca2+-channels. The expression of Na+-channels was pharmacologically identified as the Nav1.4 subtype, while the K+ and Ca2+ ion channels were identified by gene expression of KCNMA1, CACNA1C and CACNA1H which encode for the large conductance Ca2+-activated K+ channel BKCa channels, Cav1.2 L-type Ca2+ channels and Cav3.2 T-type Ca2+ channels, respectively. This protocol may be used to differentiate adult MSCs into smooth muscle-like cells with an intermediate-to-late SMC contractile phenotype exhibiting voltage-gated ion

  16. Smooth Muscle-Like Cells Generated from Human Mesenchymal Stromal Cells Display Marker Gene Expression and Electrophysiological Competence Comparable to Bladder Smooth Muscle Cells

    Science.gov (United States)

    Brun, Juliane; Lutz, Katrin A.; Neumayer, Katharina M. H.; Klein, Gerd; Seeger, Tanja; Uynuk-Ool, Tatiana; Wörgötter, Katharina; Schmid, Sandra; Kraushaar, Udo; Guenther, Elke; Rolauffs, Bernd; Aicher, Wilhelm K.; Hart, Melanie L.

    2015-01-01

    The use of mesenchymal stromal cells (MSCs) differentiated toward a smooth muscle cell (SMC) phenotype may provide an alternative for investigators interested in regenerating urinary tract organs such as the bladder where autologous smooth muscle cells cannot be used or are unavailable. In this study we measured the effects of good manufacturing practice (GMP)-compliant expansion followed by myogenic differentiation of human MSCs on the expression of a range of contractile (from early to late) myogenic markers in relation to the electrophysiological parameters to assess the functional role of the differentiated MSCs and found that differentiation of MSCs associated with electrophysiological competence comparable to bladder SMCs. Within 1–2 weeks of myogenic differentiation, differentiating MSCs significantly expressed alpha smooth muscle actin (αSMA; ACTA2), transgelin (TAGLN), calponin (CNN1), and smooth muscle myosin heavy chain (SM-MHC; MYH11) according to qRT-PCR and/or immunofluorescence and Western blot. Voltage-gated Na+ current levels also increased within the same time period following myogenic differentiation. In contrast to undifferentiated MSCs, differentiated MSCs and bladder SMCs exhibited elevated cytosolic Ca2+ transients in response to K+-induced depolarization and contracted in response to K+ indicating functional maturation of differentiated MSCs. Depolarization was suppressed by Cd2+, an inhibitor of voltage-gated Ca2+-channels. The expression of Na+-channels was pharmacologically identified as the Nav1.4 subtype, while the K+ and Ca2+ ion channels were identified by gene expression of KCNMA1, CACNA1C and CACNA1H which encode for the large conductance Ca2+-activated K+ channel BKCa channels, Cav1.2 L-type Ca2+ channels and Cav3.2 T-type Ca2+ channels, respectively. This protocol may be used to differentiate adult MSCs into smooth muscle-like cells with an intermediate-to-late SMC contractile phenotype exhibiting voltage-gated ion channel

  17. Neonatal plasmacytoid dendritic cells (pDCs display subset variation but can elicit potent anti-viral innate responses.

    Directory of Open Access Journals (Sweden)

    Xiaoming Zhang

    Full Text Available Neonates are highly susceptible to infectious diseases and defective antiviral pDC immune responses have been proposed to contribute to this phenomenon. Isolated cord blood pDCs innately responded to a variety of TLR7 and TLR9 dependent viruses, including influenza A virus (IAV, human immunodeficiency virus (HIV or herpes-simplex virus (HSV by efficiently producing IFN-α, TNF-α as well as chemokines. Interestingly, following activation by CpGA, but not viruses, cord pDCs tend to survive less efficiently. We found that a hallmark of pDCs in neonates is an extended CD2+pDCs compartment compared to adult pDCs without affecting the antiviral IFN-α response. Within CD2+pDCs, we identified a subpopulation expressing CD5 and responsible for IL-12p40 production, however this population is significantly decreased in cord blood compared to adult blood. Therefore, neonatal pDCs clearly display variation in phenotype and subset composition, but without major consequences for their antiviral responses.

  18. Are the surface layer homology domains essential for cell surface display and glycosylation of the S-layer protein from Paenibacillus alvei CCM 2051T?

    Science.gov (United States)

    Janesch, Bettina; Messner, Paul; Schäffer, Christina

    2013-02-01

    Paenibacillus alvei CCM 2051(T) cells are decorated with a two-dimensional (2D) crystalline array comprised of the glycosylated S-layer protein SpaA. At its N terminus, SpaA possesses three consecutive surface layer (S-layer) homology (SLH) domains containing the amino acid motif TRAE, known to play a key role in cell wall binding, as well as the TVEE and TRAQ variations thereof. SpaA is predicted to be anchored to the cell wall by interaction of the SLH domains with a peptidoglycan (PG)-associated, nonclassical, pyruvylated secondary cell wall polymer (SCWP). In this study, we have analyzed the role of the three predicted binding motifs within the SLH domains by mutating them into TAAA motifs, either individually, pairwise, or all of them. Effects were visualized in vivo by homologous expression of chimeras made of the mutated S-layer proteins and enhanced green fluorescent protein and in an in vitro binding assay using His-tagged SpaA variants and native PG-containing cell wall sacculi that either contained SCWP or were deprived of it. Experimental data indicated that (i) the TRAE, TVEE, and TRAQ motifs are critical for the binding function of SLH domains, (ii) two functional motifs are sufficient for cell wall binding, regardless of the domain location, (iii) SLH domains have a dual-recognition function for the SCWP and the PG, and (iv) cell wall anchoring is not necessary for SpaA glycosylation. Additionally, we showed that the SLH domains of SpaA are sufficient for in vivo cell surface display of foreign proteins at the cell surface of P. alvei.

  19. Selection of Peptide Inhibitor to Matrix Metalloproteinase-2 Using Phage Display and Its Effects on Pancreatic Cancer Cell lines PANC-1 and CFPAC-1

    Directory of Open Access Journals (Sweden)

    Gao Lu, Maqing Zheng, Yunxia Zhu, Min Sha, Yue Wu, Xiao Han

    2012-01-01

    Full Text Available Despite tremendous advances in cancer treatment and survival rates, pancreatic cancer remains one of the most deadly afflictions and the fourth leading cause of cancer deaths in the world. Matrix Metalloproteinases (MMPs are thought to be involved in cancer progression. Matrix metalloproteinase (MMP-2 is known to play a pivotal role in tumor invasion, metastasis and angiogenesis, and validated to be the anticancer target. Inhibition of MMP-2 activity is able to reduce the cancer cell invasion and suppress tumor growth in vivo. Two novel peptides, M204C4 and M205C4, which could specially inhibit MMP-2 activity, were identified by a phage display library screening. We showed that M204C4 and M205C4 inhibited the activity of MMP-2 in a dose dependent manner in vitro. Two peptides reduced MMP-2 mediated invasion of the pancreatic cancer cell lines PANC-1 and CFPAC-1, but not affected the expression and release of MMP-2. Furthermore, these two peptides could suppress tumor growth in vivo. Our results indicated that two peptides selected by phase display technology may be used as anticancer drugs in the future.

  20. Displaying gray shades in liquid crystal displays

    Indian Academy of Sciences (India)

    T N Ruckmongathan

    2003-08-01

    Quality of image in a display depends on the contrast, colour, resolution and the number of gray shades. A large number of gray shades is necessary to display images without any contour lines. These contours are due to limited number of gray shades in the display causing abrupt changes in grayness of the image, while the original image has a gradual change in brightness. Amplitude modulation has the capability to display a large number of gray shades with minimum number of time intervals [1,2]. This paper will cover the underlying principle of amplitude modulation, some variants and its extension to multi-line addressing. Other techniques for displaying gray shades in passive matrix displays are reviewed for the sake of comparison.

  1. Human umbilical cord blood-derived mesenchymal stromal cells display a novel interaction between P-selectin and galectin-1.

    Science.gov (United States)

    Suila, H; Hirvonen, T; Kotovuori, A; Ritamo, I; Kerkelä, E; Anderson, H; Natunen, S; Tuimala, J; Laitinen, S; Nystedt, J; Räbinä, J; Valmu, L

    2014-07-01

    Human multipotent mesenchymal stromal/stem cells (MSCs) have been shown to exert immunomodulatory properties that have great potential in therapies for various inflammatory and autoimmune disorders. However, intravenous delivery of these cells is followed by massive cell entrapment in the lungs and insufficient homing to target tissues or organs. In targeting to tissues, MSCs and other therapeutic cells employ similar mechanisms as leucocytes, including a cascade of rolling and adhesion steps mediated by selectins, integrins and their ligands. However, the mechanisms of MSCs homing are not well understood. We discovered that P-selectin (CD62P) binds to umbilical cord blood (UCB)-derived MSCs independently of the previously known sialyl Lewis x (sLex)-containing ligands such as P-selectin glycoprotein ligand-1 (PSGL-1, CD162). By biochemical assays, we identified galectin-1 as a novel ligand for P-selectin. Galectin-1 has previously been shown to be a key mediator of the immunosuppressive effects of human MSCs. We conclude that this novel interaction is likely to play a major role in the immunomodulatory targeting of human UCB-derived MSCs.

  2. Förster resonance energy transfer demonstrates a flavonoid metabolon in living plant cells that displays competitive interactions between enzymes

    NARCIS (Netherlands)

    Crosby, K.C.; Pietraszewska-Bogiel, A.; Gadella (jr.), T.W.J.; Winkel, B.S.J.

    2011-01-01

    We have used Förster resonance energy transfer detected by fluorescence lifetime imaging microscopy (FLIM-FRET) to provide the first evidence from living plants cells for the existence of a flavonoid metabolon. The distribution of flux within this system may be regulated by the direct competition of

  3. An Anti-EGFR IgA That Displays Improved Pharmacokinetics and Myeloid Effector Cell Engagement In Vivo

    NARCIS (Netherlands)

    Lohse, Stefan; Meyer, Saskia; Meulenbroek, Laura A P M; Jansen, J H Marco; Nederend, Maaike; Kretschmer, Anna; Klausz, Katja; Möginger, Uwe; Derer, Stefanie; Rösner, Thies; Kellner, Christian; Schewe, Denis; Sondermann, Peter; Tiwari, Sanjay; Kolarich, Daniel; Peipp, Matthias; Leusen, Jeanette H W; Valerius, Thomas

    2016-01-01

    Antibodies of IgA isotype effectively engage myeloid effector cells for cancer immunotherapy. Here, we describe preclinical studies with an Fc engineered IgA2m(1) antibody containing the variable regions of the EGFR antibody cetuximab. Compared with wild-type IgA2m(1), the engineered molecule lacked

  4. Quercetin induces cell apoptosis of myeloma and displays a synergistic effect with dexamethasone in vitro and in vivo xenograft models

    Science.gov (United States)

    Zhang, Enfan; Zi, Fuming; Chen, Jing; Chen, Qingxiao; Lin, Xuanru; Yang, Li; Li, Yi; Wu, Wenjun; Yang, Yang; He, Jingsong; Cai, Zhen

    2016-01-01

    Quercetin, a kind of dietary flavonoid, has shown its anticancer activity in many kinds of cancers including hematological malignancies (acute myelogenous leukemia, chronic myelogenous leukemia, chronic lymphocytic leukemia, and MM) in vitro and in vivo. However, its effects on MM need further investigation. In this study, MM cell lines were treated with quercetin alone or in combination with dexamethasone. In order to observe the effects in vivo, a xenograft model of human myeloma was established. Quercetin inhibited proliferation of MM cells (RPMI8226, ARP-1, and MM.1R) by inducing cell cycle arrest in the G2/M phase and apoptosis. Western blot showed that quercetin downregulated c-myc expression and upregulated p21 expression. Quercetin also activated caspase-3, caspase-9, and poly(ADP-ribose)polymerase 1. Caspase inhibitors partially blocked apoptosis induced by quercetin. Furthermore, quercetin combined with dexamethasone significantly increased MM cell apoptosis. In vivo xenograft models, quercetin obviously inhibited tumor growth. Caspase-3 was activated to a greater extent when quercetin was combined with dexamethasone. In conclusion, quercetin alone or in combination with dexamethasone may be an effective therapy for MM. PMID:27329589

  5. Identification of two linear B-cell epitopes from West Nile virus NS1 by screening a phage-displayed random peptide library

    Directory of Open Access Journals (Sweden)

    Qin Yong-Li

    2011-07-01

    Full Text Available Abstract Background The West Nile virus (WNV nonstructural protein 1 (NS1 is an important antigenic protein that elicits protective antibody responses in animals and can be used for the serological diagnosis of WNV infection. Although previous work has demonstrated the vital role of WNV NS1-specific antibody responses, the specific epitopes in the NS1 have not been identified. Results The present study describes the identification of two linear B-cell epitopes in WNV NS1 through screening a phage-displayed random 12-mer peptide library with two monoclonal antibodies (mAbs 3C7 and 4D1 that directed against the NS1. The mAbs 3C7 and 4D1 recognized phages displaying peptides with the consensus motifs LTATTEK and VVDGPETKEC, respectively. Exact sequences of both motifs were found in the NS1 (895LTATTEK901 and 925VVDGPETKEC934. Further identification of the displayed B cell epitopes were conducted using a set of truncated peptides expressed as MBP fusion proteins. The data indicated that 896TATTEK901 and925VVDGPETKEC934 are minimal determinants of the linear B cell epitopes recognized by the mAbs 3C7 and 4D1, respectively. Antibodies present in the serum of WNV-positive horses recognized the minimal linear epitopes in Western blot analysis, indicating that the two peptides are antigenic in horses during infection. Furthermore, we found that the epitope recognized by 3C7 is conserved only among WNV strains, whereas the epitope recognized by 4D1 is a common motif shared among WNV and other members of Japanese encephalitis virus (JEV serocomplex. Conclusions We identified TATTEK and VVDGPETKEC as NS1-specific linear B-cell epitopes recognized by the mAbs 3C7 and 4D1, respectively. The knowledge and reagents generated in this study may have potential applications in differential diagnosis and the development of epitope-based marker vaccines against WNV and other viruses of JEV serocomplex.

  6. Cell-Adhesive Matrices Composed of RGD Peptide-Displaying M13 Bacteriophage/Poly(lactic-co-glycolic acid) Nanofibers Beneficial to Myoblast Differentiation.

    Science.gov (United States)

    Shin, Yong Cheol; Lee, Jong Ho; Jin, Linhua; Kim, Min Jeong; Kim, Chuntae; Hong, Suck Won; Oh, Jin Woo; Han, Dong-Wook

    2015-10-01

    Recently, there has been considerable effort to develop suitable scaffolds for tissue engineering applications. Cell adhesion is a prerequisite for cells to survive. In nature, the extracellular matrix (ECM) plays this role. Therefore, an ideal scaffold should be structurally similar to the natural ECM and have biocompatibility and biodegradability. In addition, the scaffold should have biofunctionality, which provides the potent ability to enhance the cellular behaviors, such as adhesion, proliferation and differentiation. This study concentrates on fabricating cell-adhesive matrices composed of RGD peptide-displaying M13 bacteriophage (RGD-M13 phage) and poly(lactic-co-glycolic acid, PLGA) nanofibers. Long rod-shaped M13 bacteriophages are non-toxic and can express many desired proteins on their surface. A genetically engineered M13 phage was constructed to display RGD peptides on its surface. PLGA is a biodegradable polymer with excellent biocompatibility and suitable physicochemical property for adhesive matrices. In this study, RGD-M13 phage/PLGA hybrid nanofiber matrices were fabricated by electrospinning. The physicochemical properties of these matrices were characterized by scanning electron microscopy, atomic force microscopy, Raman spectroscopy, and contact angle measurement. In addition, the cellular behaviors, such as the initial attachment, proliferation and differentiation, were analyzed by a CCK-8 assay and immunofluorescence staining to evaluate the potential application of these matrices to tissue engineering scaffolds. The RGD-M13 phage/PLGA nanofiber matrices could enhance the cellular behaviors and promote the differentiation of C2C12 myoblasts. These results suggest that the RGD-M13 phage/PLGA nanofiber matrices are beneficial to myoblast differentiation and can serve as effective tissue engineering scaffolds.

  7. Surface-Displayed IL-10 by Recombinant Lactobacillus plantarum Reduces Th1 Responses of RAW264.7 Cells Stimulated with Poly(I:C) or LPS.

    Science.gov (United States)

    Cai, Ruopeng; Jiang, Yanlong; Yang, Wei; Yang, Wentao; Shi, Shaohua; Shi, Chunwei; Hu, Jingtao; Gu, Wei; Ye, Liping; Zhou, Fangyu; Gong, Qinglong; Han, Wenyu; Yang, Guilian; Wang, Chunfeng

    2016-02-01

    Recently, poly-γ-glutamic acid synthetase A (pgsA) has been applied to display exogenous proteins on the surface of Lactobacillus casei or Lactococcus lactis, which results in a surfacedisplayed component of bacteria. However, the ability of carrying genes encoded by plasmids and the expression efficiency of recombinant bacteria can be somewhat affected by the longer gene length of pgsA (1,143 bp); therefore, a truncated gene, pgsA, was generated based on the characteristics of pgsA by computational analysis. Using murine IL-10 as an exogenous gene, recombinant Lactobacillus plantarum was constructed and the capacity of the surface-displayed protein and functional differences between exogenous proteins expressed by these strains were evaluated. Surface expression of IL-10 on both recombinant bacteria with anchorins and the higher expression levels in L. plantarum-pgsA'-IL-10 were confirmed by western blot assay. Most importantly, up-regulation of IL-1β, IL-6, TNF-α, IFN-γ, and the nuclear transcription factor NF-κB p65 in RAW264.7 cells after stimulation with Poly(I:C) or LPS was exacerbated after co-culture with L. plantarum-pgsA. By contrast, IL-10 expressed by these recombinant strains could reduce these factors, and the expression of these factors was associated with recombinant strains that expressed anchorin (especially in L. plantarum-pgsA'-IL-10) and was significantly lower compared with the anchorin-free strains. These findings indicated that exogenous proteins could be successfully displayed on the surface of L. plantarum by pgsA or pgsA', and the expression of recombinant bacteria with pgsA' was superior compared with bacteria with pgsA.

  8. Ex vivo generated natural killer cells acquire typical natural killer receptors and display a cytotoxic gene expression profile similar to peripheral blood natural killer cells

    NARCIS (Netherlands)

    Lehmann, D.; Spanholtz, J.; Osl, M.; Tordoir, M.; Lipnik, K.; Bilban, M.; Schlechta, B.; Dolstra, H.; Hofer, E.

    2012-01-01

    Ex vivo differentiation systems of natural killer (NK) cells from CD34+ hematopoietic stem cells are of potential importance for adjuvant immunotherapy of cancer. Here, we analyzed ex vivo differentiation of NK cells from cord blood-derived CD34+ stem cells by gene expression profiling, real-time RT

  9. Human Pluripotent Stem Cells and Derived Neuroprogenitors Display Differential Degrees of Susceptibility to BH3 Mimetics ABT-263, WEHI-539 and ABT-199.

    Directory of Open Access Journals (Sweden)

    Carolina Paola García

    Full Text Available Human embryonic stem cells (hESCs are hypersensitive to genotoxic stress and display lower survival ability relative to their differentiated progeny. Herein, we attempted to investigate the source of this difference by comparing the DNA damage responses triggered by the topoisomerase I inhibitor camptothecin, in hESCs, human induced pluripotent stem cells (hiPSCs and hESCs-derived neuroprogenitors (NP. We observed that upon camptothecin exposure pluripotent stem cells underwent apoptosis more swiftly and at a higher rate than differentiated cells. However, the cellular response encompassing ataxia-telangiectasia mutated kinase activation and p53 phosphorylation both on serine 15 as well as on serine 46 resulted very similar among the aforementioned cell types. Importantly, we observed that hESCs and hiPSCs express lower levels of the anti-apoptotic protein Bcl-2 than NP. To assess whether Bcl-2 abundance could account for this differential response we treated cells with ABT-263, WEHI-539 and ABT-199, small molecules that preferentially target the BH3-binding pocket of Bcl-xL and/or Bcl-2 and reduce their ability to sequester pro-apoptotic factors. We found that in the absence of stress stimuli, NP exhibited a higher sensitivity to ABT- 263 and WEHI-539 than hESCs and hiPSCs. Conversely, all tested cell types appeared to be highly resistant to the Bcl-2 specific inhibitor, ABT-199. However, in all cases we determined that ABT-263 or WEHI-539 treatment exacerbated camptothecin-induced apoptosis. Importantly, similar responses were observed after siRNA-mediated down-regulation of Bcl-xL or Bcl-2. Taken together, our results suggest that Bcl-xL contrary to Bcl-2 contributes to ensure cell survival and also functions as a primary suppressor of DNA double-strand brake induced apoptosis both in pluripotent and derived NP cells. The emerging knowledge of the relative dependence of pluripotent and progenitor cells on Bcl-2 and Bcl-xL activities may help

  10. Human Pluripotent Stem Cells and Derived Neuroprogenitors Display Differential Degrees of Susceptibility to BH3 Mimetics ABT-263, WEHI-539 and ABT-199

    Science.gov (United States)

    Dimopoulos, Nicolás Alexis; Fernandez Espinosa, Damián Darío; Miriuka, Santiago Gabriel; Sevlever, Gustavo Emilio; Romorini, Leonardo; Scassa, María Elida

    2016-01-01

    Human embryonic stem cells (hESCs) are hypersensitive to genotoxic stress and display lower survival ability relative to their differentiated progeny. Herein, we attempted to investigate the source of this difference by comparing the DNA damage responses triggered by the topoisomerase I inhibitor camptothecin, in hESCs, human induced pluripotent stem cells (hiPSCs) and hESCs-derived neuroprogenitors (NP). We observed that upon camptothecin exposure pluripotent stem cells underwent apoptosis more swiftly and at a higher rate than differentiated cells. However, the cellular response encompassing ataxia-telangiectasia mutated kinase activation and p53 phosphorylation both on serine 15 as well as on serine 46 resulted very similar among the aforementioned cell types. Importantly, we observed that hESCs and hiPSCs express lower levels of the anti-apoptotic protein Bcl-2 than NP. To assess whether Bcl-2 abundance could account for this differential response we treated cells with ABT-263, WEHI-539 and ABT-199, small molecules that preferentially target the BH3-binding pocket of Bcl-xL and/or Bcl-2 and reduce their ability to sequester pro-apoptotic factors. We found that in the absence of stress stimuli, NP exhibited a higher sensitivity to ABT- 263 and WEHI-539 than hESCs and hiPSCs. Conversely, all tested cell types appeared to be highly resistant to the Bcl-2 specific inhibitor, ABT-199. However, in all cases we determined that ABT-263 or WEHI-539 treatment exacerbated camptothecin-induced apoptosis. Importantly, similar responses were observed after siRNA-mediated down-regulation of Bcl-xL or Bcl-2. Taken together, our results suggest that Bcl-xL contrary to Bcl-2 contributes to ensure cell survival and also functions as a primary suppressor of DNA double-strand brake induced apoptosis both in pluripotent and derived NP cells. The emerging knowledge of the relative dependence of pluripotent and progenitor cells on Bcl-2 and Bcl-xL activities may help to predict

  11. Identification of a panel of tumor-associated antigens from breast carcinoma cell lines, solid tumors and testis cDNA libraries displayed on lambda phage

    Directory of Open Access Journals (Sweden)

    Cianfriglia Maurizio

    2004-11-01

    Full Text Available Abstract Background Tumor-associated antigens recognized by humoral effectors of the immune system are a very attractive target for human cancer diagnostics and therapy. Recent advances in molecular techniques have led to molecular definition of immunogenic tumor proteins based on their reactivity with autologous patient sera (SEREX. Methods Several high complexity phage-displayed cDNA libraries from breast carcinomas, human testis and breast carcinoma cell lines MCF-7, MDA-MB-468 were constructed. The cDNAs were expressed in the libraries as fusion to bacteriophage lambda protein D. Lambda-displayed libraries were efficiently screened with sera from patients with breast cancer. Results A panel of 21 clones representing 18 different antigens, including eight proteins of unknown function, was identified. Three of these antigens (T7-1, T11-3 and T11-9 were found to be overexpressed in tumors as compared to normal breast. A serological analysis of the 21 different antigens revealed a strong cancer-related profile for at least five clones (T6-2, T6-7, T7-1, T9-21 and T9-27. Conclusions Preliminary results indicate that patient serum reactivity against five of the antigens is associated with tumor disease. The novel T7-1 antigen, which is overexpressed in breast tumors and recognized specifically by breast cancer patient sera, is potentially useful in cancer diagnosis.

  12. Embryonic stem cells derived from in vivo or in vitro-generated murine blastocysts display similar transcriptome and differentiation potential.

    Directory of Open Access Journals (Sweden)

    Rhodel K Simbulan

    Full Text Available The use of assisted reproductive technologies (ART such as in vitro fertilization (IVF has resulted in the birth of more than 5 million children. While children conceived by these technologies are generally healthy, there is conflicting evidence suggesting an increase in adult-onset complications like glucose intolerance and high blood pressure in IVF children. Animal models indicate similar potential risks. It remains unclear what molecular mechanisms may be operating during in vitro culture to predispose the embryo to these diseases. One of the limitations faced by investigators is the paucity of the material in the preimplantation embryo to test for molecular analysis. To address this problem, we generated mouse embryonic stem cells (mESC from blastocysts conceived after natural mating (mESCFB or after IVF, using optimal (KSOM + 5% O2; mESCKAA and suboptimal (Whitten's Medium, + 20% O2, mESCWM conditions. All three groups of embryos showed similar behavior during both derivation and differentiation into their respective mESC lines. Unsupervised hierarchical clustering of microarray data showed that blastocyst culture does not affect the transcriptome of derived mESCs. Transcriptomic changes previously observed in the inner cell mass (ICM of embryos derived in the same conditions were not present in mESCs, regardless of method of conception or culture medium, suggesting that mESC do not fully maintain a memory of the events occurring prior to their derivation. We conclude that the fertilization method or culture media used to generate blastocysts does not affect differentiation potential, morphology and transcriptome of mESCs.

  13. One-Step Recovery of scFv Clones from High-Throughput Sequencing-Based Screening of Phage Display Libraries Challenged to Cells Expressing Native Claudin-1

    Directory of Open Access Journals (Sweden)

    Emanuele Sasso

    2015-01-01

    Full Text Available Expanding the availability of monoclonal antibodies interfering with hepatitis C virus infection of hepatocytes is an active field of investigation within medical biotechnologies, to prevent graft reinfection in patients subjected to liver transplantation and to overcome resistances elicited by novel antiviral drugs. In this paper, we describe a complete pipeline for screening of phage display libraries of human scFvs against native Claudin-1, a tight-junction protein involved in hepatitis C virus infection, expressed on the cell surface of human hepatocytes. To this aim, we implemented a high-throughput sequencing approach for library screening, followed by a simple and effective strategy to recover active binder clones from enriched sublibraries. The recovered clones were successfully converted to active immunoglobulins, thus demonstrating the effectiveness of the whole procedure. This novel approach can guarantee rapid and cheap isolation of antibodies for virtually any native antigen involved in human diseases, for therapeutic and/or diagnostic applications.

  14. Direct isopropanol production from cellobiose by engineered Escherichia coli using a synthetic pathway and a cell surface display system.

    Science.gov (United States)

    Soma, Yuki; Inokuma, Kentaro; Tanaka, Tsutomu; Ogino, Chiaki; Kondo, Akihiko; Okamoto, Masahiro; Hanai, Taizo

    2012-07-01

    Efficient bio-production from lignocellulosic biomass is required for the purpose of developing an inexpensive, practical bio-refinery process. As one approach to address this problem, we genetically engineered Escherichia coli to produce isopropanol directly from cellobiose via the cellobiose degradation by Beta-Glucosidase (BGL) on the cell surface. First, we investigated the cellobiose consumption of two E. coli strains with the BGL protein from Thermobifida fusca YX (Tfu0937) fused to the anchor protein Blc (Tfu0937/Blc) using different fusion sites. Next, we introduced the synthetic pathway for isopropanol production into those strains and compared their isopropanol production in the presence of glucose. Based on the results of these assays, TA212/pTA411, which was introduced Tfu-Blc fused protein expression system and the synthetic pathway for isopropanol production, was selected for the directly isopropanol production from cellobiose. TA212/pTA411 produced 69.0±11.6mM isopropanol at 21h of fermentation, whereas TA212/pTA147, which did not introduced the BGL/anchor fused protein but was introduced the synthetic pathway for isopropanol production, showed no cellobiose consumption and no isopropanol production during fermentation. To our knowledge, this is the first report of the production of a bio-product from cellobiose using E. coli.

  15. Neuregulin1 displayed on motor axons regulates terminal Schwann cell-mediated synapse elimination at developing neuromuscular junctions.

    Science.gov (United States)

    Lee, Young Il; Li, Yue; Mikesh, Michelle; Smith, Ian; Nave, Klaus-Armin; Schwab, Markus H; Thompson, Wesley J

    2016-01-26

    Synaptic connections in the nervous system are rearranged during development and in adulthood as a feature of growth, plasticity, aging, and disease. Glia are implicated as active participants in these changes. Here we investigated a signal that controls the participation of peripheral glia, the terminal Schwann cells (SCs), at the neuromuscular junction (NMJ) in mice. Transgenic manipulation of the levels of membrane-tethered neuregulin1 (NRG1-III), a potent activator of SCs normally presented on motor axons, alters the rate of loss of motor inputs at NMJs during developmental synapse elimination. In addition, NMJs of adult transgenic mice that expressed excess axonal NRG1-III exhibited continued remodeling, in contrast to the more stable morphologies of controls. In fact, synaptic SCs of these adult mice with NRG1-III overexpression exhibited behaviors evident in wild type neonates during synapse elimination, including an affinity for the postsynaptic myofiber surface and phagocytosis of nerve terminals. Given that levels of NRG1-III expression normally peak during the period of synapse elimination, our findings identify axon-tethered NRG1 as a molecular determinant for SC-driven neuromuscular synaptic plasticity.

  16. Müller细胞在视网膜绿光损伤模型中的激活反应%Activation of Müller cells in the green light damaged retina

    Institute of Scientific and Technical Information of China (English)

    周臻; 张绍丹; 李颖; 张纯; 王薇

    2011-01-01

    Objective To investigate the reaction of Mttller cells in green light damaged retina. Methods Dark-adapted 8 week-old albino rats were exposed to green light for 3 hours at an intensity of 1 mW/cm2, and at different dark recovery intervals, ranging from 3 hours to IS days, and the the eyes were enucleated. Sections of the retina were taken vertically and the superior parts were examined by histology. Immunocytochemistry and Western blotting were performed to detect the expression and localization of glial fibrillary acidic protein (CFAP) , glutamine synthetase (GS) and phosphated signal transducer and activator of transcription 3 (pSTAT3). TdT-mediated dUTP nick-end labeling (TUNEL) was performed to detect the apoptotic cells. Results In the damaged retina, the rod outer segments (ROS) were shortened and the outer nuclear layer was thinned with the presence of apoptotic cells. According to immunocytochemistry and Western blot analysis, GFAP protein levels increased at 12 hours after injury in the observation period. GS expression remained stable after the light damage. However, during the 7-d period post-injury the GS protein appeared to translocate from the cells in the inner nuclear layer to the cells in the outer nuclear layer. pSTAT3 expression showed a transient increase at 12 hours after injury. Conclusions Green light exposure induces apoptosis in the outer nuclear layer. Mttller cells are activated with GFAP expression elevation and GS translocation. pSTAT3 may play a role in triggering GFAP expression in the Mailer cells.%目的 研究Müller细胞在大鼠视网膜绿光损伤模型中的激活反应.方法 72只出生后8周(约230~280g)的成年雄性Sprague-Dawley大鼠随机分9组.一组作为正常对照,另8组暗适应24h后置于波长为(530±1O)nm的LED灯光箱中照射3h,并分别于暗恢复3、6、12 h及1、2、3、7、15 d取眼球,光学显微镜下观察同一定位处视网膜组织形态、检测( TUNEL)凋亡细胞、免疫组织

  17. The dual specificity PI3K/mTOR inhibitor PKI-587 displays efficacy against T-cell acute lymphoblastic leukemia (T-ALL).

    Science.gov (United States)

    Gazi, Mohiuddin; Moharram, Sausan A; Marhäll, Alissa; Kazi, Julhash U

    2017-04-28

    Although significant improvements have been made in the treatment of acute lymphoblastic leukemia (ALL), there is a substantial subset of high-risk T-cell ALL (T-ALL) patients with relatively poor prognosis. Like in other leukemia types, alterations of the PI3K/mTOR pathway are predominant in ALL which is also responsible for treatment failure and relapse. In this study, we show that relapsed T-ALL patients display an enrichment of the PI3K/mTOR pathway. Using a panel of inhibitors targeting multiple components of the PI3K/mTOR pathway, we observed that the dual-specific PI3K/mTOR inhibitor PKI-587 was the most selective inhibitor for T-ALL cells dependent on the PI3K/mTOR pathway. Furthermore, we observed that PKI-587 blocked proliferation and colony formation of T-ALL cell lines. Additionally, PKI-587 selectively abrogated PI3K/mTOR signaling without affecting MAPK signaling both in in vitro and in vivo. Inhibition of the PI3K/mTOR pathway using PKI-587 delayed tumor progression, reduced tumor load and enhanced the survival rate in immune-deficient mouse xenograft models without inducing weight loss in the inhibitor treated mice. This preclinical study shows beneficial effects of PKI-587 on T-ALL that warrants further investigation in the clinical setting.

  18. Handbook of display technology

    CERN Document Server

    Castellano, Joseph A

    1992-01-01

    This book presents a comprehensive review of technical and commercial aspects of display technology. It provides design engineers with the information needed to select proper technology for new products. The book focuses on flat, thin displays such as light-emitting diodes, plasma display panels, and liquid crystal displays, but it also includes material on cathode ray tubes. Displays include a large number of products from televisions, auto dashboards, radios, and household appliances, to gasoline pumps, heart monitors, microwave ovens, and more.For more information on display tech

  19. Adolescents with clinical type 1 diabetes display reduced red blood cell glucose transporter isoform 1 (GLUT1).

    Science.gov (United States)

    Garg, Meena; Thamotharan, Manikkavasagar; Becker, Dorothy J; Devaskar, Sherin U

    2014-11-01

    Type 1 diabetic (T1D) adolescent children on insulin therapy suffer episodes of both hyper- and hypoglycemic episodes. Glucose transporter isoform GLUT1 expressed in blood-brain barrier (BBB) and red blood cells (RBC) compensates for perturbed circulating glucose toward protecting the supply to brain and RBCs. We hypothesized that RBC-GLUT1 concentration, as a surrogate for BBB-GLUT1, is altered in T1D children. To test this hypothesis, we measured RBC-GLUT1 by enzyme-linked immunosorbent assay (ELISA) in T1D children (n = 72; mean age 15.3 ± 0.2 yr) and control children (CON; n = 11; mean age 15.6 ± 0.9 yr) after 12 h of euglycemia and during a hyperinsulinemic-hypoglycemic clamp with a nadir blood glucose of ~3.3 mmol/L for 90 min (clamp I) or ~3 mmol/L for 45 min (clamp II). Reduced baseline RBC-GLUT1 was observed in T1D (2.4 ± 0.17 ng/ng membrane protein); vs. CON (4.2 ± 0.61 ng/ng protein) (p increasing during acute hypoglycemia over the durations examined, may demonstrate a vulnerability of impaired RBC glucose transport (serving as a surrogate for BBB), especially in those with the worst control. We speculate that this may contribute to the perturbed cognition seen in T1D adolescents.

  20. Alu Sequences in Undifferentiated Human Embryonic Stem Cells Display High Levels of A-to-I RNA Editing

    Science.gov (United States)

    Osenberg, Sivan; Paz Yaacov, Nurit; Safran, Michal; Moshkovitz, Sharon; Shtrichman, Ronit; Sherf, Ofra; Jacob-Hirsch, Jasmine; Keshet, Gilmor; Amariglio, Ninette; Itskovitz-Eldor, Joseph; Rechavi, Gideon

    2010-01-01

    Adenosine to Inosine (A-to-I) RNA editing is a site-specific modification of RNA transcripts, catalyzed by members of the ADAR (Adenosine Deaminase Acting on RNA) protein family. RNA editing occurs in human RNA in thousands of different sites. Some of the sites are located in protein-coding regions but the majority is found in non-coding regions, such as 3′UTRs, 5′UTRs and introns - mainly in Alu elements. While editing is found in all tissues, the highest levels of editing are found in the brain. It was shown that editing levels within protein-coding regions are increased during embryogenesis and after birth and that RNA editing is crucial for organism viability as well as for normal development. In this study we characterized the A-to-I RNA editing phenomenon during neuronal and spontaneous differentiation of human embryonic stem cells (hESCs). We identified high editing levels of Alu repetitive elements in hESCs and demonstrated a global decrease in editing levels of non-coding Alu sites when hESCs are differentiating, particularly into the neural lineage. Using RNA interference, we showed that the elevated editing levels of Alu elements in undifferentiated hESCs are highly dependent on ADAR1. DNA microarray analysis showed that ADAR1 knockdown has a global effect on gene expression in hESCs and leads to a significant increase in RNA expression levels of genes involved in differentiation and development processes, including neurogenesis. Taken together, we speculate that A-to-I editing of Alu sequences plays a role in the regulation of hESC early differentiation decisions. PMID:20574523

  1. Alu sequences in undifferentiated human embryonic stem cells display high levels of A-to-I RNA editing.

    Directory of Open Access Journals (Sweden)

    Sivan Osenberg

    Full Text Available Adenosine to Inosine (A-to-I RNA editing is a site-specific modification of RNA transcripts, catalyzed by members of the ADAR (Adenosine Deaminase Acting on RNA protein family. RNA editing occurs in human RNA in thousands of different sites. Some of the sites are located in protein-coding regions but the majority is found in non-coding regions, such as 3'UTRs, 5'UTRs and introns - mainly in Alu elements. While editing is found in all tissues, the highest levels of editing are found in the brain. It was shown that editing levels within protein-coding regions are increased during embryogenesis and after birth and that RNA editing is crucial for organism viability as well as for normal development. In this study we characterized the A-to-I RNA editing phenomenon during neuronal and spontaneous differentiation of human embryonic stem cells (hESCs. We identified high editing levels of Alu repetitive elements in hESCs and demonstrated a global decrease in editing levels of non-coding Alu sites when hESCs are differentiating, particularly into the neural lineage. Using RNA interference, we showed that the elevated editing levels of Alu elements in undifferentiated hESCs are highly dependent on ADAR1. DNA microarray analysis showed that ADAR1 knockdown has a global effect on gene expression in hESCs and leads to a significant increase in RNA expression levels of genes involved in differentiation and development processes, including neurogenesis. Taken together, we speculate that A-to-I editing of Alu sequences plays a role in the regulation of hESC early differentiation decisions.

  2. MRI displays involvement of the temporalis muscle and the deep temporal artery in patients with giant cell arteritis

    Energy Technology Data Exchange (ETDEWEB)

    Veldhoen, Simon; Bley, Thorsten A. [University Medical Center Wuerzburg, Department of Diagnostic and Interventional Radiology, Wuerzburg (Germany); Klink, Thorsten [Inselspital - University Medical Center Bern, Department of Diagnostic, Interventional and Pediatric Radiology, Bern (Switzerland); Geiger, Julia [University Medical Center Freiburg, Department of Diagnostic and Interventional Radiology, Freiburg (Germany); University Children' s Hospital Zuerich, Division of Radiology, Zuerich (Switzerland); Vaith, Peter; Glaser, Cornelia [University Medical Center Freiburg, Department of Rheumatology and Immunology, Freiburg (Germany); Ness, Thomas [University Medical Center Freiburg, Department of Ophthalmology, Freiburg (Germany); Duwendag, Dirk [University Medical Center Kiel, Department of Ophthalmology, Kiel (Germany); Both, Marcus [University Medical Center Kiel, Department of Diagnostic and Interventional Radiology, Kiel (Germany)

    2014-11-15

    To assess deep temporal artery and temporalis muscle involvement in patients with giant cell arteritis (GCA). Ninety-nine patients who received magnetic resonance imaging (MRI) and superficial temporal artery biopsy (TAB) were included in this study. Patients with positive TAB (n = 61) were defined as GCA patients, those with negative TAB (n = 38) as the GCA-negative reference group. Contrast-enhanced T1w-images were acquired utilizing 1.5 T and 3 T MRI. Two radiologists assessed the images. Mural contrast-hyperenhancement and wall thickening of the deep temporal artery and hyperenhancement of the muscle were defined as inflammation. MRI results were correlated with jaw claudication in 70 patients. The two observers found temporalis muscle involvement in 19.7 % (n = 12) and 21.3 % (n = 13) of GCA patients. It occurred bilaterally in 100 %. Specificities were 92/97 % and sensitivities were 20/21 %. Deep temporal artery involvement was found in 34.4 % (n = 21) and 49.2 % (n = 30) and occurred bilaterally in 80/90.5 %. Specificities were 84/95 % and sensitivities were 34/49 %. Both structures were affected simultaneously in 18/21.3 %. Jaw claudication correlated moderately with inflammation of the temporalis muscle (r = 0.31; p < 0.05) and the deep temporal artery (r = 0.38; p = 0.01). MRI visualizes changes in the temporalis muscle and the deep temporal artery in GCA. Moderate correlation of clinical symptoms with MRI results was observed. circle Approximately 20 % of GCA patients presented with temporalis muscle inflammation. (orig.)

  3. Tannerella forsythia strains display different cell-surface nonulosonic acids: biosynthetic pathway characterization and first insight into biological implications.

    Science.gov (United States)

    Friedrich, Valentin; Janesch, Bettina; Windwarder, Markus; Maresch, Daniel; Braun, Matthias L; Megson, Zoë A; Vinogradov, Evgeny; Goneau, Marie-France; Sharma, Ashu; Altmann, Friedrich; Messner, Paul; Schoenhofen, Ian C; Schäffer, Christina

    2016-12-16

    Tannerella forsythia is an anaerobic, Gram-negative periodontal pathogen. A unique O-linked oligosaccharide decorates the bacterium's cell surface proteins and was shown to modulate the host immune response. In our study, we investigated the biosynthesis of the nonulosonic acid (NulO) present at the terminal position of this glycan. A bioinformatic analysis of T. forsythia genomes revealed a gene locus for the synthesis of pseudaminic acid (Pse) in the type strain ATCC 43037 while strains FDC 92A2 and UB4 possess a locus for the synthesis of legionaminic acid (Leg) instead. In contrast to the NulO in ATCC 43037, which has been previously identified as a Pse derivative (5-N-acetimidoyl-7-N-glyceroyl-3,5,7,9-tetradeoxy-l-glycero-l-manno-NulO), glycan analysis of strain UB4 performed in this study indicated a 350-Da, possibly N-glycolyl Leg (3,5,7,9-tetradeoxy-d-glycero-d-galacto-NulO) derivative with unknown C5,7 N-acyl moieties. We have expressed, purified and characterized enzymes of both NulO pathways to confirm these genes' functions. Using capillary electrophoresis (CE), CE-mass spectrometry and NMR spectroscopy, our studies revealed that Pse biosynthesis in ATCC 43037 essentially follows the UDP-sugar route described in Helicobacter pylori, while the pathway in strain FDC 92A2 corresponds to Leg biosynthesis in Campylobacter jejuni involving GDP-sugar intermediates. To demonstrate that the NulO biosynthesis enzymes are functional in vivo, we created knockout mutants resulting in glycans lacking the respective NulO. Compared to the wild-type strains, the mutants exhibited significantly reduced biofilm formation on mucin-coated surfaces, suggestive of their involvement in host-pathogen interactions or host survival. This study contributes to understanding possible biological roles of bacterial NulOs.

  4. CD4~+Foxp3~+ regulatory T cells converted by rapamycin from peripheral CD4~+ CD25~-naive T cells display more potent regulatory ability in vitro

    Institute of Scientific and Technical Information of China (English)

    CHEN Jian-fei; GAO Jie; ZHANG Dong; WANG Zi-han; ZHU Ji-ye

    2010-01-01

    Background Rapamycin (RAPA) is a relatively new immunosuppressant drug that functions as a serine/threonine kinase inhibitor to prevent rejection in organ transplantation. RAPA blocks activation of T-effector (Teff) cells by inhibiting the response to interleukin-2. Recently, RAPA was also shown to selectively expand the T-regulator (Treg) cell population. To date, no studies have examined the mechanism by which RAPA converts Teff cells to Treg cells. Methods Peripheral CD4~+CD25~- naive T cells were cultivated with RAPA and B cells as antigen-presenting cells (APCs) in vitro. CD4~+CD25~- T cells were harvested after 6 days and analyzed for expression of forkhead box protein 3 (Foxp3) using flow cytometry. CD4~+CD25~+CD127~- subsets as the converted Tregs were isolated from the mixed lymphocyte reactions (MLR) with CD127 negative selection, followed by CD4 and CD25 positive selection using microbeads and magnetic separation column (MSC). Moreover, mRNA was extracted from converted Tregs and C57BL/6 naive CD4~+CD25~+ T cells and Foxp3 levels were examined by quantitative real-time polymerase chain reaction (rt-PCR). A total of 1×10~5 carboxyfluorescein succinimidyl ester (CFSE)-labeled naive CD4~+CD25~- T cells/well from C57BL/6 mice were cocultured with DBA/2 or C3H maturation of dendritic cells (mDCs) (0.25×10~5/well) in 96-well round-bottom plates for 6 days. Then 1×10~5 or 0.25×10~5 converted Treg cells were added to every well as regulatory cells. Cells were harvested after 6 days of culture and analyzed for proliferation of CFSE-labeled naive CD4~+CD25~- T cells using flow cytometry. Data were analyzed using CellQuest software.Results We found that RAPA can convert peripheral CD4~+CD25~- naive T Cells to CD4~+Foxp3~+ Treg cells using B cells as APCs, and this subtype of Treg can potently suppress Teff proliferation and maintain antigenic specificity. Conclusion Our findings provide evidence that RAPA induces Treg cell conversion from Teff cells and

  5. Displayed correlation between gene expression profiles and submicroscopic alterations in response to cetuximab, gefitinib and EGF in human colon cancer cell lines

    Directory of Open Access Journals (Sweden)

    Pezzetti Furio

    2008-08-01

    Full Text Available Abstract Background EGFR is frequently overexpressed in colon cancer. We characterized HT-29 and Caco-2, human colon cancer cell lines, untreated and treated with cetuximab or gefitinib alone and in combination with EGF. Methods Cell growth was determined using a variation on the MTT assay. Cell-cycle analysis was conducted by flow cytometry. Immunohistochemistry was performed to evaluate EGFR expression and scanning electron microscopy (SEM evidenced the ultrastructural morphology. Gene expression profiling was performed using hybridization of the microarray Ocimum Pan Human 40 K array A. Results Caco-2 and HT-29 were respectively 66.25 and 59.24 % in G0/G1. They maintained this level of cell cycle distribution after treatment, suggesting a predominantly differentiated state. Treatment of Caco-2 with EGF or the two EGFR inhibitors produced a significant reduction in their viability. SEM clearly showed morphological cellular transformations in the direction of cellular death in both cell lines treated with EGFR inhibitors. HT-29 and Caco-2 displayed an important reduction of the microvilli (which also lose their erect position in Caco-2, possibly invalidating microvilli absorption function. HT-29 treated with cetuximab lost their boundary contacts and showed filipodi; when treated with gefitinib, they showed some vesicles: generally membrane reshaping is evident. Both cell lines showed a similar behavior in terms of on/off switched genes upon treatment with cetuximab. The gefitinib global gene expression pattern was different for the 2 cell lines; gefitinib treatment induced more changes, but directly correlated with EGF treatment. In cetuximab or gefitinib plus EGF treatments there was possible summation of the morphological effects: cells seemed more weakly affected by the transformation towards apoptosis. The genes appeared to be less stimulated than for single drug cases. Conclusion This is the first study to have systematically investigated

  6. ALMS1-deficient fibroblasts over-express extra-cellular matrix components, display cell cycle delay and are resistant to apoptosis.

    Directory of Open Access Journals (Sweden)

    Elisabetta Zulato

    Full Text Available Alström Syndrome (ALMS is a rare genetic disorder (483 living cases, characterized by many clinical manifestations, including blindness, obesity, type 2 diabetes and cardiomyopathy. ALMS is caused by mutations in the ALMS1 gene, encoding for a large protein with implicated roles in ciliary function, cellular quiescence and intracellular transport. Patients with ALMS have extensive fibrosis in nearly all tissues resulting in a progressive organ failure which is often the ultimate cause of death. To focus on the role of ALMS1 mutations in the generation and maintenance of this pathological fibrosis, we performed gene expression analysis, ultrastructural characterization and functional assays in 4 dermal fibroblast cultures from ALMS patients. Using a genome-wide gene expression analysis we found alterations in genes belonging to specific categories (cell cycle, extracellular matrix (ECM and fibrosis, cellular architecture/motility and apoptosis. ALMS fibroblasts display cytoskeleton abnormalities and migration impairment, up-regulate the expression and production of collagens and despite the increase in the cell cycle length are more resistant to apoptosis. Therefore ALMS1-deficient fibroblasts showed a constitutively activated myofibroblast phenotype even if they do not derive from a fibrotic lesion. Our results support a genetic basis for the fibrosis observed in ALMS and show that both an excessive ECM production and a failure to eliminate myofibroblasts are key mechanisms. Furthermore, our findings suggest new roles for ALMS1 in both intra- and extra-cellular events which are essential not only for the normal cellular function but also for cell-cell and ECM-cell interactions.

  7. A Compressive Superresolution Display

    KAUST Repository

    Heide, Felix

    2014-06-22

    In this paper, we introduce a new compressive display architecture for superresolution image presentation that exploits co-design of the optical device configuration and compressive computation. Our display allows for superresolution, HDR, or glasses-free 3D presentation.

  8. Lunar Sample Display Locations

    Data.gov (United States)

    National Aeronautics and Space Administration — NASA provides a number of lunar samples for display at museums, planetariums, and scientific expositions around the world. Lunar displays are open to the public....

  9. A novel cell-penetrating peptide derived from WT1 enhances p53 activity, induces cell senescence and displays antimelanoma activity in xeno- and syngeneic systems

    Directory of Open Access Journals (Sweden)

    Mariana H. Massaoka

    2014-01-01

    Full Text Available The Wilms tumor protein 1 (WT1 transcription factor has been associated in malignant melanoma with cell survival and metastasis, thus emerging as a candidate for targeted therapy. A lysine–arginine rich peptide, WT1-pTj, derived from the ZF domain of WT1 was evaluated as an antitumor agent against A2058 human melanoma cells and B16F10-Nex2 syngeneic murine melanoma. Peptide WT1-pTj quickly penetrated human melanoma cells and induced senescence, recognized by increased SA-β-galactosidase activity, enhanced transcriptional activity of p53, and induction of the cell cycle inhibitors p21 and p27. Moreover, the peptide bound to p53 and competed with WT1 protein for binding to p53. WT1-pTj treatment led to sustained cell growth suppression, abrogation of clonogenicity and G2/M cell cycle arrest. Notably, in vivo studies showed that WT1-pTj inhibited both the metastases and subcutaneous growth of murine melanoma in syngeneic mice, and prolonged the survival of nude mice challenged with human melanoma cells. The 27-amino acid cell-penetrating WT1-derived peptide, depends on C3 and H16 for effective antimelanoma activity, inhibits proliferation of WT1-expressing human tumor cell lines, and may have an effective role in the treatment of WT1-expressing malignancies.

  10. Production and radioimmunoimaging of novel fully human phage display recombinant antibodies and growth inhibition of lung adenocarcinoma cell line overexpressing Prx I.

    Science.gov (United States)

    Luo, Yi; Pang, Hua; Li, Shujie; Cao, Hui; Peng, Zhiping; Fan, Chunbo; Li, Shaolin

    2009-07-01

    The Peroxiredoxin I (Prx I) is a member of the Peroxiredoxin family, which is overexpressed in many diverse tumor types and is an anti-apoptosis protein for tumor cell proliferation and survival. Therapeutic strategies targeting the Prx I may therefore be effective broad-spectrum anticancer agents. We constructed a phage display single-chain variable fragment (scFv) antibody library and sieve out the fully human, lung adenocarcinoma-sepcific monoclonal antibodies. The selection on Prx I was performed using above-mentioned lung adenocarcinoma-sepcific monoclonal antibodies with high affinity to Prx I overexpressing lung adenocarcinoma cells. The candidate scFv sequences, based on enzyme-linked immunosorbent assay (ELISA) screening data, were chosen for soluble expression, and a 30 kDa band was observed on polyacrylamide gel electrophoresis as predicted. The purified antibodies were characterized by immunoblotting and showed high specificity to Prx I-overexpressing lung adenocarcinoma cells A549. Radioimmunoimaging was taken to evaluate specificity and distribution of antibodies in vivo. The radiolocalization index (RI) of tumor/serum and tumor/muscle gradually increased, reaching its peak (4.06 +/- 0.13 and 5.17 +/- 0.97, respectively) at 48 h postadministration. Single photon emission computed tomography (SPECT) imaging showed the radioactivity was aggregated in tumor locations and tumor imaging was clearly observed. The internalized scFv resulted in antibody-mediated cell apoptosis and downregulation of Prx I expression. These results demonstrate that the scFv possesses strong antitumor activity on lung adenocarcinoma and may therefore be an effective therapeutic candidate for the treatment of cancers that are dependent on Prx I for growth and survival.

  11. Immunophenotyping of diffuse large B-cell lymphoma (DLBCL) defines multiple sub-groups of germinal centre-like tumours displaying different survival characteristics.

    Science.gov (United States)

    Anderson, John J; Fordham, Sarah; Overman, Lynne; Dignum, Helen; Wood, Katrina; Proctor, Stephen J; Crosier, Stephen; Angus, Brian; Culpin, Rachel E; Mainou-Fowler, Tryfonia

    2009-11-01

    Diffuse large B-cell lymphoma (DLBCL) forms a heterogeneous collection of aggressive non-Hodgkin's Lymphoma in which three principle classes of neoplasia have been defined according to gene expression and immunophenotyping studies. The present investigation sought to examine the immunophenotype of proposed subgroups and relate these to patient survival. A series of 155 DLBCL treated uniformly with anthracycline therapy in clinical trials, were stratified upon the basis of common biomarker expression with combination immunophenotype being related to patient overall survival. Stratification of tumours with respect to combined expression profiles of the three biological markers (CD10, Bcl-6 and MUM-1) revealed six groups showing significant differences in survival (p=0.014). The greatest difference resided between distinct populations of germinal centre (GC) cell tumours; the first being CD10-, Bcl-6+, MUM-1- and the second CD10+ Bcl-6+ MUM-1+ (p=0.002). The former group displayed median survival time of 143 months, the latter only 11 months. A third population of GC tumours (CD10+ Bcl-6+ and MUM-1-) also displayed a relative short median survival (32 months). Of the three groups presenting a non-GC or activated B cell (NGC/ABC) phenotype, only one (CD10-, Bcl-6+ and MUM-1+) presented short-term median survival (27 months) comparable with poor prognosis GC sub-populations. Within the remaining ABC tumour groups (CD10- Bcl-6- MUM-1- and CD10- Bcl-6- MUM-1+) patients presented intermediate median survival times of 54 and 58 months, respectively. Thus, the GC phenotype did not act as a universal indicator of good clinical prognosis, but rather multiple groups of GC tumours were associated with distinct overall survival profiles. Ultimately, the data allowed definition of a predictive algorithm defining three groups predicting poor, intermediate and good clinical prognosis. The first of these comprised two patient sub-populations with GC-like tumours together with one sub

  12. pRV-Display-NY-ESO-1真核表达质粒的构建及其在Myc-CaP细胞株中的稳定表达%Construction of pRV-Display-NY-ESO-1 eukaryotic plasmid and its stable transfected Myc-CaP cell line

    Institute of Scientific and Technical Information of China (English)

    谢冲; 王国民

    2014-01-01

    Objective To construct the pRV-Display-NY-ESO-1 eukaryotic plasmid,and establish Myc-CaP cell line transfected with NY-ESO-1.Methods The full sequence of NY-ESO-1 was get from Genbank and amplified by polymerase chain reaction (PCR).Insert the fragments into p-Display eukaryotic expression vector.The recombinant plasmid with targeted gene was identified by digestion.Display-NY-ESO-1 was amplified by PCR and inserted into pRV retroviral vector.Retrovirus was produced by Phoenix cell line,and transfected into Myc-CaP cell line.Results The recombinant plasmid pRV-Display-NY-ESO-1 was successfully constructed and identified by PCR and DNA sequencing.Green fluorescence was observed on Phoenix cells and Myc-CaP cells encoding with GFP.The purity of Myc-CaP cells encoded with NY-ESO-1 was more than 80% according to flow cytometry.Conclusion Myc-CaP cell line was stably transfected with pRV-Display-NY-ESO-1,which is the base for further experiments.%目的 构建pRV-Display-NY-ESO-1真核表达质粒,建立并检测稳定表达纽约人食管鳞状细胞癌抗原(NY-ESO-1)的Myc-CaP细胞株.方法 从Genbank查找NY-ESO-1基因全长序列,聚合酶链反应(PCR)扩增目的片段,克隆至pDisplay载体,构建pDisplay-NY-ESO-1真核表达质粒,双酶切证实NY-ESO-1插入后,PCR扩增并定向连入pRV载体,构建pRV-Display-NY-ESO-1真核表达质粒,同时构建pDisplay-绿色荧光蛋白(GFP)作为阳性对照.经酶切、测序鉴定后,用Phoenix细胞产生逆转录病毒,脂质体转染法将重组质粒转入Myc-CaP细胞,并用Western blot和流式细胞术鉴定稳定转染的Myc-CaP细胞中NY-ESO-1的表达.结果 经PCR、测序鉴定成功构建pRV-Display-NY-ESO-1真核重组质粒.表达GFP的Phoenix细胞和Myc-CaP细胞在倒置荧光显微镜下均可见绿色荧光.Western blot检测Myc-CaP细胞中NY-ESO-1表达阳性,流式细胞术鉴定携带目的基因的Myc-CaP细胞纯度均大于80%.结论 pRV-Display-NY-ESO-1真核重组质粒稳定高表达于Myc-CaP细胞.

  13. Donor-matched mesenchymal stem cells from knee infrapatellar and subcutaneous adipose tissue of osteoarthritic donors display differential chondrogenic and osteogenic commitment

    Directory of Open Access Journals (Sweden)

    S Lopa

    2014-04-01

    Full Text Available Cell-based therapies have recently been proposed for the treatment of degenerative articular pathologies, such as early osteoarthritis, with an emphasis on autologous mesenchymal stem cells (MSCs, as an alternative to terminally differentiated cells. In this study, we performed a donor-matched comparison between infrapatellar fat pad MSCs (IFP-MSCs and knee subcutaneous adipose tissue stem cells (ASCs, as appealing candidates for cell-based therapies that are easily accessible during surgery. IFP-MSCs and ASCs were obtained from 25 osteoarthritic patients undergoing total knee replacement and compared for their immunophenotype and differentiative potential. Undifferentiated IFP-MSCs and ASCs displayed the same immunophenotype, typical of MSCs (CD13+/CD29+/CD44+/CD73+/CD90+/CD105+/CD166+/CD31-/CD45-. IFP-MSCs and ASCs showed similar adipogenic potential, though undifferentiated ASCs had higher LEP expression compared to IFP-MSCs (p < 0.01. Higher levels of calcified matrix (p < 0.05 and alkaline phosphatase (p < 0.05 in ASCs highlighted their superior osteogenic commitment compared to IFP-MSCs. Conversely, IFP-MSCs pellets showed greater amounts of glycosaminoglycans (p < 0.01 and superior expression of ACAN (p < 0.001, SOX9, COMP (p < 0.001 and COL2A1 (p < 0.05 compared to ASCs pellets, revealing a superior chondrogenic potential. This was also supported by lower COL10A1 (p < 0.05 and COL1A1 (p < 0.01 expression and lower alkaline phosphatase release (p < 0.05 by IFP-MSCs compared to ASCs. The observed dissimilarities between IFP-MSCs and ASCs show that, despite expressing similar surface markers, MSCs deriving from different fat depots in the same surgical site possess specific features. Furthermore, the in vitro peculiar commitment of IFP-MSCs and ASCs from osteoarthritic donors towards the chondrogenic or osteogenic lineage may suggest a preferential use for cartilage and bone cell-based treatments, respectively.

  14. Estrogen Receptor Beta Displays Cell Cycle-Dependent Expression and Regulates the G1 Phase through a Non-Genomic Mechanism in Prostate Carcinoma Cells

    Directory of Open Access Journals (Sweden)

    Antoni Hurtado

    2008-01-01

    Full Text Available Background: It is well known that estrogens regulate cell cycle progression, but the specific contributions and mechanisms of action of the estrogen receptor beta (ERβ remain elusive.

  15. Offspring of mothers fed a high fat diet display hepatic cell cycle inhibition and associated changes in gene expression and DNA methylation.

    Directory of Open Access Journals (Sweden)

    Kevin J Dudley

    Full Text Available The association between an adverse early life environment and increased susceptibility to later-life metabolic disorders such as obesity, type 2 diabetes and cardiovascular disease is described by the developmental origins of health and disease hypothesis. Employing a rat model of maternal high fat (MHF nutrition, we recently reported that offspring born to MHF mothers are small at birth and develop a postnatal phenotype that closely resembles that of the human metabolic syndrome. Livers of offspring born to MHF mothers also display a fatty phenotype reflecting hepatic steatosis and characteristics of non-alcoholic fatty liver disease. In the present study we hypothesised that a MHF diet leads to altered regulation of liver development in offspring; a derangement that may be detectable during early postnatal life. Livers were collected at postnatal days 2 (P2 and 27 (P27 from male offspring of control and MHF mothers (n = 8 per group. Cell cycle dynamics, measured by flow cytometry, revealed significant G0/G1 arrest in the livers of P2 offspring born to MHF mothers, associated with an increased expression of the hepatic cell cycle inhibitor Cdkn1a. In P2 livers, Cdkn1a was hypomethylated at specific CpG dinucleotides and first exon in offspring of MHF mothers and was shown to correlate with a demonstrable increase in mRNA expression levels. These modifications at P2 preceded observable reductions in liver weight and liver∶brain weight ratio at P27, but there were no persistent changes in cell cycle dynamics or DNA methylation in MHF offspring at this time. Since Cdkn1a up-regulation has been associated with hepatocyte growth in pathologic states, our data may be suggestive of early hepatic dysfunction in neonates born to high fat fed mothers. It is likely that these offspring are predisposed to long-term hepatic dysfunction.

  16. Invisible Display in Aluminum

    DEFF Research Database (Denmark)

    Prichystal, Jan Phuklin; Hansen, Hans Nørgaard; Bladt, Henrik Henriksen

    2005-01-01

    Bang & Olufsen a/s has been working with ideas for invisible integration of displays in metal surfaces. Invisible integration of information displays traditionally has been possible by placing displays behind transparent or semitransparent materials such as plastic or glass. The wish for an integ...... be obtained by shining light from the backside of the workpiece. When there is no light from the backside, the front surface seems totally untouched. This was achieved by laser ablation with ultra-short pulses.......Bang & Olufsen a/s has been working with ideas for invisible integration of displays in metal surfaces. Invisible integration of information displays traditionally has been possible by placing displays behind transparent or semitransparent materials such as plastic or glass. The wish...... for an integrated display in a metal surface is often ruled by design and functionality of a product. The integration of displays in metal surfaces requires metal removal in order to clear the area of the display to some extent. The idea behind an invisible display in Aluminum concerns the processing of a metal...

  17. Effects of leptin on human breast cancer cell lines in relationship to estrogen receptor and HER2 status.

    Science.gov (United States)

    Ray, Amitabha; Nkhata, Katai J; Cleary, Margot P

    2007-06-01

    Obesity is a risk factor for postmenopausal breast cancer and is associated with poor prognosis. Leptin, a cytokine synthesized in adipose tissue, has been implicated as a link between obesity and breast cancer. In the present study, the effects of leptin on cell proliferation and proteins associated with leptin signaling and/or breast cell growth were investigated in ER-positive, MCF-7, T47-D and MDA-MB-361, and ER-negative, MDA-MB-231 and SK-BR-3, breast cancer cell lines. MDA-MB-361 and SK-BR-3 also overexpress HER2/neu. For proliferation assays, 96-well plates were used and for protein determinations cells were synchronized in 6-well plates for 18-24 h in serum-free medium. Leptin was added at 0, 5, 10, 25, 50 and 100 ng/ml for 24 and 48 h. For Western blot analyses, protein extracts were probed for Ob-Rb, Ob-R, leptin, Jak2, PI3K, Stat3, p-Stat3, PCNA, cyclin D1, Cox-2, VEGF, Bcl-2, Bcl-xL, Bax, insulin, IGF-I, IGFBP3, IGF-IRalpha, aromatase, CYP1A1 and CYP1B1. Overall, except for MCF-7 cells, leptin stimulated proliferation in all lines. MCF-7 cells expressed higher levels of Ob-Rb, Jak2, PI3K, Stat3 and p-Stat3 in a dose-dependent manner to 50 ng/ml at 24 h; and IGF-IRalpha increased at 24 h. Cyclin D1 and Cox-2 levels increased with leptin treatment. Higher CYP1B1 expression was observed at both 24 and 48 h. In MDA-MB-231 cells, p-Stat3 and Bcl-xL were increased at 48 h; whereas PCNA and cyclin D1 expression increased in leptin-treated cells at 24 and 48 h. In T47-D cells, Jak2 and Stat3 were elevated at higher leptin concentrations at 24 and 48 h. However, p-Stat3 and PCNA demonstrated an increase only in 48-h leptin-treated cells. Furthermore, cyclin D1 exhibited higher expression at both 24 and 48 h, while Bcl-xL expression was lower with increasing concentrations of leptin at 48 h. In MDA-MB-361 cells, Ob-Rb and VEGF increased at 24 and 48 h; whereas PI3K, Stat3, PCNA and insulin levels increased in leptin-treated MDA-MB-361 cells after 48 h. Bcl-2 and

  18. Targeted silencing of DNA-specific B cells combined with partial plasma cell depletion displays additive effects on delaying disease onset in lupus-prone mice.

    Science.gov (United States)

    Nikolova-Ganeva, K A; Gesheva, V V; Todorov, T A; Voll, R E; Vassilev, T L

    2013-11-01

    Targeting autoreactive B lymphocytes at any stage of their differentiation could yield viable therapeutic strategies for treating autoimmunity. All currently used drugs, including the most recently introduced biological agents, lack target specificity. Selective silencing of double-stranded DNA-specific B cells in animals with spontaneous lupus has been achieved previously by the administration of a chimeric antibody molecule that cross-links their DNA-reactive B cell immunoglobulin receptors with inhibitory FcγIIb (CD32) receptors. However, long-lived plasmacytes are resistant to this chimeric antibody as well as to all conventional treatments. Bortezomib (a proteasome inhibitor) depletes most plasma cells and has been shown recently to suppress disease activity in lupus mice. We hypothesized that the co-administration of non-toxic doses of bortezomib, that partially purge long-lived plasma cells, together with an agent that selectively silences DNA-specific B cells, should have additive effects in an autoantibody-mediated disease. Indeed, our data show that the simultaneous treatment of lupus-prone MRL/lpr mice with suboptimal doses of bortezomib plus the chimeric antibody resulted in the prevention or the delayed appearance of the disease manifestations as well as in a prolonged survival. The effect of the combination therapy was significantly stronger than that of the respective monotherapies and was comparable to that observed after cyclophosphamide administration.

  19. Polyplanar optic display

    Energy Technology Data Exchange (ETDEWEB)

    Veligdan, J.; Biscardi, C.; Brewster, C.; DeSanto, L. [Brookhaven National Lab., Upton, NY (United States). Dept. of Advanced Technology; Beiser, L. [Leo Beiser Inc., Flushing, NY (United States)

    1997-07-01

    The Polyplanar Optical Display (POD) is a unique display screen which can be used with any projection source. This display screen is 2 inches thick and has a matte black face which allows for high contrast images. The prototype being developed is a form, fit and functional replacement display for the B-52 aircraft which uses a monochrome ten-inch display. The new display uses a 100 milliwatt green solid state laser (532 nm) as its optical source. In order to produce real-time video, the laser light is being modulated by a Digital Light Processing (DLP{trademark}) chip manufactured by Texas Instruments, Inc. A variable astigmatic focusing system is used to produce a stigmatic image on the viewing face of the POD. In addition to the optical design, the authors discuss the electronic interfacing to the DLP{trademark} chip, the opto-mechanical design and viewing angle characteristics.

  20. OLED displays and lighting

    CERN Document Server

    Koden, Mitsuhiro

    2017-01-01

    Organic light-emitting diodes (OLEDs) have emerged as the leading technology for the new display and lighting market. OLEDs are solid-state devices composed of thin films of organic molecules that create light with the application of electricity. OLEDs can provide brighter, crisper displays on electronic devices and use less power than conventional light-emitting diodes (LEDs) or liquid crystal displays (LCDs) used today. This book covers both the fundamentals and practical applications of flat and flexible OLEDs.

  1. Scalable Resolution Display Walls

    KAUST Repository

    Leigh, Jason

    2013-01-01

    This article will describe the progress since 2000 on research and development in 2-D and 3-D scalable resolution display walls that are built from tiling individual lower resolution flat panel displays. The article will describe approaches and trends in display hardware construction, middleware architecture, and user-interaction design. The article will also highlight examples of use cases and the benefits the technology has brought to their respective disciplines. © 1963-2012 IEEE.

  2. Essential Oil from Berries of Lebanese Juniperus excelsa M. Bieb Displays Similar Antibacterial Activity to Chlorhexidine but Higher Cytocompatibility with Human Oral Primary Cells

    Directory of Open Access Journals (Sweden)

    Barbara Azzimonti

    2015-05-01

    Full Text Available Chlorhexidine (CHX, one of the most effective drugs administered for periodontal treatment, presents collateral effects including toxicity when used for prolonged periods; here, we have evaluated the bactericidal potency and the cytocompatibility of Juniperus excelsa M. Bieb essential oil (EO in comparison with 0.05% CHX. The EO was extracted from berries by hydrodistillation and components identified by gas chromatography and mass spectrometry. Bacterial inhibition halo analysis, quantitative cell viability 2,3-bis(2-methoxy-4-nitro-5-sulphophenyl-5-[(phenyl amino carbonyl]-2H-tetrazolium hydroxide assay (XTT, and colony forming unit (CFU count were evaluated against the two biofilm formers Aggregatibacter actinomycetemcomitans and Streptococcus mutans. Finally, cytocompatibility was assessed with human primary gingival fibroblasts (HGF and mucosal keratinocytes (HK. The resulting EO was mainly composed of monoterpene hydrocarbons and oxygenated monoterpenes. An inhibition halo test demonstrated that both bacteria were sensitive to the EO; XTT analysis and CFU counts confirmed that 10-fold-diluted EO determined a statistically significant (p < 0.05 reduction in bacteria count and viability towards both biofilm and planktonic forms in a comparable manner to those obtained with CHX. Moreover, EO displayed higher cytocompatibility than CHX (p < 0.05. In conclusion, EO exhibited bactericidal activity similar to CHX, but a superior cytocompatibility, making it a promising antiseptic alternative to CHX.

  3. JAVA Stereo Display Toolkit

    Science.gov (United States)

    Edmonds, Karina

    2008-01-01

    This toolkit provides a common interface for displaying graphical user interface (GUI) components in stereo using either specialized stereo display hardware (e.g., liquid crystal shutter or polarized glasses) or anaglyph display (red/blue glasses) on standard workstation displays. An application using this toolkit will work without modification in either environment, allowing stereo software to reach a wider audience without sacrificing high-quality display on dedicated hardware. The toolkit is written in Java for use with the Swing GUI Toolkit and has cross-platform compatibility. It hooks into the graphics system, allowing any standard Swing component to be displayed in stereo. It uses the OpenGL graphics library to control the stereo hardware and to perform the rendering. It also supports anaglyph and special stereo hardware using the same API (application-program interface), and has the ability to simulate color stereo in anaglyph mode by combining the red band of the left image with the green/blue bands of the right image. This is a low-level toolkit that accomplishes simply the display of components (including the JadeDisplay image display component). It does not include higher-level functions such as disparity adjustment, 3D cursor, or overlays all of which can be built using this toolkit.

  4. Helmet-Mounted Displays (HMD)

    Data.gov (United States)

    Federal Laboratory Consortium — The Helmet-Mounted Display labis responsible for monocular HMD day display evaluations; monocular HMD night vision performance processes; binocular HMD day display...

  5. Standardizing visual display quality

    NARCIS (Netherlands)

    Besuijen, Ko; Spenkelink, Gerd P.J.

    1998-01-01

    The current ISO 9241–3 standard for visual display quality and the proposed user performance tests are reviewed. The standard is found to be more engineering than ergonomic and problems with system configuration, software applications, display settings, user behaviour, wear and physical environment

  6. Polyplanar optical display electronics

    Energy Technology Data Exchange (ETDEWEB)

    DeSanto, L.; Biscardi, C. [Brookhaven National Lab., Upton, NY (United States). Dept. of Advanced Technology

    1997-07-01

    The Polyplanar Optical Display (POD) is a unique display screen which can be used with any projection source. The prototype ten inch display is two inches thick and has a matte black face which allows for high contrast images. The prototype being developed is a form, fit and functional replacement display for the B-52 aircraft which uses a monochrome ten-inch display. In order to achieve a long lifetime, the new display uses a 100 milliwatt green solid-state laser (10,000 hr. life) at 532 nm as its light source. To produce real-time video, the laser light is being modulated by a Digital Light Processing (DLP{trademark}) chip manufactured by Texas Instruments. In order to use the solid-state laser as the light source and also fit within the constraints of the B-52 display, the Digital Micromirror Device (DMD{trademark}) circuit board is removed from the Texas Instruments DLP light engine assembly. Due to the compact architecture of the projection system within the display chassis, the DMD{trademark} chip is operated remotely from the Texas Instruments circuit board. The authors discuss the operation of the DMD{trademark} divorced from the light engine and the interfacing of the DMD{trademark} board with various video formats (CVBS, Y/C or S-video and RGB) including the format specific to the B-52 aircraft. A brief discussion of the electronics required to drive the laser is also presented.

  7. Visual merchandising window display

    Directory of Open Access Journals (Sweden)

    Opris (Cas. Stanila M.

    2013-12-01

    Full Text Available Window display plays a major part in the selling strategies; it does not only include the simple display of goods, nowadays it is a form of art, also having the purpose of sustaining the brand image. This article wants to reveal the tools that are essential in creating a fabulous window display. Being a window designer is not an easy job, you have to always think ahead trends, to have a sense of colour, to know how to use light to attract customers in the store after only one glance at the window. The big store window displays are theatre scenes: with expensive backgrounds, special effects and high fashion mannequins. The final role of the displays is to convince customers to enter the store and trigger the purchasing act which is the final goal of the retail activity.

  8. Scaling-up the synthesis of myristate glucose ester catalyzed by a CALB-displaying Pichia pastoris whole-cell biocatalyst.

    Science.gov (United States)

    Guo, DongHeng; Jin, Zi; Xu, YanShan; Wang, Ping; Lin, Ying; Han, ShuangYan; Zheng, SuiPing

    2015-01-01

    The novel whole-cell biocatalyst Candida antarctica lipase B displaying-Pichia pastoris (Pp-CALB) is characterized by its low preparation cost and could be an alternative to the commercial immobilized Candida antarctica lipase B (CALB). This study addresses the feasibility of using Pp-CALB in large scale glucose fatty acid esters production. 1,2-O-Isopropylidene-α-D-glucofuranose (IpGlc) was used as the acyl acceptor to overcome the low solubility of glucose in an organic solvent and to avoid the addition of toxic co-solvents. IpGlc significantly improved the Pp-CALB catalyzing esterification efficiency when using long chain fatty acids as the acyl donor. Under the preferred operating conditions (50 °C, 40 g/L molecular sieve dosage and 200 rpm mixing intensity), 60.5% of IpGlc converted to 6-O-myristate-1, 2-O-isopropylidene-α-D-glucofuranose (C14-IpGlc) after a 96-h reaction in a 2-L stirred reactor. In a 5-L pilot scale test, Pp-CALB also showed a similar substrate conversion rate of 55.4% and excellent operational stability. After C14-IpGlc was collected, 70% trifluoroacetic acid was adopted to hydrolyze C14-IpGlc to myristate glucose ester (C14-Glc) with a high yield of 95.3%. In conclusion, Pp-CALB is a powerful biocatalyst available for industrial synthesis, and this study describes an applicable and economical process for the large scale production of myristate glucose ester.

  9. AarF Domain Containing Kinase 3 (ADCK3 Mutant Cells Display Signs of Oxidative Stress, Defects in Mitochondrial Homeostasis and Lysosomal Accumulation.

    Directory of Open Access Journals (Sweden)

    Jason K Cullen

    Full Text Available Autosomal recessive ataxias are a clinically diverse group of syndromes that in some cases are caused by mutations in genes with roles in the DNA damage response, transcriptional regulation or mitochondrial function. One of these ataxias, known as Autosomal Recessive Cerebellar Ataxia Type-2 (ARCA-2, also known as SCAR9/COQ10D4; OMIM: #612016, arises due to mutations in the ADCK3 gene. The product of this gene (ADCK3 is an atypical kinase that is thought to play a regulatory role in coenzyme Q10 (CoQ10 biosynthesis. Although much work has been performed on the S. cerevisiae orthologue of ADCK3, the cellular and biochemical role of its mammalian counterpart, and why mutations in this gene lead to human disease is poorly understood. Here, we demonstrate that ADCK3 localises to mitochondrial cristae and is targeted to this organelle via the presence of an N-terminal localisation signal. Consistent with a role in CoQ10 biosynthesis, ADCK3 deficiency decreased cellular CoQ10 content. In addition, endogenous ADCK3 was found to associate in vitro with recombinant Coq3, Coq5, Coq7 and Coq9, components of the CoQ10 biosynthetic machinery. Furthermore, cell lines derived from ARCA-2 patients display signs of oxidative stress, defects in mitochondrial homeostasis and increases in lysosomal content. Together, these data shed light on the possible molecular role of ADCK3 and provide insight into the cellular pathways affected in ARCA-2 patients.

  10. Panoramic projection avionics displays

    Science.gov (United States)

    Kalmanash, Michael H.

    2003-09-01

    Avionics projection displays are entering production in advanced tactical aircraft. Early adopters of this technology in the avionics community used projection displays to replace or upgrade earlier units incorporating direct-view CRT or AMLCD devices. Typical motivation for these upgrades were the alleviation of performance, cost and display device availability concerns. In these systems, the upgraded (projection) displays were one-for-one form / fit replacements for the earlier units. As projection technology has matured, this situation has begun to evolve. The Lockheed-Martin F-35 is the first program in which the cockpit has been specifically designed to take advantage of one of the more unique capabilities of rear projection display technology, namely the ability to replace multiple small screens with a single large conformal viewing surface in the form of a panoramic display. Other programs are expected to follow, since the panoramic formats enable increased mission effectiveness, reduced cost and greater information transfer to the pilot. Some of the advantages and technical challenges associated with panoramic projection displays for avionics applications are described below.

  11. Behaviour of one-step spray-coated carbon nanotube supercapacitor in ambient light harvester circuit with printed organic solar cell and electrochromic display.

    Science.gov (United States)

    Tuukkanen, Sampo; Välimäki, Marja; Lehtimäki, Suvi; Vuorinen, Tiina; Lupo, Donald

    2016-03-09

    A printed energy harvesting and storage circuit powered by ambient office lighting and its use to power a printed display is reported. The autonomous device is composed of three printed electronic components: an organic photovoltaic module, a carbon-nanotubes-only supercapacitor and an electrochromic display element. Components are fabricated from safe and environmentally friendly materials, and have been fabricated using solution processing methods, which translate into low-cost and high-throughput manufacturing. A supercapacitor made of spray-coated carbon nanotube based ink and aqueous NaCl electrolyte was charged using a printed organic photovoltaic module exposed to office lighting conditions. The supercapacitor charging rate, self-discharge rate and display operation were studied in detail. The supercapacitor self-discharge rate was found to depend on the charging rate. The fully charged supercapacitor was used as a power source to run the electrochromic display over 50 times.

  12. Behaviour of one-step spray-coated carbon nanotube supercapacitor in ambient light harvester circuit with printed organic solar cell and electrochromic display

    Science.gov (United States)

    Tuukkanen, Sampo; Välimäki, Marja; Lehtimäki, Suvi; Vuorinen, Tiina; Lupo, Donald

    2016-03-01

    A printed energy harvesting and storage circuit powered by ambient office lighting and its use to power a printed display is reported. The autonomous device is composed of three printed electronic components: an organic photovoltaic module, a carbon-nanotubes-only supercapacitor and an electrochromic display element. Components are fabricated from safe and environmentally friendly materials, and have been fabricated using solution processing methods, which translate into low-cost and high-throughput manufacturing. A supercapacitor made of spray-coated carbon nanotube based ink and aqueous NaCl electrolyte was charged using a printed organic photovoltaic module exposed to office lighting conditions. The supercapacitor charging rate, self-discharge rate and display operation were studied in detail. The supercapacitor self-discharge rate was found to depend on the charging rate. The fully charged supercapacitor was used as a power source to run the electrochromic display over 50 times.

  13. Functional characterization and localization of a Bacillus subtilis sortase and its substrate and use of this sortase system to covalently anchor a heterologous protein to the B. subtilis cell wall for surface display.

    Science.gov (United States)

    Liew, Pei Xiong; Wang, Christopher L C; Wong, Sui-Lam

    2012-01-01

    Sortases catalyze the covalent anchoring of proteins to the cell surface on Gram-positive bacteria. Bioinformatic analysis suggests the presence of structural genes encoding sortases and their substrates in the Bacillus subtilis genome. In this study, a β-lactamase reporter was fused to the cell wall anchoring domain from a putative sortase substrate, YhcR. Covalent anchoring of this fusion protein to the cell wall was confirmed by using the eight-protease-deficient B. subtilis strain WB800 as the host. Inactivation of yhcS abolished the cell wall anchoring reaction. The amounts of fusion protein anchored to the cell wall were proportional to the levels of YhcS. These data demonstrate that YhcS and YhcR are the sortase and sortase substrate, respectively, in B. subtilis. Furthermore, yhcS is not essential for the survival of B. subtilis under the cultivation condition tested. YhcR fusions were distributed helically in the lateral cell wall. Interestingly, when viewed with an epifluorescence microscope, YhcS also appeared to form short helical arcs. This is the first report to illustrate such distribution of sortases in a rod-shaped bacterium. Models for the spatial distribution of both the sortase and its substrate are discussed. The amount of the reporters displayed on the surface was unambiguously quantified via a unique strategy. Under optimal conditions with the overproduction of YhcS, 47,300 YhcR fusions could be displayed per cell. Displayed reporters were biologically functional and surface accessible. Characterization of the sortase-substrate system allowed the successful development of a YhcR-based covalent surface display system. This system may have various biotechnological applications.

  14. Small - Display Cartography

    DEFF Research Database (Denmark)

    Nissen, Flemming; Hvas, Anders; Münster-Swendsen, Jørgen

    This report comprises the work carried out in the work-package of small display cartography. The work-package has aimed at creating a general framework for the small-display cartography. A solid framework facilitates an increased use of spatial data in mobile devices - thus enabling, together...... with the rapidly evolving positioning techniques, a new category of position-dependent, map-based services to be introduced. The report consists of the following parts: Part I: Categorization of handheld devices, Part II: Cartographic design for small-display devices, Part III: Study on the GiMoDig Client ? Portal...... Appendix E consisting of confidential material and Appendix F with examples.   The work has resulted in a small device categorisation, a cartographic design specification for a small-display device for a specified navigation task as well as a comparison between OpenLS and an in house developed protocol...

  15. Flexible displays, rigid designs?

    DEFF Research Database (Denmark)

    Hornbæk, Kasper

    2015-01-01

    Rapid technological progress has enabled a wide range of flexible displays for computing devices, but the user experience--which we're only beginning to understand--will be the key driver for successful designs....

  16. Military display performance parameters

    Science.gov (United States)

    Desjardins, Daniel D.; Meyer, Frederick

    2012-06-01

    The military display market is analyzed in terms of four of its segments: avionics, vetronics, dismounted soldier, and command and control. Requirements are summarized for a number of technology-driving parameters, to include luminance, night vision imaging system compatibility, gray levels, resolution, dimming range, viewing angle, video capability, altitude, temperature, shock and vibration, etc., for direct-view and virtual-view displays in cockpits and crew stations. Technical specifications are discussed for selected programs.

  17. Liquid Crystal Airborne Display

    Science.gov (United States)

    1977-08-01

    81/2X 11- 10 -9 .8 display using a large advertising alphanimeric ( TCI ) has been added to the front of the optical box used in the F-4 aircraft for HUD...properties over a wide range of tempera - tures, including normal room temperature. What are Liquid Crystals? Liquid crystals have been classified in three...natic fanctions and to present data needed for the semi- automatic and manual control of system functions. Existing aircraft using CRT display

  18. Raster graphics display library

    Science.gov (United States)

    Grimsrud, Anders; Stephenson, Michael B.

    1987-01-01

    The Raster Graphics Display Library (RGDL) is a high level subroutine package that give the advanced raster graphics display capabilities needed. The RGDL uses FORTRAN source code routines to build subroutines modular enough to use as stand-alone routines in a black box type of environment. Six examples are presented which will teach the use of RGDL in the fastest, most complete way possible. Routines within the display library that are used to produce raster graphics are presented in alphabetical order, each on a separate page. Each user-callable routine is described by function and calling parameters. All common blocks that are used in the display library are listed and the use of each variable within each common block is discussed. A reference on the include files that are necessary to compile the display library is contained. Each include file and its purpose are listed. The link map for MOVIE.BYU version 6, a general purpose computer graphics display system that uses RGDL software, is also contained.

  19. A case of mistaken identity: CD11c-eYFP(+) cells in the normal mouse brain parenchyma and neural retina display the phenotype of microglia, not dendritic cells.

    Science.gov (United States)

    Dando, Samantha J; Naranjo Golborne, Cecilia; Chinnery, Holly R; Ruitenberg, Marc J; McMenamin, Paul G

    2016-08-01

    Under steady-state conditions the central nervous system (CNS) is traditionally thought to be devoid of antigen presenting cells; however, putative dendritic cells (DCs) expressing enhanced yellow fluorescent protein (eYFP) are present in the retina and brain parenchyma of CD11c-eYFP mice. We previously showed that these mice carry the Crb1(rd8) mutation, which causes retinal dystrophic lesions; therefore we hypothesized that the presence of CD11c-eYFP(+) cells within the CNS may be due to pathology associated with the Crb1(rd8) mutation. We generated CD11c-eYFP Crb1(wt/wt) mice and compared the distribution and immunophenotype of CD11c-eYFP(+) cells in CD11c-eYFP mice with and without the Crb1(rd8) mutation. The number and distribution of CD11c-eYFP(+) cells in the CNS was similar between CD11c-eYFP Crb1(wt/wt) and CD11c-eYFP Crb1(rd8/rd8) mice. CD11c-eYFP(+) cells were distributed throughout the inner retina, and clustered in brain regions that receive input from the external environment or lack a blood-brain barrier. CD11c-eYFP(+) cells within the retina and cerebral cortex of CD11c-eYFP Crb1(wt/wt) mice expressed CD11b, F4/80, CD115 and Iba-1, but not DC or antigen presentation markers, whereas CD11c-eYFP(+) cells within the choroid plexus and pia mater expressed CD11c, I-A/I-E, CD80, CD86, CD103, DEC205, CD8α and CD135. The immunophenotype of CD11c-eYFP(+) cells and microglia within the CNS was similar between CD11c-eYFP Crb1(wt/wt) and CD11c-eYFP Crb1(rd8/rd8) mice; however, CD11c and I-A/I-E expression was significantly increased in CD11c-eYFP Crb1(rd8/rd8) mice. This study demonstrates that the overwhelming majority of CNS CD11c-eYFP(+) cells do not display the phenotype of DCs or their precursors and are most likely a subpopulation of microglia. GLIA 2016. GLIA 2016;64:1331-1349.

  20. Discovery of high affinity anti-ricin antibodies by B cell receptor sequencing and by yeast display of combinatorial VH:VL libraries from immunized animals.

    Science.gov (United States)

    Wang, Bo; Lee, Chang-Han; Johnson, Erik L; Kluwe, Christien A; Cunningham, Josephine C; Tanno, Hidetaka; Crooks, Richard M; Georgiou, George; Ellington, Andrew D

    2016-01-01

    Ricin is a toxin that could potentially be used as a bioweapon. We identified anti-ricin A chain antibodies by sequencing the antibody repertoire from immunized mice and by selecting high affinity antibodies using yeast surface display. These methods led to the isolation of multiple antibodies with high (sub-nanomolar) affinity. Interestingly, the antibodies identified by the 2 independent approaches are from the same clonal lineages, indicating for the first time that yeast surface display can identify native antibodies. The new antibodies represent well-characterized reagents for biodefense diagnostics and therapeutics development.

  1. The Ultimate Display

    CERN Document Server

    Fluke, C J

    2016-01-01

    Astronomical images and datasets are increasingly high-resolution and multi-dimensional. The vast majority of astronomers perform all of their visualisation and analysis tasks on low-resolution, two-dimensional desktop monitors. If there were no technological barriers to designing the ultimate stereoscopic display for astronomy, what would it look like? What capabilities would we require of our compute hardware to drive it? And are existing technologies even close to providing a true 3D experience that is compatible with the depth resolution of human stereoscopic vision? We consider the CAVE2 (an 80 Megapixel, hybrid 2D and 3D virtual reality environment directly integrated with a 100 Tflop/s GPU-powered supercomputer) and the Oculus Rift (a low- cost, head-mounted display) as examples at opposite financial ends of the immersive display spectrum.

  2. Virtual Auditory Displays

    Science.gov (United States)

    2000-01-01

    timbre , intensity, distance, room modeling, radio communication Virtual Environments Handbook Chapter 4 Virtual Auditory Displays Russell D... musical note “A” as a pure sinusoid, there will be 440 condensations and rarefactions per second. The distance between two adjacent condensations or...and complexity are pitch, loudness, and timbre respectively. This distinction between physical and perceptual measures of sound properties is an

  3. Virtual acoustic displays

    Science.gov (United States)

    Wenzel, Elizabeth M.

    1991-01-01

    A 3D auditory display can potentially enhance information transfer by combining directional and iconic information in a quite naturalistic representation of dynamic objects in the interface. Another aspect of auditory spatial clues is that, in conjunction with other modalities, it can act as a potentiator of information in the display. For example, visual and auditory cues together can reinforce the information content of the display and provide a greater sense of presence or realism in a manner not readily achievable by either modality alone. This phenomenon will be particularly useful in telepresence applications, such as advanced teleconferencing environments, shared electronic workspaces, and monitoring telerobotic activities in remote or hazardous situations. Thus, the combination of direct spatial cues with good principles of iconic design could provide an extremely powerful and information-rich display which is also quite easy to use. An alternative approach, recently developed at ARC, generates externalized, 3D sound cues over headphones in realtime using digital signal processing. Here, the synthesis technique involves the digital generation of stimuli using Head-Related Transfer Functions (HRTF's) measured in the two ear-canals of individual subjects. Other similar approaches include an analog system developed by Loomis, et. al., (1990) and digital systems which make use of transforms derived from normative mannikins and simulations of room acoustics. Such an interface also requires the careful psychophysical evaluation of listener's ability to accurately localize the virtual or synthetic sound sources. From an applied standpoint, measurement of each potential listener's HRTF's may not be possible in practice. For experienced listeners, localization performance was only slightly degraded compared to a subject's inherent ability. Alternatively, even inexperienced listeners may be able to adapt to a particular set of HRTF's as long as they provide adequate

  4. New role of JAK2/STAT3 signaling in endothelial cell oxidative stress injury and protective effect of melatonin.

    Directory of Open Access Journals (Sweden)

    Weixun Duan

    Full Text Available Previous studies have shown that the JAK2/STAT3 signaling pathway plays a regulatory role in cellular oxidative stress injury (OSI. In this study, we explored the role of the JAK2/STAT3 signaling pathway in hydrogen peroxide (H2O2-induced OSI and the protective effect of melatonin against (H2O2-induced injury in human umbilical vein endothelial cells (HUVECs. AG490 (a specific inhibitor of the JAK2/STAT3 signaling pathway and JAK2 siRNA were used to manipulate JAK2/STAT3 activity, and the results showed that AG490 and JAK2 siRNA inhibited OSI and the levels of p-JAK2 and p-STAT3. HUVECs were then subjected to H2O2 in the absence or presence of melatonin, the main secretory product of the pineal gland. Melatonin conferred a protective effect against H2O2, which was evidenced by improvements in cell viability, adhesive ability and migratory ability, decreases in the apoptotic index and reactive oxygen species (ROS production and several biochemical parameters in HUVECs. Immunofluorescence and Western blotting showed that H2O2 treatment increased the levels of p-JAK2, p-STAT3, Cytochrome c, Bax and Caspase3 and decreased the levels of Bcl2, whereas melatonin treatment partially reversed these effects. We, for the first time, demonstrate that the inhibition of the JAK2/STAT3 signaling pathway results in a protective effect against endothelial OSI. The protective effects of melatonin against OSI, at least partially, depend upon JAK2/STAT3 inhibition.

  5. Identification of Hitherto Undefined B-Cell Epitopes by Antibodies in the Sera of Vitiligo Patients Using Phage-Display Peptide Library

    Directory of Open Access Journals (Sweden)

    Zohreh Jadali

    2003-12-01

    Full Text Available A random 12 mers phage library was used to screen a pool of immunoglo¬bulin fractions obtained from vitiligo patients. Subsequent to panning experiments, a panel of affinity selected phage from vitiligo patients were obtained. This panel was tested using an ELIS A for their reactivity with pooled sera from patients and normal controls. Among the 16 randomly selected clones, two of clones showed distinct positive reactivity with the patient's sera compared with controls. The peptides displayed by these phages expressed the following amino acid sequences: SHMPLANQYQWA and NHVQAWEQFWDS. Thus, screening with phage-displayed random peptide library of vitiligo sera can reveal peptide sequences that mimic vitiligo-related self-antigen.

  6. Chronic lymphocytic leukemia nurse-like cells express hepatocyte growth factor receptor (c-MET) and indoleamine 2,3-dioxygenase and display features of immunosuppressive type 2 skewed macrophages

    Science.gov (United States)

    Giannoni, Paolo; Pietra, Gabriella; Travaini, Giorgia; Quarto, Rodolfo; Shyti, Genti; Benelli, Roberto; Ottaggio, Laura; Mingari, Maria Cristina; Zupo, Simona; Cutrona, Giovanna; Pierri, Ivana; Balleari, Enrico; Pattarozzi, Alessandra; Calvaruso, Marco; Tripodo, Claudio; Ferrarini, Manlio; de Totero, Daniela

    2014-01-01

    Hepatocyte growth factor, produced by stromal and follicular dendritic cells, and present at high concentrations in the sera of patients with chronic lymphocytic leukemia, prolongs the survival of leukemic B cells by interacting with their receptor, c-MET. It is, however, unknown whether hepatocyte growth factor influences microenvironmental cells, such as nurse-like cells, which deliver survival signals to the leukemic clone. We evaluated the expression of c-MET on nurse-like cells and monocytes from patients with chronic lymphocytic leukemia and searched for phenotypic/functional features supposed to be influenced by the hepatocyte growth factor/c-MET interaction. c-MET is expressed at high levels on nurse-like cells and at significantly higher levels than normal on monocytes from patients. Moreover, the hepatocyte growth factor/c-MET interaction activates STAT3TYR705 phosphorylation in nurse-like cells. Indoleamine 2,3-dioxygenase, an enzyme modulating T-cell proliferation and induced on normal monocytes after hepatocyte growth factor treatment, was detected together with interleukin-10 on nurse-like cells, and on freshly-prepared patients’ monocytes. Immunohistochemical/immunostaining analyses demonstrated the presence of c-MET+ and indoleamine 2,3-dioxygenase+ cells in lymph node biopsies, co-expressed with CD68 and vimentin. Furthermore nurse-like cells and chronic lymphocytic monocytes significantly inhibited T-cell proliferation, prevented by anti-transforming growth factor beta and interleukin-10 antibodies and indoleamine 2,3-dioxygenase inhibitors, and supported CD4+CD25high+/FOXP3+ T regulatory cell expansion. We suggest that nurse-like cells display features of immunosuppressive type 2 macrophages: higher hepatocyte growth factor levels, produced by leukemic or other microenvironmental surrounding cells, may cooperate to induce M2 polarization. Hepatocyte growth factor may thus have a dual pathophysiological role: directly through enhancement of

  7. Application of whole-cell subtractive panning in phage display library screening%全细胞差减筛选法在噬菌体展示文库筛选中的应用

    Institute of Scientific and Technical Information of China (English)

    吴红珍; 张林波

    2012-01-01

    全细胞差减筛选法是近年来在细胞筛选的基础上发展起来的一项筛选技术,其利用成对的细胞,即两种不同状态的细胞对噬菌体展示文库进行差减筛选,主要用于筛选新的抗原表位、受体、配体、新型疫苗、肿瘤细胞的靶向活性肽、靶向基因载体及受体或酶的激动剂或抑制剂等.本文就其筛选的原理、优缺点、优化及其在噬菌体展示文库筛选中的应用作一综述.%Whole-cell subtractive panning is a screening technique developed based on whole-cell screening in recent years, which performs subtractive panning on phage display library by using couple cells, I.e. Two states of cells. It is mainly used for screening of novel antigenic epitopes, receptors, ligands, novel vaccines, the targeted active peptides of tumor cells, targeted gene vectors, and stimulants or antagonists of receptors or enzyme etc. The principle, advantages, disadvantages and optimization of the technique as well as its application in phage display library screening are reviewed in this paper.

  8. Refrigerated display cabinets; Butikskyla

    Energy Technology Data Exchange (ETDEWEB)

    Fahlen, Per

    2000-07-01

    This report summarizes experience from SP research and assignments regarding refrigerated transport and storage of food, mainly in the retail sector. It presents the fundamentals of heat and mass transfer in display cabinets with special focus on indirect systems and secondary refrigerants. Moreover, the report includes a brief account of basic food hygiene and the related regulations. The material has been compiled for educational purposes in the Masters program at Chalmers Technical University.

  9. Decorating microbes : surface display of proteins on Escherichia coli

    NARCIS (Netherlands)

    van Bloois, Edwin; Winter, Remko T.; Kolmar, Harald; Fraaije, Marco W.

    2011-01-01

    Bacterial surface display entails the presentation of recombinant proteins or peptides on the surface of bacterial cells. Escherichia coil is the most frequently used bacterial host for surface display and, as such, a variety of E. coil display systems have been described that primarily promote the

  10. Book Display as Adult Service

    Directory of Open Access Journals (Sweden)

    Matthew S. Moore

    1997-03-01

    Full Text Available 無Book display as an adult service is defined as choosing and positioning adult books from the collection to increase their circulation. The author contrasts bookstore arrangement for sales versus library arrangement for access. The paper considers the library-as-a-whole as a display, examines the right size for an in-library display, and discusses mass displays, end-caps, on-shelf displays, and the Tiffany approach. The author proposes that an effective display depends on an imaginative, unifying theme, and that book displays are part of the joy of libraries.

  11. Handbook of Visual Display Technology

    CERN Document Server

    Cranton, Wayne; Fihn, Mark

    2012-01-01

    The Handbook of Visual Display Technology is a unique work offering a comprehensive description of the science, technology, economic and human interface factors associated with the displays industry. An invaluable compilation of information, the Handbook will serve as a single reference source with expert contributions from over 150 international display professionals and academic researchers. All classes of display device are covered including LCDs, reflective displays, flexible solutions and emissive devices such as OLEDs and plasma displays, with discussion of established principles, emergent technologies, and particular areas of application. The wide-ranging content also encompasses the fundamental science of light and vision, image manipulation, core materials and processing techniques, display driving and metrology.

  12. Identification of genes coding for B cell antigens of Mycoplasma mycoides subsp. mycoides Small Colony (MmmSC by using phage display

    Directory of Open Access Journals (Sweden)

    Vilei Edy M

    2009-10-01

    Full Text Available Abstract Background Contagious bovine pleuropneumonia (CBPP is a mycoplasmal disease caused by Mycoplasma mycoides subsp. mycoides SC (MmmSC. Since the disease is a serious problem that can affect cattle production in parts of Africa, there is a need for an effective and economical vaccine. Identifying which of the causative agent's proteins trigger potentially protective immune responses is an important step towards developing a subunit vaccine. Accordingly, the purpose of this study was to determine whether phage display combined with bioinformatics could be used to narrow the search for genes that code for potentially immunogenic proteins of MmmSC. Since the production of IgG2 and IgA are associated with a Th1 cellular immune response which is implicated in protection against CBPP, antigens which elicit these immunoglobulin subclasses may be useful in developing a subunit vaccine. Results A filamentous phage library displaying a repertoire of peptides expressed by fragments of the genome of MmmSC was constructed. It was subjected to selection using antibodies from naturally- and experimentally-infected cattle. Mycoplasmal genes were identified by matching the nucleotide sequences of DNA from immunoselected phage particles with the mycoplasmal genome. This allowed a catalogue of genes coding for the proteins that elicited an immune response to be compiled. Using this method together with computer algorithms designed to score parameters that influence surface accessibility and hence potential antigenicity, five genes (abc, gapN, glpO, lppB and ptsG were chosen to be expressed in Escherichia coli. After appropriate site-directed mutagenesis, polypeptides representing portions of each of these proteins were tested for immunoreactivity. Of these five, polypeptides representing expression products of abc and lppB were recognised on immunoblots by sera obtained from cattle during a natural outbreak of the disease. Conclusion Since phage display

  13. Lycorine, the Main Phenanthridine Amaryllidaceae Alkaloid, Exhibits Significant Anti-Tumor Activity in Cancer Cells that Display Resistance to Proapoptotic Stimuli: an Investigation of Structure-Activity Relationship and Mechanistic Insight

    Science.gov (United States)

    Lamoral-Theys, Delphine; Andolfi, Anna; Van Goietsenoven, Gwendoline; Cimmino, Alessio; Le Calvé, Benjamin; Wauthoz, Nathalie; Mégalizzi, Véronique; Gras, Thierry; Bruyère, Céline; Dubois, Jacques; Mathieu, Véronique; Kornienko, Alexander; Kiss, Robert; Evidente, Antonio

    2011-01-01

    Twenty-two lycorine-related compounds were investigated for in vitro anti-tumor activity using four cancer cell lines displaying different levels of resistance to pro-apoptotic stimuli and two cancer cell lines sensitive to pro-apoptotic stimuli. Lycorine and six of its congeners exhibited potency in the single-digit micromolar range, while no compound appeared more active than lycorine. Lycorine also displayed the highest potential (in vitro) therapeutic ratio, being at least 15 times more active against cancer than normal cells. Our studies also showed that lycorine exerts its in vitro anti-tumor activity through cytostatic rather than cytotoxic effects. Furthermore, lycorine provided significant therapeutic benefit in mice bearing brain grafts of the B16F10 melanoma model at non-toxic doses. Thus, the results of the current study make lycorine an excellent lead for the generation of compounds able to combat cancers, which are naturally resistant to pro-apoptotic stimuli, such as glioblastoma, melanoma, non-small-cell-lung cancers, metastatic cancers, among others. PMID:19788245

  14. Lycorine, the main phenanthridine Amaryllidaceae alkaloid, exhibits significant antitumor activity in cancer cells that display resistance to proapoptotic stimuli: an investigation of structure-activity relationship and mechanistic insight.

    Science.gov (United States)

    Lamoral-Theys, Delphine; Andolfi, Anna; Van Goietsenoven, Gwendoline; Cimmino, Alessio; Le Calvé, Benjamin; Wauthoz, Nathalie; Mégalizzi, Véronique; Gras, Thierry; Bruyère, Céline; Dubois, Jacques; Mathieu, Véronique; Kornienko, Alexander; Kiss, Robert; Evidente, Antonio

    2009-10-22

    Twenty-two lycorine-related compounds were investigated for in vitro antitumor activity using four cancer cell lines displaying different levels of resistance to proapoptotic stimuli and two cancer cell lines sensitive to proapoptotic stimuli. Lycorine and six of its congeners exhibited potency in the single-digit micromolar range, while no compound appeared more active than lycorine. Lycorine also displayed the highest potential (in vitro) therapeutic ratio, being at least 15 times more active against cancer than normal cells. Our studies also showed that lycorine exerts its in vitro antitumor activity through cytostatic rather than cytotoxic effects. Furthermore, lycorine provided significant therapeutic benefit in mice bearing brain grafts of the B16F10 melanoma model at nontoxic doses. Thus, the results of the current study make lycorine an excellent lead for the generation of compounds able to combat cancers, which are naturally resistant to proapoptotic stimuli, such as glioblastoma, melanoma, non-small-cell-lung cancers, and metastatic cancers, among others.

  15. Adult Human Pancreatic Islet Beta-Cells Display Limited Turnover and Long Lifespan as Determined by In-Vivo Thymidine Analog Incorporation and Radiocarbon Dating

    Energy Technology Data Exchange (ETDEWEB)

    Perl, S; Kushner, J A; Buchholz, B A; Meeker, A K; Stein, G M; Hsieh, M; Kirby, M; Pechhold, S; Liu, E H; Harlan, D M; Tisdale, J F

    2010-03-15

    Diabetes mellitus results from an absolute or relative deficiency of insulin producing pancreatic beta-cells. The adult human beta-cell's turnover rate remains unknown. We employed novel techniques to examine adult human islet beta-cell turnover and longevity in vivo. Subjects enrolled in NIH clinical trials received thymidine analogues [iododeoxyuridine (IdU) or bromodeoxyuridine (BrdU)] 8-days to 4-years prior to death. Archival autopsy samples from ten patients (aged 17-74 years) were employed to assess beta-cell turnover by scoring nuclear analog labeling within insulin staining cells. Human adult beta-cell longevity was determined by estimating the cells genomic DNA integration of atmospheric carbon-14 ({sup 14}C). DNA was purified from pancreatic islets isolated from cadaveric donors; whole islet prep DNA was obtained from a 15 year old donor, and purified beta-cell DNA was obtained from two donors (age 48 and 80 years). {sup 14}C levels were then determined using accelerator mass spectrometry (AMS). Cellular 'birth date' was determined by comparing the subject's DNA {sup 14}C content relative to a well-established {sup 14}C atmospheric prevalence curve. In the two subjects less than age 20 years, 1-2% of the beta-cell nuclei co-stained for BrdU/IdU. No beta-cell nuclei co-stained in the eight patients more than 30 years old. Consistent with the BrdU/IdU turnover data, beta-cell DNA {sup 14}C content indicated the cells 'birth date' occurred within the subject's first 30 years of life. Under typical circumstances, adult human beta-cells and their cellular precursors are established by young adulthood.

  16. [Peptide phage display in biotechnology and biomedicine].

    Science.gov (United States)

    Kuzmicheva, G A; Belyavskaya, V A

    2016-07-01

    To date peptide phage display is one of the most common combinatorial methods used for identifying specific peptide ligands. Phage display peptide libraries containing billions different clones successfully used for selection of ligands with high affinity and selectivity toward wide range of targets including individual proteins, bacteria, viruses, spores, different kind of cancer cells and variety of nonorganic targets (metals, alloys, semiconductors etc.) Success of using filamentous phage in phage display technologies relays on the robustness of phage particles and a possibility to genetically modify its DNA to construct new phage variants with novel properties. In this review we are discussing characteristics of the most known non-commercial peptide phage display libraries of different formats (landscape libraries in particular) and their successful applications in several fields of biotechnology and biomedicine: discovery of peptides with diagnostic values against different pathogens, discovery and using of peptides recognizing cancer cells, trends in using of phage display technologies in human interactome studies, application of phage display technologies in construction of novel nano materials.

  17. ZnT8-Specific CD4+ T cells display distinct cytokine expression profiles between type 1 diabetes patients and healthy adults.

    Directory of Open Access Journals (Sweden)

    Daisuke Chujo

    Full Text Available Determination of antigen-specific T cell repertoires in human blood has been a challenge. Here, we show a novel integrated approach that permits determination of multiple parameters of antigen-specific T cell repertoires. The approach consists of two assays: the Direct assay and the Cytokine-driven assay. Briefly, human PBMCs are first stimulated with overlapping peptides encoding a given antigen for 48 hours to measure cytokine secretion (Direct assay. Peptide-reactive T cells are further expanded by IL-2 for 5 days; and after overnight starvation, expanded cells are stimulated with the same peptides from the initial culture to analyze cytokine secretion (Cytokine-driven assay. We first applied this integrated approach to determine the type of islet-antigen-specific T cells in healthy adults. Out of ten donors, the Direct assay identified GAD65-specific CD4(+ T cells in three adults and zinc transporter 8 (ZnT8-specific CD4(+ T cells in five adults. The intracytoplasmic cytokine staining assay showed that these islet-antigen-specific CD4(+ T cells belonged to the CD45RO(+ memory compartment. The Cytokine-driven assay further revealed that islet-antigen-specific CD4(+ T cells in healthy adults were capable of secreting various types of cytokines including type 1 and type 2 cytokines as well as IL-10. We next applied our integrated assay to determine whether the type of ZnT8-specific CD4(+ T cells is different between Type 1 diabetes patients and age/gender/HLA-matched healthy adults. We found that ZnT8-specific CD4(+ T cells were skewed towards Th1 cells in T1D patients, while Th2 and IL-10-producing cells were prevalent in healthy adults. In conclusion, the Direct assay and the Cytokine-driven assay complement each other, and the combination of the two assays provides information of antigen-specific T cell repertoires on the breadth, type, and avidity. This strategy is applicable to determine the differences in the quality of antigen-specific T

  18. Display of a thermostable lipase on the surface of a solvent-resistant bacterium, Pseudomonas putida GM730, and its applications in whole-cell biocatalysis

    Directory of Open Access Journals (Sweden)

    Kwon Seok-Joon

    2006-04-01

    Full Text Available Abstract Background Whole-cell biocatalysis in organic solvents has been widely applied to industrial bioprocesses. In two-phase water-solvent processes, substrate conversion yields and volumetric productivities can be limited by the toxicity of solvents to host cells and by the low mass transfer rates of the substrates from the solvent phase to the whole-cell biocatalysts in water. Results To solve the problem of solvent toxicity, we immobilized a thermostable lipase (TliA from Pseudomonas fluorescens on the cell surface of a solvent-resistant bacterium, Pseudomonas putida GM730. Surface immobilization of enzymes eliminates the mass-transfer limitation imposed by the cell wall and membranes. TliA was successfully immobilized on the surface of P. putida cells using the ice-nucleation protein (INP anchoring motif from Pseudomonas syrinage. The surface location was confirmed by flow cytometry, protease accessibility and whole-cell enzyme activity using a membrane-impermeable substrate. Three hundred and fifty units of whole-cell hydrolytic activity per gram dry cell mass were obtained when the enzyme was immobilized with a shorter INP anchoring motif (INPNC. The surface-immobilized TliA retained full enzyme activity in a two-phase water-isooctane reaction system after incubation at 37°C for 12 h, while the activity of the free form enzyme decreased to 65% of its initial value. Whole cells presenting immobilized TliA were shown to catalyze three representative lipase reactions: hydrolysis of olive oil, synthesis of triacylglycerol and chiral resolution. Conclusion In vivo surface immobilization of enzymes on solvent-resistant bacteria was demonstrated, and appears to be useful for a variety of whole-cell bioconversions in the presence of organic solvents.

  19. Reduced Plasmodium Parasite Burden Associates with CD38+ CD4+ T Cells Displaying Cytolytic Potential and Impaired IFN-γ Production

    Science.gov (United States)

    Burel, Julie G.; Apte, Simon H.; Groves, Penny L.; Klein, Kerenaftali; McCarthy, James S.; Doolan, Denise L.

    2016-01-01

    Using a unique resource of samples from a controlled human malaria infection (CHMI) study, we identified a novel population of CD4+ T cells whose frequency in the peripheral blood was inversely correlated with parasite burden following P. falciparum infection. These CD4+ T cells expressed the multifunctional ectoenzyme CD38 and had unique features that distinguished them from other CD4+ T cells. Specifically, their phenotype was associated with proliferation, activation and cytotoxic potential as well as significantly impaired production of IFN-γ and other cytokines and reduced basal levels of activated STAT1. A CD38+ CD4+ T cell population with similar features was identified in healthy uninfected individuals, at lower frequency. CD38+ CD4+ T cells could be generated in vitro from CD38- CD4+ T cells after antigenic or mitogenic stimulation. This is the first report of a population of CD38+ CD4+ T cells with a cytotoxic phenotype and markedly impaired IFN-γ capacity in humans. The expansion of this CD38+ CD4+ T population following infection and its significant association with reduced blood-stage parasite burden is consistent with an important functional role for these cells in protective immunity to malaria in humans. Their ubiquitous presence in humans suggests that they may have a broad role in host-pathogen defense. Trial Registration ClinicalTrials.gov clinical trial numbers ACTRN12612000814875, ACTRN12613000565741 and ACTRN12613001040752 PMID:27662621

  20. Latest development of display technologies

    Science.gov (United States)

    Gao, Hong-Yue; Yao, Qiu-Xiang; Liu, Pan; Zheng, Zhi-Qiang; Liu, Ji-Cheng; Zheng, Hua-Dong; Zeng, Chao; Yu, Ying-Jie; Sun, Tao; Zeng, Zhen-Xiang

    2016-09-01

    In this review we will focus on recent progress in the field of two-dimensional (2D) and three-dimensional (3D) display technologies. We present the current display materials and their applications, including organic light-emitting diodes (OLEDs), flexible OLEDs quantum dot light emitting diodes (QLEDs), active-matrix organic light emitting diodes (AMOLEDs), electronic paper (E-paper), curved displays, stereoscopic 3D displays, volumetric 3D displays, light field 3D displays, and holographic 3D displays. Conventional 2D display devices, such as liquid crystal devices (LCDs) often result in ambiguity in high-dimensional data images because of lacking true depth information. This review thus provides a detailed description of 3D display technologies.

  1. Camphene isolated from essential oil of Piper cernuum (Piperaceae) induces intrinsic apoptosis in melanoma cells and displays antitumor activity in vivo.

    Science.gov (United States)

    Girola, Natalia; Figueiredo, Carlos R; Farias, Camyla F; Azevedo, Ricardo A; Ferreira, Adilson K; Teixeira, Sarah F; Capello, Tabata M; Martins, Euder G A; Matsuo, Alisson L; Travassos, Luiz R; Lago, João H G

    2015-11-27

    Natural monoterpenes were isolated from the essential oil of Piper cernuum Vell. (Piperaceae) leaves. The crude oil and the individual monoterpenes were tested for cytotoxicity in human tumor cell lineages and B16F10-Nex2 murine melanoma cells. In the present work we demonstrate the activity of camphene against different cancer cells, with its mechanism of action being investigated in vitro and in vivo in murine melanoma. Camphene induced apoptosis by the intrinsic pathway in melanoma cells mainly by causing endoplasmic reticulum (ER) stress, with release of Ca(2+) together with HmgB1 and calreticulin, loss of mitochondrial membrane potential and up regulation of caspase-3 activity. Importantly, camphene exerted antitumor activity in vivo by inhibiting subcutaneous tumor growth of highly aggressive melanoma cells in a syngeneic model, suggesting a promising role of this compound in cancer therapy.

  2. Breast Cancer Cells in Three-dimensional Culture Display an Enhanced Radioresponse after Coordinate Targeting of Integrin ?5?1 and Fibronectin

    Energy Technology Data Exchange (ETDEWEB)

    Nam, Jin-Min; Onodera, Yasuhito; Bissell, Mina J; Park, Catherine C

    2010-04-07

    Tactics to selectively enhance cancer radioresponse are of great interest. Cancer cells actively elaborate and remodel their extracellular matrix (ECM) to aid in survival and progression. Previous work has shown that {beta}1-integrin inhibitory antibodies can enhance the growth-inhibitory and apoptotic responses of human breast cancer cell lines to ionizing radiation, either when cells are cultured in three-dimensional laminin-rich ECM (3D lrECM) or grown as xenografts in mice. Here, we show that a specific {alpha} heterodimer of {beta}1-integrin preferentially mediates a prosurvival signal in human breast cancer cells that can be specifically targeted for therapy. 3D lrECM culture conditions were used to compare {alpha}-integrin heterodimer expression in malignant and nonmalignant cell lines. Under these conditions, we found that expression of {alpha}5{beta}1-integrin was upregulated in malignant cells compared with nonmalignant breast cells. Similarly, we found that normal and oncofetal splice variants of fibronectin, the primary ECM ligand of {alpha}5{beta}1-integrin, were also strikingly upregulated in malignant cell lines compared with nonmalignant acini. Cell treatment with a peptide that disrupts the interactions of {alpha}5{beta}1-integrin with fibronectin promoted apoptosis in malignant cells and further heightened the apoptotic effects of radiation. In support of these results, an analysis of gene expression array data from breast cancer patients revealed an association of high levels of {alpha}5-integrin expression with decreased survival. Our findings offer preclinical validation of fibronectin and {alpha}5{beta}1-integrin as targets for breast cancer therapy.

  3. Synthesis of diisooctyl adipate catalyzed by lipase-displaying Pichia pastoris whole-cell biocatalysts%酵母表面展示脂肪酶合成己二酸二异辛酯

    Institute of Scientific and Technical Information of China (English)

    张娜; 金子; 林影; 郑穗平; 韩双艳

    2013-01-01

    An enzyme-displaying yeast as a whole-cell biocatalyst is an alternative to immobihzed enzyme,due to its low-cost preparation and simple recycle course.Here,lipase-displaying Pichia pastoris whole-cell was used as a biocatalyst to synthesize diisooctyl adipate in the non-aqueous system.The maximum productivity of diisooctyl adipate was obtained as 85.0% in a 10 mL reaction system.The yield could be reached as high as 97.8% when the reaction system was scaled up to 200 mL.The purity obtained is 98.2% after vacuum distillation.Thus,the lipase-displaying P.pastoris whole-cell biocatalyst was promising in commercial application for diisooctyl adipate synthesis in non-aqueous phase.%展示酶的酵母细胞既具有固定化酶的优点,又有制备简单、成本较低的特点.采用表面展示南极假丝酵母脂肪酶B (Candida antarctica lipase B,CALB)的毕赤酵母细胞催化合成己二酸二异辛酯(Diisooctyl adipate,DIOA),对该反应体系进行优化,并实现了初步工艺放大制备.经条件优化后,在10mL反应体系中,DIOA的产率可达85.0%.该工艺放大到200mL反应体系时,DIOA产率可达97.8%.经减压蒸馏,DIOA纯度可达到98.2%.该酵母表面展示脂肪酶在合成绿色润滑油己二酸二异辛酯中具有良好应用前景.

  4. Generation of a phage-display library of single-domain camelid VH H antibodies directed against Chlamydomonas reinhardtii antigens, and characterization of VH Hs binding cell-surface antigens.

    Science.gov (United States)

    Jiang, Wenzhi; Rosenberg, Julian N; Wauchope, Akelia D; Tremblay, Jacqueline M; Shoemaker, Charles B; Weeks, Donald P; Oyler, George A

    2013-11-01

    Single-domain antibodies (sdAbs) are powerful tools for the detection, quantification, purification and subcellular localization of proteins of interest in biological research. We have generated camelid (Lama pacos) heavy chain-only variable VH domain (VH H) libraries against antigens in total cell lysates from Chlamydomonas reinhardtii. The sdAbs in the sera from immunized animals and VH H antibody domains isolated from the library show specificity to C. reinhardtii and lack of reactivity to antigens from four other algae: Chlorella variabilis, Coccomyxa subellipsoidea, Nannochloropsis oceanica and Thalassiosira pseudonana. Antibodies were produced against a diverse representation of antigens as evidenced by sera ELISA and protein-blot analyses. A phage-display library consisting of the VH H region contained at least 10(6) individual transformants, and thus should represent a wide range of C. reinhardtii antigens. The utility of the phage library was demonstrated by using live C. reinhardtii cells to pan for VH H clones with specific recognition of cell-surface epitopes. The lead candidate VH H clones (designated B11 and H10) bound to C. reinhardtii with EC50 values ≤ 0.5 nm. Treatment of cells with VH H B11 fused to the mCherry or green fluorescent proteins allowed brilliant and specific staining of the C. reinhardtii cell wall and analysis of cell-wall genesis during cell division. Such high-complexity VH H antibody libraries for algae will be valuable tools for algal researchers and biotechnologists.

  5. Intrinsic and extrinsic mechanisms contribute to maintain the JAK/STAT pathway aberrantly activated in T-type large granular lymphocyte leukemia.

    Science.gov (United States)

    Teramo, Antonella; Gattazzo, Cristina; Passeri, Francesca; Lico, Albana; Tasca, Giulia; Cabrelle, Anna; Martini, Veronica; Frezzato, Federica; Trimarco, Valentina; Ave, Elisa; Boscaro, Elisa; Piazza, Francesco; Facco, Monica; Trentin, Livio; Semenzato, Gianpietro; Zambello, Renato

    2013-05-09

    The JAK/STAT pathway is altered in T-cell large granular lymphocytic leukemia. In all patients, leukemic LGLs display upregulation of phosphorylated STAT3 (P-STAT3) that activates expression of many antiapoptotic genes. To investigate the mechanisms maintaining STAT3 aberrantly phosphorylated using transcriptional protein and functional assays, we analyzed interleukin (IL)-6 and suppressor of cytokine signaling-3 (SOCS3), 2 key factors of the JAK/STAT pathway that induce and inhibit STAT3 activation, respectively. We showed that IL-6 was highly expressed and released by the patients' peripheral blood LGL-depleted population, accounting for a trans-signaling process. By neutralizing IL-6 or its specific receptor with specific antibodies, a significant reduction of P-STAT3 levels and, consequently, LGL survival was demonstrated. In addition, we found that SOCS3 was down-modulated in LGL and unresponsive to IL-6 stimulation. By treating neoplastic LGLs with a demethylating agent, IL-6-mediated SOCS3 expression was restored with consequent P-STAT3 and myeloid cell leukemia-1 down-modulation. Methylation in the SOCS3 promoter was not detectable, suggesting that an epigenetic inhibition mechanism occurs at a different site. Our data indicate that loss of the inhibitor SOCS3 cooperates with IL-6 to maintain JAK/STAT pathway activation, thus contributing to leukemic LGL survival, and suggest a role of demethylating agents in the treatment of this disorder.

  6. Displaying the researched body

    DEFF Research Database (Denmark)

    Whiteley, Louise; Tybjerg, Karin; Pedersen, Bente Vinge

    2017-01-01

    In Heirloom, artist Gina Czarnecki and scientist John Hunt grow portraits of the artist’s daughters from their own cells, onto glass casts of their faces. This required the development of novel scien- tific techniques to allow the growth of human cells in a gallery. Heirloom was exhibited at Medi...

  7. Doxorubicin-resistant LoVo adenocarcinoma cells display resistance to apoptosis induction by some but not all inhibitors of ser/thr phosphatases 1 and 2A.

    Science.gov (United States)

    Sieder, S; Richter, E; Becker, K; Heins, R; Steinfelder, H J

    1999-06-15

    LoVo adenocarcinoma cells are fairly sensitive to cytostatic drugs, e.g. doxorubicin, but can develop drug resistance by expression of a P-glycoprotein-mediated MDR1 phenotype. LoVo cells respond with apoptosis to nanomolar concentrations of okadaic acid and micromolar concentrations of cantharidic acid. Interestingly, LoVoDx cells which had become about 10-fold less sensitive to doxorubicin by incubation in increasing concentrations of this cytostatic drug were also less sensitive to the toxicity of okadaic acid. Resistance to both agents was lost or significantly reduced by incubation in drug-free medium for about 4 months. On the other hand, LoVoDx cells did not lose responsiveness to the structurally different phosphatase inhibitor cantharidic acid but were about twofold more sensitive to the cytotoxic effect of this agent. Thus, MDR expression protects LoVo cells from the toxicity of phosphatase inhibitors that presumably are substrates of the P-glycoprotein, e.g. okadaic acid and its derivatives but not cantharidic acid, despite the fact that both agents are potent inducers of apoptotic cell death via ser/thr phosphatase inhibition.

  8. A novel anti-alpha-fetoprotein single-chain variable fragment displays anti-tumor effects in HepG2 cells as a single agent or in combination with paclitaxel.

    Science.gov (United States)

    Ji, Xiaonan; Shen, Yanli; Sun, Hao; Gao, Xiangdong

    2016-08-01

    Human hepatocellular carcinoma (HCC) has a high rate of tumor recurrence and metastasis, resulting in shortened survival time. The function of alpha-fetoprotein (AFP) as a regulatory factor in the growth of HCC cells has been well defined. The aim of this study was to investigate the use of a novel AFP-specific single-chain variable fragment that blocked AFP and inhibited HCC cell growth. The results indicated that the anti-AFP single-chain variable fragment (scFv) induced growth inhibition of AFP-expressing HCC cell lines in vitro through induction of G1 cell cycle arrest and apoptosis. The mechanism of apoptosis probably involved with blocking AFP internalization and regulation of the PTEN/PI3K/Akt signaling network. Moreover, the anti-AFP-scFv also effectively sensitized the HepG2 cells to paclitaxel (PTX) at a lower concentration. The combination effect of PTX and anti-AFP-scFv displayed a synergistic effect on HepG2 cells both in vitro and in vivo. Our results demonstrated that targeting AFP by specific antibodies has potential immunotherapeutic efficacy in human HCC.

  9. Ras-mutant cancer cells display B-Raf binding to Ras that activates extracellular signal-regulated kinase and is inhibited by protein kinase A phosphorylation.

    Science.gov (United States)

    Li, Yanping; Takahashi, Maho; Stork, Philip J S

    2013-09-20

    The small G protein Ras regulates proliferation through activation of the mitogen-activated protein (MAP) kinase (ERK) cascade. The first step of Ras-dependent activation of ERK signaling is Ras binding to members of the Raf family of MAP kinase kinase kinases, C-Raf and B-Raf. Recently, it has been reported that in melanoma cells harboring oncogenic Ras mutations, B-Raf does not bind to Ras and does not contribute to basal ERK activation. For other types of Ras-mutant tumors, the relative contributions of C-Raf and B-Raf are not known. We examined non-melanoma cancer cell lines containing oncogenic Ras mutations and express both C-Raf and B-Raf isoforms, including the lung cancer cell line H1299 cells. Both B-Raf and C-Raf were constitutively bound to oncogenic Ras and contributed to Ras-dependent ERK activation. Ras binding to B-Raf and C-Raf were both subject to inhibition by the cAMP-dependent protein kinase PKA. cAMP inhibited the growth of H1299 cells and Ras-dependent ERK activation via PKA. PKA inhibited the binding of Ras to both C-Raf and B-Raf through phosphorylations of C-Raf at Ser-259 and B-Raf at Ser-365, respectively. These studies demonstrate that in non-melanocytic Ras-mutant cancer cells, Ras signaling to B-Raf is a significant contributor to ERK activation and that the B-Raf pathway, like that of C-Raf, is a target for inhibition by PKA. We suggest that cAMP and hormones coupled to cAMP may prove useful in dampening the effects of oncogenic Ras in non-melanocytic cancer cells through PKA-dependent actions on B-Raf as well as C-Raf.

  10. Human bone marrow mesenchymal stem cells display anti-cancer activity in SCID mice bearing disseminated non-Hodgkin's lymphoma xenografts.

    Directory of Open Access Journals (Sweden)

    Paola Secchiero

    Full Text Available BACKGROUND: Although multimodality treatment can induce high rate of remission in many subtypes of non-Hodgkin's lymphoma (NHL, significant proportions of patients relapse with incurable disease. The effect of human bone marrow (BM mesenchymal stem cells (MSC on tumor cell growth is controversial, and no specific information is available on the effect of BM-MSC on NHL. METHODOLOGY/PRINCIPAL FINDINGS: The effect of BM-MSC was analyzed in two in vivo models of disseminated non-Hodgkin's lymphomas with an indolent (EBV(- Burkitt-type BJAB, median survival = 46 days and an aggressive (EBV(+ B lymphoblastoid SKW6.4, median survival = 27 days behavior in nude-SCID mice. Intra-peritoneal (i.p. injection of MSC (4 days after i.p. injection of lymphoma cells significantly increased the overall survival at an optimal MSC:lymphoma ratio of 1:10 in both xenograft models (BJAB+MSC, median survival = 58.5 days; SKW6.4+MSC, median survival = 40 days. Upon MSC injection, i.p. tumor masses developed more slowly and, at the histopathological observation, exhibited a massive stromal infiltration coupled to extensive intra-tumor necrosis. In in vitro experiments, we found that: i MSC/lymphoma co-cultures modestly affected lymphoma cell survival and were characterized by increased release of pro-angiogenic cytokines with respect to the MSC, or lymphoma, cultures; ii MSC induce the migration of endothelial cells in transwell assays, but promoted endothelial cell apoptosis in direct MSC/endothelial cell co-cultures. CONCLUSIONS/SIGNIFICANCE: Our data demonstrate that BM-MSC exhibit anti-lymphoma activity in two distinct xenograft SCID mouse models of disseminated NHL.

  11. Rare Circulating Cells in Familial Waldenstrom Macroglobulinemia Displaying the MYD88 L265P Mutation Are Enriched by Epstein-Barr Virus Immortalization.

    Directory of Open Access Journals (Sweden)

    Maroulio Pertesi

    Full Text Available The MYD88 L265P is a recurrent somatic mutation in neoplastic cells from patients with Waldenström Macroglobulinemia (WM. We identified the MYD88 L265P mutation in three individuals from unrelated families, but its presence did not explain the disease segregation within these WM pedigrees. We observed the mutation in these three individuals at high allele fractions in DNA extracted from EBV-immortalized Lymphoblastoid cell lines established from peripheral blood (LCL, but at much lower allele fractions in DNA extracted directly from peripheral blood, suggesting that this mutation is present in a clonal cell subpopulation rather than of germ-line origin. Furthermore, we observed that the MYD88 L265P mutation is enriched in WM families, detected in 40.5% of patients with familial WM or MGUS (10/22 WM, 5/15 MGUS, compared to 3.5% of patients with familial MM or MGUS (0/72 MM, 4/41 MGUS (p = 10-7. The mutant allele frequency increased with passages in vitro after immortalization with Epstein-Barr virus (EBV consistent with the MYD88 L265P described gain-of-function proposed for this mutation. The MYD88 L265P mutation appears to be frequently present in circulating cells in patients with WM, and MGUS, and these cells are amenable to immortalization by EBV.

  12. Rare Circulating Cells in Familial Waldenström Macroglobulinemia Displaying the MYD88 L265P Mutation Are Enriched by Epstein-Barr Virus Immortalization.

    Science.gov (United States)

    Pertesi, Maroulio; Galia, Perrine; Nazaret, Nicolas; Vallée, Maxime; Garderet, Laurent; Leleu, Xavier; Avet-Loiseau, Hervé; Foll, Matthieu; Byrnes, Graham; Lachuer, Joel; McKay, James D; Dumontet, Charles

    2015-01-01

    The MYD88 L265P is a recurrent somatic mutation in neoplastic cells from patients with Waldenström Macroglobulinemia (WM). We identified the MYD88 L265P mutation in three individuals from unrelated families, but its presence did not explain the disease segregation within these WM pedigrees. We observed the mutation in these three individuals at high allele fractions in DNA extracted from EBV-immortalized Lymphoblastoid cell lines established from peripheral blood (LCL), but at much lower allele fractions in DNA extracted directly from peripheral blood, suggesting that this mutation is present in a clonal cell subpopulation rather than of germ-line origin. Furthermore, we observed that the MYD88 L265P mutation is enriched in WM families, detected in 40.5% of patients with familial WM or MGUS (10/22 WM, 5/15 MGUS), compared to 3.5% of patients with familial MM or MGUS (0/72 MM, 4/41 MGUS) (p = 10-7). The mutant allele frequency increased with passages in vitro after immortalization with Epstein-Barr virus (EBV) consistent with the MYD88 L265P described gain-of-function proposed for this mutation. The MYD88 L265P mutation appears to be frequently present in circulating cells in patients with WM, and MGUS, and these cells are amenable to immortalization by EBV.

  13. Ovarian teratoma displaying a wide variety of tissue components in a broiler chicken (Gallus Domesticus: morphological heterogeneity of pluripotential germ cell during tumorigenesis

    Directory of Open Access Journals (Sweden)

    S. Ohfuji

    2016-05-01

    Full Text Available Spontaneous ovarian teratoma was found in a seven-week-old female Chunky broiler chicken that was slaughtered for food. On post-mortem inspection, a spherical tumor mass attaching to a juvenile ovary was found in the abdominal cavity. Histopathologically, the tumor was comprised of immature mesenchymal stroma and a variety of mature tissue elements of mesodermal and ectodermal origin. In addition, there were multiple indistinguishable tissue elements, which showed no malignant cytological features but were unidentifiable as to corresponding embryological layer of origin. These heterogeneous teratoma tissues consisted of a variety of glandular, cystic, duct-like, and tubular structures, some of which exhibited a lining by a mixture of both keratinizing/non-keratinizing stratified squamous epithelial cells and cuboidal/columnar epithelial cells. The ovarian tetatoma was considered a benign and congenital one. The highly diverse differentiation of the teratoma might have manifested a morphological aspect of intrinsic character of the pluripotential germ cells during tumorigenesis.

  14. PrP{sup C} displays an essential protective role from oxidative stress in an astrocyte cell line derived from PrP{sup C} knockout mice

    Energy Technology Data Exchange (ETDEWEB)

    Bertuchi, Fernanda R. [Center for Natural Sciences and Humanities, Federal University of ABC - UFABC, Avenida dos Estados, 5001, Bloco B, 09210-170, Santo Andre, SP (Brazil); Bourgeon, Dominique M.G.; Landemberger, Michele C.; Martins, Vilma R. [International Center for Research and Education, A.C. Camargo Hospital, Rua Tagua 440, 01505-010 Sao Paulo, SP (Brazil); Cerchiaro, Giselle, E-mail: giselle.cerchiaro@ufabc.edu.br [Center for Natural Sciences and Humanities, Federal University of ABC - UFABC, Avenida dos Estados, 5001, Bloco B, 09210-170, Santo Andre, SP (Brazil)

    2012-02-03

    Highlights: Black-Right-Pointing-Pointer PrP{sup C} in solution acts as a radical scavenger. Black-Right-Pointing-Pointer PrP{sup C} reduces hydrogen peroxide toxicity in astrocytes. Black-Right-Pointing-Pointer Increase in ROS disrupted the cell cycle in the PrP{sup C}-knockout astrocytes. Black-Right-Pointing-Pointer PrP{sup C} prevents the cell death independently of an SOD-like activity. -- Abstract: The PrP{sup C} protein, which is especially present in the cellular membrane of nervous system cells, has been extensively studied for its controversial antioxidant activity. In this study, we elucidated the free radical scavenger activity of purified murine PrP{sup C} in solution and its participation as a cell protector in astrocytes that were subjected to treatment with an oxidant. In vitro and using an EPR spin-trapping technique, we observed that PrP{sup C} decreased the oxidation of the DMPO trap in a Fenton reaction system (Cu{sup 2+}/ascorbate/H{sub 2}O{sub 2}), which was demonstrated by approximately 70% less DMPO/OH{sup {center_dot}}. In cultured PrP{sup C}-knockout astrocytes from mice, the absence of PrP{sup C} caused an increase in intracellular ROS (reactive oxygen species) generation during the first 3 h of H{sub 2}O{sub 2} treatment. This rapid increase in ROS disrupted the cell cycle in the PrP{sup C}-knockout astrocytes, which increased the population of cells in the sub-G1 phase when compared with cultured wild-type astrocytes. We conclude that PrP{sup C} in solution acts as a radical scavenger, and in astrocytes, it is essential for protection from oxidative stress caused by an external chemical agent, which is a likely condition in human neurodegenerative CNS disorders and pathological conditions such as ischemia.

  15. Endoplasmic reticulum-directed recombinant mRNA displays subcellular localization equal to endogenous mRNA during transient expression in CHO cells

    DEFF Research Database (Denmark)

    Beuchert Kallehauge, Thomas; Kol, Stefan; Andersen, Mikael Rørdam;

    2016-01-01

    When expressing pharmaceutical recombinant proteins in mammalian cells, the protein is commonly directed through the secretory pathway, in a signal peptide-dependent manner, to acquire specific post-translational modifications and to facilitate secretion into the culture medium. One key premise...... for this is the direction of the mRNA encoding the recombinant protein to the surface of the endoplasmic reticulum (ER) for subsequent protein translocation into the secretory pathway. To evaluate the efficiency of this process in Chinese hamster ovary (CHO) cells, the subcellular localization of recombinant mRNA encoding...

  16. Dengue virus-reactive CD8+ T cells display quantitative and qualitative differences in their response to variant epitopes of heterologous viral serotypes.

    Science.gov (United States)

    Bashyam, Hema S; Green, Sharone; Rothman, Alan L

    2006-03-01

    Reactivation of serotype cross-reactive CD8+ memory T lymphocytes is thought to contribute to the immunopathogenesis of dengue disease during secondary infection by a heterologous serotype. Using cytokine flow cytometry, we have defined four novel HLA-A*02-restricted dengue viral epitopes recognized by up to 1.5% of circulating CD8+ T cells in four donors after primary vaccination. All four donors had the highest cytokine response to the epitope NS4b 2353. We also studied the effect of sequence differences in heterologous dengue serotypes on dengue-reactive CD8+ memory T cell cytokine and proliferative responses. The D3 variant of a different NS4b epitope 2423 and the D2 variant of the NS4a epitope 2148 induced the largest cytokine response, compared with their respective heterologous sequences in all donors regardless of the primary vaccination serotype. Stimulation with variant peptides also altered the relative frequencies of the various subsets of cells that expressed IFN-gamma, TNF-alpha, MIP-1beta, and combinations of these cytokines. These results indicate that the prior infection history of the individual as well as the serotypes of the primary and heterologous secondary viruses influence the nature of the secondary response. These differences in the effector functions of serotype cross-reactive memory T cells induced by heterologous variant epitopes, which are both quantitative and qualitative, may contribute to the clinical outcome of secondary dengue infection.

  17. Colorimetry for CRT displays.

    Science.gov (United States)

    Golz, Jürgen; MacLeod, Donald I A

    2003-05-01

    We analyze the sources of error in specifying color in CRT displays. These include errors inherent in the use of the color matching functions of the CIE 1931 standard observer when only colorimetric, not radiometric, calibrations are available. We provide transformation coefficients that prove to correct the deficiencies of this observer very well. We consider four different candidate sets of cone sensitivities. Some of these differ substantially; variation among candidate cone sensitivities exceeds the variation among phosphors. Finally, the effects of the recognized forms of observer variation on the visual responses (cone excitations or cone contrasts) generated by CRT stimuli are investigated and quantitatively specified. Cone pigment polymorphism gives rise to variation of a few per cent in relative excitation by the different phosphors--a variation larger than the errors ensuing from the adoption of the CIE standard observer, though smaller than the differences between some candidate cone sensitivities. Macular pigmentation has a larger influence, affecting mainly responses to the blue phosphor. The estimated combined effect of all sources of observer variation is comparable in magnitude with the largest differences between competing cone sensitivity estimates but is not enough to disrupt very seriously the relation between the L and M cone weights and the isoluminance settings of individual observers. It is also comparable with typical instrumental colorimetric errors, but we discuss these only briefly.

  18. LHCb Event display

    CERN Document Server

    Trisovic, Ana

    2014-01-01

    The LHCb Event Display was made for educational purposes at the European Organization for Nuclear Research, CERN in Geneva, Switzerland. The project was implemented as a stand-alone application using C++ and ROOT, a framework developed by CERN for data analysis. This paper outlines the development and architecture of the application in detail, as well as the motivation for the development and the goals of the exercise. The application focuses on the visualization of events recorded by the LHCb detector, where an event represents a set of charged particle tracks in one proton-proton collision. Every particle track is coloured by its type and can be selected to see its essential information such as mass and momentum. The application allows students to save this information and calculate the invariant mass for any pair of particles. Furthermore, the students can use additional calculating tools in the application and build up a histogram of these invariant masses. The goal for the students is to find a $D^0$ par...

  19. Toxicological Responses of Environmental Mixtures: Environmental Metals Mixtures Display Synergistic Induction of Metal-Responsive and Oxidative Stress Genes in Placental Cells

    Science.gov (United States)

    Adebambo, Oluwadamilare A.; Ray, Paul D.; Shea, Damian; Fry, Rebecca C.

    2016-01-01

    Exposure to elevated levels of the toxic metals inorganic arsenic (iAs) and cadmium (Cd) represents a major global health problem. These metals often occur as mixtures in the environment, creating the potential for interactive or synergistic biological effects different from those observed in single exposure conditions. In the present study, environmental mixtures collected from two waste sites in China and comparable mixtures prepared in the laboratory were tested for toxicogenomic response in placental JEG-3 cells. These cells serve as a model for evaluating cellular responses to exposures during pregnancy. One of the mixtures was predominated by iAs and one by Cd. Six gene biomarkers were measured in order to evaluate the effects from the metals mixtures using dose and time-course experiments including: heme oxygenase 1 (HO-1) and metallothionein isoforms (MT1A, MT1F and MT1G) previously shown to be preferentially induced by exposure to either iAs or Cd, and metal transporter genes aquaporin-9 (AQP9) and ATPase, Cu2+ transporting, beta polypeptide (ATP7B). There was a significant increase in the mRNA expression levels of ATP7B, HO-1, MT1A, MT1F, and MT1G in mixture-treated cells compared to the iAs or Cd only-treated cells. Notably, the genomic responses were observed at concentrations significantly lower than levels found at the environmental collection sites. These data demonstrate that metal mixtures increase the expression of gene biomarkers in placental JEG-3 cells in a synergistic manner. Taken together, the data suggest that toxic metals that co-occur may induce detrimental health effects that are currently underestimated when analyzed as single metals. PMID:26472158

  20. Toxicological responses of environmental mixtures: Environmental metal mixtures display synergistic induction of metal-responsive and oxidative stress genes in placental cells.

    Science.gov (United States)

    Adebambo, Oluwadamilare A; Ray, Paul D; Shea, Damian; Fry, Rebecca C

    2015-12-15

    Exposure to elevated levels of the toxic metals inorganic arsenic (iAs) and cadmium (Cd) represents a major global health problem. These metals often occur as mixtures in the environment, creating the potential for interactive or synergistic biological effects different from those observed in single exposure conditions. In the present study, environmental mixtures collected from two waste sites in China and comparable mixtures prepared in the laboratory were tested for toxicogenomic response in placental JEG-3 cells. These cells serve as a model for evaluating cellular responses to exposures during pregnancy. One of the mixtures was predominated by iAs and one by Cd. Six gene biomarkers were measured in order to evaluate the effects from the metal mixtures using dose and time-course experiments including: heme oxygenase 1 (HO-1) and metallothionein isoforms (MT1A, MT1F and MT1G) previously shown to be preferentially induced by exposure to either iAs or Cd, and metal transporter genes aquaporin-9 (AQP9) and ATPase, Cu(2+) transporting, beta polypeptide (ATP7B). There was a significant increase in the mRNA expression levels of ATP7B, HO-1, MT1A, MT1F, and MT1G in mixture-treated cells compared to the iAs or Cd only-treated cells. Notably, the genomic responses were observed at concentrations significantly lower than levels found at the environmental collection sites. These data demonstrate that metal mixtures increase the expression of gene biomarkers in placental JEG-3 cells in a synergistic manner. Taken together, the data suggest that toxic metals that co-occur may induce detrimental health effects that are currently underestimated when analyzed as single metals.

  1. Data Display in Qualitative Research

    Directory of Open Access Journals (Sweden)

    Susana Verdinelli PsyD

    2013-02-01

    Full Text Available Visual displays help in the presentation of inferences and conclusions and represent ways of organizing, summarizing, simplifying, or transforming data. Data displays such as matrices and networks are often utilized to enhance data analysis and are more commonly seen in quantitative than in qualitative studies. This study reviewed the data displays used by three prestigious qualitative research journals within a period of three years. The findings include the types of displays used in these qualitative journals, the frequency of use, and the purposes for using visual displays as opposed to presenting data in text.

  2. Unique interactive projection display screen

    Energy Technology Data Exchange (ETDEWEB)

    Veligdan, J.T.

    1997-11-01

    Projection systems continue to be the best method to produce large (1 meter and larger) displays. However, in order to produce a large display, considerable volume is typically required. The Polyplanar Optic Display (POD) is a novel type of projection display screen, which for the first time, makes it possible to produce a large projection system that is self-contained and only inches thick. In addition, this display screen is matte black in appearance allowing it to be used in high ambient light conditions. This screen is also interactive and can be remotely controlled via an infrared optical pointer resulting in mouse-like control of the display. Furthermore, this display need not be flat since it can be made curved to wrap around a viewer as well as being flexible.

  3. A major population of mucosal memory CD4+ T cells, coexpressing IL-18Rα and DR3, display innate lymphocyte functionality

    DEFF Research Database (Denmark)

    Holmkvist, P.; Roepstorff, K.; Uronen-Hansson, H.

    2015-01-01

    , IL-5, IL-13, granulocyte-macrophage colony-stimulating factor (GM-CSF), and IL-22 in the presence of IL-12/ IL-18. TL1a synergized with IL-15 to enhance this response, while suppressing IL-15-induced IL-10 production. TL1a- and IL-15-mediated cytokine induction required the presence of IL-18, whereas...... induction of IL-5, IL-13, GM-CSF, and IL-22 was IL-12 independent. IL-18Rα+DR3+CD4+ T cells with similar functionality were present in human skin, nasal polyps, and, in particular, the intestine, where in chronic inflammation they localized with IL-18-producing cells in lymphoid aggregates. Collectively...

  4. HY-Specific Induced Regulatory T Cells Display High Specificity and Efficacy in the Prevention of Acute Graft-versus-Host Disease

    OpenAIRE

    Li, Jun; Heinrichs, Jessica; Haarberg, Kelley; Semple, Kenrick; Veerapathran, Anandharaman; Liu, Chen; Anasetti, Claudio; Yu, Xue-Zhong

    2015-01-01

    Naturally derived regulatory T cells (nTregs) may prevent graft-versus-host disease (GVHD) while preserving graft-versus-leukemia (GVL) activity. However, clinical application of nTregs has been severely hampered by their scarce availability and non-selectivity. To overcome these limitations, we took alternative approaches to generate Ag-specific induced Tregs (iTregs) and tested their efficacy and selectivity in the prevention of GVHD in pre-clinical models of bone marrow transplantation (BM...

  5. Resistin and interleukin-6 exhibit racially-disparate expression in breast cancer patients, display molecular association and promote growth and aggressiveness of tumor cells through STAT3 activation.

    Science.gov (United States)

    Deshmukh, Sachin K; Srivastava, Sanjeev K; Bhardwaj, Arun; Singh, Ajay P; Tyagi, Nikhil; Marimuthu, Saravanakumar; Dyess, Donna L; Dal Zotto, Valeria; Carter, James E; Singh, Seema

    2015-05-10

    African-American (AA) women with breast cancer (BC) are diagnosed with more aggressive disease, have higher risk of recurrence and poorer prognosis as compared to Caucasian American (CA) women. Therefore, it is imperative to define the factors associated with such disparities to reduce the unequal burden of cancer. Emerging data suggest that inherent differences exist in the tumor microenvironment of AA and CA BC patients, however, its molecular bases and functional impact have remained poorly understood. Here, we conducted cytokine profiling in serum samples from AA and CA BC patients and identified resistin and IL-6 to be the most differentially-expressed cytokines with relative greater expression in AA patients. Resistin and IL-6 exhibited positive correlation in serum levels and treatment of BC cells with resistin led to enhanced production of IL-6. Moreover, resistin also enhanced the expression and phosphorylation of STAT3, and treatment of BC cells with IL-6-neutralizing antibody prior to resistin stimulation abolished STAT3 phosphorylation. In addition, resistin promoted growth and aggressiveness of BC cells, and these effects were mediated through STAT3 activation. Together, these findings suggest a crucial role of resistin, IL-6 and STAT3 in BC racial disparity.

  6. A novel shogaol analog suppresses cancer cell invasion and inflammation, and displays cytoprotective effects through modulation of NF-κB and Nrf2-Keap1 signaling pathways

    Energy Technology Data Exchange (ETDEWEB)

    Gan, Fei-Fei; Ling, Hui; Ang, Xiaohui; Reddy, Shridhivya A.; Lee, Stephanie S-H.; Yang, Hong; Tan, Sock-Hoon [Department of Pharmacy, Faculty of Science, National University of Singapore (Singapore); Hayes, John D. [Jacqui Wood Cancer Centre, Division of Cancer Research, Ninewells Hospital and Medical School, University of Dundee, Dundee, Scotland (United Kingdom); Chui, Wai-Keung [Department of Pharmacy, Faculty of Science, National University of Singapore (Singapore); Chew, Eng-Hui, E-mail: phaceh@nus.edu.sg [Department of Pharmacy, Faculty of Science, National University of Singapore (Singapore)

    2013-11-01

    Natural compounds containing vanilloid and Michael acceptor moieties appear to possess anti-cancer and chemopreventive properties. The ginger constituent shogaol represents one such compound. In this study, the anti-cancer potential of a synthetic novel shogaol analog 3-phenyl-3-shogaol (3-Ph-3-SG) was assessed by evaluating its effects on signaling pathways. At non-toxic concentrations, 3-Ph-3-SG suppressed cancer cell invasion in MDA-MB-231 and MCF-7 breast carcinoma cells through inhibition of PMA-activated MMP-9 expression. At similar concentrations, 3-Ph-3-SG reduced expression of the inflammatory mediators nitric oxide (NO), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and prostanglandin-E{sub 2} (PGE{sub 2}) in RAW 264.7 macrophage-like cells. Inhibition of cancer cell invasion and inflammation by 3-Ph-3-SG were mediated through suppression of the nuclear factor-kappaB (NF-κB) signaling pathway. The 3-Ph-3-SG also demonstrated cytoprotective effects by inducing the antioxidant response element (ARE)-driven genes NAD(P)H quinone oxidoreductase-1 (NQO1) and heme oxygenase-1 (HO-1). Cytoprotection by 3-Ph-3-SG was achieved at least partly through modification of cysteine residues in the E3 ubiquitin ligase substrate adaptor Kelch-like ECH-associated protein 1 (Keap1), which resulted in accumulation of transcription factor NF-E2 p45-related factor 2 (Nrf2). The activities of 3-Ph-3-SG were comparable to those of 6-shogaol, the most abundant naturally-occurring shogaol, and stronger than those of 4-hydroxyl-null deshydroxy-3-phenyl-3-shogaol, which attested the importance of the 4-hydroxy substituent in the vanilloid moiety for bioactivity. In summary, 3-Ph-3-SG is shown to possess activities that modulate stress-associated pathways relevant to multiple steps in carcinogenesis. Therefore, it warrants further investigation of this compound as a promising candidate for use in chemotherapeutic and chemopreventive strategies. - Highlights:

  7. Baculovirus display of functional antibody Fab fragments.

    Science.gov (United States)

    Takada, Shinya; Ogawa, Takafumi; Matsui, Kazusa; Suzuki, Tasuku; Katsuda, Tomohisa; Yamaji, Hideki

    2015-08-01

    The generation of a recombinant baculovirus that displays antibody Fab fragments on the surface was investigated. A recombinant baculovirus was engineered so that the heavy chain (Hc; Fd fragment) of a mouse Fab fragment was expressed as a fusion to the N-terminus of baculovirus gp64, while the light chain of the Fab fragment was simultaneously expressed as a secretory protein. Following infection of Sf9 insect cells with the recombinant baculovirus, the culture supernatant was analyzed by enzyme-linked immunosorbent assay using antigen-coated microplates and either an anti-mouse IgG or an anti-gp64 antibody. A relatively strong signal was obtained in each case, showing antigen-binding activity in the culture supernatant. In western blot analysis of the culture supernatant using the anti-gp64 antibody, specific protein bands were detected at an electrophoretic mobility that coincided with the molecular weight of the Hc-gp64 fusion protein as well as that of gp64. Flow cytometry using a fluorescein isothiocyanate-conjugated antibody specific to mouse IgG successfully detected the Fab fragments on the surface of the Sf9 cells. These results suggest that immunologically functional antibody Fab fragments can be displayed on the surface of baculovirus particles, and that a fluorescence-activated cell sorter with a fluorescence-labeled antigen can isolate baculoviruses displaying specific Fab fragments. This successful baculovirus display of antibody Fab fragments may offer a novel approach for the efficient selection of specific antibodies.

  8. [Novel bioconversion systems using a yeast molecular display system].

    Science.gov (United States)

    Shibasaki, Seiji

    2010-11-01

    The budding yeast Saccharomyces cerevisiae has been used for the process of fermentation as well as for studies in biochemistry and molecular biology as a eukaryotic model cell or tool for the analysis of gene functions. Thus, yeast is essential in industries and researches. Yeast cells have a cell wall, which is one characteristic that helps distinguish yeast cells from other eukaryotic cells such as mammalian cells. We have developed a molecular display system using the protein of the yeast cell wall as an anchor for foreign proteins. Yeast cells have been designed for use in sensing and metal adsorption, and have been used in vaccines and for screening novel proteins. Currently, yeast is used not only as a tool for analyzing gene or protein function but also in molecular display technology. The phage display system, which is at the forefront of molecular display technologies, is a powerful tool for screening ligands bound to a target molecule and for analyzing protein-protein interactions; however, in some cases, eukaryotic proteins are not easily expressed by this system. On the other hand, yeast cells have the ability to express eukaryotic proteins and proliferate; thus, these cells display various proteins. Yeast cells are more appropriate for white biotechnology. In this review, displays of enzymes that are important in bioconversion, such as lipases and β-glucosidases, are going to be introduced.

  9. Augmenting digital displays with computation

    Science.gov (United States)

    Liu, Jing

    As we inevitably step deeper and deeper into a world connected via the Internet, more and more information will be exchanged digitally. Displays are the interface between digital information and each individual. Naturally, one fundamental goal of displays is to reproduce information as realistically as possible since humans still care a lot about what happens in the real world. Human eyes are the receiving end of such information exchange; therefore it is impossible to study displays without studying the human visual system. In fact, the design of displays is rather closely coupled with what human eyes are capable of perceiving. For example, we are less interested in building displays that emit light in the invisible spectrum. This dissertation explores how we can augment displays with computation, which takes both display hardware and the human visual system into consideration. Four novel projects on display technologies are included in this dissertation: First, we propose a software-based approach to driving multiview autostereoscopic displays. Our display algorithm can dynamically assign views to hardware display zones based on multiple observers' current head positions, substantially reducing crosstalk and stereo inversion. Second, we present a dense projector array that creates a seamless 3D viewing experience for multiple viewers. We smoothly interpolate the set of viewer heights and distances on a per-vertex basis across the arrays field of view, reducing image distortion, crosstalk, and artifacts from tracking errors. Third, we propose a method for high dynamic range display calibration that takes into account the variation of the chrominance error over luminance. We propose a data structure for enabling efficient representation and querying of the calibration function, which also allows user-guided balancing between memory consumption and the amount of computation. Fourth, we present user studies that demonstrate that the ˜ 60 Hz critical flicker fusion

  10. Carriers of loss-of-function mutations in EXT display impaired pancreatic beta-cell reserve due to smaller pancreas volume.

    Directory of Open Access Journals (Sweden)

    Sophie J Bernelot Moens

    Full Text Available Exotosin (EXT proteins are involved in the chain elongation step of heparan sulfate (HS biosynthesis, which is intricately involved in organ development. Loss of function mutations (LOF in EXT1 and EXT2 result in hereditary exostoses (HME. Interestingly, HS plays a role in pancreas development and beta-cell function, and genetic variations in EXT2 are associated with an increased risk for type 2 diabetes mellitus. We hypothesized that loss of function of EXT1 or EXT2 in subjects with hereditary multiple exostoses (HME affects pancreatic insulin secretion capacity and development. We performed an oral glucose tolerance test (OGTT followed by hyperglycemic clamps to investigate first-phase glucose-stimulated insulin secretion (GSIS in HME patients and age and gender matched non-affected relatives. Pancreas volume was assessed with magnetic resonance imaging (MRI. OGTT did not reveal significant differences in glucose disposal, but there was a markedly lower GSIS in HME subjects during hyperglycemic clamp (iAUC HME: 0.72 [0.46-1.16] vs. controls 1.53 [0.69-3.36] nmol·l-1·min-1, p<0.05. Maximal insulin response following arginine challenge was also significantly attenuated (iAUC HME: 7.14 [4.22-10.5] vs. controls 10.2 [7.91-12.70] nmol·l-1·min-1 p<0.05, indicative of an impaired beta-cell reserve. MRI revealed a significantly smaller pancreatic volume in HME subjects (HME: 72.0±15.8 vs. controls 96.5±26.0 cm3 p = 0.04. In conclusion, loss of function of EXT proteins may affect beta-cell mass and insulin secretion capacity in humans, and render subjects at a higher risk of developing type 2 diabetes when exposed to environmental risk factors.

  11. Low Doses of Cadmium Chloride and Methallothionein-1-Bound Cadmium Display Different Accumulation Kinetics and Induce Different Genes in Cells of the Human Nephron

    Directory of Open Access Journals (Sweden)

    Dana Cucu

    2011-08-01

    Full Text Available Background/Aims: The present study was conducted to investigate the renal tubular handling of inorganic cadmium (Cd2+ by exposing primary human tubular cell cultures to physiologically relevant doses of cadmium chloride (CdCl2. Furthermore, the cellular accumulation of Cd2+ was compared to that of metallothionein-1-bound Cd (Cd7MT-1. Finally, this study aimed to investigate the effect of the accumulation of Cd (both Cd2+ and Cd7MT-1 in renal cells on the expression of genes relevant to nephrotoxic processes. Methods: Cd concentration was measured using atomic absorption spectrometry. mRNA expression was evaluated by quantitative real-time RT-PCR. Results: Cd2+ accumulated into human tubular cells in a concentration- and time-dependent way. Furthermore, cellular accumulation of Cd2+ was different from the cellular accumulation of Cd7MT-1, indicative for different uptake routes. Finally, mRNA expression of the genes encoding the anti-oxidative proteins metallothionein-1 (MT-1 and heme-oxygenase-1 (HO-1 as well as the pro-apoptotic Bcl-2-associated X protein (Bax were upregulated by CdCl2 and not by Cd7MT1. Conclusion: In the presence of physiologically relevant Cd concentrations, tubular accumulation of the element in its inorganic form is different from that of Cd7MT-1. Furthermore, the tubular accumulation of inorganic Cd induces mRNA expression of genes of which the protein products may play a role in Cd-associated renal toxicity.

  12. Adipose tissue-derived mesenchymal stem cells in long-term dialysis patients display downregulation of PCAF expression and poor angiogenesis activation.

    Directory of Open Access Journals (Sweden)

    Shuichiro Yamanaka

    Full Text Available We previously demonstrated that mesenchymal stem cells (MSCs differentiate into functional kidney cells capable of urine and erythropoietin production, indicating that they may be used for kidney regeneration. However, the viability of MSCs from dialysis patients may be affected under uremic conditions. In this study, we isolated MSCs from the adipose tissues of end-stage kidney disease (ESKD patients undergoing long-term dialysis (KD-MSCs; mean: 72.3 months and from healthy controls (HC-MSCs to compare their viability. KD-MSCs and HC-MSCs were assessed for their proliferation potential, senescence, and differentiation capacities into adipocytes, osteoblasts, and chondrocytes. Gene expression of stem cell-specific transcription factors was analyzed by PCR array and confirmed by western blot analysis at the protein level. No significant differences of proliferation potential, senescence, or differentiation capacity were observed between KD-MSCs and HC-MSCs. However, gene and protein expression of p300/CBP-associated factor (PCAF was significantly suppressed in KD-MSCs. Because PCAF is a histone acetyltransferase that mediates regulation of hypoxia-inducible factor-1α (HIF-1α, we examined the hypoxic response in MSCs. HC-MSCs but not KD-MSCs showed upregulation of PCAF protein expression under hypoxia. Similarly, HIF-1α and vascular endothelial growth factor (VEGF expression did not increase under hypoxia in KD-MSCs but did so in HC-MSCs. Additionally, a directed in vivo angiogenesis assay revealed a decrease in angiogenesis activation of KD-MSCs. In conclusion, long-term uremia leads to persistent and systematic downregulation of PCAF gene and protein expression and poor angiogenesis activation of MSCs from patients with ESKD. Furthermore, PCAF, HIF-1α, and VEGF expression were not upregulated by hypoxic stimulation of KD-MSCs. These results suggest that the hypoxic response may be blunted in MSCs from ESKD patients.

  13. Phage display derived human monoclonal antibodies isolated by binding to the surface of live primary breast cancer cells recognize GRP78

    DEFF Research Database (Denmark)

    Jakobsen, Charlotte G; Rasmussen, Nicolaj; Laenkholm, Anne-Vibeke

    2007-01-01

    Clinical trials using monoclonal antibodies (mAb) against cell-surface markers have yielded encouraging therapeutic results in several cancer types. Generally, however, anticancer antibodies are only efficient against a subpopulation of cancers, and there is a strong need for identification of no...... therapies including mAb-based immunotherapy. Our results suggest that the human antibody Ab39 may be a useful starting point for further genetic optimization that could render it a useful diagnostic and therapeutic reagent for a variety of cancers...

  14. Inecalcitol, an analog of 1,25D₃, displays enhanced antitumor activity through the induction of apoptosis in a squamous cell carcinoma model system

    Science.gov (United States)

    Ma, Yingyu; Yu, Wei-Dong; Hidalgo, Alejandro A.; Luo, Wei; Delansorne, Remi; Johnson, Candace S.; Trump, Donald L.

    2013-01-01

    Epidemiological data suggest an important role of vitamin D signaling in cancer development and progression, and experimental studies demonstrate that the active vitamin D metabolite 1α, 25-dihydroxyvitamin D₃ (1,25D₃) has broad spectrum antitumor activity. Hypercalcemia has often been suggested to limit the clinical application of these data. The 14-epi-analog of 1,25D₃, inecalcitol [19-nor-14-epi-23-yne-1,25-(OH)₂D₃; TX522], was developed to have superagonistic antitumor activities but low hypercalcemia potential. We examined the antitumor activity of inecalcitol and the underlying mechanisms in a murine squamous cell carcinoma (SCC) model system. In vitro, compared with 1,25D₃, inecalcitol showed enhanced vitamin D receptor (VDR)-mediated transcriptional activity. Inecalcitol suppressed SCC cell proliferation in a dose-dependent manner with an IC₅₀ value 30 times lower than that of 1,25D₃. Both inecalcitol and 1,25D₃ induced a comparable level of G₀/G₁ cell cycle arrest in SCC cells. The level of apoptosis induced by inecalcitol was markedly higher than that of 1,25D₃. Apoptosis was mediated through the activation of the caspase 8/10- caspase 3 pathway. Further, inecalcitol markedly inhibited the mRNA and protein expression of c-IAP1 and XIAP compared with 1,25D₃. In vivo, inecalcitol inhibits SCC tumor growth in a dose-dependent fashion. Notably, inecalcitol induced a significantly higher level of apoptosis in the SCC xenograft model. While in vitro inecalcitol demonstrates apparent enhanced VDR binding and antiproliferative effects compared to 1,25D₃, in vivo these advantages disappear; at doses of inecalcitol that have equivalent antitumor effects, similar hypercalcemia is seen. This may be explained by the pharmacokinetics of 1,25D₃ vs. inecalcitol and attributed to the much shorter serum half-life of inecalcitol.We show that inecalcitol has potent antitumor activity in the SCC model system, and this is associated with a

  15. A novel shogaol analog suppresses cancer cell invasion and inflammation, and displays cytoprotective effects through modulation of NF-κB and Nrf2-Keap1 signaling pathways.

    Science.gov (United States)

    Gan, Fei-Fei; Ling, Hui; Ang, Xiaohui; Reddy, Shridhivya A; Lee, Stephanie S-H; Yang, Hong; Tan, Sock-Hoon; Hayes, John D; Chui, Wai-Keung; Chew, Eng-Hui

    2013-11-01

    Natural compounds containing vanilloid and Michael acceptor moieties appear to possess anti-cancer and chemopreventive properties. The ginger constituent shogaol represents one such compound. In this study, the anti-cancer potential of a synthetic novel shogaol analog 3-phenyl-3-shogaol (3-Ph-3-SG) was assessed by evaluating its effects on signaling pathways. At non-toxic concentrations, 3-Ph-3-SG suppressed cancer cell invasion in MDA-MB-231 and MCF-7 breast carcinoma cells through inhibition of PMA-activated MMP-9 expression. At similar concentrations, 3-Ph-3-SG reduced expression of the inflammatory mediators nitric oxide (NO), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and prostanglandin-E2 (PGE2) in RAW 264.7 macrophage-like cells. Inhibition of cancer cell invasion and inflammation by 3-Ph-3-SG were mediated through suppression of the nuclear factor-kappaB (NF-κB) signaling pathway. The 3-Ph-3-SG also demonstrated cytoprotective effects by inducing the antioxidant response element (ARE)-driven genes NAD(P)H quinone oxidoreductase-1 (NQO1) and heme oxygenase-1 (HO-1). Cytoprotection by 3-Ph-3-SG was achieved at least partly through modification of cysteine residues in the E3 ubiquitin ligase substrate adaptor Kelch-like ECH-associated protein 1 (Keap1), which resulted in accumulation of transcription factor NF-E2 p45-related factor 2 (Nrf2). The activities of 3-Ph-3-SG were comparable to those of 6-shogaol, the most abundant naturally-occurring shogaol, and stronger than those of 4-hydroxyl-null deshydroxy-3-phenyl-3-shogaol, which attested the importance of the 4-hydroxy substituent in the vanilloid moiety for bioactivity. In summary, 3-Ph-3-SG is shown to possess activities that modulate stress-associated pathways relevant to multiple steps in carcinogenesis. Therefore, it warrants further investigation of this compound as a promising candidate for use in chemotherapeutic and chemopreventive strategies.

  16. [16S rDNA diversity analysis of 30 Streptomycetes isolates displaying significant cytotoxic activity against B16 cell from near-shore sediments of Hainan Island].

    Science.gov (United States)

    Yan, Li-Ping; Hong, Kui; Hu, Shen-cai; Liu, Li-hua

    2005-04-01

    A total of 354 isolates of actinomycetes, of which 76 were detected cytotoxic activity was isolated from near-shore marine samples collected at Wenchang mangrove, DanZhou harbor and YanPu harbor. Four isolation methods were employed, which are SDS pretreatment, phenol pretreatment, heating pretreatment and potassium dichromate selection culture, and media such as'Yeast extract-Malt extract (YE), Glucose-Asprine (GA), Starch-Casin (SC), Starch-KNO3 (Gause) were used. It was showed that heating pretreatment and potassium dichromate selection culture were more considerable methods for extensive isolation of actinomycetes. Medium YE and Gause showed best results in both the total number of actinomycetes and the number of active isolates against tumor cell B16. The genotypic diversity of 30 strains of Streptomycetes possessing strong cytotoxic activity against B16 cell (ID50 > or =200) was analyzed by 16S ARDRA, which resulted in 17 RFLP types, and indicated relatively rich genotypic diversity among these Streptomycetes. 16S rDNA sequence analysis of three strains, 050642, 060386 and 060524 (ID50 > or = 1200) further confirmed that they all belong to Streptomyces genus and strain 050642 was suggested a novel Streptomyces. Spp with the highest similarity of 95% to Streptomyces cattleya.

  17. X-1 on display

    Science.gov (United States)

    1949-01-01

    A Bell Aircraft Corporation X-1 series aircraft on display at an Open House at NACA Muroc Flight Test Unit or High-Speed Flight Research Station hangar on South Base of Edwards Air Force Base, California. (The precise date of the photo is uncertain, but it is probably before 1948.) The instrumentation that was carried aboard the aircraft to gather data is on display. The aircraft data was recorded on oscillograph film that was read, calibrated, and converted into meaningful parameters for the engineers to evaluate from each research flight. In the background of the photo are several early U.S. jets. These include several Lockheed P-80 Shooting Stars, which were used as chase planes on X-1 flights; two Bell P-59 Airacomets, the first U.S. jet pursuit aircraft (fighter in later parlance); and a prototype Republic XP-84 Thunderjet. There were five versions of the Bell X-1 rocket-powered research aircraft that flew at the NACA High-Speed Flight Research Station, Edwards, California. The bullet-shaped X-1 aircraft were built by Bell Aircraft Corporation, Buffalo, N.Y. for the U.S. Army Air Forces (after 1947, U.S. Air Force) and the National Advisory Committee for Aeronautics (NACA). The X-1 Program was originally designated the XS-1 for eXperimental Sonic. The X-1's mission was to investigate the transonic speed range (speeds from just below to just above the speed of sound) and, if possible, to break the 'sound barrier.' Three different X-1s were built and designated: X-1-1, X-1-2 (later modified to become the X-1E), and X-1-3. The basic X-1 aircraft were flown by a large number of different pilots from 1946 to 1951. The X-1 Program not only proved that humans could go beyond the speed of sound, it reinforced the understanding that technological barriers could be overcome. The X-1s pioneered many structural and aerodynamic advances including extremely thin, yet extremely strong wing sections; supersonic fuselage configurations; control system requirements; powerplant

  18. Laser illuminated flat panel display

    Energy Technology Data Exchange (ETDEWEB)

    Veligdan, J.T.

    1995-12-31

    A 10 inch laser illuminated flat panel Planar Optic Display (POD) screen has been constructed and tested. This POD screen technology is an entirely new concept in display technology. Although the initial display is flat and made of glass, this technology lends itself to applications where a plastic display might be wrapped around the viewer. The display screen is comprised of hundreds of planar optical waveguides where each glass waveguide represents a vertical line of resolution. A black cladding layer, having a lower index of refraction, is placed between each waveguide layer. Since the cladding makes the screen surface black, the contrast is high. The prototype display is 9 inches wide by 5 inches high and approximately I inch thick. A 3 milliwatt HeNe laser is used as the illumination source and a vector scanning technique is employed.

  19. Immunogenicity of adenovirus-derived porcine parvovirus-like particles displaying B and T cell epitopes of foot-and-mouth disease.

    Science.gov (United States)

    Pan, Qunxing; Wang, Hui; Ouyang, Wei; Wang, Xiaoli; Bi, Zhenwei; Xia, Xingxia; Wang, Yongshan; He, Kongwang

    2016-01-20

    Virus-like particles (VLPs) vaccines combine many of the advantages of whole-virus vaccines and recombinant subunit vaccines, integrating key features that underlay their immunogenicity, safety and protective potential. We have hypothesized here the effective insertion of the VP1 epitopes (three amino acid residues 21-40, 141-160 and 200-213 in VP1, designated VPe) of foot-and-mouth disease (FMDV) within the external loops of PPV VP2 could be carried out without altering assembly based on structural and antigenic data. To investigate the possibility, development of two recombinant adenovirus rAd-PPV:VP2-FMDV:VPe a or rAd-PPV:VP2-FMDV:VPe b were expressed in HEK-293 cells. Out of the two insertion strategies tested, one of them tolerated an insert of 57 amino acids in one of the four external loops without disrupting the VLPs assembly. Mice were inoculated with the two recombinant adenoviruses, and an immunogenicity study showed that the highest levels of FMDV-specific humoral responses and T cell proliferation could be induced by rAd-PPV:VP2-FMDV:VPe b expressing hybrid PPV:VLPs (FMDV) in the absence of an adjuvant. Then, the protective efficacy of inoculating swine with rAd-PPV:VP2-FMDV:VPe b was tested. All pigs inoculated with rAd-PPV:VP2-FMDV:VPe b were protected from viral challenge, meanwhile the neutralizing antibody titers were significantly higher than those in the group inoculated with swine FMD type O synthetic peptide vaccine. Our results clearly demonstrate the potential usefulness of adenovirus-derived PPV VLPs as a vaccine strategy in prevention of FMDV.

  20. Miniature information displays: primary applications

    Science.gov (United States)

    Alvelda, Phillip; Lewis, Nancy D.

    1998-04-01

    Positioned to replace current liquid crystal display technology in many applications, miniature information displays have evolved to provide several truly portable platforms for the world's growing personal computing and communication needs. The technology and functionality of handheld computer and communicator systems has finally surpassed many of the standards that were originally established for desktop systems. In these new consumer electronics, performance, display size, packaging, power consumption, and cost have always been limiting factors for fabricating genuinely portable devices. The rapidly growing miniature information display manufacturing industry is making it possible to bring a wide range of highly anticipated new products to new markets.

  1. Maintenance Procedure Display: Head Mounted Display (HMD) Evaluations

    Science.gov (United States)

    Whitmore, Milrian; Litaker, Harry L., Jr.; Solem, Jody A.; Holden, Kritina L.; Hoffman, Ronald R.

    2007-01-01

    A viewgraph presentation describing maintenance procedures for head mounted displays is shown. The topics include: 1) Study Goals; 2) Near Eye Displays (HMDs); 3) Design; 4) Phase I-Evaluation Methods; 5) Phase 1 Results; 6) Improved HMD Mounting; 7) Phase 2 -Evaluation Methods; 8) Phase 2 Preliminary Results; and 9) Next Steps.

  2. Research on Screening Peptides Specifically Targeting Laryngeal Squamous Cell Carcinoma by Phage Display Technique%用噬菌体展示技术筛选喉癌细胞靶向肽的研究

    Institute of Scientific and Technical Information of China (English)

    冯俊; 李丽; 杨洪斌; 刘世喜

    2011-01-01

    目的 筛选人源喉癌Hep-2细胞株特异结合的短肽,作为喉癌靶向治疗的载体.方法 体外培养Hep-2细胞株作为靶细胞,人正常喉黏膜上皮细胞为吸附细胞;用噬菌体展示十二肽库进行3轮差减筛选,随机挑取10个噬菌体克隆进行测序;采用酶联免疫吸附(enzyme linked immunosorbent assay,ELISA)法鉴定噬菌体与Hep-2细胞的结合活性;通过免疫荧光鉴定喉癌细胞特异性结合肽(F2)噬菌体阳性克隆与喉癌细胞结合的特异性.结果 经过3轮筛选后,噬菌体在靶细胞Hep-2上出现明显富集;ELISA分析鉴定显示5个阳性克隆能与Hep-2细胞特异结合,其中F2噬菌体克隆对喉癌细胞的结合靶向性明显高于对照细胞(P<0.05);免疫荧光显色显示,F2能特异性地与喉癌细胞结合.结论 利用噬菌体展示肽库技术,可以成功筛选到F2,其可能成为喉癌靶向治疗的载体.%Objective To obtain the polypeptides specifically bound to laryngeal squamous cell carcinoma line (Hep-2) and use it as a potential therapeutic vector targeting laryngeal squamous cell carcinoma patients. Methods With the Hep-2 cells as the target cells and human normal laryngeal squamous epithelial cells (HNLE cells) as the absorber cells, 3 rounds of panning from a Ph. D. -12TM phage-display peptide library were carried out. Ten randomly selected phage clones were sent for sequence detection. The affinity of phage clones was detected by enzyme-linked immunosorbent assay (ELISA). The positive phage clones (F2) specifically bound to Hep-2 were identified by immunofluorescence detection. Results After 3 rounds of screening, 5 positive phage clones showed specific binding to Hep-2 cells and the affinity of positive phage clones (F2) was significantly higher than that of the control groups (P<0. 05). The results of immunofluorescence detection indicated that F2 could be specifically bound to Hep-2. Conclusion Phage display peptide libraries technique can

  3. Recombinant baculovirus displayed vaccine

    Science.gov (United States)

    Prabakaran, Mookkan; Kwang, Jimmy

    2014-01-01

    The rapid evolution of new sublineages of H5N1 influenza in Asia poses the greatest challenge in vaccine development for pre-pandemic preparedness. To overcome the antigenic diversity of H5N1 strains, multiple vaccine strains can be designed based on the distribution of neutralizing epitopes in the globular head of H5 hemagglutinin (HA). Recently, we selected two different HAs of H5N1 strains based on the neutralizing epitopes and reactivity with different neutralizing antibodies. The HAs of selected vaccine strains were individually expressed on the baculovirus envelope (bivalent-BacHA) with its native antigenic configuration. Further, oral delivery of live bivalent-BacHA elicited broadly reactive humoral, mucosal and cell-mediated immune responses and showed complete protection against antigenically distinct H5N1 strains in mice. The strategy for the vaccine strain selection, vaccine design and route of administration will provide an idea for development of a widely protective vaccine against highly pathogenic H5N1 for pre-pandemic preparedness. PMID:23941989

  4. Three-dimensional display technologies.

    Science.gov (United States)

    Geng, Jason

    2013-01-01

    The physical world around us is three-dimensional (3D), yet traditional display devices can show only two-dimensional (2D) flat images that lack depth (i.e., the third dimension) information. This fundamental restriction greatly limits our ability to perceive and to understand the complexity of real-world objects. Nearly 50% of the capability of the human brain is devoted to processing visual information [Human Anatomy & Physiology (Pearson, 2012)]. Flat images and 2D displays do not harness the brain's power effectively. With rapid advances in the electronics, optics, laser, and photonics fields, true 3D display technologies are making their way into the marketplace. 3D movies, 3D TV, 3D mobile devices, and 3D games have increasingly demanded true 3D display with no eyeglasses (autostereoscopic). Therefore, it would be very beneficial to readers of this journal to have a systematic review of state-of-the-art 3D display technologies.

  5. Flat panel display - Impurity doping technology for flat panel displays

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Toshiharu [Advanced Technology Planning, Sumitomo Eaton Nova Corporation, SBS Tower 9F, 10-1, Yoga 4-chome, Setagaya-ku, 158-0097 Tokyo (Japan)]. E-mail: suzuki_tsh@senova.co.jp

    2005-08-01

    Features of the flat panel displays (FPDs) such as liquid crystal display (LCD) and organic light emitting diode (OLED) display, etc. using low temperature poly-Si (LTPS) thin film transistors (TFTs) are briefly reviewed comparing with other FPDs. The requirements for fabricating TFTs used for high performance FPDs and system on glass (SoG) are addressed. This paper focuses on the impurity doping technology, which is one of the key technologies together with crystallization by laser annealing, formation of high quality gate insulator and gate-insulator/poly-Si interface. The issues to be solved in impurity doping technology for state of the art and future TFTs are clarified.

  6. Tone compatibility between HDR displays

    Science.gov (United States)

    Bist, Cambodge; Cozot, Rémi; Madec, Gérard; Ducloux, Xavier

    2016-09-01

    High Dynamic Range (HDR) is the latest trend in television technology and we expect an in ux of HDR capable consumer TVs in the market. Initial HDR consumer displays will operate on a peak brightness of about 500-1000 nits while in the coming years display peak brightness is expected to go beyond 1000 nits. However, professionally graded HDR content can range from 1000 to 4000 nits. As with Standard Dynamic Range (SDR) content, we can expect HDR content to be available in variety of lighting styles such as low key, medium key and high key video. This raises concerns over tone-compatibility between HDR displays especially when adapting to various lighting styles. It is expected that dynamic range adaptation between HDR displays uses similar techniques as found with tone mapping and tone expansion operators. In this paper, we survey simple tone mapping methods of 4000 nits color-graded HDR content for 1000 nits HDR displays. We also investigate tone expansion strategies when HDR content graded in 1000 nits is displayed on 4000 nits HDR monitors. We conclude that the best tone reproduction technique between HDR displays strongly depends on the lighting style of the content.

  7. A Cinnamon-Derived Procyanidin Compound Displays Anti-HIV-1 Activity by Blocking Heparan Sulfate- and Co-Receptor- Binding Sites on gp120 and Reverses T Cell Exhaustion via Impeding Tim-3 and PD-1 Upregulation

    Science.gov (United States)

    Connell, Bridgette Janine; Chang, Sui-Yuan; Prakash, Ekambaranellore; Yousfi, Rahima; Mohan, Viswaraman; Posch, Wilfried; Wilflingseder, Doris; Moog, Christiane; Kodama, Eiichi N.; Clayette, Pascal; Lortat-Jacob, Hugues

    2016-01-01

    Amongst the many strategies aiming at inhibiting HIV-1 infection, blocking viral entry has been recently recognized as a very promising approach. Using diverse in vitro models and a broad range of HIV-1 primary patient isolates, we report here that IND02, a type A procyanidin polyphenol extracted from cinnamon, that features trimeric and pentameric forms displays an anti-HIV-1 activity against CXCR4 and CCR5 viruses with 1–7 μM ED50 for the trimer. Competition experiments, using a surface plasmon resonance-based binding assay, revealed that IND02 inhibited envelope binding to CD4 and heparan sulphate (HS) as well as to an antibody (mAb 17b) directed against the gp120 co-receptor binding site with an IC50 in the low μM range. IND02 has thus the remarkable property of simultaneously blocking gp120 binding to its major host cell surface counterparts. Additionally, the IND02-trimer impeded up-regulation of the inhibitory receptors Tim-3 and PD-1 on CD4+ and CD8+ cells, thereby demonstrating its beneficial effect by limiting T cell exhaustion. Among naturally derived products significantly inhibiting HIV-1, the IND02-trimer is the first component demonstrating an entry inhibition property through binding to the viral envelope glycoprotein. These data suggest that cinnamon, a widely consumed spice, could represent a novel and promising candidate for a cost-effective, natural entry inhibitor for HIV-1 which can also down-modulate T cell exhaustion markers Tim-3 and PD-1. PMID:27788205

  8. Comparative study of antitumor effects of bromelain and papain in human cholangiocarcinoma cell lines.

    Science.gov (United States)

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

    2016-05-01

    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.

  9. Tactile display device using an electrorheological fluid

    Science.gov (United States)

    Garner, H. Douglas

    1994-08-01

    A tactile display device utilizes an electrorheological fluid to activate a plurality of tactile dots. A voltage is selectively produced uniformly across an electrorheological fluid flowing between a common ground electrode and a plurality of conductive dot electrodes, thereby producing an increase in the fluid's viscosity to the extent that fluid flow between the two electrodes is restricted. The flow restriction produces a build-up of electrorheological fluid in a corresponding dot actuator chamber. The resulting pressure increase in the chamber displaces an elastic diaphragm fixed to a display surface to form a lump which can be perceived by the reader as one dot in a Braille character cell. A flow regulation system provides a continually pressurized flow system and provides for free flow of the electrorheological fluid through the plurality of dot actuator chambers when they are not activated. The device is adaptable to printed circuit techniques and can simultaneously display tactile dots representative of a full page of Braille characters stored on a medium such as a tape cassette or to display tactile dots representative of non-Braille data appearing on a computer monitor or contained on another data storage medium. In an alternate embodiment, the elastic diaphragm drives a plurality of spring-loaded pins provided with positive stops to maintain consistent displacements of the pins in both their actuated and nonactuated positions.

  10. Ultraminiature, Micropower Multipurpose Display Project

    Data.gov (United States)

    National Aeronautics and Space Administration — High information content electronic displays remain the most difficult element of the human-machine interface to effectively miniaturize. Mobile applications need a...

  11. Color speckle in laser displays

    Science.gov (United States)

    Kuroda, Kazuo

    2015-07-01

    At the beginning of this century, lighting technology has been shifted from discharge lamps, fluorescent lamps and electric bulbs to solid-state lighting. Current solid-state lighting is based on the light emitting diodes (LED) technology, but the laser lighting technology is developing rapidly, such as, laser cinema projectors, laser TVs, laser head-up displays, laser head mounted displays, and laser headlamps for motor vehicles. One of the main issues of laser displays is the reduction of speckle noise1). For the monochromatic laser light, speckle is random interference pattern on the image plane (retina for human observer). For laser displays, RGB (red-green-blue) lasers form speckle patterns independently, which results in random distribution of chromaticity, called color speckle2).

  12. ENERGY STAR Certified Displays - Deprecated

    Data.gov (United States)

    U.S. Environmental Protection Agency — This dataset is up-to-date but newer better data can be retrieved at: https://data.energystar.gov/dataset/ENERGY-STAR-Certified-Displays/xsyb-v8gs Certified models...

  13. Ten inch Planar Optic Display

    Energy Technology Data Exchange (ETDEWEB)

    Beiser, L. [Beiser (Leo) Inc., Flushing, NY (United States); Veligdan, J. [Brookhaven National Lab., Upton, NY (United States)

    1996-04-01

    A Planar Optic Display (POD) is being built and tested for suitability as a high brightness replacement for the cathode ray tube, (CRT). The POD display technology utilizes a laminated optical waveguide structure which allows a projection type of display to be constructed in a thin (I to 2 inch) housing. Inherent in the optical waveguide is a black cladding matrix which gives the display a black appearance leading to very high contrast. A Digital Micromirror Device, (DMD) from Texas Instruments is used to create video images in conjunction with a 100 milliwatt green solid state laser. An anamorphic optical system is used to inject light into the POD to form a stigmatic image. In addition to the design of the POD screen, we discuss: image formation, image projection, and optical design constraints.

  14. Tyrosine Kinase Display of Prostate Cancer Cells

    Science.gov (United States)

    2001-10-01

    Swanson, P.E., Ratliff, T.L., Vollmer, R.T., Day, M.L. (1995) HGF and its receptor c-met in prostatic carcinoma. Am. J. Pathol. 147:386-396. 4. Levitzki , A...alpha and erbB-3 receptor in human prostatic adenocarcinoma. Br. J. Urol. 79: 212-216. 78. Levitzki , A. and A. Gazit. 1995. Tyrosine kinase inhibition

  15. Hydrogen inhibits cytotrophoblast cells apoptosis in hypertensive disorders complicating pregnancy.

    Science.gov (United States)

    Guo, L; Guan, Z; Li, H; Yang, X

    2016-05-30

    Hypertensive disorders complicating pregnancy (HDCP) is one of the most serious medical disorders during pregnancy. Hydrogen is a therapeutic antioxidant and used to treat HDCP effectively. However, the molecular mechanism about the effect of hydrogen on HDCP still remains unclear. In this study, we found ROS content in HDCP group was significantly higher than that in the control and was reduced markedly in the presence of 100μmol/L hydrogen. IL6, Caspase3, Bax1, P-JAK2, P-Stat3 and P-p38 expression was much higher than the control, and was notably decreasedby the application of 100μmol/L hydrogen. Bcl2 expression in HDCP group was notably lower than the control and was increased by 100 μmol/L hydrogen. The apoptosis rate of cytotrophoblast cells was decreased, andratio of cytotrophoblast cells at G1 and G2 phase was increased and decreased by hydrogen, respectively. All those data indicated a potential molecular mechanism of hydrogen-mediated treatment in HDCP.

  16. Memory CD8(+) T cells elicited by HIV-1 lipopeptide vaccines display similar phenotypic profiles but differences in term of magnitude and multifunctionality compared with FLU- or EBV-specific memory T cells in humans.

    Science.gov (United States)

    Figueiredo, Suzanne; Charmeteau, Benedicte; Surenaud, Mathieu; Salmon, Dominique; Launay, Odile; Guillet, Jean-Gérard; Hosmalin, Anne; Gahery, Hanne

    2014-01-16

    Differentiation marker, multifunctionality and magnitude analyses of specific-CD8(+) memory T cells are crucial to improve development of HIV vaccines designed to generate cell-mediated immunity. Therefore, we fully characterized the HIV-specific CD8(+) T cell responses induced in volunteers vaccinated with HIV lipopeptide vaccines for phenotypic markers, tetramer staining, cytokine secretion, and cytotoxic activities. The frequency of ex vivo CD8(+) T cells elicited by lipopeptide vaccines is very rare and central-memory phenotype and functions of these cells were been shown to be important in AIDS immunity. So, we expanded them using specific peptides to compare the memory T cell responses induced in volunteers by HIV vaccines with responses to influenza (FLU) or Epstein Barr virus (EBV). By analyzing the differentiation state of IFN-γ-secreting CD8(+) T cells, we found a CCR7(-)CD45RA(-)CD28(+int)/CD28(-) profile (>85%) belonging to a subset of intermediate-differentiated effector T cells for HIV, FLU, and EBV. We then assessed the quality of the response by measuring various T cell functions. The percentage of single IFN-γ T cell producers in response to HIV was 62% of the total of secreting T cells compared with 35% for FLU and EBV, dual and triple (IFN-γ/IL-2/CD107a) T cell producers could also be detected but at lower levels (8% compared with 37%). Finally, HIV-specific T cells secreted IFN-γ and TNF-α, but not the dual combination like FLU- and EBV-specific T cells. Thus, we found that the functional profile and magnitude of expanded HIV-specific CD8(+) T precursors were more limited than those of to FLU- and EBV-specific CD8(+) T cells. These data show that CD8(+) T cells induced by these HIV vaccines have a similar differentiation profile to FLU and EBV CD8(+) T cells, but that the vaccine potency to induce multifunctional T cells needs to be increased in order to improve vaccination strategies.

  17. Modulation of cell signaling networks after CTLA4 blockade in patients with metastatic melanoma.

    Directory of Open Access Journals (Sweden)

    Begoña Comin-Anduix

    Full Text Available BACKGROUND: The effects on cell signalling networks upon blockade of cytotoxic T lymphocyte-associated antigen-4 (CTLA4 using the monoclonal antibody tremelimumab were studied in peripheral blood mononuclear cell (PBMC samples from patients with metastatic melanoma. METHODOLOGY/PRINCIPAL: Findings Intracellular flow cytometry was used to detect phosphorylated (p signaling molecules downstream of the T cell receptor (TCR and cytokine receptors. PBMC from tremelimumab-treated patients were characterized by increase in pp38, pSTAT1 and pSTAT3, and decrease in pLck, pERK1/2 and pSTAT5 levels. These changes were noted in CD4 and CD8 T lymphocytes but also in CD14 monocytes. A divergent pattern of phosphorylation of Zap70, LAT, Akt and STAT6 was noted in patients with or without an objective tumor response. CONCLUSIONS/SIGNIFICANCE: The administration of the CTLA4-blocking antibody tremelimumab to patients with metastatic melanoma influences signaling networks downstream of the TCR and cytokine receptors both in T cells and monocytes. The strong modulation of signaling networks in monocytes suggests that this cell subset may be involved in clinical responses to CTLA4 blockade. CLINICAL TRIAL REGISTRATION: clinicaltrials.gov; Registration numbers NCT00090896 and NCT00471887.

  18. Protective Effects and Mechanisms of Salvianolic Acid B Against H₂O₂-Induced Injury in Induced Pluripotent Stem Cell-Derived Neural Stem Cells.

    Science.gov (United States)

    Shu, Tao; Pang, Mao; Rong, Limin; Liu, Chang; Wang, Juan; Zhou, Wei; Wang, Xuan; Liu, Bin

    2015-06-01

    Induced pluripotent stem cells (iPSCs) have the potential to differentiate into neural lineages. Salvianolic acid B (Sal B) is a commonly used, traditional Chinese medicine for enhancing neuroprotective effects, and has antioxidant, anti-inflammatory, and antiapoptotic properties. Here, we explore the potential mechanism of Sal B in protecting iPSC-derived neural stem cells (NSCs) against H2O2-induced injury. iPSCs were induced into NSCs, iPSC-derived NSCs were treated with 50 μM Sal B for 24.5 h and 500 μM H2O2 for 24 h. The resulting effects were examined by flow cytometry analysis, quantitative reverse-transcription polymerase chain reaction, and western blotting. Upon H2O2 exposure, Sal B significantly promoted cell viability and stabilization of the mitochondrial membrane potential. Sal B also visibly decreased the cell apoptotic ratio. In addition, Sal B markedly reduced expression of matrix metalloproteinase (MMP)-2 and -9, and phosphospecific signal transducer and activator of transcription 3 (p-STAT3), and increased the level of tissue inhibitor of metalloproteinase (TIMP)-2 in iPSC-derived NSCs induced by H2O2. These results suggest that Sal B protects iPSC-derived NSCs against H2O2-induced oxidative stress. The mechanisms of this stress tolerance may be attributed to modulation of the MMP/TIMP system and inhibition of the STAT3 signaling pathway.

  19. A nanobody:GFP bacterial platform that enables functional enzyme display and easy quantification of display capacity

    DEFF Research Database (Denmark)

    Wendel, Sofie; Christian Fischer, Emil; Martinez, Virginia;

    2016-01-01

    Background: Bacterial surface display is an attractive technique for the production of cell-anchored, functional proteins and engineering of whole-cell catalysts. Although various outer membrane proteins have been used for surface display, an easy and versatile high-throughput-compatible assay...... for evaluating and developing surface display systems is missing.Results: Using a single domain antibody (also called nanobody) with high affinity for green fluorescent protein (GFP), we constructed a system that allows for fast, fluorescence-based detection of displayed proteins. The outer membrane hybrid...... protein LppOmpA and the autotransporter C-IgAP exposed the nanobody on the surface of Escherichia coli with very different efficiency. Both anchors were capable of functionally displaying the enzyme Chitinase A as a fusion with the nanobody, and this considerably increased expression levels compared...

  20. Peripheral T-cell lymphomas of follicular helper T-cell type frequently display an aberrant CD3(-/dim)CD4(+) population by flow cytometry: an important clue to the diagnosis of a Hodgkin lymphoma mimic.

    Science.gov (United States)

    Alikhan, Mir; Song, Joo Y; Sohani, Aliyah R; Moroch, Julien; Plonquet, Anne; Duffield, Amy S; Borowitz, Michael J; Jiang, Liuyan; Bueso-Ramos, Carlos; Inamdar, Kedar; Menon, Madhu P; Gurbuxani, Sandeep; Chan, Ernest; Smith, Sonali M; Nicolae, Alina; Jaffe, Elaine S; Gaulard, Philippe; Venkataraman, Girish

    2016-10-01

    Nodal follicular helper T-cell-derived lymphoproliferations (specifically the less common peripheral T-cell lymphomas of follicular type) exhibit a spectrum of histologic features that may mimic reactive hyperplasia or Hodgkin lymphoma. Even though angioimmunoblastic T-cell lymphoma and peripheral T-cell lymphoma of follicular type share a common biologic origin from follicular helper T-cells and their morphology has been well characterized, flow cytometry of peripheral T-cell lymphomas of follicular type has not been widely discussed as a tool for identifying this reactive hyperplasia/Hodgkin lymphoma mimic. We identified 10 peripheral T-cell lymphomas of follicular type with available flow cytometry data from five different institutions, including two cases with peripheral blood evaluation. For comparison, we examined flow cytometry data for 8 classical Hodgkin lymphomas (including 1 lymphocyte-rich classical Hodgkin lymphoma), 15 nodular lymphocyte predominant Hodgkin lymphomas, 15 angioimmunoblastic T-cell lymphomas, and 26 reactive nodes. Lymph node histology and flow cytometry data were reviewed, specifically for the presence of a CD3(-/dim)CD4(+) aberrant T-cell population (described in angioimmunoblastic T-cell lymphomas), besides other T-cell aberrancies. Nine of 10 (90%) peripheral T-cell lymphomas of follicular type showed a CD3(-/dim)CD4(+) T-cell population constituting 29.3% (range 7.9-62%) of all lymphocytes. Five of 10 (50%) had nodular lymphocyte predominant Hodgkin lymphoma or lymphocyte-rich classical Hodgkin lymphoma-like morphology with scattered Hodgkin-like cells that expressed CD20, CD30, CD15, and MUM1. Three cases had a nodular growth pattern and three others exhibited a perifollicular growth pattern without Hodgkin-like cells. Epstein-Barr virus was positive in 1 of 10 cases (10%). PCR analysis showed clonal T-cell receptor gamma gene rearrangement in all 10 peripheral T-cell lymphomas of follicular type. By flow cytometry, 11 of 15 (73

  1. Phosphors for flat panel emissive displays

    Energy Technology Data Exchange (ETDEWEB)

    Anderson, M.T.; Walko, R.J.; Phillips, M.L.F.

    1995-07-01

    An overview of emissive display technologies is presented. Display types briefly described include: cathode ray tubes (CRTs), field emission displays (FEDs), electroluminescent displays (ELDs), and plasma display panels (PDPs). The critical role of phosphors in further development of the latter three flat panel emissive display technologies is outlined. The need for stable, efficient red, green, and blue phosphors for RGB fall color displays is emphasized.

  2. Copper(II) complexes of salicylic acid combining superoxide dismutase mimetic properties with DNA binding and cleaving capabilities display promising chemotherapeutic potential with fast acting in vitro cytotoxicity against cisplatin sensitive and resistant cancer cell lines.

    Science.gov (United States)

    O'Connor, Mark; Kellett, Andrew; McCann, Malachy; Rosair, Georgina; McNamara, Mary; Howe, Orla; Creaven, Bernadette S; McClean, Siobhán; Kia, Agnieszka Foltyn-Arfa; O'Shea, Denis; Devereux, Michael

    2012-03-08

    The complexes [Cu(salH)(2)(H(2)O)] (1), [Cu(dipsH)(2)(H(2)O)] (2), {Cu(3-MeOsal)(H(2)O)(0.75)}(n) (3), [Cu(dipsH)(2)(BZDH)(2)] (4), [Cu(dipsH)(2)(2-MeOHBZDH)(2)]·EtOH (5), [Cu(sal)(phen)] (6), [Cu(dips)(phen)]·H(2)O (7), and [Cu(3-MeOsal)(phen)]·H(2)O (8) (salH(2) = salicylic acid; dipsH(2) = 3,5-diisopropylsalicylic acid; 3-MeOsalH(2) = 3-methoxysalicylic acid; BZDH = benzimidazole; 2-MeOHBZDH = 2 methanolbenzimidazole and phen =1,10-phenanthroline) were prepared and characterized. Structures of 4, 5, and 8 were determined by X-ray crystallography. Compounds 1-8 are potent superoxide dismutase mimetics, and they are inactive as inhibitors of COX-2 activity. Compounds 1, 4, and 5 exhibit moderate inhibition of COX-1. Complexes 6-8 display rapid micromolar cytotoxicity against cisplatin sensitive (breast (MCF-7), prostate (DU145), and colon (HT29)) and cisplatin resistant (ovarian (SK-OV-3)) cell lines compared to 1-5, and they exhibit potent in vitro DNA binding and cleavage capabilities.

  3. Naturally processed peptides spanning the HPA-1a polymorphism are efficiently generated and displayed from platelet glycoprotein by HLA-DRB3*0101-positive antigen-presenting cells.

    Science.gov (United States)

    Anani Sarab, Gholamreza; Moss, Michael; Barker, Robert N; Urbaniak, Stanislaw J

    2009-08-27

    In neonatal alloimmune thrombocytopenia, almost all human platelet antigen (HPA)-1b1b mothers who produce anti-HPA-1a antibody through carrying an HPA-1a fetus are human histocompatibility leukocyte antigen (HLA)-DRB3*0101 positive. It is predicted that the HPA-1a Leu(33) polymorphism forms part of an HLA-DRB3*0101-restricted T-helper epitope, and acts as an anchor residue for binding this class II molecule. However, it is not known whether any corresponding peptides are naturally processed and presented from platelet glycoprotein. In this study, peptides displayed by a homozygous HLA-DRB3*0101 antigen-presenting cell line were identified after pulsing with recombinant HPA-1a (Leu(33) plexin-semaphorin-integrin domain). The peptides were eluted from HLA-DR molecules, fractionated by high performance liquid chromatography, and analyzed by tandem mass spectrometry. A "nested set" of naturally presented HPA-1a-derived peptides, each containing the Trp(25)-Leu(33) core epitope, was identified, with the most abundant member being the 16-mer Met(22)-Arg(37). These peptides may provide the basis for novel treatments to tolerize the corresponding T-helper response in women at risk of neonatal alloimmune thrombocytopenia.

  4. BES Monitoring & Displaying System

    Institute of Scientific and Technical Information of China (English)

    MengWANG; BingyunZHANG; 等

    2001-01-01

    BES1 Monitoring & Displaying System(BESMDS)is projected to monitor and display the running status of DAQ and Slow Control systems of BES through the Web for worldwide accessing.It provides a real-time remote means of monitoring as well as an approach to study the environmental influence upon physical data taking.The system collects real-time data separately from BES online subsystems by network sockets and stores the data into a database.People can access the system through its web site.which retrieves data on request from the database and can display results in dynamically created images.Its web address in http:// besmds,ihep.ac.cn/

  5. Engineering antibodies by yeast display.

    Science.gov (United States)

    Boder, Eric T; Raeeszadeh-Sarmazdeh, Maryam; Price, J Vincent

    2012-10-15

    Since its first application to antibody engineering 15 years ago, yeast display technology has been developed into a highly potent tool for both affinity maturing lead molecules and isolating novel antibodies and antibody-like species. Robust approaches to the creation of diversity, construction of yeast libraries, and library screening or selection have been elaborated, improving the quality of engineered molecules and certainty of success in an antibody engineering campaign and positioning yeast display as one of the premier antibody engineering technologies currently in use. Here, we summarize the history of antibody engineering by yeast surface display, approaches used in its application, and a number of examples highlighting the utility of this method for antibody engineering.

  6. PROGRAMMABLE DISPLAY PUSHBUTTON LEGEND EDITOR

    Science.gov (United States)

    Busquets, A. M.

    1994-01-01

    The Programmable Display Pushbutton (PDP) is a pushbutton device available from Micro Switch which has a programmable 16 x 35 matrix of LEDs on the pushbutton surface. Any desired legends can be displayed on the PDPs, producing user-friendly applications which greatly reduce the need for dedicated manual controls. Because the PDP can interact with the operator, it can call for the correct response before transmitting its next message. It is both a simple manual control and a sophisticated programmable link between the operator and the host system. The Programmable Display Pushbutton Legend Editor, PDPE, is used to create the LED displays for the pushbuttons. PDPE encodes PDP control commands and legend data into message byte strings sent to a Logic Refresh and Control Unit (LRCU). The LRCU serves as the driver for a set of four PDPs. The legend editor (PDPE) transmits to the LRCU user specified commands that control what is displayed on the LED face of the individual pushbuttons. Upon receiving a command, the LRCU transmits an acknowledgement that the message was received and executed successfully. The user then observes the effect of the command on the PDP displays and decides whether or not to send the byte code of the message to a data file so that it may be called by an applications program. The PDPE program is written in FORTRAN for interactive execution. It was developed on a DEC VAX 11/780 under VMS. It has a central memory requirement of approximately 12800 bytes. It requires four Micro Switch PDPs and two RS-232 VAX 11/780 terminal ports. The PDPE program was developed in 1985.

  7. Computational multi-projection display.

    Science.gov (United States)

    Moon, Seokil; Park, Soon-Gi; Lee, Chang-Kun; Cho, Jaebum; Lee, Seungjae; Lee, Byoungho

    2016-04-18

    A computational multi-projection display is proposed by employing a multi-projection system combining with compressive light field displays. By modulating the intensity of light rays from a spatial light modulator inside a single projector, the proposed system can offer several compact views to observer. Since light rays are spread to all directions, the system can provide flexible positioning of viewpoints without stacking projectors in vertical direction. Also, if the system is constructed properly, it is possible to generate view images with inter-pupillary gap and satisfy the super multi-view condition. We explain the principle of the proposed system and verify its feasibility with simulations and experimental results.

  8. Display standards for commercial flight decks

    Science.gov (United States)

    Lamberth, Larry S.; Penn, Cecil W.

    1994-06-01

    SAE display standards are used as guidelines for certifying commercial airborne electronic displays. The SAE document generation structure and approval process is described. The SAE committees that generate display standards are described. Three SAE documents covering flat panel displays (AS-8034, ARP-4256, and ARP-4260) are discussed with their current status. Head-Up Display documents are also in work.

  9. Autostereoscopic display with eye tracking

    Science.gov (United States)

    Tomono, Takao; Hoon, Kyung; Ha, Yong Soo; Kim, Sung-Sik; Son, Jung-Young

    2002-05-01

    Auto-stereoscopic 21-inch display with eye tracking having wide viewing zone and bright image was fabricated. The image of display is projected to retinal through several optical components. We calculated optical system for wider viewing zone by using Inverse-Ray Trace Method. The viewing zone of first model is 155mm (theoretical value: 161mm). We could widen viewing zone by controlling paraxial radius of curvature of spherical mirror, the distance between lenses and so on. The viewing zone of second model is 208mm. We used two spherical mirrors to obtain twice brightness. We applied eye-tracking system to the display system. Eye recognition is based on neural network card based on ZICS technology. We fabricated Auto-stereoscopic 21-inch display with eye tracking. We measured viewing zone based on illumination area. The viewing zone was 206mm, which was close to theoretical value. We could get twice brightness also. We could see 3D image according to position without headgear.

  10. Graphics Display of Foreign Scripts.

    Science.gov (United States)

    Abercrombie, John R.

    1987-01-01

    Describes Graphics Project for Foreign Language Learning at the University of Pennsylvania, which has developed ways of displaying foreign scripts on microcomputers. Character design on computer screens is explained; software for graphics, printing, and language instruction is discussed; and a text editor is described that corrects optically…

  11. Real Time Sonic Boom Display

    Science.gov (United States)

    Haering, Ed

    2014-01-01

    This presentation will provide general information about sonic boom mitigation technology to the public in order to supply information to potential partners and licensees. The technology is a combination of flight data, atmospheric data and terrain information implemented into a control room real time display for flight planning. This research is currently being performed and as such, any results and conclusions are ongoing.

  12. Verbal Modification via Visual Display

    Science.gov (United States)

    Richmond, Edmun B.; Wallace-Childers, La Donna

    1977-01-01

    The inability of foreign language students to produce acceptable approximations of new vowel sounds initiated a study to devise a real-time visual display system whereby the students could match vowel production to a visual pedagogical model. The system used amateur radio equipment and a standard oscilloscope. (CHK)

  13. Information retrieval and display system

    Science.gov (United States)

    Groover, J. L.; King, W. L.

    1977-01-01

    Versatile command-driven data management system offers users, through simplified command language, a means of storing and searching data files, sorting data files into specified orders, performing simple or complex computations, effecting file updates, and printing or displaying output data. Commands are simple to use and flexible enough to meet most data management requirements.

  14. Colour displays for categorical images

    NARCIS (Netherlands)

    Glasbey, C.; Heijden, van der G.W.A.M.; Toh, V.F.K.; Gray, A.J.

    2007-01-01

    We propose a method for identifying a set of colours for displaying 2D and 3D categorical images when the categories are unordered labels. The principle is to find maximally distinct sets of colours. We either generate colours sequentially, to maximize the dissimilarity or distance between a new col

  15. Book Display as Adult Service.

    Science.gov (United States)

    Moore, Matthew S.

    1997-01-01

    Defines book display as an adult service as choosing and positioning adult books from the library collection to increase their circulation. The author contrasts bookstore arrangement for sales versus library arrangement for access, including contrasting missions, genre grouping, weeding, problems, and dimensions. (Author/LRW)

  16. The molecular mechanism of curcumol on inducing cell growth arrest and apoptosis in Jurkat cells, a model of CD4⁺ T cells.

    Science.gov (United States)

    Wang, Heng; Wang, Yong; Jiang, Xiaoji; Wang, Zhizhong; Zhong, Bing; Fang, Yongfei

    2014-08-01

    CD4(+) T cells in rheumatoid arthritis (RA) express growth signaling pathway in association with deregulated growth and resistance to apoptosis. The janus kinase (Jak) 3 and signal transducer and activator of transcription (STAT) pathway play a critical role in interleukin-2 (IL-2)-induced CD4(+) T cell proliferation. The present study aimed to explore the anti-cell proliferation mechanism of curcumol, a pure monomer extracted from Chinese medical plant Rhizoma curcumae. Cell proliferation was determined using WST-1 assay after curcumol treatment. The cell cycle distribution and Bcl-2 protein expression were assessed by flow cytometry. The cellular morphology of apoptosis was evaluated by Hoechst 33258 staining. The expressions of phosphorylated-Jak3 (p-Jak3), p-STAT3, and p-STAT5a following IL-2 stimulation were determined by western blot analysis. The Electrophoretic Mobility Shift Assay was used to detect the DNA binding activities of transcription factors STAT3 and STAT5. The study results showed that curcumol could inhibit the IL-2-induced Jurkat cell proliferation in a concentration- and time-dependent manner in vitro. Curcumol could cause cell cycle arrest at the S phase, induce cell apoptosis, and inhibit the expression of Bcl-2 in a dose-dependent manner. Curcumol at 50μg/mL and Jak3 inhibitor ZM39923 could inhibit the phosphorylation of Jak3 and STAT5a. In conclusion, the underlying mechanism of curcumol on suppressing CD4(+) T cell proliferation and inducing apoptosis might partly be mediated by inhibition of Jak3-STAT5-related molecular activities and Bcl-2 expression, respectively; further studies are required in vivo to test the use of curcumol as a promising therapeutic option for RA.

  17. Versatile microbial surface-display for environmental remediation and biofuels production

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Cindy H.; Mulchandani, Ashok; Chen, wilfred

    2008-02-14

    Surface display is a powerful technique that utilizes natural microbial functional components to express proteins or peptides on the cell exterior. Since the reporting of the first surface-display system in the mid-1980s, a variety of new systems have been reported for yeast, Gram-positive and Gram-negative bacteria. Non-conventional display methods are emerging, eliminating the generation of genetically modified microorganisms. Cells with surface display are used as biocatalysts, biosorbents and biostimulants. Microbial cell-surface display has proven to be extremely important for numerous applications ranging from combinatorial library screening and protein engineering to bioremediation and biofuels production.

  18. Saw Palmetto Extract Inhibits Metastasis and Antiangiogenesis through STAT3 Signal Pathway in Glioma Cell

    Directory of Open Access Journals (Sweden)

    Hong Ding

    2015-01-01

    Full Text Available Signal transducer and activator of transcription factor 3 (STAT3 plays an important role in the proliferation and angiogenesis in human glioma. Previous research indicated that saw palmetto extract markedly inhibited the proliferation of human glioma cells through STAT3 signal pathway. But its effect on tumor metastasis and antiangiogenesis is not clear. This study is to further clear the impact of saw palmetto extract on glioma cell metastasis, antiangiogenesis, and its mechanism. TUNEL assay indicated that the apoptotic cells in the saw palmetto treated group are higher than that in the control group (p<0.05. The apoptosis related protein is detected and the results revealed that saw palmetto extract inhibits the proliferation of human glioma. Meanwhile pSTAT3 is lower in the experimental group and CD34 is also inhibited in the saw palmetto treated group. This means that saw palmetto extract could inhibit the angiogenesis in glioma. We found that saw palmetto extract was an important phytotherapeutic drug against the human glioma through STAT3 signal pathway. Saw palmetto extract may be useful as an adjunctive therapeutic agent for treatment of individuals with glioma and other types of cancer in which STAT3 signaling is activated.

  19. Phage display library screening for identification of interacting protein partners.

    Science.gov (United States)

    Addepalli, Balasubrahmanyam; Rao, Suryadevara; Hunt, Arthur G

    2015-01-01

    Phage display is a versatile high-throughput screening method employed to understand and improve the chemical biology, be it production of human monoclonal antibodies or identification of interacting protein partners. A majority of cell proteins operate in a concerted fashion either by stable or transient interactions. Such interactions can be mediated by recognition of small amino acid sequence motifs on the protein surface. Phage display can play a crucial role in identification of such motifs. This report describes the use of phage display for the identification of high affinity sequence motifs that could be responsible for interactions with a target (bait) protein.

  20. Peroxiredoxin II Is Essential for Maintaining Stemness by Redox Regulation in Liver Cancer Cells.

    Science.gov (United States)

    Kwon, Taeho; Bak, Yesol; Park, Young-Ho; Jang, Gyu-Beom; Nam, Jeong-Seok; Yoo, Jeong Eun; Park, Young Nyun; Bak, In Seon; Kim, Jin-Man; Yoon, Do-Young; Yu, Dae-Yeul

    2016-05-01

    Redox regulation in cancer stem cells (CSCs) is viewed as a good target for cancer therapy because redox status plays an important role in cancer stem-cell maintenance. Here, we investigated the role of Peroxiredoxin II (Prx II), an antioxidant enzyme, in association with maintenance of liver CSCs. Our study demonstrates that Prx II overexpressed in liver cancer cells has high potential for self-renewal activity. Prx II expression significantly corelated with expression of epithelial-cell adhesion molecules (EpCAM) and cytokerain 19 in liver cancer tissues of hepatocellular carcinoma (HCC) patients. Downregulation of Prx II in Huh7 cells with treatment of siRNA reduced expression of EpCAM and CD133 as well as Sox2 in accordance with increased ROS and apoptosis, which were reversed in Huh7-hPrx II cells. Huh7-hPrx II cells exhibited strong sphere-formation activity compared with mock cells. Vascular endothelial growth factor (VEGF) exposure enhanced sphere formation, cell-surface expression of EpCAM and CD133, and pSTAT3 along with activation of VEGF receptor 2 in Huh7-hPrx II cells. The result also emerged in Huh7-H-ras(G12V) and SK-HEP-1-H-ras(G12V) cells with high-level expression of Prx II. Prx II was involved in regulation of VEGF driving cancer stem cells through VEGFR-2/STAT3 signaling to upregulate Bmi1 and Sox2. In addition, knockdown of Prx II in Huh7-H-ras(G12V) cells showed significant reduction in cell migration in vitro and in tumorigenic potential in vivo. Taken together, all the results demonstrated that Prx II plays a key role in the CSC self-renewal of HCC cells through redox regulation. Stem Cells 2016;34:1188-1197.

  1. Game engines and immersive displays

    Science.gov (United States)

    Chang, Benjamin; Destefano, Marc

    2014-02-01

    While virtual reality and digital games share many core technologies, the programming environments, toolkits, and workflows for developing games and VR environments are often distinct. VR toolkits designed for applications in visualization and simulation often have a different feature set or design philosophy than game engines, while popular game engines often lack support for VR hardware. Extending a game engine to support systems such as the CAVE gives developers a unified development environment and the ability to easily port projects, but involves challenges beyond just adding stereo 3D visuals. In this paper we outline the issues involved in adapting a game engine for use with an immersive display system including stereoscopy, tracking, and clustering, and present example implementation details using Unity3D. We discuss application development and workflow approaches including camera management, rendering synchronization, GUI design, and issues specific to Unity3D, and present examples of projects created for a multi-wall, clustered, stereoscopic display.

  2. Characterization of the rotating display.

    Science.gov (United States)

    Keyes, J W; Fahey, F H; Harkness, B A; Eggli, D F; Balseiro, J; Ziessman, H A

    1988-09-01

    The rotating display is a useful method for reviewing single photon emission computed tomography (SPECT) data. This study evaluated the requirements for a subjectively pleasing and useful implementation of this technique. Twelve SPECT data sets were modified and viewed by several observers who recorded the minimum framing rates for apparent smooth rotation, 3D effect, effects of image size, and other parameters. The results showed that a minimum of 16 frames was needed for a useful display. Smaller image sizes and more frames were preferred. The recommended minimal framing rate for a 64-frame study is 16-17 frames per second and for a 32-frame study, 12-13 frames per second. Other enhancements also were useful.

  3. Displays for future intermediate UAV

    Science.gov (United States)

    Desjardins, Daniel; Metzler, James; Blakesley, David; Rister, Courtney; Nuhu, Abdul-Razak

    2008-04-01

    The Dedicated Autonomous Extended Duration Airborne Long-range Utility System (DAEDALUS) is a prototype Unmanned Aerial Vehicle (UAV) that won the 2007 AFRL Commander's Challenge. The purpose of the Commander's Challenge was to find an innovative solution to urgent warfighter needs by designing a UAV with increased persistence for tactical employment of sensors and communication systems. DAEDALUS was chosen as a winning prototype by AFRL, AFMC and SECAF. Follow-on units are intended to fill an intermediate role between currently fielded Tier I and Tier II UAV's. The UAV design discussed in this paper, including sensors and displays, will enter Phase II for Rapid Prototype Development with the intent of developing the design for eventual production. This paper will discuss the DAEDALUS UAV prototype system, with particular focus on its communications, to include the infrared sensor and electro-optical camera, but also displays, specifically man-portable.

  4. Interactive display of polygonal data

    Energy Technology Data Exchange (ETDEWEB)

    Wood, P.M.

    1977-10-01

    Interactive computer graphics is an excellent approach to many types of applications. It is an exciting method of doing geographic analysis when desiring to rapidly examine existing geographically related data or to display specially prepared data and base maps for publication. One such program is the interactive thematic mapping system called CARTE, which combines polygonal base maps with statistical data to produce shaded maps using a variety of shading symbolisms on a variety of output devices. A polygonal base map is one where geographic entities are described by points, lines, or polygons. It is combined with geocoded data to produce special subject or thematic maps. Shading symbolisms include texture shading for areas, varying widths for lines, and scaled symbols for points. Output devices include refresh and storage CRTs and auxiliary Calcomp or COM hardcopy. The system is designed to aid in the quick display of spatial data and in detailed map design.

  5. 毕赤酵母表面展示米黑根毛霉脂肪酶发酵条件%Production of Rhizomucor miehei Lipase Displayed on Pichia pastoris Cell Surface Under Lab-scale Culture

    Institute of Scientific and Technical Information of China (English)

    李华珍; 陈明祥; 胡辉; 叶燕锐; 余永强; 林影

    2011-01-01

    Several key factors which affect the production of Rhizomucor miehei lipase (RML) displayed on cell surface of Pichia pastoris were studied by 50 L scale fermentation. Results indicated that the optimum initial induction cell density ( OD6oo) was around 250 ; the optimum pH during induction phase was 6.0; reducing temperature during induction phase resulted in increase of specific productivity by 1.56 and 1.32 folds at 20℃and 25℃ respectively, compared to that of 30℃. Under the above optimum conditions, the RML activity and productivity reached 275.0 U/ g and 462.5 U/(L· h), increased by 54.5% compared to that of initial conditions. Methanol/Sorbitol co -feeding strategy was also studied. By implementing this strategy and setting the sorbitol and methanol mix rate at 1:4 and 1: 5, the RML productivity at 12h was 1.7 times of that by sole carbon resource.%在50L发酵罐水平对影响毕赤酵母表面展示米黑根毛霉脂肪酶(Rhizomucor miehei lipase。RML)发酵因素进行了研究。最适初始菌体密度(OD600)约为250,最佳诱导pH为6.0;降低诱导温度能有效提高单位甲醇转化率,诱导温度为20℃和25℃时的转化率分别是30℃的1.56倍和1.32倍。在以上最适条件下单位于菌体展示酶活及生产强度分别达275.0U/g和462.5U/(L·h),比优化前提高54.5%。研究发现,山梨醇和甲醇混合补料有效地提高了诱导前期(t=12h)的产酶效率,当山梨醇和甲醇混合比例为1:4及1:5是以甲醇为单一碳源时酶活力的1.7倍。

  6. Polydendrocytes display large lineage plasticity following focal cerebral ischemia.

    Directory of Open Access Journals (Sweden)

    Pavel Honsa

    Full Text Available Polydendrocytes (also known as NG2 glial cells constitute a fourth major glial cell type in the adult mammalian central nervous system (CNS that is distinct from other cell types. Although much evidence suggests that these cells are multipotent in vitro, their differentiation potential in vivo under physiological or pathophysiological conditions is still controversial.To follow the fate of polydendrocytes after CNS pathology, permanent middle cerebral artery occlusion (MCAo, a commonly used model of focal cerebral ischemia, was carried out on adult NG2creBAC:ZEG double transgenic mice, in which enhanced green fluorescent protein (EGFP is expressed in polydendrocytes and their progeny. The phenotype of the EGFP(+ cells was analyzed using immunohistochemistry and the patch-clamp technique 3, 7 and 14 days after MCAo. In sham-operated mice (control, EGFP(+ cells in the cortex expressed protein markers and displayed electrophysiological properties of polydendrocytes and oligodendrocytes. We did not detect any co-labeling of EGFP with neuronal, microglial or astroglial markers in this region, thus proving polydendrocyte unipotent differentiation potential under physiological conditions. Three days after MCAo the number of EGFP(+ cells in the gliotic tissue dramatically increased when compared to control animals, and these cells displayed properties of proliferating cells. However, in later phases after MCAo a large subpopulation of EGFP(+ cells expressed protein markers and electrophysiological properties of astrocytes that contribute to the formation of glial scar. Importantly, some EGFP(+ cells displayed membrane properties typical for neural precursor cells, and moreover these cells expressed doublecortin (DCX--a marker of newly-derived neuronal cells. Taken together, our data indicate that polydendrocytes in the dorsal cortex display multipotent differentiation potential after focal ischemia.

  7. Striations in Plasma Display Panel

    Institute of Scientific and Technical Information of China (English)

    OUYANG Ji-Ting; CAO Jing; MIAO Jin-Song

    2005-01-01

    @@ The phenomenon of striation has been investigated experimentally in a macroscopic ac-plasma display panel (PDP). The relationship between the characteristics of striation and the operation conditions including voltage, frequency, rib, and electrode configuration, etc is obtained experimentally. The origin of the striations is considered to be the ionization waves in the transient positive column near the dielectric surface in the anode area during the discharge, and the perturbation is caused by resonance kinetic effects in inert gas.

  8. Modern Display Technologies and Applications

    Science.gov (United States)

    1982-01-01

    conventional tubes, LSI circuitry offers the possibility of correcting some of the deficiencies in electron-optic perform- ance and may lead to acceptable...certain ceramic materials such as PLZT (lead lanthanum zirconate titanate) can be utilized for display applications. PLZT is transparent in the visible...consuming power (3.8.12). 3.8.4.2 State of development, Magnetic particles have been made of polyethylene with powdered Strontium ferrite as a filler

  9. GridOrbit public display

    DEFF Research Database (Denmark)

    Ramos, Juan David Hincapie; Tabard, Aurélien; Bardram, Jakob

    2010-01-01

    We introduce GridOrbit, a public awareness display that visualizes the activity of a community grid used in a biology laboratory. This community grid executes bioin-formatics algorithms and relies on users to donate CPU cycles to the grid. The goal of GridOrbit is to create a shared awareness about...... people comment on projects. Our work explores the usage of interactive technologies as enablers for the appropriation of an otherwise invisible infrastructure....

  10. Proof nets for display logic

    CERN Document Server

    Moot, Richard

    2007-01-01

    This paper explores several extensions of proof nets for the Lambek calculus in order to handle the different connectives of display logic in a natural way. The new proof net calculus handles some recent additions to the Lambek vocabulary such as Galois connections and Grishin interactions. It concludes with an exploration of the generative capacity of the Lambek-Grishin calculus, presenting an embedding of lexicalized tree adjoining grammars into the Lambek-Grishin calculus.

  11. Multiview synthesis for autostereoscopic displays

    Science.gov (United States)

    Dane, Gökçe.; Bhaskaran, Vasudev

    2013-09-01

    Autostereoscopic (AS) displays spatially multiplex multiple views, providing a more immersive experience by enabling users to view the content from different angles without the need of 3D glasses. Multiple views could be captured from multiple cameras at different orientations, however this could be expensive, time consuming and not applicable to some applications. The goal of multiview synthesis in this paper is to generate multiple views from a stereo image pair and disparity map by using various video processing techniques including depth/disparity map processing, initial view interpolation, inpainting and post-processing. We specifically emphasize the need for disparity processing when there is no depth information is available that is associated with the 2D data and we propose a segmentation based disparity processing algorithm to improve disparity map. Furthermore we extend the texture based 2D inpainting algorithm to 3D and further improve the hole-filling performance of view synthesis. The benefit of each step of the proposed algorithm is demonstrated with comparison to state of the art algorithms in terms of visual quality and PSNR metric. Our system is evaluated in an end-to-end multi view synthesis framework where only stereo image pair is provided as input to the system and 8 views are outputted and displayed in 8-view Alioscopy AS display.

  12. Screening of p53 Promoter Binding Proteins by Phage Display in Liver Cells%筛选肝细胞p53启动子结合蛋白的噬菌体表面展示技术

    Institute of Scientific and Technical Information of China (English)

    吴顺华; 钟彦伟; 成军; 郑玉建

    2011-01-01

    目的 筛选p53启动子结合蛋白,探讨p53对肝细胞调节的分子生物学机制.方法 应用噬菌体表面展示技术,以p53作为固相筛选分子,对噬菌体人肝细胞cDNA文库进行5轮"吸附-洗脱-扩增"过程,经噬斑的PCR扩增后,构建克隆载体,最后对所筛选克隆进行DNA测序和生物信息学分析.结果 噬菌体经5轮富集后,成功构建了噬菌体p53展示文库,该文库包含与p53结合的肝细胞蛋白有17种.其中2个克隆编码未知功能蛋白;其余的克隆分别编码人α2HS糖蛋白(AHSG)、人D4、锌指蛋白家族2(DPF2)、凋亡反应锌指基因(REQ)、Ⅰ型纤溶酶原激活物抑制因子结合蛋白、氨基甲酰磷酸合成酶1、磷酸化酶激酶、酪氨酸氨基转移酶、酪蛋白激酶细胞周期素激酶、谷胱甘肽过氧化物酶、组蛋白结合蛋白3、跨膜蛋白2、血清黏蛋白2、有核红细胞白血病病毒癌基因2、毛细管扩张失调症突变基因,涉及到细胞发育和代谢、细胞外基质、细胞周期调控、信号转导、细胞凋亡以及原癌基因.结论 噬菌体表面展示技术成功筛选出了肝细胞p53启动子结合的多种类型和功能蛋白.%Objective To investigate the p53 regulation mechanisms in liver by screening binding proteins of p53 promoter using phage display. Methods Biotin p53 promoter DNA was incubated with a phage display cDNA library that expressed a library of human liver proteins on the surface of bacteriophage T7. Unbound phages were washed off and the phages bound to p53 were amplified. Positive plaqe were amplified by PCR and cloned into pGEM-Teasy vector in order to perform DNA sequence analysis and subsequently computer blasting analysis. Results Positive phage-displayed proteins binding with p53 promoter were enriched after five rounds of biopanning . Seventeen kinds of proteins might have relevance to p53,including Homo sapiens alpha-2-HS-glycoprotein (AHSG);Homo sapiens D4, zinc and double PHD

  13. Gestures to Intuitively Control Large Displays

    NARCIS (Netherlands)

    Fikkert, F.W.; Vet, van der P.E.; Rauwerda, H.; Breit, T.; Nijholt, A.; Sales Dias, M.; Gibet, S.; Wanderley, M.W.; Bastos, R.

    2009-01-01

    Large displays are highly suited to support discussions in empirical science. Such displays can display project results on a large digital surface to feed the discussion. This paper describes our approach to closely involve multidisciplinary omics scientists in the design of an intuitive display con

  14. 利用细菌表面展示技术构建镉耐受性的重组根瘤菌%Construction of Recombinant Rhizobial Strains with Enhanced Tolerance to Cadmium Using Bacterial Cell Surface Display

    Institute of Scientific and Technical Information of China (English)

    方彩云; 谢福莉; 付玮; 李友国

    2011-01-01

    This study aims to construct recombinant thizobia strains with enhanced tolerance to cadmium by using bacterial cell surface displaying technique, and explore their application in bioremediation of Ca2+ contamintated soil under symbiosis with the host leguminous plants.Two recombinant plasmids, pTNIM and pBLIM, were constructed based on the components of bacterial surface display functional gene InaXN, monkey metallothionein tetramer MT4α and two promoters of PnifH and Plac.By tri-parent matting technique, two recombinant plasmids were transferred into recipient Sinorhizobim fredii HN01.An inducible expression type of recombinant thizobium HNOI (pTNIM) and a constitutive expression type recombinant thizobium HN01 (pBLIM) were obtained, respectively.Under pure culture conditions, the adsorption capacity and tolerance ability of the recombinant and recipient strains to cadmium were comparatively examined, which showed the capacities for both aspects of HN01 (pBLIM) were significantly improved, as such their cadmium-adsorption capacities were increased by over two folds.The results also indicated that the recombinant thizobia demonstrated very good static-adsorption characteristics.Lastly, pot plant experiments showed that the amount of Cd2+ absorbed in root nodules and roots of soybean plants inoculated with HN01(pTNIM) was remarkably higher than that with wild-type strain HN01.Fig 5, Tab 1, Ref 22%通过细菌表面展示功能基因InaXN、猴金属硫蛋白α亚基四聚体MT4α和两种启动子PnifH及Plac元件,分别构建两个重组质粒pTNIM和pBLIM.进一步通过三亲本杂交,将重组质粒分别导入费氏中华根瘤菌HN01,分别构建获得诱导表达型重组菌HN01(pTNIM)和组成表达型重组菌HN01(pBLIM).在培养条件下比较测定重组菌和出发菌对镉的吸附能力和耐受性,结果表明,组成表达型重组菌HN01(pBLIM)在上述两个方面的能力得到明显增强,对镉的吸附富集能力提高了2倍.研究还发

  15. Review of Defense Display Research Programs

    Science.gov (United States)

    2001-01-01

    Programs Flat Panel Autostereoscopic N-perspective 3D High Definition DMD Digital Projector Light Piping & Quantum Cavity Displays Solid State Laser...Megapixel Displays • Size Commonality • 67 % Weight Reduction • > 200 sq. in. per Display 20-20 Vision Simulators True 3D , sparse symbols Foldable Display...megapixel 2D and True 3D Display Technology 25M & T3D FY02-FY06 New service thrusts

  16. Recent Trend in Development of Olfactory Displays

    Science.gov (United States)

    Yanagida, Yasuyuki

    An olfactory display is a device that generates scented air with desired concentration of aroma, and delivers it to the user's olfactory organ. In this article, the nature of olfaction is briefly described from the view point of how to configure olfactory displays. Next, component technologies to compose olfactory displays, i.e., making scents and delivering scents, are categorized. Several existing olfactory display systems are introduced to show the current status of research and development of olfactory displays.

  17. A novel screening system for claudin binder using baculoviral display.

    Directory of Open Access Journals (Sweden)

    Hideki Kakutani

    Full Text Available Recent progress in cell biology has provided new insight into the claudin (CL family of integral membrane proteins, which contains more than 20 members, as a target for pharmaceutical therapy. Few ligands for CL have been identified because it is difficult to prepare CL in an intact form. In the present study, we developed a method to screen for CL binders by using the budded baculovirus (BV display system. CL4-displaying BV interacted with a CL4 binder, the C-terminal fragment of Clostridium perfringens enterotoxin (C-CPE, but it did not interact with C-CPE that was mutated in its CL4-binding region. C-CPE did not interact with BV and CL1-displaying BV. We used CL4-displaying BV to select CL4-binding phage in a mixture of a scFv-phage and C-CPE-phage. The percentage of C-CPE-phage in the phage mixture increased from 16.7% before selection to 92% after selection, indicating that CL-displaying BV may be useful for the selection of CL binders. We prepared a C-CPE phage library by mutating the functional amino acids. We screened the library for CL4 binders by affinity to CL4-displaying BV, and we found that the novel CL4 binders modulated the tight-junction barrier. These findings indicate that the CL-displaying BV system may be a promising method to produce a novel CL binder and modulator.

  18. Optical display for radar sensing

    Science.gov (United States)

    Szu, Harold; Hsu, Charles; Willey, Jefferson; Landa, Joseph; Hsieh, Minder; Larsen, Louis V.; Krzywicki, Alan T.; Tran, Binh Q.; Hoekstra, Philip; Dillard, John T.; Krapels, Keith A.; Wardlaw, Michael; Chu, Kai-Dee

    2015-05-01

    Boltzmann headstone S = kB Log W turns out to be the Rosette stone for Greek physics translation optical display of the microwave sensing hieroglyphics. The LHS is the molecular entropy S measuring the degree of uniformity scattering off the sensing cross sections. The RHS is the inverse relationship (equation) predicting the Planck radiation spectral distribution parameterized by the Kelvin temperature T. Use is made of the conservation energy law of the heat capacity of Reservoir (RV) change T Δ S = -ΔE equals to the internal energy change of black box (bb) subsystem. Moreover, an irreversible thermodynamics Δ S > 0 for collision mixing toward totally larger uniformity of heat death, asserted by Boltzmann, that derived the so-called Maxwell-Boltzmann canonical probability. Given the zero boundary condition black box, Planck solved a discrete standing wave eigenstates (equation). Together with the canonical partition function (equation) an average ensemble average of all possible internal energy yielded the celebrated Planck radiation spectral (equation) where the density of states (equation). In summary, given the multispectral sensing data (equation), we applied Lagrange Constraint Neural Network (LCNN) to solve the Blind Sources Separation (BSS) for a set of equivalent bb target temperatures. From the measurements of specific value, slopes and shapes we can fit a set of Kelvin temperatures T's for each bb targets. As a result, we could apply the analytical continuation for each entropy sources along the temperature-unique Planck spectral curves always toward the RGB color temperature display for any sensing probing frequency.

  19. Simulated monitor display for CCTV

    Energy Technology Data Exchange (ETDEWEB)

    Steele, B.J.

    1982-01-01

    Two computer programs have been developed which generate a two-dimensional graphic perspective of the video output produced by a Closed Circuit Television (CCTV) camera. Both programs were primarily written to produce a graphic display simulating the field-of-view (FOV) of a perimeter assessment system as seen on a CCTV monitor. The original program was developed for use on a Tektronix 4054 desktop computer; however, the usefulness of this graphic display program led to the development of a similar program for a Hewlett-Packard 9845B desktop computer. After entry of various input parameters, such as, camera lens and orientation, the programs automatically calculate and graphically plot the locations of various items, e.g., fences, an assessment zone, running men, and intrusion detection sensors. Numerous special effects can be generated to simulate such things as roads, interior walls, or sides of buildings. Other objects can be digitized and entered into permanent memory similar to the running men. With this type of simulated monitor perspective, proposed camera locations with respect to fences and a particular assessment zone can be rapidly evaluated without the costly time delays and expenditures associated with field evaluation.

  20. Induction of cell cycle arrest at G1 and S phases and cAMP-dependent differentiation in C6 glioma by low concentration of cycloheximide

    Directory of Open Access Journals (Sweden)

    Zhang Samuel S

    2010-12-01

    Full Text Available Abstract Background Differentiation therapy has been shown effective in treatment of several types of cancer cells and may prove to be effective in treatment of glioblastoma multiforme, the most common and most aggressive primary brain tumor. Although extensively used as a reagent to inhibit protein synthesis in mammalian cells, whether cycloheximide treatment leads to glioma cell differentiation has not been reported. Methods C6 glioma cell was treated with or without cycloheximide at low concentrations (0.5-1 μg/ml for 1, 2 and 3 days. Cell proliferation rate was assessed by direct cell counting and colony formation assays. Apoptosis was assessed by Hoechst 33258 staining and FACS analysis. Changes in several cell cycle regulators such as Cyclins D1 and E, PCNA and Ki67, and several apoptosis-related regulators such as p53, p-JNK, p-AKT, and PARP were determined by Western blot analysis. C6 glioma differentiation was determined by morphological characterization, immunostaining and Western blot analysis on upregulation of GFAP and o p-STAT3 expression, and upregulation of intracellular cAMP. Results Treatment of C6 cell with low concentration of cycloheximide inhibited cell proliferation and depleted cells at both G2 and M phases, suggesting blockade at G1 and S phases. While no cell death was observed, cells underwent profound morphological transformation that indicated cell differentiation. Western blotting and immunostaining analyses further indicated that changes in expression of several cell cycle regulators and the differentiation marker GFAP were accompanied with cycloheximide-induced cell cycle arrest and cell differentiation. Increase in intracellular cAMP, a known promoter for C6 cell differentiation, was found to be elevated and required for cycloheximide-promoted C6 cell differentiation. Conclusion Our results suggest that partial inhibition of protein synthesis in C6 glioma by low concentration of cycloheximide induces cell cycle

  1. Reconfigurable Braille display with phase change locking

    Science.gov (United States)

    Soule, Cody W.; Lazarus, Nathan

    2016-07-01

    Automatically updated signs and displays for sighted people are common in today’s world. However, there is no cheap, low power equivalent available for the blind. This work demonstrates a reconfigurable Braille cell using the solid-to-liquid phase change of a low melting point alloy as a zero holding power locking mechanism. The device is actuated with the alloy in the liquid state, and is then allowed to solidify to lock the Braille dot in the actuated position. A low-cost manufacturing process is developed that includes molding of a rigid silicone to create pneumatic channels, and bonding of a thin membrane of a softer silicone on the surface for actuation. A plug of Field’s metal (melting point 62 °C) is placed in the pneumatic channels below each Braille dot to create the final device. The device is well suited for low duty cycle operation in applications such as signs, and is able to maintain its state indefinitely without additional power input. The display requires a pneumatic pressure of only 24 kPa for actuation, and reconfiguration has been demonstrated in less than a minute and a half.

  2. Optimal screening of surface-displayed polypeptide libraries.

    Science.gov (United States)

    Boder, E T; Wittrup, K D

    1998-01-01

    Cell surface display of polypeptide libraries combined with flow cytometric cell sorting presents remarkable potential for enhancement of protein-ligand recognition properties. To maximize the utility of this approach, screening and purification conditions must be optimized to take full advantage of the quantitative feature of this technique. In particular, discrimination of improved library mutants from an excess of wild-type polypeptides is dependent upon an effective screening methodology. Fluorescence discrimination profiles for improved library mutants were derived from a mathematical model of expected cell fluorescence intensities for polypeptide libraries screened with fluorescent ligand. Profiles for surface-displayed libraries under equilibrium or kinetic screening conditions demonstrate distinct discrimination optima from which optimal equilibrium and kinetic screening parameters were derived. In addition, a statistical model of low cytometrically analyzed cell populations indicates the importance of low-stringency sorting followed by amplification through regrowth and resorting at increased stringency. This analysis further yields quantitative recommendations for cell-sorting stringency.

  3. Simplified Night Sky Display System

    Science.gov (United States)

    Castellano, Timothy P.

    2010-01-01

    A document describes a simple night sky display system that is portable, lightweight, and includes, at most, four components in its simplest configuration. The total volume of this system is no more than 10(sup 6) cm(sup 3) in a disassembled state, and weighs no more than 20 kilograms. The four basic components are a computer, a projector, a spherical light-reflecting first surface and mount, and a spherical second surface for display. The computer has temporary or permanent memory that contains at least one signal representing one or more images of a portion of the sky when viewed from an arbitrary position, and at a selected time. The first surface reflector is spherical and receives and reflects the image from the projector onto the second surface, which is shaped like a hemisphere. This system may be used to simulate selected portions of the night sky, preserving the appearance and kinesthetic sense of the celestial sphere surrounding the Earth or any other point in space. These points will then show motions of planets, stars, galaxies, nebulae, and comets that are visible from that position. The images may be motionless, or move with the passage of time. The array of images presented, and vantage points in space, are limited only by the computer software that is available, or can be developed. An optional approach is to have the screen (second surface) self-inflate by means of gas within the enclosed volume, and then self-regulate that gas in order to support itself without any other mechanical support.

  4. A decade of yeast surface display technology: where are we now?

    Science.gov (United States)

    Pepper, Lauren R; Cho, Yong Ku; Boder, Eric T; Shusta, Eric V

    2008-02-01

    Yeast surface display has become an increasingly popular tool for protein engineering and library screening applications. Recent advances have greatly expanded the capability of yeast surface display, and are highlighted by cell-based selections, epitope mapping, cDNA library screening, and cell adhesion engineering. In this review, we discuss the state-of-the-art yeast display methodologies and the rapidly expanding set of applications afforded by this technology.

  5. A simplified procedure for antibody engineering by yeast surface display: Coupling display levels and target binding by ribosomal skipping.

    Science.gov (United States)

    Grzeschik, Julius; Hinz, Steffen C; Könning, Doreen; Pirzer, Thomas; Becker, Stefan; Zielonka, Stefan; Kolmar, Harald

    2017-02-01

    Yeast surface display is a valuable, widely used method for protein engineering. However, current yeast display applications rely on the staining of epitope tags in order to verify full-length presentation of the protein of interest on the cell surface. We aimed at developing a modified yeast display approach that relies on ribosomal skipping, thereby enabling the translation of two proteins from one open reading frame and, in that manner, generating an intracellular fluorescence signal. This improved setup is based on a 2A sequence that is encoded between the protein to be displayed and a gene for green fluorescent protein (GFP). The intracellular GFP fluorescence signal of yeast cells correlates with full-length protein presentation and omits the need for the immunofluorescence detection of epitope tags. For method validation, shark-derived IgNAR variable domains (vNAR) were subjected to affinity maturation using the 2A-GFP system. Yeast library screening of full-length vNAR variants which were detected via GFP expression yielded the same high-affinity binder that had previously been isolated by our group using the conventional epitope tag-based display format. The presented method obviates the need for additional immunofluorescence cell staining, offering an easy and cost-friendly alternative to conventional epitope tag detections.

  6. Full resolution hologram-like autostereoscopic display

    Science.gov (United States)

    Eichenlaub, Jesse B.; Hutchins, Jamie

    1995-01-01

    Under this program, Dimension Technologies Inc. (DTI) developed a prototype display that uses a proprietary illumination technique to create autostereoscopic hologram-like full resolution images on an LCD operating at 180 fps. The resulting 3D image possesses a resolution equal to that of the LCD along with properties normally associated with holograms, including change of perspective with observer position and lack of viewing position restrictions. Furthermore, this autostereoscopic technique eliminates the need to wear special glasses to achieve the parallax effect. Under the program a prototype display was developed which demonstrates the hologram-like full resolution concept. To implement such a system, DTI explored various concept designs and enabling technologies required to support those designs. Specifically required were: a parallax illumination system with sufficient brightness and control; an LCD with rapid address and pixel response; and an interface to an image generation system for creation of computer graphics. Of the possible parallax illumination system designs, we chose a design which utilizes an array of fluorescent lamps. This system creates six sets of illumination areas to be imaged behind an LCD. This controlled illumination array is interfaced to a lenticular lens assembly which images the light segments into thin vertical light lines to achieve the parallax effect. This light line formation is the foundation of DTI's autostereoscopic technique. The David Sarnoff Research Center (Sarnoff) was subcontracted to develop an LCD that would operate with a fast scan rate and pixel response. Sarnoff chose a surface mode cell technique and produced the world's first large area pi-cell active matrix TFT LCD. The device provided adequate performance to evaluate five different perspective stereo viewing zones. A Silicon Graphics' Iris Indigo system was used for image generation which allowed for static and dynamic multiple perspective image rendering

  7. 4-Methoxydalbergione suppresses growth and induces apoptosis in human osteosarcoma cells in vitro and in vivo xenograft model through down-regulation of the JAK2/STAT3 pathway.

    Science.gov (United States)

    Park, Kyung-Ran; Yun, Hyung-Mun; Quang, Tran-Hong; Oh, Hyuncheol; Lee, Dong-Sung; Auh, Q-Schick; Kim, Eun-Cheol

    2016-02-09

    Although the heartwood of Dalbergia odorifera T. Chen (Leguminosae) is an important source of traditional Korean and Chinese medicines, the effects of novel compound methoxydalbergione (4-MD) isolated from Dalbergia odorifera was not reported. Herein, we investigated the effects of the 4-MD in vitro and in vivo against osteosarcoma cells and its molecular mechanisms. 4-MD inhibited the proliferation of osteosarcoma cells and induced apoptosis as evidenced by Annexin V + and TUNEL + cells. This apoptosis was accompanied by upregulation of apoptotic proteins (procaspase-3 and PARP), but downregulation of anti-apoptotic proteins (Bcl-2, Bcl-xL, and Survivin). 4-MD inhibited phosphorylation of JAK2 and STAT3 with the inactivation of mitogen-activated protein kinases (MAPKs) and CREB, and the upregulation of PTEN in osteosarcoma cells. Importantly, 4-MD reduced colony formation in soft agar and inhibited tumor growth in mice xenograft model in association with the reduced expression of PCNA, Ki67, p-STAT3, and Survivin. Taken together, the present study for the first time demonstrates that 4-MD exerts in vitro and in vivo anti-proliferative effects against osteosarcoma cells through the inhibition of the JAK2/STAT3 pathway, and suggest the potential for therapeutic application of 4-MD in the treatment of osteosarcoma.

  8. IL-22 promoted CD3+ T cell infiltration by IL-22R induced STAT3 phosphorylation in murine acute graft versus host disease target organs after allogeneic bone marrow transplantation.

    Science.gov (United States)

    Zhao, Kai; Ruan, Suhong; Tian, Yu; Zhao, Dongmei; Chen, Chong; Pan, Bin; Yan, Zhiling; Yin, Lingling; Zhu, Shengyun; Xu, Kailin

    2016-10-01

    Graft versus host disease (GVHD) is a life threatening complication of bone marrow stem cell transplantation, in which considerable numbers of proinflammatory cytokines secreted by allo-reactive donor T cells are involved. We and other previous studies have found that interleukin-22 (IL-22) was able to aggravate the target organs damage of GVHD. However, the mechanism and the signal pathway of IL-22 in murine acute GVHD was not clear. Here, we observed that compared with GVHD group, more serious pathological damage and more CD3(+) T cells infiltrated in GVHD target organs were detected in the mice injected with IL-22. Meanwhile, transcription factor T-bet, RORγt and AhR respectively associated with Th1, Th17 and Th22 cells changed in varying degrees in different GVHD target organs. Furthermore, the increased expression of IL-22R and its downstream protein P-STAT3 were detected in GVHD mice with IL-22 treated. These results suggested that the pathological role of IL-22 in GVHD target organs contribute to exogenous injected IL-22 as well as secreted IL-22 from the infiltrated allo-reactive effector T cells. In addition, the IL-22R-STAT3 pathway may play important role in GVHD tissue injury and target this way may yield new approaches for reduction of GVHD.

  9. OZ: An Innovative Primary Flight Display Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The proposed SBIR project will develop OZ, an innovative primary flight display for aircraft. The OZ display, designed from "first principles" of vision science,...

  10. Holographic Waveguided See-Through Display Project

    Data.gov (United States)

    National Aeronautics and Space Administration — To address the NASA need for lightweight, space suit-mounted displays, Luminit proposes a novel Holographic Waveguided See-Through Display. Our proposed Holographic...

  11. ProPath Display Process Overview

    Data.gov (United States)

    Department of Veterans Affairs — This API displays an overview of the process including the description, goals, associated roles (linked to their detailed information). It also displays all of the...

  12. Projection/Reflection Heads-up Display

    Data.gov (United States)

    National Aeronautics and Space Administration — To address the NASA need for an EVA information display device, Physical Optics Corporation (POC) proposes to develop a new Projection/Reflection Heads-up Display...

  13. Heterologous surface display on lactic acid bacteria: non-GMO alternative?

    Science.gov (United States)

    Zadravec, Petra; Štrukelj, Borut; Berlec, Aleš

    2015-01-01

    Lactic acid bacteria (LAB) are food-grade hosts for surface display with potential applications in food and therapy. Alternative approaches to surface display on LAB would avoid the use of recombinant DNA technology and genetically-modified organism (GMO)-related regulatory requirements. Non-covalent surface display of proteins can be achieved by fusing them to various cell-wall binding domains, of which the Lysine motif domain (LysM) is particularly well studied. Fusion proteins have been isolated from recombinant bacteria or from their growth medium and displayed on unmodified bacteria, enabling heterologous surface display. This was demonstrated on non-viable cells devoid of protein content, termed bacteria-like particles, and on various species of genus Lactobacillus. Of the latter, Lactobacillus salivarius ATCC 11741 was recently shown to be particularly amenable for LysM-mediated display. Possible regulatory implications of heterologous surface display are discussed, particularly those relevant for the European Union.

  14. Miniature Color Display Phase 4

    Science.gov (United States)

    1993-05-01

    36 Two Notch Filters Figure 22 Experimental Setup for Thermal Tests of MCD .................... 37 Figure 23 Thermal Test Results with all Cells...in the Transparent ............... 38 Figure 24 Thermal Test Results with All (Cells in the Absorbing State ........... 38 Figure 25 Angular...filter. C4-W r -- - - D, -- . ° . NIm Figure 22 Experimental setup for thermal tests of MCD. Thermocouples were cemented to the input and output surfaces

  15. Hyperglycemia enhances the proliferation of non-tumorigenic and malignant mammary epithelial cells through increased leptin/IGF1R signaling and activation of AKT/mTOR.

    Directory of Open Access Journals (Sweden)

    Rebecca Lopez

    Full Text Available Obesity and diabetes are associated with increased breast cancer risk and worse disease progression once cancer is diagnosed; however, the exact etiology behind these observations remains to be fully elucidated. Due to the global obesity/diabetes pandemic, it is imperative to understand how these diseases promote and enhance breast cancer and other common cancers. In this study we demonstrate that hyperglycemia promotes breast cancer by altering leptin/IGF1R and AKT/mTOR signaling. To our knowledge, we show for the first time that in breast epithelial cells, hyperglycemia alone directly impacts leptin signaling. Hyperglycemia increased proliferation of both non-tumorigenic and malignant mammary epithelial cells. These observations coincided with increased leptin receptor and IGF1R receptor, as well as, increased levels of GRB2, pJAK2, pSTAT3, pIRS1/2, pAKT, and p-mTOR. Moreover, pJAK2 was almost completely colocalized with leptin receptor under high glucose conditions. These results demonstrate how hyperglycemia can potentially increase the risk of breast cancer in premalignant lesions and enhance cancer progression in malignant cells.

  16. 27 CFR 6.83 - Product displays.

    Science.gov (United States)

    2010-04-01

    ... industry member of giving or selling product displays to a retailer does not constitute a means to induce... costs are excluded. (2) All product displays must bear conspicuous and substantial advertising matter on... address of the retailer may appear on the product displays. (3) The giving or selling of such...

  17. Emerging Large-Screen Display Technology

    Science.gov (United States)

    1992-11-01

    1255, Santa Clara, CA. 25. Williams, R. D., and F. Garcia, 1988, "A Real Time Autostereoscopic Multiplanar 3D Display System," Society for Information...K. Miyaji, 1989, " 3D Display using Laser and Moving Screen, Japan Display 1989, Paper P3-5. 27. Sterling, R. D., R. D. TeKolste, J. M. Haggerty, T. C

  18. Thermal modeling of laser-addressed liquid-crystal displays

    Science.gov (United States)

    Evans, K. E.; Nkansah, M. A.

    1990-06-01

    Optical-absorption calculations and finite-element methods are used to calculate time-dependent temperature profiles in two contrasting laser-addressed liquid-crystal displays. It is shown that the presence of conducting electrode layers has a significant effect on the temperature profiles both by affecting the optical-absorption characteristics of the cell and the resulting thermal conductivity. It is shown that efficient optical absorption does not necessarily result in the best cell-addressing performance.

  19. Advancement and applications of peptide phage display technology in biomedical science.

    Science.gov (United States)

    Wu, Chien-Hsun; Liu, I-Ju; Lu, Ruei-Min; Wu, Han-Chung

    2016-01-01

    Combinatorial phage library is a powerful research tool for high-throughput screening of protein interactions. Of all available molecular display techniques, phage display has proven to be the most popular approach. Screening phage-displayed random peptide libraries is an effective means of identifying peptides that can bind target molecules and regulate their function. Phage-displayed peptide libraries can be used for (i) B-cell and T-cell epitope mapping, (ii) selection of bioactive peptides bound to receptors or proteins, disease-specific antigen mimics, peptides bound to non-protein targets, cell-specific peptides, or organ-specific peptides, and (iii) development of peptide-mediated drug delivery systems and other applications. Targeting peptides identified using phage display technology may be useful for basic research and translational medicine. In this review article, we summarize the latest technological advancements in the application of phage-displayed peptide libraries to applied biomedical sciences.

  20. CERN students display their work

    CERN Multimedia

    Anaïs Schaeffer

    2011-01-01

    The first poster session by students working on the LHC experiments, organised by the LPCC, was a great success. Showcasing the talents of over a hundred young physicists from all over the world, it was an opportunity for everyone at CERN to check out the wide range of research work being done by the new generation of physicists at CERN.   At 5.30 p.m. on Wednesday 23 March, the first poster session by CERN students took place in Restaurant No.1, where no fewer than 87 posters went on public display. The students were split into 8 groups according to their research field* and all were on hand to answer the questions of an inquisitive audience. TH Department's Michelangelo Mangano, who is head of the LHC Physics Centre at CERN (LPCC) and is responsible for the initiative, confirms that nothing was left to chance, even the choice of date: "We wanted to make the most of the general enthusiasm around the winter conferences and the meeting of the LHC Experiments Committee to present the stud...

  1. Citizenship displayed by disabled people

    Directory of Open Access Journals (Sweden)

    Eliana Prado Carlino

    2010-12-01

    Full Text Available By investigating the processes by which successful teachers become activate citizens and by listening to the diversity and richness of their life and formation stories, this work became possible. Its aim is to display some of the utterances of two Down Syndrome individuals and their active-citizenship activities. Their stories were told in the reports of two teachers when describing their personal and professional history, and were considered to be an integral part of it. Thus, some of the utterances and perceptions with which these two individuals elaborate their references, their worldview and their active-citizenship activity are evidenced in this paper. This article is based on the language conceptions of Vygotsky and Bakhtin who defend the idea that the group and the social mentality are ingrain in the individual. Hence, the history of one person reveals that of many others, since there is a deep link between the individual and the social in the formation of a subjective worldview. As a result, it can be easily seen that the utterances expressed by the participants in this research cannot be considered strictly individual because enunciation is social in nature. Despite the fact that the utterances are those of individuals, they manifest a collective reality. This demonstrates the real advantages and possibilities that deficient people get from their participation and intervention in society.

  2. Inhibition of Stat3 activation suppresses caspase-3 and the ubiquitin-proteasome system, leading to preservation of muscle mass in cancer cachexia.

    Science.gov (United States)

    Silva, Kleiton Augusto Santos; Dong, Jiangling; Dong, Yanjun; Dong, Yanlan; Schor, Nestor; Tweardy, David J; Zhang, Liping; Mitch, William E

    2015-04-24

    Cachexia occurs in patients with advanced cancers. Despite the adverse clinical impact of cancer-induced muscle wasting, pathways causing cachexia are controversial, and clinically reliable therapies are not available. A trigger of muscle protein loss is the Jak/Stat pathway, and indeed, we found that conditioned medium from C26 colon carcinoma (C26) or Lewis lung carcinoma cells activates Stat3 (p-Stat3) in C2C12 myotubes. We identified two proteolytic pathways that are activated in muscle by p-Stat3; one is activation of caspase-3, and the other is p-Stat3 to myostatin, MAFbx/Atrogin-1, and MuRF-1 via CAAT/enhancer-binding protein δ (C/EBPδ). Using sequential deletions of the caspase-3 promoter and CHIP assays, we determined that Stat3 activation increases caspase-3 expression in C2C12 cells. Caspase-3 expression and proteolytic activity were stimulated by p-Stat3 in muscles of tumor-bearing mice. In mice with cachexia caused by Lewis lung carcinoma or C26 tumors, knock-out of p-Stat3 in muscle or with a small chemical inhibitor of p-Stat3 suppressed muscle mass losses, improved protein synthesis and degradation in muscle, and increased body weight and grip strength. Activation of p-Stat3 stimulates a pathway from C/EBPδ to myostatin and expression of MAFbx/Atrogin-1 and increases the ubiquitin-proteasome system. Indeed, C/EBPδ KO decreases the expression of MAFbx/Atrogin-1 and myostatin, while increasing muscle mass and grip strength. In conclusion, cancer stimulates p-Stat3 in muscle, activating protein loss by stimulating caspase-3, myostatin, and the ubiquitin-proteasome system. These results could lead to novel strategies for preventing cancer-induced muscle wasting.

  3. TWS119 upregulates CCR5 expression of γδT cells by inhibiting STAT3 phos-phorylation%TWS119通过抑制STAT3磷酸化促进γδT细胞CCR5的表达

    Institute of Scientific and Technical Information of China (English)

    徐晶; 孙蕾清; 陈永强; 郑璐; 吕小婷; 陈复兴; 刘军权; 周忠海

    2016-01-01

    Objective:To investigate the mechanisms of TWS119 induced CCR5 expression in hunman γδT cells. Methods:After treatment with various concentrations of TWS119 for 48h, the expression of CCR5 in γδT cells were detected by flow cytometry. The p-STAT3 and GAPDH expression were examined by Western blot analysis. Results: TWS119 could upregulate the expression of CCR5 in dose dependent manner. Western blot analysis revealed that TWS119 inhibit phosphorylation of STAT3,but had no significant impact on GAPDH. In addition, pretreatment of γδT cells with 0. 5 μmol/L STAT3 specific phosphorylation inhibitor Stattic could upregulate the expression of CCR5 and enhance the TWS119 induced CCR5 expression. Conclusion: TWS119 could upregulate CCR5 expression of γδT cells by inhibiting STAT3 phosphorylation in vitro.%目的:研究糖原合酶激酶-3β抑制剂4,6-二取代吡咯并嘧啶(TWS119)促进γδT细胞趋化因子受体CCR5表达的分子机制。方法:用TWS119诱导从健康人外周血中分离培养获得γδT细胞48 h后,用流式细胞仪检测γδT细胞CCR5的表达;Western blot检测GAPDH和p-STAT3的表达。结果:培养10 d后的γδT细胞纯度达到85.79%±5.01%。 TWS119浓度在0~8.0μmol/L范围内能显著促进趋化因子受体CCR5的表达,且剂量依赖性抑制STAT3的磷酸化。同时,用STAT3磷酸化抑制剂Stattic (0.5μmol/L)预处理γδT细胞也可以促进CCR5的表达,并能协同增强TWS119促进趋化因子受体CCR5的表达。结论:TWS119通过抑制STAT3磷酸化促进γδT细胞CCR5的表达。

  4. Military display market segment: wearable and portable

    Science.gov (United States)

    Desjardins, Daniel D.; Hopper, Darrel G.

    2003-09-01

    The military display market (MDM) is analyzed in terms of one of its segments, wearable and portable displays. Wearable and portable displays are those embedded in gear worn or carried by warfighters. Categories include hand-mobile (direct-view and monocular/binocular), palm-held, head/helmet-mounted, body-strapped, knee-attached, lap-born, neck-lanyard, and pocket/backpack-stowed. Some 62 fielded and developmental display sizes are identified in this wearable/portable MDM segment. Parameters requiring special consideration, such as weight, luminance ranges, light emission, viewing angles, and chromaticity coordinates, are summarized and compared. Ruggedized commercial versus commercial off-the-shelf designs are contrasted; and a number of custom displays are also found in this MDM category. Display sizes having aggregate quantities of 5,000 units or greater or having 2 or more program applications are identified. Wearable and portable displays are also analyzed by technology (LCD, LED, CRT, OLED and plasma). The technical specifications and program history of several high-profile military programs are discussed to provide a systems context for some representative displays and their function. As of August 2002 our defense-wide military display market study has documented 438,882 total display units distributed across 1,163 display sizes and 438 weapon systems. Wearable and portable displays account for 202,593 displays (46% of total DoD) yet comprise just 62 sizes (5% of total DoD) in 120 weapons systems (27% of total DoD). Some 66% of these wearable and portable applications involve low information content displays comprising just a few characters in one color; however, there is an accelerating trend towards higher information content units capable of showing changeable graphics, color and video.

  5. Microspheres in Plasma Display Panels

    Science.gov (United States)

    2006-01-01

    Filling small bubbles of molten glass with gases is just as difficult as it sounds, but the technical staff at NASA is not known to shy away from a difficult task. When Microsphere Systems, Inc. (MSI), of Ypsilanti, Michigan, and Imaging Systems Technology, Inc. (IST), of Toledo, Ohio, were trying to push the limits of plasma displays but were having difficulty with the designs, NASA s Glenn Garrett Morgan Commercialization Initiative (GMCI) assembled key personnel at Glenn Research Center and Ohio State University for a brainstorming session to come up with a solution for the companies. They needed a system that could produce hollow, glass micro-sized spheres (microspheres) that could be filled with a variety of gasses. But the extremely high temperature required to force the micro-sized glass bubbles to form at the tip of a metal nozzle resulted in severe discoloration of the microspheres. After countless experiments on various glass-metal combinations, they had turned to the GMCI for help. NASA experts in advanced metals, ceramics, and glass concluded that a new design approach was necessary. The team determined that what was needed was a phosphate glass composition that would remain transparent, and they went to work on a solution. Six weeks later, using the design tips from the NASA team, Tim Henderson, president of MSI, had designed a new system in which all surfaces in contact with the molten glass would be ceramic instead of metal. Meanwhile, IST was able to complete a Phase I Small Business Innovation Research (SBIR) grant supported by the National Science Foundation (NSF) and supply a potential customer with samples of the microspheres for evaluation as filler materials for high-performance insulations.

  6. STAT3 signaling pathway is necessary for cell survival and tumorsphere forming capacity in ALDH{sup +}/CD133{sup +} stem cell-like human colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Li, E-mail: lin.796@osu.edu [Center for Childhood Cancer, The Research Institute at Nationwide Children' s Hospital, Department of Pediatrics, Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43205 (United States); Division of Cardiology, Department of Internal Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030 (China); Fuchs, James; Li, Chenglong [Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, Columbus, OH 43210 (United States); Olson, Veronica [Center for Childhood Cancer, The Research Institute at Nationwide Children' s Hospital, Department of Pediatrics, Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43205 (United States); Bekaii-Saab, Tanios [Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43210 (United States); Lin, Jiayuh, E-mail: lin.674@osu.edu [Center for Childhood Cancer, The Research Institute at Nationwide Children' s Hospital, Department of Pediatrics, Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43205 (United States)

    2011-12-16

    Highlights: Black-Right-Pointing-Pointer The phosphorylated or activated form of STAT3 was expressed in colon cancer stem-like cells. Black-Right-Pointing-Pointer STAT3 inhibitor, FLLL32 inhibits P-STAT3 and STAT3 target genes in colon cancer stem-like cells. Black-Right-Pointing-Pointer Inhibition of STAT3 resulted in decreased cell viability and reduced numbers of tumorspheres. Black-Right-Pointing-Pointer STAT3 is required for survival and tumorsphere forming capacity in colon cancer stem-like cells. Black-Right-Pointing-Pointer Targeting STAT3 in cancer stem-like cells may offer a novel treatment approach for colon cancer. -- Abstract: Persistent activation of Signal Transducers and Activators of Transcription 3 (STAT3) is frequently detected in colon cancer. Increasing evidence suggests the existence of a small population of colon cancer stem or cancer-initiating cells may be responsible for tumor initiation, metastasis, and resistance to chemotherapy and radiation. Whether STAT3 plays a role in colon cancer-initiating cells and the effect of STAT3 inhibition is still unknown. Flow cytometry was used to isolate colon cancer stem-like cells from three independent human colon cancer cell lines characterized by both aldehyde dehydrogenase (ALDH)-positive and CD133-positive subpopulation (ALDH{sup +}/CD133{sup +}). The effects of STAT3 inhibition in colon cancer stem-like cells were examined. The phosphorylated or activated form of STAT3 was expressed in colon cancer stem-like cells and was reduced by a STAT3-selective small molecular inhibitor, FLLL32. FLLL32 also inhibited the expression of potential STAT3 downstream target genes in colon cancer stem-like cells including survivin, Bcl-XL, as well as Notch-1, -3, and -4, which may be involved in stem cell function. Furthermore, FLLL32 inhibited cell viability and tumorsphere formation as well as induced cleaved caspase-3 in colon cancer stem-like cells. FLLL32 is more potent than curcumin as evidenced with lower

  7. Cancer stem cells display extremely large evolvability: alternating plastic and rigid networks as a potential Mechanism: network models, novel therapeutic target strategies, and the contributions of hypoxia, inflammation and cellular senescence.

    Science.gov (United States)

    Csermely, Peter; Hódsági, János; Korcsmáros, Tamás; Módos, Dezső; Perez-Lopez, Áron R; Szalay, Kristóf; Veres, Dániel V; Lenti, Katalin; Wu, Ling-Yun; Zhang, Xiang-Sun

    2015-02-01

    Cancer is increasingly perceived as a systems-level, network phenomenon. The major trend of malignant transformation can be described as a two-phase process, where an initial increase of network plasticity is followed by a decrease of plasticity at late stages of tumor development. The fluctuating intensity of stress factors, like hypoxia, inflammation and the either cooperative or hostile interactions of tumor inter-cellular networks, all increase the adaptation potential of cancer cells. This may lead to the bypass of cellular senescence, and to the development of cancer stem cells. We propose that the central tenet of cancer stem cell definition lies exactly in the indefinability of cancer stem cells. Actual properties of cancer stem cells depend on the individual "stress-history" of the given tumor. Cancer stem cells are characterized by an extremely large evolvability (i.e. a capacity to generate heritable phenotypic variation), which corresponds well with the defining hallmarks of cancer stem cells: the possession of the capacity to self-renew and to repeatedly re-build the heterogeneous lineages of cancer cells that comprise a tumor in new environments. Cancer stem cells represent a cell population, which is adapted to adapt. We argue that the high evolvability of cancer stem cells is helped by their repeated transitions between plastic (proliferative, symmetrically dividing) and rigid (quiescent, asymmetrically dividing, often more invasive) phenotypes having plastic and rigid networks. Thus, cancer stem cells reverse and replay cancer development multiple times. We describe network models potentially explaining cancer stem cell-like behavior. Finally, we propose novel strategies including combination therapies and multi-target drugs to overcome the Nietzschean dilemma of cancer stem cell targeting: "what does not kill me makes me stronger".

  8. Lambda-Display: A Powerful Tool for Antigen Discovery

    Directory of Open Access Journals (Sweden)

    Nicola Gargano

    2011-04-01

    Full Text Available Since its introduction in 1985, phage display technology has been successfully used in projects aimed at deciphering biological processes and isolating molecules of practical value in several applications. Bacteriophage lambda, representing a classical molecular cloning and expression system has also been exploited for generating large combinatorial libraries of small peptides and protein domains exposed on its capsid. More recently, lambda display has been consistently and successfully employed for domain mapping, antigen discovery and protein interaction studies or, more generally, in functional genomics. We show here the results obtained by the use of large libraries of cDNA and genomic DNA for the molecular dissection of the human B-cell response against complex pathogens, including protozoan parasites, bacteria and viruses. Moreover, by reviewing the experimental work performed in recent investigations we illustrate the potential of lambda display in the diagnostics field and for identifying antigens useful as targets for vaccine development.

  9. Evaluating Ambient Displays in the Wild

    DEFF Research Database (Denmark)

    Messeter, Jörn; Molenaar, Daryn

    A prominent issue for evaluating ambient displays has been the conflict between the relative intrusiveness of evaluation methods and the intention to keep the display at the periphery of the user’s attention. There is a general lack of research discussing the difficulties of evaluating ambient...... displays in the wild, and in particular social aspects of use has received little attention. This paper presents a case study of an ambient light display designed for a public setting. Based on results from a non-intrusive in situ evaluation, we argue that viewing ambient displays as features of a broader...... social setting may aid our understanding of issues regarding the evaluation of ambient displays in the wild....

  10. Laser-driven polyplanar optic display

    Energy Technology Data Exchange (ETDEWEB)

    Veligdan, J.T.; Biscardi, C.; Brewster, C.; DeSanto, L. [Brookhaven National Lab., Upton, NY (United States). Dept. of Advanced Technology; Beiser, L. [Leo Beiser Inc., Flushing, NY (United States)

    1998-01-01

    The Polyplanar Optical Display (POD) is a unique display screen which can be used with any projection source. This display screen is 2 inches thick and has a matte-black face which allows for high contrast images. The prototype being developed is a form, fit and functional replacement display for the B-52 aircraft which uses a monochrome ten-inch display. The new display uses a 200 milliwatt green solid-state laser (532 nm) as its optical source. In order to produce real-time video, the laser light is being modulated by a Digital Light Processing (DLP) chip manufactured by Texas Instruments, Inc. A variable astigmatic focusing system is used to produce a stigmatic image on the viewing face of the POD. In addition to the optical design, the authors discuss the DLP chip, the optomechanical design and viewing angle characteristics.

  11. Spectroradiometric characterization of autostereoscopic 3D displays

    Science.gov (United States)

    Rubiño, Manuel; Salas, Carlos; Pozo, Antonio M.; Castro, J. J.; Pérez-Ocón, Francisco

    2013-11-01

    Spectroradiometric measurements have been made for the experimental characterization of the RGB channels of autostereoscopic 3D displays, giving results for different measurement angles with respect to the normal direction of the plane of the display. In the study, 2 different models of autostereoscopic 3D displays of different sizes and resolutions were used, making measurements with a spectroradiometer (model PR-670 SpectraScan of PhotoResearch). From the measurements made, goniometric results were recorded for luminance contrast, and the fundamental hypotheses have been evaluated for the characterization of the displays: independence of the RGB channels and their constancy. The results show that the display with the lower angle variability in the contrast-ratio value and constancy of the chromaticity coordinates nevertheless presented the greatest additivity deviations with the measurement angle. For both displays, when the parameters evaluated were taken into account, lower angle variability consistently resulted in the 2D mode than in the 3D mode.

  12. Conceptual Design of Industrial Process Displays

    DEFF Research Database (Denmark)

    Pedersen, C.R.; Lind, Morten

    1999-01-01

    by a simple example from a plant with batch processes. Later the method is applied to develop a supervisory display for a condenser system in a nuclear power plant. The differences between the continuous plant domain of power production and the batch processes from the example are analysed and broad...... categories of display types are proposed. The problems involved in specification and invention of a supervisory display are analysed and conclusions from these problems are made. It is concluded that the design method proposed provides a framework for the progress of the display design and is useful in pin......-pointing the actual problems. The method was useful in reducing the number of existing displays that could fulfil the requirements of the supervision task. The method provided at the same time a framework for dealing with the problems involved in inventing new displays based on structured analysis. However...

  13. Three-dimensional Imaging, Visualization, and Display

    CERN Document Server

    Javidi, Bahram; Son, Jung-Young

    2009-01-01

    Three-Dimensional Imaging, Visualization, and Display describes recent developments, as well as the prospects and challenges facing 3D imaging, visualization, and display systems and devices. With the rapid advances in electronics, hardware, and software, 3D imaging techniques can now be implemented with commercially available components and can be used for many applications. This volume discusses the state-of-the-art in 3D display and visualization technologies, including binocular, multi-view, holographic, and image reproduction and capture techniques. It also covers 3D optical systems, 3D display instruments, 3D imaging applications, and details several attractive methods for producing 3D moving pictures. This book integrates the background material with new advances and applications in the field, and the available online supplement will include full color videos of 3D display systems. Three-Dimensional Imaging, Visualization, and Display is suitable for electrical engineers, computer scientists, optical e...

  14. Laser Based 3D Volumetric Display System

    Science.gov (United States)

    1993-03-01

    Literature, Costa Mesa, CA July 1983. 3. "A Real Time Autostereoscopic Multiplanar 3D Display System", Rodney Don Williams, Felix Garcia, Jr., Texas...8217 .- NUMBERS LASER BASED 3D VOLUMETRIC DISPLAY SYSTEM PR: CD13 0. AUTHOR(S) PE: N/AWIU: DN303151 P. Soltan, J. Trias, W. Robinson, W. Dahlke 7...laser generated 3D volumetric images on a rotating double helix, (where the 3D displays are computer controlled for group viewing with the naked eye

  15. Volumetric Three-Dimensional Display Systems

    Science.gov (United States)

    Blundell, Barry G.; Schwarz, Adam J.

    2000-03-01

    A comprehensive study of approaches to three-dimensional visualization by volumetric display systems This groundbreaking volume provides an unbiased and in-depth discussion on a broad range of volumetric three-dimensional display systems. It examines the history, development, design, and future of these displays, and considers their potential for application to key areas in which visualization plays a major role. Drawing substantially on material that was previously unpublished or available only in patent form, the authors establish the first comprehensive technical and mathematical formalization of the field, and examine a number of different volumetric architectures. System level design strategies are presented, from which proposals for the next generation of high-definition predictable volumetric systems are developed. To ensure that researchers will benefit from work already completed, they provide: * Descriptions of several recent volumetric display systems prepared from material supplied by the teams that created them * An abstract volumetric display system design paradigm * An historical summary of 90 years of development in volumetric display system technology * An assessment of the strengths and weaknesses of many of the systems proposed to date * A unified presentation of the underlying principles of volumetric display systems * A comprehensive bibliography Beautifully supplemented with 17 color plates that illustrate volumetric images and prototype displays, Volumetric Three-Dimensional Display Systems is an indispensable resource for professionals in imaging systems development, scientific visualization, medical imaging, computer graphics, aerospace, military planning, and CAD/CAE.

  16. New ultraportable display technology and applications

    Science.gov (United States)

    Alvelda, Phillip; Lewis, Nancy D.

    1998-08-01

    MicroDisplay devices are based on a combination of technologies rooted in the extreme integration capability of conventionally fabricated CMOS active-matrix liquid crystal display substrates. Customized diffraction grating and optical distortion correction technology for lens-system compensation allow the elimination of many lenses and systems-level components. The MicroDisplay Corporation's miniature integrated information display technology is rapidly leading to many new defense and commercial applications. There are no moving parts in MicroDisplay substrates, and the fabrication of the color generating gratings, already part of the CMOS circuit fabrication process, is effectively cost and manufacturing process-free. The entire suite of the MicroDisplay Corporation's technologies was devised to create a line of application- specific integrated circuit single-chip display systems with integrated computing, memory, and communication circuitry. Next-generation portable communication, computer, and consumer electronic devices such as truly portable monitor and TV projectors, eyeglass and head mounted displays, pagers and Personal Communication Services hand-sets, and wristwatch-mounted video phones are among the may target commercial markets for MicroDisplay technology. Defense applications range from Maintenance and Repair support, to night-vision systems, to portable projectors for mobile command and control centers.

  17. Microencapsulated Electrophoretic Films for Electronic Paper Displays

    Science.gov (United States)

    Amundson, Karl

    2003-03-01

    Despite the dominance of liquid crystal displays, they do not perform some functions very well. While backlit liquid crystal displays can offer excellent color performance, they wash out in bright lighting and suffer from high power consumption. Reflective liquid crystal displays have limited brightness, making these devices challenging to read for long periods of time. Flexible liquid crystal displays are difficult to manufacture and keep stable. All of these attributes (long battery lifetime, bright reflective appearance, compatibility with flexible substrates) are traits that would be found in an ideal electronic paper display - an updateable substitute for paper that could be employed in electronic books, newspapers, and other applications. I will discuss technologies that are being developed for electronic-paper-like displays, and especially on particle-based technologies. A microencapsulated electrophoretic display technology is being developed at the E Ink corporation. This display film offers offer high brightness and an ink-on-paper appearance, compatibility with flexible substrates, and image stability that can lead to very low power consumption. I will present some of the physical and chemical challenges associated with making display films with high performance.

  18. Pengaruh Display Produk pada Keputusan Pembelian Konsumen

    Directory of Open Access Journals (Sweden)

    Ina Melati

    2012-10-01

    Full Text Available Most of ritel outlet recently using product display as a one of their best marketing strategy, the reason is quiet easy to be understood, since consumers are too easy to be teased by those kind of beautiful product display that is being displayed by the retail outlet. The good retail outlets are trying their best to design and make the very good product display, so they can attract more consumers and make them not thinking twice to visit their store and purchase lots of thing. Clearly seeing that an attractive product design is able to influence a consumer to make a buying decision.

  19. Framework for effective use of multiple displays

    Science.gov (United States)

    Liu, Qiong; Kimber, Don; Zhao, Frank; Huang, Jeffrey

    2005-10-01

    Meeting environments, such as conference rooms, executive briefing centers, and exhibition spaces, are now commonly equipped with multiple displays, and will become increasingly display-rich in the future. Existing authoring/presentation tools such as PowerPoint, however, provide little support for effective utilization of multiple displays. Even using advanced multi-display enabled multimedia presentation tools, the task of assigning material to displays is tedious and distracts presenters from focusing on content. This paper describes a framework for automatically assigning presentation material to displays, based on a model of the quality of views of audience members. The framework is based on a model of visual fidelity which takes into account presentation content, audience members' locations, the limited resolution of human eyes, and display location, orientation, size, resolution, and frame rate. The model can be used to determine presentation material placement based on average or worst case audience member view quality, and to warn about material that would be illegible. By integrating this framework with a previous system for multi-display presentation [PreAuthor, others], we created a tool that accepts PowerPoint and/or other media input files, and automatically generates a layout of material onto displays for each state of the presentation. The tool also provides an interface allowing the presenter to modify the automatically generated layout before or during the actual presentation. This paper discusses the framework, possible application scenarios, examples of the system behavior, and our experience with system use.

  20. Refreshable Braille Displays Using EAP Actuators

    Science.gov (United States)

    Bar-Cohen, Yoseph

    2010-01-01

    Refreshable Braille can help visually impaired persons benefit from the growing advances in computer technology. The development of such displays in a full screen form is a great challenge due to the need to pack many actuators in small area without interferences. In recent years, various displays using actuators such as piezoelectric stacks have become available in commercial form but most of them are limited to one line Braille code. Researchers in the field of electroactive polymers (EAP) investigated methods of using these materials to form full screen displays. This manuscript reviews the state of the art of producing refreshable Braille displays using EAP-based actuators..

  1. Refreshable Braille displays using EAP actuators

    Science.gov (United States)

    Bar-Cohen, Yoseph

    2010-04-01

    Refreshable Braille can help visually impaired persons benefit from the growing advances in computer technology. The development of such displays in a full screen form is a great challenge due to the need to pack many actuators in small area without interferences. In recent years, various displays using actuators such as piezoelectric stacks have become available in commercial form but most of them are limited to one line Braille code. Researchers in the field of electroactive polymers (EAP) investigated methods of using these materials to form full screen displays. This manuscript reviews the state of the art of producing refreshable Braille displays using EAP-based actuators.

  2. Simultaneous expression of displayed and secreted antibodies for antibody screen.

    Directory of Open Access Journals (Sweden)

    Yuanping Zhou

    Full Text Available The display of full-length antibody on the cell surface was achieved by fusing a transmembrane domain of the platelet-derived growth factor receptor (PDGFR to the C-terminus of the heavy chain constant region. We also incorporated a furin cleavage site between the constant region and PDGFR transmembrane domain to obtain secreted antibodies. As a result, antibodies can be expressed simultaneously on the cell surface in a membrane-anchored version for screening and selecting through fluorescence-activated cell sorting (FACS analysis, as well as in conditioned medium in a secreted version for function analysis.

  3. Heterologous surface display on lactic acid bacteria: non-GMO alternative?

    OpenAIRE

    Zadravec, Petra; Štrukelj, Borut; Berlec, Aleš

    2015-01-01

    Lactic acid bacteria (LAB) are food-grade hosts for surface display with potential applications in food and therapy. Alternative approaches to surface display on LAB would avoid the use of recombinant DNA technology and genetically-modified organism (GMO)-related regulatory requirements. Non-covalent surface display of proteins can be achieved by fusing them to various cell-wall binding domains, of which the Lysine motif domain (LysM) is particularly well studied. Fusion proteins have been is...

  4. Effects of sevoflurane and isoflurane on proliferation of neural stem cells in rat cerebral cortex%七氟醚和异氟醚对大鼠大脑皮质神经干细胞增殖的影响

    Institute of Scientific and Technical Information of China (English)

    林函; 刘劲; 李纯; 王佩芳; 高雅静; 马晓晓; 王春满; 梅虹霞; 连庆泉

    2013-01-01

    Objective To evaluate the effects of sevoflurane and isoflurane on the proliferation of neural stem cells in rat cerebral cortex.Methods The neural stem cells were isolated from Sprague-Dawley rats at 15 days of gestation and cultured at a density of (1-2) × 106 cells/ml.The cells of 3rd generation were seeded in 6-well plates coated with poly-lysine and randomly divided into 3 groups (n =24 wells each) using a random number table:control group (group C),sevoflurane group (group S) and isoflurane group (group Ⅰ).In S and Ⅰ groups,the cells were exposed to 4.9% sevoflurane and 2.8% isoflurane in a mixture of 5% CO2-95% O2 for 6 h,respectively.The cells were exposed to a mixture of 5 % CO2-95 % O2 for 6 h in group C.After 6 h of exposure,the plates were removed and the cells were continuously incubated for 2 h in an incubator at 37 ℃.The proliferation of cells was detected by immunocytochemistry and microplate method.The expression of proliferation-related genes such as signal transducers and activators of transcription 3 (STAT3),Sox2,Notchl and P21 mRNA was detected using quantitative real-time PCR.The expression of total STAT3 protein and phosphorylated STAT3 protein (p-STAT3) was determined using Western blot.Results Compared with C group,the rate of proliferation was significantly decreased,and the expression of p-STAT3 was down-regulated in I and S groups,and the expression of STAT3 mRNA was down-regulated in Ⅰ group (P < 0.05),and no significant change was found in the expression of STAT3 mRNA in S group (P > 0.05).There was no significant difference in the expression of Sox2,Notch1 and P21 mRNA and total STAT3 protein between the three groups (P > 0.05).Conclusion Sevoflurane and isoflurane both can inhibit the proliferation of neural stem cells in the rat cerebral cortex,and the mechanism may be that sevoflurane inhibits activation of STAT3 protein,however,isoflurane not only inhibits the activation of STAT3 protein,but also inhibit

  5. TLR4 signaling promotes a COX-2/PGE2/STAT3 positive feedback loop in hepatocellular carcinoma (HCC) cells

    Science.gov (United States)

    Lin, Ang; Wang, Guan; Zhao, Huajun; Zhang, Yuyi; Han, Qiuju; Zhang, Cai; Tian, Zhigang; Zhang, Jian

    2016-01-01

    ABSTRACT Toll-like receptors (TLRs) can be expressed by tumor cells, and each TLR exhibits different biological functions. Evidences showed the activation of some certain TLRs could promote tumor progression. One of which TLR4 has been found to promote hepatocellular carcinoma (HCC) cells proliferation, but the detailed mechanism is still unknown. In the present study, we verified that TLR4 was functionally expressed on HCC cells, and TLR4 agonist lipopolysaccharide (LPS) could stimulate the proliferation and clone formation of HCC cells. Most importantly, we found a COX-2/PGE2/STAT3 positive feedback loop exists in HCC cells, which could be provoked by TLR4 activation. Consistently, the expression of TLR4, COX-2 and p-STAT3Y705 was positively correlated with each other in liver tumor tissues from patients with primary HCC. Further investigation demonstrated this loop played a dominant role in TLR4-induced HCC cell proliferation and multidrug resistance (MDR) to chemotherapy. Inhibition of TLR4 or COX-2/PGE2/STAT3 loop would attenuate LPS-induced inflammation and proliferation of HCC cells, and enhance the sensitivity of HCC cells to chemotherapeutics in vitro. By using a primary HCC model, we observed COX-2/PGE2/STAT3 loop was significantly blocked in TLR4−/− mice compared to wild type mice, and there was no obvious tumorgenesis sign in TLR4−/− mice. Therefore, these findings provided the precise molecular mechanism of TLR4 signaling pathway involved in HCC progress, and suggested that TLR4 may be a promising target for HCC treatment. PMID:27057441

  6. Anandamide drives cell cycle progression through CB1 receptors in a rat model of synchronized liver regeneration.

    Science.gov (United States)

    Pisanti, Simona; Picardi, Paola; Pallottini, Valentina; Martini, Chiara; Petrosino, Stefania; Proto, Maria Chiara; Vitale, Mario; Laezza, Chiara; Gazzerro, Patrizia; Di Marzo, Vincenzo; Bifulco, Maurizio

    2015-12-01

    The endocannabinoid system, through cannabinoid receptor signaling by endocannabinoids, is involved in a wide range of functions and physiopathological conditions. To date, very little is known concerning the role of the endocannabinoids in the control and regulation of cell proliferation. An anti-proliferative action of CB1 signaling blockade in neurogenesis and angiogenesis argues in favor of proliferation-promoting functions of endocannabinoids through CB1 receptors when pro-growth signals are present. Furthermore, liver regeneration, a useful in vivo model of synchronized cell proliferation, is characterized by a peak of anandamide that elicits through CB1 receptor, the expression of critical mitosis genes. The aim of this study was to focus on the timing of endocannabinoid signaling changes during the different phases of the cell cycle, exploiting the rat liver regeneration model following partial hepatectomy, the most useful to study synchronized cell cycle in vivo. Hepatic regeneration led to increased levels of anandamide and endocannabinoid-like molecules oleoylethanolamide (OEA) and palmitoylethanolamide (PEA) in the G1 phase of the cell cycle, with a concomitant increase in CB1 mRNA levels, whose protein expression peaked later during the S phase. Blocking of CB1 receptor with a low dose of the selective antagonist/inverse agonist SR141716 (0.7 mg/kg/dose) affected cell cycle progression reducing the expression of PCNA, and through the inhibition of pERK and pSTAT3 pathways. These results support the notion that the signaling mediated by anandamide through CB1 receptor may be important for the entry and progression of cells into the cell cycle and hence for their proliferation under mitogenic signals.

  7. Methods for Selecting Phage Display Antibody Libraries.

    Science.gov (United States)

    Jara-Acevedo, Ricardo; Diez, Paula; Gonzalez-Gonzalez, Maria; Degano, Rosa Maria; Ibarrola, Nieves; Gongora, Rafael; Orfao, Alberto; Fuentes, Manuel

    2016-01-01

    The selection process aims sequential enrichment of phage antibody display library in clones that recognize the target of interest or antigen as the library undergoes successive rounds of selection. In this review, selection methods most commonly used for phage display antibody libraries have been comprehensively described.

  8. Teacher Portfolios: Displaying the Art of Teaching

    Science.gov (United States)

    Reese, Susan

    2004-01-01

    A portfolio can convey a teacher's beliefs, knowledge and skills. An artist uses a portfolio to display artistic talent, and a teacher can use his or her portfolio to display teaching talent and teaching style. A teacher's portfolio may be used to obtain new employment, to document teaching accomplishments in order to receive a promotion or tenure…

  9. Display Developer for Firing Room Applications

    Science.gov (United States)

    Bowman, Elizabeth A.

    2013-01-01

    The firing room at Kennedy Space Center (KSC) is responsible for all NASA human spaceflight launch operations, therefore it is vital that all displays within the firing room be properly tested, up-to-date, and user-friendly during a launch. The Ground Main Propulsion System (GMPS) requires a number of remote displays for Vehicle Integration and Launch (VIL) Operations at KSC. My project is to develop remote displays for the GMPS using the Display Services and Framework (DSF) editor. These remote displays will be based on model images provided by GMPS through PowerPoint. Using the DSF editor, the PowerPoint images can be recreated with active buttons associated with the correct Compact Unique Identifiers (CUIs). These displays will be documented in the Software Requirements and Design Specifications (SRDS) at the 90% GMPS Design Review. In the future, these remote displays will be available for other developers to improve, edit, or add on to so that the display may be incorporated into the firing room to be used for launches.

  10. Interruption and Pausing of Public Display Games

    DEFF Research Database (Denmark)

    Feuchtner, Tiare; Walter, Robert; Müller, Jörg

    2016-01-01

    We present a quantitative and qualitative analysis of interruptions of interaction with a public display game, and explore the use of a manual pause mode in this scenario. In previous public display installations we observed users frequently interrupting their interaction. To explore ways of supp...

  11. Additive and subtractive transparent depth displays

    NARCIS (Netherlands)

    Kooi, F.L.; Toet, A.

    2003-01-01

    Image fusion is the generally preferred method to combine two or more images for visual display on a single screen. We demonstrate that perceptual image separation may be preferable over perceptual image fusion for the combined display of enhanced and synthetic imagery. In this context image separat

  12. Extraction and Analysis of Display Data

    Science.gov (United States)

    Land, Chris; Moye, Kathryn

    2008-01-01

    The Display Audit Suite is an integrated package of software tools that partly automates the detection of Portable Computer System (PCS) Display errors. [PCS is a lap top computer used onboard the International Space Station (ISS).] The need for automation stems from the large quantity of PCS displays (6,000+, with 1,000,000+ lines of command and telemetry data). The Display Audit Suite includes data-extraction tools, automatic error detection tools, and database tools for generating analysis spread sheets. These spread sheets allow engineers to more easily identify many different kinds of possible errors. The Suite supports over 40 independent analyses, 16 NASA Tech Briefs, November 2008 and complements formal testing by being comprehensive (all displays can be checked) and by revealing errors that are difficult to detect via test. In addition, the Suite can be run early in the development cycle to find and correct errors in advance of testing.

  13. 2D/3D switchable displays

    Science.gov (United States)

    Dekker, T.; de Zwart, S. T.; Willemsen, O. H.; Hiddink, M. G. H.; IJzerman, W. L.

    2006-02-01

    A prerequisite for a wide market acceptance of 3D displays is the ability to switch between 3D and full resolution 2D. In this paper we present a robust and cost effective concept for an auto-stereoscopic switchable 2D/3D display. The display is based on an LCD panel, equipped with switchable LC-filled lenticular lenses. We will discuss 3D image quality, with the focus on display uniformity. We show that slanting the lenticulars in combination with a good lens design can minimize non-uniformities in our 20" 2D/3D monitors. Furthermore, we introduce fractional viewing systems as a very robust concept to further improve uniformity in the case slanting the lenticulars and optimizing the lens design are not sufficient. We will discuss measurements and numerical simulations of the key optical characteristics of this display. Finally, we discuss 2D image quality, the switching characteristics and the residual lens effect.

  14. Three-dimensional hologram display system

    Science.gov (United States)

    Mintz, Frederick (Inventor); Chao, Tien-Hsin (Inventor); Bryant, Nevin (Inventor); Tsou, Peter (Inventor)

    2009-01-01

    The present invention relates to a three-dimensional (3D) hologram display system. The 3D hologram display system includes a projector device for projecting an image upon a display medium to form a 3D hologram. The 3D hologram is formed such that a viewer can view the holographic image from multiple angles up to 360 degrees. Multiple display media are described, namely a spinning diffusive screen, a circular diffuser screen, and an aerogel. The spinning diffusive screen utilizes spatial light modulators to control the image such that the 3D image is displayed on the rotating screen in a time-multiplexing manner. The circular diffuser screen includes multiple, simultaneously-operated projectors to project the image onto the circular diffuser screen from a plurality of locations, thereby forming the 3D image. The aerogel can use the projection device described as applicable to either the spinning diffusive screen or the circular diffuser screen.

  15. An interactive multiview 3D display system

    Science.gov (United States)

    Zhang, Zhaoxing; Geng, Zheng; Zhang, Mei; Dong, Hui

    2013-03-01

    The progresses in 3D display systems and user interaction technologies will help more effective 3D visualization of 3D information. They yield a realistic representation of 3D objects and simplifies our understanding to the complexity of 3D objects and spatial relationship among them. In this paper, we describe an autostereoscopic multiview 3D display system with capability of real-time user interaction. Design principle of this autostereoscopic multiview 3D display system is presented, together with the details of its hardware/software architecture. A prototype is built and tested based upon multi-projectors and horizontal optical anisotropic display structure. Experimental results illustrate the effectiveness of this novel 3D display and user interaction system.

  16. Interruption and Pausing of Public Display Games

    DEFF Research Database (Denmark)

    Feuchtner, Tiare; Walter, Robert; Müller, Jörg

    We present a quantitative and qualitative analysis of interruptions of interaction with a public display game, and explore the use of a manual pause mode in this scenario. In previous public display installations we observed users frequently interrupting their interaction. To explore ways...... of supporting such behavior, we implemented a gesture controlled multiuser game with four pausing techniques. We evaluated them in a field study analyzing 704 users and found that our pausing techniques were eagerly explored, but rarely used with the intention to pause the game. Our study shows...... that interactions with public displays are considerably intermissive, and that users mostly interrupt interaction to socialize and mainly approach public displays in groups. We conclude that, as a typical characteristic of public display interaction, interruptions deserve consideration. However, manual pause modes...

  17. BRCA1 deficient embryonic stem cells display a decreased homologous recombination frequency and an increased frequency of non-homologous recombination that is corrected by expression of a brca1 transgene.

    Science.gov (United States)

    Snouwaert, J N; Gowen, L C; Latour, A M; Mohn, A R; Xiao, A; DiBiase, L; Koller, B H

    1999-12-20

    BRCA1 is a nuclear phosphoprotein that has been classified as a tumor suppressor based on the fact that women carrying a mutated copy of the BRCA1 gene are at increased risk of developing breast and ovarian cancer. The association of BRCA1 with RAD51 has led to the hypothesis that BRCA1 is involved in DNA repair. We describe here the generation and analysis of murine embryonic stem (ES) cell lines in which both copies of the murine homologue of the human BRCA1 gene have been disrupted by gene targeting. We show that exogenous DNA introduced into these BRCA1 deficient cells by electroporation is randomly integrated into the genome at a significantly higher rate than in wild type ES cells. In contrast, integration of exogenous DNA by homologous recombination occurs in BRCA1 deficient cells at a significantly lower rate than in wild type controls. When BRCA1 expression is re-established at 5-10% of normal levels by introduction of a Brca1 transgene into BRCA1 deficient ES cells, the frequency of random integration is reduced to wild type levels, although the frequency of homologous recombination is not significantly improved. These results suggest that BRCA1 plays a role in determining the response of cells to double stranded DNA breaks.

  18. Profiling of liquid crystal displays with Raman spectroscopy: Preprocessing of spectra.

    NARCIS (Netherlands)

    O. Stanimirovic; H.F.M. Boelens; A.J.G. Mank; H.C.J. Hoefsloot; A.K. Smilde

    2005-01-01

    Raman spectroscopy is applied for characterizing paintable displays. Few other options than Raman spectroscopy exist for doing so because of the liquid nature of functional materials. The challenge is to develop a method that can be used for estimating the composition of a single display cell on the

  19. Organic Thin Film Devices for Displays and Lighting

    Science.gov (United States)

    Weiss, Oliver J.; Krause, Ralf; Paetzold, Ralph

    Organic materials can be used for fabrication of, e.g., electronic circuits, solar cells, light sensors, memory cells and light emitting diodes. Especially organic light emitting diodes (OLEDs) are increasingly attractive because of their huge market potential. The feasibility of efficient OLEDs was first shown in 1987 [3]. Only about ten years later the first product, a display for car radios, entered the market. Today monochrome and full colour OLED-displays can be found in many applications replacing established flat panel display technologies like TFT-LCDs. This substitution is a consequence of the outstanding attributes of OLED technology: Organic light emitting displays are self-emissive, thin, video capable and in addition they show a wide temperature operation range and allow a viewing angle of nearly 180 degree in conjunction with a low power consumption. As performance has steadily increased over the last years, today OLEDs are also under investigation as next generation light source. In contrast to inorganic LEDs, they can be built as flat 2-dimensional light sources that are lightweight, colour tunable, and potentially cheap. This will open up new degrees of freedom in design leading also to completely new applications. In this contribution we will have a brief view on the history of organic electroluminescent materials before we introduce the basic principles of OLEDs with a focus on the physical processes leading to light generation in thin organic films. Along with an overview of different concepts and technologies used to build OLEDs, the current status of OLED development will be illustrated. The last part focuses on the challenges that have to be overcome to enable a sustainable success in the display and lighting markets.

  20. Fast-response liquid crystal display by the VA-IPS display mode with nematic liquid crystal and polymer networks

    Science.gov (United States)

    Chen, Tien-Jung; Lin, Guan-Jhong; Chen, Bo-Yu; Wu, Jin-Jei; Yang, Ying-Jay

    2012-10-01

    To improve electrooptical characteristics of the vertical aligned (VA) liquid crystal displays (LCDs), the monomer material and in-plane switching (IPS) field produced by interdigital electrodes are employed in LC cells. The fast switching response and well optical transmittance of the VA-IPS display mode are successfully achieved by mixing the nematic LC with polymer networks, attributed to the surface anchoring, and the molecular orientation of the LC cell will be further governed, especially under the greater applied voltage. Furthermore, the high concentration doping of the monomer can effectively improve the response behavior, but it also results in the transmittance sacrificed due to the light scattering, and the threshold voltage (Vth) increased.

  1. Touch sensitive electrorheological fluid based tactile display

    Science.gov (United States)

    Liu, Yanju; Davidson, Rob; Taylor, Paul

    2005-12-01

    A tactile display is programmable device whose controlled surface is intended to be investigated by human touch. It has a great number of potential applications in the field of virtual reality and elsewhere. In this research, a 5 × 5 tactile display array including electrorheological (ER) fluid has been developed and investigated. Force responses of the tactile display array have been measured while a probe was moved across the upper surface. The purpose of this was to simulate the action of touch performed by human finger. Experimental results show that the sensed surface information could be controlled effectively by adjusting the voltage activation pattern imposed on the tactels. The performance of the tactile display is durable and repeatable. The touch sensitivity of this ER fluid based tactile display array has also been investigated in this research. The results show that it is possible to sense the touching force normal to the display's surface by monitoring the change of current passing through the ER fluid. These encouraging results are helpful for constructing a new type of tactile display based on ER fluid which can act as both sensor and actuator at the same time.

  2. Crosstalk in stereoscopic displays: a review

    Science.gov (United States)

    Woods, Andrew J.

    2012-10-01

    Crosstalk, also known as ghosting or leakage, is a primary factor in determining the image quality of stereoscopic three dimensional (3D) displays. In a stereoscopic display, a separate perspective view is presented to each of the observer's two eyes in order to experience a 3D image with depth sensation. When crosstalk is present in a stereoscopic display, each eye will see a combination of the image intended for that eye, and some of the image intended for the other eye-making the image look doubled or ghosted. High levels of crosstalk can make stereoscopic images hard to fuse and lack fidelity, so it is important to achieve low levels of crosstalk in the development of high-quality stereoscopic displays. Descriptive and mathematical definitions of these terms are formalized and summarized. The mechanisms by which crosstalk occurs in different stereoscopic display technologies are also reviewed, including micropol 3D liquid crystal displays (LCDs), autostereoscopic (lenticular and parallax barrier), polarized projection, anaglyph, and time-sequential 3D on LCDs, plasma display panels and cathode ray tubes. Crosstalk reduction and crosstalk cancellation are also discussed along with methods of measuring and simulating crosstalk.

  3. [Odor sensing system and olfactory display].

    Science.gov (United States)

    Nakamoto, Takamichi

    2014-01-01

    In this review, an odor sensing system and an olfactory display are introduced into people in pharmacy. An odor sensing system consists of an array of sensors with partially overlapping specificities and pattern recognition technique. One of examples of odor sensing systems is a halitosis sensor which quantifies the mixture composition of three volatile sulfide compounds. A halitosis sensor was realized using a preconcentrator to raise sensitivity and an electrochemical sensor array to suppress the influence of humidity. Partial least squares (PLS) method was used to quantify the mixture composition. The experiment reveals that the sufficient accuracy was obtained. Moreover, the olfactory display, which present scents to human noses, is explained. A multi-component olfactory display enables the presentation of a variety of smells. The two types of multi-component olfactory display are described. The first one uses many solenoid valves with high speed switching. The valve ON frequency determines the concentration of the corresponding odor component. The latter one consists of miniaturized liquid pumps and a surface acoustic wave (SAW) atomizer. It enables the wearable olfactory display without smell persistence. Finally, the application of the olfactory display is demonstrated. Virtual ice cream shop with scents was made as a content of interactive art. People can enjoy harmony among vision, audition and olfaction. In conclusion, both odor sensing system and olfactory display can contribute to the field of human health care.

  4. Memory effect in ac plasma displays

    Science.gov (United States)

    Szlenk, K.; Obuchowicz, E.

    1993-10-01

    The bistable or `memory' mode of operation of an ac plasma display panel is presented. The difference between dc and ac plasma panel operation from the point of view of memory function is discussed. The graphic ac plasma display with thin film Cr-Cu-Cr electrodes was developed in OBREP and its basic parameters are described. It consists of 36 X 59 picture elements, its outer dimensions are: 76 X 52 mm2 and the screen size is: 49 X 30 mm2. The different dielectric glass materials were applied as dielectric layers and the influence of the properties of these materials on display parameters and memory function was investigated.

  5. Embedded systems for controlling LED matrix displays

    Science.gov (United States)

    Marghescu, Cristina; Drumea, Andrei

    2016-12-01

    LED matrix displays are a common presence in everyday life - they can be found in trains, buses, tramways, office information tables or outdoor media. The structure of the display unit is similar for all these devices, a matrix of light emitting diodes coupled between row and column lines, but there are many options for the display controller that switches these lines. Present paper analyzes different types of embedded systems that can control the LED matrix, based on single board computers, on microcontrollers with different peripheral devices or with programmable logic devices like field programmable gate arrays with implemented soft processor cores. Scalability, easiness of implementation and costs are analyzed for all proposed solutions.

  6. Human cytomegalovirus gene expression in long-term infected glioma stem cells.

    Directory of Open Access Journals (Sweden)

    Estefania Fiallos

    Full Text Available The most common adult primary brain tumor, glioblastoma (GBM, is characterized by fifteen months median patient survival and has no clear etiology. We and others have identified the presence of human cytomegalovirus (HCMV gene products endogenously expressed in GBM tissue and primary cells, with a subset of viral genes being consistently expressed in most samples. Among these viral genes, several have important oncomodulatory properties, regulating tumor stemness, proliferation, immune evasion, invasion and angiogenesis. These findings lead us to hypothesize that a specific HCMV gene signature may be associated with GBM pathogenesis. To investigate this hypothesis, we used glioma cell lines and primary glioma stem-like cells (GSC infected with clinical and laboratory HCMV strains and measured relative viral gene expression levels along several time points up to 15 weeks post-infection. While HCMV gene expression was detected in several infected glioma lines through week 5 post-infection, only HCMV-infected GSC expressed viral gene products 15 weeks post-infection. Efficiency of infection across time was higher in GSC compared to cell lines. Importantly, HCMV-infected GSC outlived their uninfected counterparts, and this extended survival was paralleled by increased tumorsphere frequency and upregulation of stemness regulators, such as SOX2, p-STAT3, and BMX (a novel HCMV target identified in this study. Interleukin 6 (IL-6 treatment significantly upregulated HCMV gene expression in long-term infected glioma cultures, suggesting that pro-inflammatory signaling in the tumor milieu may further augment HCMV gene expression and subsequent tumor progression driven by viral-induced cellular signaling. Together, our data support a critical role for long-term, low-level HCMV infection in promoting survival, stemness, and proliferation of GSC that could significantly contribute to GBM pathogenesis.

  7. microRNA-200a silencing protects neural stem cells against cerebral ischemia/reperfusion injury

    Science.gov (United States)

    Ma, Ji; Shui, Shaofeng; Han, Xinwei; Guo, Dong; Li, Tengfei; Yan, Lei

    2017-01-01

    Neural stem cells (NSCs) play major roles in neurological recovery after cerebral infarction (CI). This study was trying to investigate whether miR-200a, a vital regulator in cell proliferation, migration and apoptosis, also has a role in oxygen-glucose deprivation/reperfusion (OGD/R) injured NSCs. In this study, primary NSCs were subjected to OGD/R conditions to mimic an in vitro CI model. Before OGD/R induction, NSCs were transfected with vector or shRNA against miR-200a to overexpress or suppress miR-200a expression. The changes in cell viability, apoptosis, migration, the expression of c-Myc, and the phosphorylation of STAT1, STAT3 and MAPK were respectively assessed. Inhibitors of STAT1/3 and MAPK, i.e., Nifuroxazide and BIRB 796, were used to administrate miR-200a-silenced NSCs, and the expressions of above mentioned proteins were detected. After OGD/R exposure, miR-200a was up-regulated in NSCs (P < 0.001). miR-200a silencing alleviated OGD/R-induced the decrease of cell viability and migration (P < 0.01); meanwhile, alleviated OGD/R-induced apoptosis via reducing Bax/Bcl-2 ratio and down-regulating p53 and cytochrome c (P < 0.01 or P < 0.001). c-Myc, p-STAT1, p-STAT3, p-MAPK were all negatively regulated by miR-200a (P < 0.01 or P < 0.001); more important, the increase of c-Myc induced by miR-200a silencing was abolished by Nifuroxazide or BIRB 796 (P < 0.01 or P < 0.001). These data indicate miR-200a silencing protects NSCs from OGD/R-induced injury, possibly via regulating the STATs/c-Myc and MAPK/c-Myc signalings. PMID:28222148

  8. A cationic vaccine adjuvant based on a saturated quaternary ammonium lipid have different in vivo distribution kinetics and display a distinct CD4 T cell-inducing capacity compared to its unsaturated analog

    DEFF Research Database (Denmark)

    Christensen, Dennis; Henriksen-Lacey, Malou; Kamath, Arun T;

    2012-01-01

    Adjuvants are often composed of different constituents that can be divided into two groups based on their primary activity: the delivery system which carries and presents the vaccine antigen to antigen-presenting cells, and the immunostimulator that activates and modulates the ensuing immune...

  9. Efficient biological conversion of carbon monoxide (CO) to carbon dioxide (CO2) and for utilization in bioplastic production by Ralstonia eutropha through the display of an enzyme complex on the cell surface.

    Science.gov (United States)

    Hyeon, Jeong Eun; Kim, Seung Wook; Park, Chulhwan; Han, Sung Ok

    2015-06-25

    An enzyme complex for biological conversion of CO to CO2 was anchored on the cell surface of the CO2-utilizing Ralstonia eutropha and successfully resulted in a 3.3-fold increase in conversion efficiency. These results suggest that this complexed system may be a promising strategy for CO2 utilization as a biological tool for the production of bioplastics.

  10. Phage display of peptide / major histocompatibility class I complexes

    DEFF Research Database (Denmark)

    Vest Hansen, N; Ostergaard Pedersen, L; Stryhn, A;

    2001-01-01

    Major histocompatibility complex class I (MHC-I) molecules sample peptides from the intracellular environment and present them to cytotoxic T cells (CTL). To establish a selection system, and, thereby, enable a library approach to identify the specificities involved (that of the MHC-I for peptides...... and subsequently that ot the T cell receptor for peptide-MHC-I complex), we have fused a single chain peptide-MHC-I complex to the phage minor coat protein, gpIII, and displayed it on filamentous phage. Expression of peptide-MHC-I complexes was shown with relevant conformation-specific monoclonal antibodies and...

  11. A faster switching regime for zenithal bistable nematic displays

    Energy Technology Data Exchange (ETDEWEB)

    Rudin, J

    1997-12-01

    A simpler and faster switching regime for Zenithal Bistable Nematic displays is reported. A cell, based on homeotropic alignment of nematic liquid crystal over a continuous blazed monograting on one surface, can be switched using bipolar pulses an order of magnitude faster than monopolar pulses of the same voltage. We propose that this regime relies on simple dielectric coupling to drive the cell into a higher energy state with a long pulse time, and the relaxation into a lower energy state after the creation of surface defects from a shorter applied pulse. Although flexoelectric effects are observed, they do not form the basis of state selection as was proposed for the monopolar pulses

  12. Yeast surface display for screening combinatorial polypeptide libraries.

    Science.gov (United States)

    Boder, E T; Wittrup, K D

    1997-06-01

    Display on the yeast cell wall is well suited for engineering mammalian cell-surface and secreted proteins (e.g., antibodies, receptors, cytokines) that require endoplasmic reticulum-specific post-translational processing for efficient folding and activity. C-terminal fusion to the Aga2p mating adhesion receptor of Saccharomyces cerevisiae has been used for the selection of scFv antibody fragments with threefold decreased antigen dissociation rate from a randomly mutated library. A eukaryotic host should alleviate expression biases present in bacterially propagated combinatorial libraries. Quantitative flow cytometric analysis enables fine discrimination of kinetic parameters for protein binding to soluble ligands.

  13. Display depth analyses with the wave aberration for the auto-stereoscopic 3D display

    Science.gov (United States)

    Gao, Xin; Sang, Xinzhu; Yu, Xunbo; Chen, Duo; Chen, Zhidong; Zhang, Wanlu; Yan, Binbin; Yuan, Jinhui; Wang, Kuiru; Yu, Chongxiu; Dou, Wenhua; Xiao, Liquan

    2016-07-01

    Because the aberration severely affects the display performances of the auto-stereoscopic 3D display, the diffraction theory is used to analyze the diffraction field distribution and the display depth through aberration analysis. Based on the proposed method, the display depth of central and marginal reconstructed images is discussed. The experimental results agree with the theoretical analyses. Increasing the viewing distance or decreasing the lens aperture can improve the display depth. Different viewing distances and the LCD with two lens-arrays are used to verify the conclusion.

  14. rhG-CSF促进体外培养神经干细胞的增殖%rhG-CSF promoting the proliferation of neural stem cells in vitro

    Institute of Scientific and Technical Information of China (English)

    李延兰; 贾德永; 付洁; 刘慧娟

    2011-01-01

    Objective To investigate whether granulocyte colony-stimulating factor ( G-CSF ) has a direct effect on the proliferation of mouse neural stem cells in vitro. Methods Primary mouse neural stem cells( NSCs ) were isolated from Kunming mice and cultured in serum free medium. The third passage NSCs were used in the experiments. NSCs were treated with G-CSF ( 10, 30, 60 , 100 , and 200μg/L ). The proliferation of NSCs was detected by MTT colorimetric assay and bromodeoxyuridine ( BrdU ) incorporation assay. The expressions of STAT3 and p-STAT3 were examined by Western blotting. Results There was the expression of G-CSF receptor on the neural stem cells. G-CSF obviously improved the viabilities of NSCs in a dose-dependent manner. Furthermore, 100μg/L G-CSF yielded the optimal effect. The BrdU-positive cells against total NSCs treated with G-CSF were markedly enhanced. Time course experiments demonstrated that STAT3 phosphorylation by G-CSF was evident after 5 minutes, reaching the maximum after 30 minutes. At 75 minutes, the effect on STAT3 returned to the basal value. Pretreatment with the G-CSF receptor antibody significantly reduced the phosphorylation of STAT3 induced by G-CSF. BrdU incorporation showed that cell proliferation in the presence of G-CSF and neutralizing antibody for G-CSF receptor was similar to that of the reduced growth medium group. Conclusion G-CSF may stimulate the proliferation of NSCs in vitro. Our results provide the cellular basis of the role of G-CSF in NSCs in vitro ,which may serve as a useful reference for future studies of the powerful neuroprotective effect of G-CSF in various types of neurological disorders.%目的 探讨重组人粒细胞集落刺激因子(rhG-CSF)对小鼠胚胎神经干细胞(NSCs)增殖的作用.方法 分离培养小鼠NSCs,通过向培养基中添加G-CSF(10、30、60、100和200μg/L),四甲基偶氮唑蓝(MTT)比色法以及5-溴-2′-脱氧尿苷(BrdU) 免疫荧光染色标记检测NSCs的增殖.免疫印

  15. In vitro affinity screening of protein and peptide binders by megavalent bead surface display.

    Science.gov (United States)

    Diamante, Letizia; Gatti-Lafranconi, Pietro; Schaerli, Yolanda; Hollfelder, Florian

    2013-10-01

    The advent of protein display systems has provided access to tailor-made protein binders by directed evolution. We introduce a new in vitro display system, bead surface display (BeSD), in which a gene is mounted on a bead via strong non-covalent (streptavidin/biotin) interactions and the corresponding protein is displayed via a covalent thioether bond on the DNA. In contrast to previous monovalent or low-copy bead display systems, multiple copies of the DNA and the protein or peptide of interest are displayed in defined quantities (up to 10(6) of each), so that flow cytometry can be used to obtain a measure of binding affinity. The utility of the BeSD in directed evolution is validated by library selections of randomized peptide sequences for binding to the anti-hemagglutinin (HA) antibody that proceed with enrichments in excess of 10(3) and lead to the isolation of high-affinity HA-tags within one round of flow cytometric screening. On-bead K(d) measurements suggest that the selected tags have affinities in the low nanomolar range. In contrast to other display systems (such as ribosome, mRNA and phage display) that are limited to affinity panning selections, BeSD possesses the ability to screen and rank binders by their affinity in vitro, a feature that hitherto has been excl